Posts Tagged ‘Cell Biology’

New Generation of Platinated Compounds to Circumvent Resistance

Curator/Writer: Stephen J. Williams, Ph.D.

Resistance to chemotherapeutic drugs continues to be a major hurdle in the treatment of neoplastic disorders, irregardless if the drug is a member of the cytotoxic “older” drugs or the cytostatic “newer” personalized therapies like the tyrosine kinase inhibitors.  For the platinatum compounds such as cisplatin and carboplatin, which are mainstays in therapeutic regimens for ovarian and certain head and neck cancers, development of resistance is often regarded as the final blow, as new options for these diseases have been limited.

Although there are many mechanisms by which resistance to platinated compounds may develop the purpose of this posting is not to do an in-depth review of this area except to refer the reader to the book   Ovarian Cancer and just to summarize the well accepted mechanisms of cisplatin resistance including:

  • Decreased cellular cisplatin influx
  • Increased cellular cisplatin efflux
  • Increased cellular glutathione and subsequent conjugation, inactivation
  • Increased glutathione-S-transferase activity (GST) and subsequent inactivation, conjugation
  • Increased γ-GGT
  • Increased metallothionenes with subsequent conjugation, inactivation
  • Increased DNA repair: increased excision repair
  • DNA damage tolerance: loss of mismatch repair (MMR)
  • altered cell signaling activities and cell cycle protein expression

Williams, S.J., and Hamilton, T.C. Chemotherapeutic resistance in ovarian cancer. In: S.C. Rubin, and G.P. Sutton (eds.), Ovarian Cancer, pp.34-44. Lippincott, Wilkins, and Williams, New York, 2000.

Also for a great review on clinical platinum resistance by Drs. Maritn, Hamilton and Schilder please see the following Clinical Cancer Research link here.

This curation represents the scientific rationale for the development of a new class of platinated compounds which are meant to circumvent mechanisms of resistance, in this case the loss of mismatch repair (MMR) and increased tolerance to DNA damage.

An early step in the production of cytotoxicity by the important anticancer drug cisplatin and its analog carboplatin is the formation of intra- and inter-strand adducts with tumor cell DNA 1-3. This damage triggers a cascade of events, best characterized by activation of damage-sensing kinases (reviewed in 4), p53 stabilization, and induction of p53-related genes involved in apoptosis and cell cycle arrest, such as bax and the cyclin-dependent kinase inhibitor p21waf1/cip1/sdi1 (p21), respectively 5,6. DNA damage significantly induces p21 in various p53 wild-type tumor cell lines, including ovarian carcinoma cells, and this induction is responsible for the cell cycle arrest at G1/S and G2/M borders, allowing time for repair 7,8.  DNA lesions have the ability of  to result in an opening of chromatin structure, allowing for transcription factors to enter 56-58.  Therefore the anti-tumoral ability of cisplatin and other DNA damaging agents is correlated to their ability to bind to DNA and elicit responses, such as DNA breaks or DNA damage responses which ultimately lead to cell cycle arrest and apoptosis.  Therefore either repair of such lesions, the lack of recognition of such lesions, or the cellular tolerance of such lesions can lead to resistance of these agents.


Mechanisms of Cisplatin Sensitivity and Resistance. Red arrows show how a DNA lesion results in chemo-sensitivity while the beige arrow show common mechanisms of resistance including increased repair of the lesion, effects on expression patterns, and increased inactivation of the DNA damaging agent by conjugation reactions


















Increased DNA Repair Mechanisms of Platinated Lesion Lead to ChemoResistance



Description of Different Types of Cellular DNA Repair Pathways. Nucleotide Excision Repair is commonly up-regulated in highly cisplatin resistant cells












Loss of Mismatch Repair Can Lead to DNA Damage Tolerance

dnadamage tolerance









In the following Cancer Research paper Dr. Vaisman in the lab of Dr. Steve Chaney at North Carolina (and in collaboration with Dr. Tom Hamilton) describe how cisplatin resistance may arise from loss of mismatch repair and how oxaliplatin lesions are not recognized by the mismatch repair system.
Cancer Res. 1998 Aug 15;58(16):3579-85.

The role of hMLH1, hMSH3, and hMSH6 defects in cisplatin and oxaliplatin resistance: correlation with replicative bypass of platinum-DNA adducts.


Defects in mismatch repair are associated with cisplatin resistance, and several mechanisms have been proposed to explain this correlation. It is hypothesized that futile cycles of translesion synthesis past cisplatin-DNA adducts followed by removal of the newly synthesized DNA by an active mismatch repair system may lead to cell death. Thus, resistance to platinum-DNA adducts could arise through loss of the mismatch repair pathway. However, no direct link between mismatch repair status and replicative bypass ability has been reported. In this study, cytotoxicity and steady-state chain elongation assays indicate that hMLH1 or hMSH6 defects result in 1.5-4.8-fold increased cisplatin resistance and 2.5-6-fold increased replicative bypass of cisplatin adducts. Oxaliplatin adducts are not recognized by the mismatch repair complex, and no significant differences in bypass of oxaliplatin adducts in mismatch repair-proficient and -defective cells were found. Defects in hMSH3 did not alter sensitivity to, or replicative bypass of, either cisplatin or oxaliplatin adducts. These observations support the hypothesis that mismatch repair defects in hMutL alpha and hMutS alpha, but not in hMutS beta, contribute to increased net replicative bypass of cisplatin adducts and therefore to drug resistance by preventing futile cycles of translesion synthesis and mismatch correction.



The following are slides I had co-prepared with my mentor Dr. Thomas C. Hamilton, Ph.D. of Fox Chase Cancer Center on DNA Mismatch Repair, Oxaliplatin and Ovarina Cancer.








Multiple Platinum Analogs of Cisplatin (like Oxaliplatin )Had Been Designed to be Sensitive in MMR Deficient Tumors












































Please see below video on 2015 Nobel Laureates and their work to elucidate the celluar DNA repair mechanisms.

Clinical genetics expert Kenneth Offit gives an overview of Lynch syndrome, a genetic disorder that can cause colon (HNPCC) and other cancers by defects in the MSH2 DNA mismatch repair gene. (View Video)




  1. Johnson, S. W. et al. Relationship between platinum-DNA adduct formation, removal, and cytotoxicity in cisplatin sensitive and resistant human ovarian cancer cells. Cancer Res 54, 5911-5916 (1994).
  2. Eastman, A. The formation, isolation and characterization of DNA adducts produced by anticancer platinum complexes. Pharmacology and Therapeutics 34, 155-166 (1987).
  3. Zhen, W. et al. Increased gene-specific repair of cisplatin interstrand cross-links in cisplatin-resistant human ovarian cancer cell lines. Molecular and Cellular Biology 12, 3689-3698 (1992).
  4. Durocher, D. & Jackson, S. P. DNA-PK, ATM and ATR as sensors of DNA damage: variations on a theme? Curr Opin Cell Biol 13, 225-231 (2001).
  5. el-Deiry, W. S. p21/p53, cellular growth control and genomic integrity. Curr Top Microbiol Immunol 227, 121-37 (1998).
  6. Ewen, M. E. & Miller, S. J. p53 and translational control. Biochim Biophys Acta 1242, 181-4 (1996).
  7. Gartel, A. L., Serfas, M. S. & Tyner, A. L. p21–negative regulator of the cell cycle. Proc Soc Exp Biol Med 213, 138-49 (1996).
  8. Chang, B. D. et al. p21Waf1/Cip1/Sdi1-induced growth arrest is associated with depletion of mitosis-control proteins and leads to abnormal mitosis and endoreduplication in recovering cells. Oncogene 19, 2165-70 (2000).
  9. Davies, N. P., Hardman, L. C. & Murray, V. The effect of chromatin structure on cisplatin damage in intact human cells. Nucleic Acids Res 28, 2954-2958 (2000).
  10. Vichi, P. et al. Cisplatin- and UV-damaged DNA lure the basal transcription factor TFIID/TBP. Embo J 16, 7444-7456 (1997).
  11. Xiao, G. et al. A DNA damage signal is required for p53 to activate gadd45. Cancer Res 60, 1711-9 (2000).

Other articles in this Open Access Journal on ChemoResistance Include:

Cancer Stem Cells as a Mechanism of Resistance

An alternative approach to overcoming the apoptotic resistance of pancreatic cancer

Mutation D538G – a novel mechanism conferring acquired Endocrine Resistance causes a change in the Estrogen Receptor and Treatment of Breast Cancer with Tamoxifen

Can IntraTumoral Heterogeneity Be Thought of as a Mechanism of Resistance?

Nitric Oxide Mitigates Sensitivity of Melanoma Cells to Cisplatin

Heroes in Medical Research: Barnett Rosenberg and the Discovery of Cisplatin


Read Full Post »

Cancer Stem Cells as a Mechanism of Resistance


Curator: Stephen J. Williams, Ph.D.

The cancer stem-cell hypothesis proposes the existence of a subset of cells within a heterogeneous tumor cell population that have stem-cell like properties [1], and may be essential for the progression and metastases of epithelial malignancies, by providing a reservoir of cells that self-renew and differentiate into the bulk of the tumor [2]. The stem-cell hypothesis implies that similar genetic regulatory pathways might define critical stem-cell like functions, such as self-renewal and pluripotency, in both normal and cancer stem-cells. Indeed, cancer stem-cells have been identified in many tumor types, such as breast [3], pancreas [4] and ovarian [5], based on screening with cellular markers typically found in normal stem-cells such as CD44, ALDH1, and CD133 (reviewed in [2]). A number of studies have suggested that the expression of these stem-cell markers is correlated with poor prognosis [6-9]. The ability to identify and isolate these populations may have a significant impact on design of individualized therapies.

Great general posts and good review on this site about Cancer Stem Cells, their markers, and ability to target them with chemotherapy can be seen here.

In Focus: Identity of Cancer Stem Cells

In Focus: Targeting of Cancer Stem Cells

Stem Cells and Cancer


However, there has been growing acknowledgement of the ability of cancer stem cell populations to resist the cytotoxic effects of most chemotherapeutic agents, including cisplatin, topoisomerase inhibitors, DNA damaging agents, and even tyrosine kinase inhibitors (TKI). Indeed, some feel that intrinsic resistance to cytotoxic drugs may be a biological feature of cancer stem cells.


Acquired resistance: a resistance to a particular drug which results following continued exposure to said drug. Can take days (in cases of some TKIs) or months to develop. Acquired resistant cells lines are developed by exposure to increasing drug concentration over a time period (either intermittent exposure or continuous exposure)

Intrinsic resistance: a pre-existing resistance usually termed refractory where cancer cells THAT HAVE NOT BEEN EXPOSED to drug, do not respond to initial drug exposure. Can be seen experimentally in panels of unrelated cancer cells lines isolated from untreated patients which show no cytotoxicity to drug exposure in vitro.

Below is one of the first reports which described the drug resistant phenotype of cancer stem cells in an in vivo (mouse) model of breast cancer with videos.

Cancer Res. 2008 May 1;68(9):3243-50. doi: 10.1158/0008-5472.CAN-07-5480.

Cancer stem cells contribute to cisplatin resistance in Brca1/p53-mediated mouse mammary tumors.

Shafee N1, Smith CR, Wei S, Kim Y, Mills GB, Hortobagyi GN, Stanbridge EJ, Lee EY.

Author information


The majority of BRCA1-associated breast cancers are basal cell-like, which is associated with a poor outcome. Using a spontaneous mouse mammary tumor model, we show that platinum compounds, which generate DNA breaks during the repair process, are more effective than doxorubicin in Brca1/p53-mutated tumors. At 0.5 mg/kg of daily cisplatin treatment, 80% primary tumors (n = 8) show complete pathologic response. At greater dosages, 100% show complete response (n = 19). However, after 2 to 3 months of complete remission following platinum treatment, tumors relapse and become refractory to successive rounds of treatment. Approximately 3.8% to 8.0% (mean, 5.9%) of tumor cells express the normal mammary stem cell markers, CD29(hi)24(med), and these cells are tumorigenic, whereas CD29(med)24(-/lo) and CD29(med)24(hi) cells have diminished tumorigenicity or are nontumorigenic, respectively. In partially platinum-responsive primary transplants, 6.6% to 11.0% (mean, 8.8%) tumor cells are CD29(hi)24(med); these populations significantly increase to 16.5% to 29.2% (mean, 22.8%; P < 0.05) in platinum-refractory secondary tumor transplants. Further, refractory tumor cells have greater colony-forming ability than the primary transplant-derived cells in the presence of cisplatin. Expression of a normal stem cell marker, Nanog, is decreased in the CD29(hi)24(med) populations in the secondary transplants. Top2A expression is also down-regulated in secondary drug-resistant tumor populations and, in one case, was accompanied by genomic deletion of Top2A. These studies identify distinct cancer cell populations for therapeutic targeting in breast cancer and implicate clonal evolution and expansion of cancer stem-like cells as a potential cause of chemoresistance.

Please Watch Videos


Below is a curation of talks and abstracts from the 2015 Annual AACR Meeting in Philadelphia, PA.

The Talk by Dr. Cheresh is an example of this school of thought; that inducing cancer cell stemness can result in development of drug resistance, in this case to a TKI. (For a press release on this finding see here.)

SY27-04: Induction of cancer stemness and drug resistance by EGFR blockade
Tuesday, Apr 21, 2015, 12:00 PM -12:15 PM
David A. Cheresh. UCSD Moores Cancer Center, La Jolla, CA

Presentation Title: Induction of cancer stemness and drug resistance by EGFR blockade
Presentation Time: Tuesday, Apr 21, 2015, 12:00 PM -12:15 PM
Abstract Body: Tumor drug resistance is often accompanied by genetic and biological changes in the tumor cell population reflecting the acquisition of a stem-like state. However, it is not clear whether cancer therapies select for the growth of drug resistance cancer stem cells and/or directly induce the reprograming of tumor cells to a cancer stem-like, drug resistance state. We provide evidence that breast, pancreas and lung carcinomas in the presence of prolonged exposure to EGFR inhibitors undergo an epigenetic reprogramming resulting in a drug resistant stem-like tumor population expressing the cell surface marker CD61 (b3 integrin). In fact, CD61 in the context of KRAS, is necessary and sufficient to account for drug resistance, tumor initiation, self-renewal and expression of the pluripotent genes Oct 4 and Nanog. Once expressed, CD61 in the unligated state recruits KRAS to the plasma membrane leading to the activation of RalB, TBK1 and c-Rel driving both stemness and EGFR inhibitor resistance. Pharmacological targeting this pathway with drugs such as bortezomib or revlimid not only reverses stemness but resensitizes these epithelial tumors to EGFR inhibition. This epigenetic pathway can also be initiated by range of cellular stresses found within the tumor microenvironment such as hypoxia, nutrient deprivation, low pH, and oxidative stress. In normal tissues CD61 is induced during tissue remodeling and repair. For example, CD61 was found to be critical for mammary gland remodeling during pregnancy and as a mediator of pathological neovascularization. Together these findings reveal a stress-induced epigenetic pathway characterized by the upregulation of CD61 that promotes the remodeling of normal tissues but in tumors contributes to EGFR inhibitor resistance and tumor progression.

Selected Abstracts

  1. Abstract 1
  2. Molecular and Cellular Biology – Poster Presentations – Proffered Abstracts – Poster Presentations – Cell Death Mechanisms: Abstract 4: ABT-263 is effective in a subset of non-small cell lung cancer cell lines
    • Aoi Kuroda,
    • Keiko Ohgino,
    • Hiroyuki Yasuda,
    • Junko Hamamoto,
    • Daisuke Arai,
    • Kota Ishioka,
    • Tetsuo Tani,
    • Shigenari Nukaga,
    • Ichiro Kawada,
    • Katsuhiko Naoki,
    • Kenzo Soejima,
    • and Tomoko Betsuyaku

Cancer Res August 1, 2015 75:4; doi:10.1158/1538-7445.AM2015-4

  1. Abstract 2
  2. Molecular and Cellular Biology – Poster Presentations – Proffered Abstracts – Poster Presentations – Cell Death Mechanisms: Abstract 6: Quantitative assessment of BCL-2:BIM complexes as a pharmacodynamic marker for venetoclax (ABT-199)
    • Sha Jin,
    • Paul Tapang,
    • Donald J. Osterling,
    • Wenqing Gao,
    • Daniel H. Albert,
    • Andrew J. Souers,
    • Joel D. Leverson,
    • Darren C. Phillips,
    • and Jun Chen

Cancer Res August 1, 2015 75:6; doi:10.1158/1538-7445.AM2015-6

  1. Molecular and Cellular Biology – Poster Presentations – Proffered Abstracts – Poster Presentations – Cell Death Mechanisms: Abstract 24: The phosphorylation of p53 at serine 46 is essential to induce cell death through palmdelphin in response to DNA damage
    • Nurmaa Khund Dashzeveg and
    • Kiyotsugu Yoshida

Cancer Res August 1, 2015 75:24; doi:10.1158/1538-7445.AM2015-24

  1. Abstract 5
  2. Molecular and Cellular Biology – Poster Presentations – Proffered Abstracts – Poster Presentations – Cell Signaling in Cancer 1: Abstract 48: Identification of a novel binding protein playing a critical role in HER2 activation in lung cancer cells
    • Tomoaki Ohtsuka,
    • Masakiyo Sakaguchi,
    • Katsuyoshi Takata,
    • Shinsuke Hashida,
    • Mototsugu Watanabe,
    • Ken Suzawa,
    • Yuho Maki,
    • Hiromasa Yamamoto,
    • Junichi Soh,
    • Hiroaki Asano,
    • Kazunori Tsukuda,
    • Shinichiro Miyoshi,
    • and Shinichi Toyooka

Cancer Res August 1, 2015 75:48; doi:10.1158/1538-7445.AM2015-48

  1. Abstract 1 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Death Mechanisms

Abstract 4: ABT-263 is effective in a subset of non-small cell lung cancer cell lines

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA


ABT-263 (Navitoclax) is one of the BH3 mimetics targeting anti-apoptotic B-cell lymphoma-2 (Bcl-2) family proteins such as Bcl-2, Bcl-XL, and Bcl-w, thereby inducing apoptosis. It has been reported that the response to ABT-263 is associated with expressions of myeloid cell leukemia-1 (Mcl-1), an anti-apoptotic protein. Given its effectiveness as a single agent in preclinical studies, ABT-263 is currently being evaluated in clinical trials for small cell lung cancer (SCLC) and leukemia. However, the efficacy of ABT-263 in non-small cell lung cancer (NSCLC) has not been fully evaluated. We examined the effect of ABT-263 on cell proliferation of NSCLC cell lines and investigated the underlying mechanisms.


The following 9 NSCLC cell lines were examined: SK-LU-1, A549, H358, Calu3, H3122, H1975, H460, H441, and BID007. The effects of ABT-263 in NSCLC cell lines were evaluated by MTS assay. Apoptosis was examined by flowcytometry using staining for annexin V and propidium iodide (PI), and also western blotting for cleaved PARP. Quantitative RT-PCR was carried out to assess the mRNA expression levels of anti-apoptotic genes and pro-apoptotic genes. Immunoprecipitation and western blotting were performed to compare the levels of anti-apoptotic and pro-apoptotic proteins between the sensitive and resistant cell lines. In addition, knockdown of Mcl-1 was performed by siRNA.


By screening 9 NSCLC cell lines using MTS assay, we found Calu3 and BID007were sensitive to ABT-263. We also confirmed that apoptosis was induced only in the ABT-263 sensitive lines but not in the ABT-263 resistant cell lines after ABT-263 treatment. However, the expression levels of Bcl-2 family proteins, including Mcl-1, did not differ significantly among the ABT-263 sensitive and resistant cell lines. Unlike the results in previous reports regarding SCLC, Mcl-1 was not decreased in the sensitive cell lines. The ABT-263 resistant cell lines became sensitive to ABT-263 after knockdown of Mcl-1 by siRNA, while the ABT-263 sensitive cell lines maintained the same sensitivity.


We found that Calu3 and BID007 were sensitive to ABT-263. In the sensitive NSCLC cell lines, ABT-263 induces apoptosis irrespective of Mcl-1 expression levels.

Citation Format: Aoi Kuroda, Keiko Ohgino, Hiroyuki Yasuda, Junko Hamamoto, Daisuke Arai, Kota Ishioka, Tetsuo Tani, Shigenari Nukaga, Ichiro Kawada, Katsuhiko Naoki, Kenzo Soejima, Tomoko Betsuyaku. ABT-263 is effective in a subset of non-small cell lung cancer cell lines. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4. doi:10.1158/1538-7445.AM2015-4

    • ©2015 American Association for Cancer Research.
  1. Abstract 2 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Death Mechanisms

Abstract 6: Quantitative assessment of BCL-2:BIM complexes as a pharmacodynamic marker for venetoclax (ABT-199)

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

The BCL-2-selective inhibitor venetoclax (ABT-199) binds with high affinity to the BH3-binding groove of BCL-2, thereby competing for binding with the BH3-only protein BIM (Souers et al., 2013). Venetoclax is currently being evaluated in clinical trials for CLL, AML, multiple myeloma and NHL. To facilitate these studies, we developed and validated a 384-well electrochemiluminescent ELISA (MSD, Gaithersburg, MD,USA) that quantifies expression of BCL-2, BCL-XL, and MCL-1protein alone or in complex with BIM. We subsequently quantified expression of BCL-2 and BCL-2:BIM complexes in 16 hematologic tumor cell lines. We found the EC50 of venetoclax in these tumor cell lines to correlate strongly with baseline BCL-2:BIM complex levels. This correlation was superior to the correlation between venetoclax EC50 and absolute BCL-2 expression. We also applied the assay to measure disruption of BCL-2:BIM complexes in vivo. Treatment of the Non-Hodgkin’s Lymphoma (NHL) xenograft model SU-DHL-4 with a BCL-2-selective inhibitor resulted in disruption of tumor BCL-2:BIM complexes that aligned with serum and tumor concentrations of inhibitor. Collectively, these data demonstrate that quantifying BCL-2:BIM complexes offers an accurate means of assessing target engagement by venetoclax and, potentially, predicting its efficacy. The utility of this assay is currently being assessed in clinical trials.

Citation Format: Sha Jin, Paul Tapang, Donald J. Osterling, Wenqing Gao, Daniel H. Albert, Andrew J. Souers, Joel D. Leverson, Darren C. Phillips, Jun Chen. Quantitative assessment of BCL-2:BIM complexes as a pharmacodynamic marker for venetoclax (ABT-199). [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 6. doi:10.1158/1538-7445.AM2015-6

    • ©2015 American Association for Cancer Research.
  1. Abstract 3 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Death Mechanisms

Abstract 19: Antitumor activity of selective inhibitors of XPO1/CRM1-mediated nuclear export in diffuse malignant peritoneal mesothelioma: the role of survivin

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

Survivin, which is highly expressed and promotes cell survival in diffuse malignant peritoneal mesothelioma (DMPM), exclusively relies on the nuclear exportin 1 (XPO1/CRM1) to be released in the cytoplasm and perform its anti-apoptotic function. Here, we explored the efficacy of selective inhibitors of nuclear export (SINEs) in patient-derived DMPM preclinical models. Exposure to individual SINE (KPT-251, KPT-276, KPT-330) was able to induce a time- and dose-dependent inhibition of the growth of two DMPM cell lines without affecting normal cell proliferation. Such a cell growth inhibition was preceded by a decline in the nuclear XPO1/CRM1 levels and an increase in the nuclear accumulation of its cargo proteins p53 and p21, which led to a cell cycle arrest at G1-phase. Our results also indicated that survivin is an essential component of the downstream signaling pathway of XPO1/CRM1 inhibition in DMPM cells. In fact, in both cell lines, exposure to SINEs led to a time-dependent reduction of cytoplasmic survivin levels and, after an initial survivin nuclear accumulation, also to a progressive decrease in the nuclear protein abundance, through the ubiquitin-proteasomal degradation pathway, leading to the complete depletion of total survivin levels. In both DMPM cell models, according to survivin anti-apoptotic activity, drug-induced reduction of cytoplasmic survivin levels correlated with the onset of caspase-dependent apoptosis. We further observed that SINEs can be combined with other survivin inhibitors, such as the survivin suppressant YM155 to achieve enhanced growth inhibition in DMPM cells. Initial in vivo experiments with orally administered KPT-251, KPT-276 and the orally available, clinical stage KPT-330 (selinexor) indicated that each compound was able to significantly reduce the growth of early-stage subcutaneous DMPM xenografts. Interestingly, additional experiments carry out with selinexor demonstrated that the compound was also able to inhibit the growth of late-stage subcutaneous DMPM xenografts in nude mice. Most importantly, oral administration of selinexor to SCID mice reduced the growth of orthotopic DMPM xenografts, which properly recapitulate the dissemination pattern in the peritoneal cavity of human DMPM and, for this reason, represent a valuable model for investigating novel therapeutic approaches for the disease. Consistent with an important role of survivin as a determinant of anti-cancer activity of SINE compounds, a reduction of the protein expression was observed in tumor specimens obtained from selinexor treated mice. Overall, our results (i) demonstrate a marked efficacy of SINEs in DMPM preclinical models, which is, at least in part, dependent on the interference with survivin intracellular distribution and function, and (ii) suggest SINE-mediated XPO1/CRM1 inhibition as a novel therapeutic option for the disease.

Citation Format: Nadia Zaffaroni, Michelandrea De Cesare, Denis Cominetti, Valentina Doldi, Alessia Lopergolo, Marcello Deraco, Paolo Gandellini, Yosef Landesman, Sharon Friedlander, Michael G. Kauffman, Sharon Shacham, Marzia Pennati. Antitumor activity of selective inhibitors of XPO1/CRM1-mediated nuclear export in diffuse malignant peritoneal mesothelioma: the role of survivin. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 19. doi:10.1158/1538-7445.AM2015-19

    • ©2015 American Association for Cancer Research.
  1. Abstract 4 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Death Mechanisms

Abstract 24: The phosphorylation of p53 at serine 46 is essential to induce cell death through palmdelphin in response to DNA damage

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

Tumor suppressor p53 plays a pivotal role in cell cycle arrest, DNA repair, and apoptosis in response to DNA damage. Promoter selectivity of p53 depends mainly on post-translational modification. Notably, the apoptotic function of p53 is related to its phosphorylation at serine-46 (ser46) to promote pro-apoptotic genes. However, little is known about the pro-apoptotic genes induced by Ser46 phosphorylation. Our research achieved to investigate the pro-apoptotic genes induced by p53 in a phospho-ser46-specific manner using microarray and ChIP sequencing in human cancer cell lines. As a result, palmdelphin (PALMD), an isoform of paralemmin protein, was strongly elicited from the phosphorylation of ser46. The mRNA and protein expression of PALMD increased only in wild type p53 transfected cells, but not in ser46-mutated cells. Importantly, PALMD moved to the nucleus in response to DNA damage and the apoptotic function of PALMD was tightly exerted with localization into nucleus. Interestingly, down-regulation of PALMD by siRNA resulted in necroptosis-like cell death through ATP depletion. Moreover, we found vimentin as a PALMD interacting protein and the depletion of vimentin increased PALMD level to accelerate apoptosis. These results demonstrate that p53 regulates cell death fate (apoptosis or necroptosis-like cell death) through promoting PALMD expression in a phospho-ser46-specific manner in response to DNA damage.

Citation Format: Nurmaa Khund Dashzeveg, Kiyotsugu Yoshida. The phosphorylation of p53 at serine 46 is essential to induce cell death through palmdelphin in response to DNA damage. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 24. doi:10.1158/1538-7445.AM2015-24

    • ©2015 American Association for Cancer Research.
  1. Abstract 5 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Signaling in Cancer 1

Abstract 48: Identification of a novel binding protein playing a critical role in HER2 activation in lung cancer cells

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

Human epidermal growth factor receptor 2 (HER2) is a member of epidermal growth factor receptor (EGFR) family. Previous studies have revealed that many kinds of malignant tumors have genetic mutations or amplification of HER2, indicating that HER2 alterations are oncogenic. Many kinds of HER2 targeted therapies are effective to HER2 positive tumors, but those treated tumors often get resistance to drugs. Thus, to elucidate HER2 related pathway in cancer biology is important to develop new therapeutic strategy for cancers.

Recently, we newly identified a protein X (a temporary name) as a novel binding protein to HER2 with immunoprecipitation and following LC-Ms/Ms analysis. The protein generally expressed in lung and breast cancers at remarkable level.

We constructed plasmid vectors carrying wild type HER2 and gene X. These vectors were simultaneously introduced to HEK293T cells to examine the binding ability of protein X and HER2 as well as the effect of gene X on HER2-mediated signal-transduction pathway. The approach clearly showed that the expression of gene X, resulted in phosphorylation of HER2 and subsequent activation of oncogenic effector molecules.

We next constructed several kinds of gene X-truncated variants and subjected to the binding assay to look for the binding domain of gene X to HER2. The analysis showed that N-terminal head domain of gene X was essential for the HER2 binding. This domain has an ability to induce HER2 phosphorylation and subsequent activation of the effector kinase, ERK.

In conclusion, we found that gene X is a novel binding protein to HER2 and has a role in HER2 activation.

Citation Format: Tomoaki Ohtsuka, Masakiyo Sakaguchi, Katsuyoshi Takata, Shinsuke Hashida, Mototsugu Watanabe, Ken Suzawa, Yuho Maki, Hiromasa Yamamoto, Junichi Soh, Hiroaki Asano, Kazunori Tsukuda, Shinichiro Miyoshi, Shinichi Toyooka. Identification of a novel binding protein playing a critical role in HER2 activation in lung cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 48. doi:10.1158/1538-7445.AM2015-48

    • ©2015 American Association for Cancer Research.
  1. Abstract 6 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Signaling in Cancer 1

Abstract 54: Ezrin enhances signaling and nuclear translocation of the epidermal growth factor receptor in non-small cell lung cancer cells

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

The cytoskeletal cross linker protein ezrin is a member of the ezrin-radixin-moesin (ERM) family and plays important roles not only in cell motility, cell adhesion, and apoptosis, but also in various cell-signaling pathways. Ezrin interacts with EGFR in the cell membrane and involves in cell motility events, but little is known about the effects of this interaction on the EGFR signaling pathway. We investigated the role of Ezrin in EGFR signaling and nuclear trafficking in non-small cell lung cancer (NSCLC) cell lines. The ligand induced interaction between Ezrin and EGFR was evaluated by immunoprecipitation (IP) and immunofluorescence (IF) in H292 and A549 cells. Ezrin levels were reduced using siRNA in these two cell lines. Downstream signaling protein phosphorylation and nuclear localization of EGFR were detected after EGF treatment. Expressions of nuclear EGFR target genes were evaluated by qPCR. Endogenous Ezrin was found in a complex with EGFR in IP and IF. When Ezrin protein expression was inhibited, phosphorylation levels of EGFR at Y1068, Y1101 and Y845 were reduced as well as phosphorylation levels of downstream signaling pathway proteins ERK and STAT3. Cell fractionation revealed that EGFR nuclear translocation after EGF treatment significantly reduced in Ezrin-knockdown cells. Further, mRNA levels of EGFR target genes AuroraK-A, COX2, Cyclin D1 and iNOS were decreased in Ezrin-knockdown A549 cells. Small molecule ezrin inhibitors showed strong synergy with EGFR inhibitors in cytotoxicity assays. These results suggest that Ezrin has a role as an enhancer in the EGFR pathway and targeting ezrin may potentiate anti-EGFR based therapies in NSCLC.

Citation Format: Yasemin Saygideger Kont, Haydar Celik, Hayriye V. Erkizan, Tsion Minas, Jenny Han, Jeffrey Toretsky, Aykut Uren. Ezrin enhances signaling and nuclear translocation of the epidermal growth factor receptor in non-small cell lung cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 54. doi:10.1158/1538-7445.AM2015-54

    • ©2015 American Association for Cancer Research.
  1. Abstract 7 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Cell Signaling in Cancer 1

Abstract 57: Substrates of protein kinase C drive cell rac1-dependent motility

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

This laboratory has identified and/or characterized substrates of PKC that upon phosphorylation give rise to motility, an aspect of metastasis. By use of the traceable kinase method, we discovered that alpha-tubulin and Cdc42 effector protein-4 (CEP4) are PKC substrates. Phosphorylation of alpha-tubulin stimulates its incorporation into microtubules (MTs), consequently increasing the stability and prolonged growth of MTs and leading to the activation of the small GTPase Rac1. CEP4 undergoes phosphorylation by PKC that results in its release from Cdc42, whereupon CEP4 binds a guanine nucleotide exchange factor (GEF) that in turn activates Rac1 GTPase. These results imply that Rac1 acts as a node in pathways driven by phosphorylated PKC substrates. Since translocation of IQGAP to the membrane is known to be promoted by Rac1, a role is explored in non-transformed human MCF-10A cells that express a specific phospho-mimetic mutant substrate. In addition, the phospho-mimetic mutant for each substrate expressed in human metastatic MDA-MB-231 cells produces different morphologies in 3-D growth assays. This research is being supported by NIH CA125632.

Citation Format: Susan A. Rotenberg, Xin Zhao, Shatarupa De. Substrates of protein kinase C drive cell rac1-dependent motility. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 57. doi:10.1158/1538-7445.AM2015-57

    • ©2015 American Association for Cancer Research.
  1. Abstract 8 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Deregulation of Gene Expression in Prostate Cancer and Sarcoma

Abstract 88: The Nkx3.1 homeobox gene maintains prostatic identity while its loss leads to prostate cancer initiation

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA


Maintenance of epithelial cell identity is tightly coordinated by tissue-specific gene expression programs, which are often deregulated during tumorigenesis. The homeodomain-containing transcription factor, Nkx3.1, is a key regulator of normal prostatic development and is frequently lost at early stages of prostate cancer initiation. In this study, we aim to elucidate detailed mechanisms governing Nkx3.1-driven maintenance of prostate identity and how deregulation of such can lead to prostate tumorigenesis.

Models and Methods

We evaluated the consequences of Nkx3.1 loss or gain of function in vivo using genetically-engineered mouse models and cell-recombination assays. RNA sequencing was performed to generate gene expression profiles, which were analyzed using Gene Set Enrichment analysis (GSEA), and validated by quantitative real-time PCR. In parallel, protein expression was assessed by immunofluorescence and western blot. Immunoprecipitation (IP) and chromatin-immunoprecipitation (ChIP) assays were performed using RWPE1 prostate epithelial cells.


Here, we show that loss of function of Nkx3.1 leads to the progressive down-regulation of a prostate-specific gene expression program and to aberrant expression of genes that are not typically expressed in the prostate epithelium. Conversely, gain of function of Nkx3.1 in non-prostatic epithelium leads to the acquisition of a prostate-like morphology and expression of prostate-related genes. Our findings indicate that the underlying mechanism by which Nkx3.1 promotes prostatic identity is via epigenetic regulation of gene expression. In particular, we show that Nkx3.1 interacts with the histone methyl-transferase complex G9a/Glp. Finally, we demonstrate that this interaction is necessary for maintenance of prostate identity in vivo and that Nkx3.1 and G9a cooperate to control expression of genes that coordinate prostatic epithelial integrity.


Our results suggest that Nkx3.1 promotes prostatic identity by interacting with histone modifying enzymes to coordinate the expression of prostate-specific genes and that the loss of this function results in a failure to maintain prostate identity associated with early stages of prostate tumorigenesis.

Citation Format: Clémentine Le Magnen, Aditya Dutta, Cory Abate-Shen. The Nkx3.1 homeobox gene maintains prostatic identity while its loss leads to prostate cancer initiation. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 88. doi:10.1158/1538-7445.AM2015-88

    • ©2015 American Association for Cancer Research.
  1. Abstract 9 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Deregulation of Gene Expression in Prostate Cancer and Sarcoma

Abstract 90: K63-linked JARID1B ubiquitination by TRAF6 contributes to aberrant elevation of JARID1B in prostate cancer

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

Aberrant elevation of JARID1B and histone H3 Lys4 trimethylations (H3K4me3) is frequently observed in many diseases including prostate cancer (PCa), yet the mechanisms on the regulations of JARID1B and H3K4me3 through epigenetic modifications still remain poorly understood. In this study we performed immunohistochemistry staining, immunofluorescence imaging, immunoprecipitation, shRNA and Western blotting analysis in mouse embryonic fibroblasts (MEFs), mouse models, and cultured human prostate cancer cells. As a result, we discovered that SKP2 modulates JARID1B and H3K4me3 levels in vitro in PTEN null prostate cancer cells and in vivo in Pten/Trp53 mouse models. We demonstrated that levels of SKP2, JARID1B and H3K4me3 are strikingly elevated in vitro and in vivo when both PTEN and P53 are inactivated. Importantly, SKP2 inactivation resulted in a reduction of cell growth, cell migration and malignant transformation of Pten/Trp53 double null MEFs, and further restrained prostate tumorigenesis of Pten/Trp53 mutant mice. Mechanistically, JARID1B is ubiquitinated by E3 ligase TRAF6 through the K63-linkage in prostate cancer cells. Interestingly, SKP2 contributes to JARID1B ubiquitination machinery as a non-E3 ligase regulator by decreasing TRAF6-mediated ubiquitination of JARID1B. SKP2 deficiency resulted in an increase of JARID1B ubiquitination and in turn a reduction of H3K4me3, and induced senescence through JARID1B accumulation in nucleoli of PCa cells and prostate tumors of mice. Furthermore, we showed that the aberrant levels of SKP2, JARID1B, and H3K4me3 are associated with malignant features of castration-resistant prostate cancer (CRPC) in mice. Overall, our findings reveal a novel network of SKP2- JARID1B, and targeting SKP2 and JARID1B may be a potential strategy for PCa control.

Citation Format: Wenfu Lu, Shenji Liu, Bo Li, Yingqiu Xie, Christine Adhiambo, Qing Yang, Billy R. Ballard, Keiichi I. Nakayama, Robert J. Matusik, Zhenbang Chen. K63-linked JARID1B ubiquitination by TRAF6 contributes to aberrant elevation of JARID1B in prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 90. doi:10.1158/1538-7445.AM2015-90

    • ©2015 American Association for Cancer Research.
  1. Abstract 10 of 10Molecular and Cellular Biology / Poster Presentations – Proffered Abstracts / Poster Presentations – Histone Methylation and Acetylation

Abstract 97: CARM1 preferentially methylates H3R17 over H3R26 through a random kinetic mechanism

Proceedings: AACR 106th Annual Meeting 2015; April 18-22, 2015; Philadelphia, PA

CARM1 (PRMT4) is a type I arginine methyltransferase involved in the regulation of transcription, pre-mRNA splicing, cell cycle progression and the DNA damage response. Overexpression of CARM1 has been implicated in breast, prostate, and colorectal cancers. Since CARM1 appears to be a good target for the development of therapies against these cancers, we studied the substrate specificity and kinetic mechanism of the full-length human enzyme. CARM1 has been shown to methylate both residues R17 and R26 of histone H3. Substrate specificity was examined by testing CARM1 activity with several H3-based peptide substrates using a radiometric assay. Comparison of kcat/KM values reveal that methylation of H3R17 is preferred over H3R26. An R17/R26K peptide produced 8-fold greater kcat/KM value compared to the corresponding R17K/R26 peptide. These effects are KM-driven as kcat values remain relatively constant for the peptides tested. Shortening the peptide at the C-terminus by 5 amino acid residues greatly reduced the specificity (16-24-fold), demonstrating the contribution of distal residues to substrate binding. In contrast, adding residues to the N-terminus of the shortened peptide had a negative effect on activity. CARM1 displays little preference for monomethylated over unmethylated H3R17 (2-5-fold by kcat/KM) suggesting that it operates through a distributive mechanism. Previous crystallographic studies with mouse CARM1 showed that part of the substrate binding groove was formed by cofactor binding, thereby suggesting an ordered kinetic mechanism (Yue et al., EMBO J., 2007). Our results from dead-end and product inhibition studies performed with human CARM1, however, are consistent with a random kinetic mechanism. SAH and sinefungin demonstrate competitive inhibition with respect to SAM and produced noncompetitive inhibition patterns with respect to peptide. Both dimethylated R17 product peptide and dead-end R17K peptide exhibited noncompetitive inhibition patterns with respect to SAM. Furthermore, binding of SAM and peptide substrates were shown to be independent of each other in initial velocity experiments where both substrates were varied. Together, these results elucidate the kinetic mechanism of CARM1 and highlight elements important for binding affinity.

Citation Format: Suzanne L. Jacques, Katrina P. Aquino, Jodi Gureasko, P Ann Boriack-Sjodin, Robert A. Copeland, Thomas V. Riera. CARM1 preferentially methylates H3R17 over H3R26 through a random kinetic mechanism. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 97. doi:10.1158/1538-7445.AM2015-97

    • ©2015 American Association for Cancer Research.



  1. Bonnet D, Dick JE: Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell. Nat Med 1997, 3(7):730-737.
  2. Al-Hajj M, Clarke MF: Self-renewal and solid tumor stem cells. Oncogene 2004, 23(43):7274-7282.
  3. Hughes L, Malone C, Chumsri S, Burger AM, McDonnell S: Characterisation of breast cancer cell lines and establishment of a novel isogenic subclone to study migration, invasion and tumourigenicity. Clin Exp Metastasis 2008, 25(5):549-557.
  4. Li C, Lee CJ, Simeone DM: Identification of human pancreatic cancer stem cells. Methods Mol Biol 2009, 568:161-173.
  5. Zhang S, Balch C, Chan MW, Lai HC, Matei D, Schilder JM, Yan PS, Huang TH, Nephew KP: Identification and characterization of ovarian cancer-initiating cells from primary human tumors. Cancer Res 2008, 68(11):4311-4320.
  6. Kakarala M, Wicha MS: Implications of the cancer stem-cell hypothesis for breast cancer prevention and therapy. J Clin Oncol 2008, 26(17):2813-2820.
  7. Ginestier C, Hur MH, Charafe-Jauffret E, Monville F, Dutcher J, Brown M, Jacquemier J, Viens P, Kleer CG, Liu S et al: ALDH1 is a marker of normal and malignant human mammary stem cells and a predictor of poor clinical outcome. Cell Stem Cell 2007, 1(5):555-567.
  8. Dontu G: Breast cancer stem cell markers – the rocky road to clinical applications. Breast Cancer Res 2008, 10(5):110.
  9. Ferrandina G, Bonanno G, Pierelli L, Perillo A, Procoli A, Mariotti A, Corallo M, Martinelli E, Rutella S, Paglia A et al: Expression of CD133-1 and CD133-2 in ovarian cancer. Int J Gynecol Cancer 2008, 18(3):506-514.


Additional Articles on this Open Access Journal on Cancer Stem Cells Include

Nonhematologic Cancer Stem Cells [11.2.3]

In Focus: Identity of Cancer Stem Cells

In Focus: Targeting of Cancer Stem Cells

Stem Cells and Cancer

Positron Emission Tomography (PET) and Near-Infrared Fluorescence Imaging: Noninvasive Imaging of Cancer Stem Cells (CSCs) monitoring of AC133+ glioblastoma in subcutaneous and intracerebral xenograft tumors

“To Die or Not To Die” – Time and Order of Combination drugs for Triple Negative Breast Cancer cells: A Systems Level Analysis

Can IntraTumoral Heterogeneity Be Thought of as a Mechanism of Resistance?



Read Full Post »

New Topoisomerase Inhibitors in Clinical Trials

Curator: Stephen J. Williams, Ph.D.

Below is a great review of topoisomerase in cancer, approved inhibitors as well as some in clinical trials.

Biomolecules 2015, 5, 1652-1670; doi:10.3390/biom5031652



ISSN 2218-273X


Inhibition of Topoisomerase (DNA) I (TOP1): DNA Damage Repair and Anticancer Therapy

Yang Xu and Chengtao Her *

School of Molecular Biosciences, College of Veterinary Medicine, Washington State University, Mail Drop 64-7520, Pullman, WA 99164, USA; E-Mail:

* Author to whom correspondence should be addressed; E-Mail:; Tel.: +1-509-335-7537; Fax: +1-509-335-4159.

Academic Editors: Wolf-Dietrich Heyer, Thomas Helleday and Fumio Hanaoka Received: 22 May 2015 / Accepted: 14 July 2015 / Published: 22 July 2015

Abstract: Most chemotherapy regimens contain at least one DNA-damaging agent that preferentially affects the growth of cancer cells. This strategy takes advantage of the differences in cell proliferation between normal and cancer cells. Chemotherapeutic drugs are usually designed to target rapid-dividing cells because sustained proliferation is a common feature of cancer [1,2]. Rapid DNA replication is essential for highly proliferative cells, thus blocking of DNA replication will create numerous mutations and/or chromosome rearrangements—ultimately triggering cell death [3]. Along these lines, DNA topoisomerase inhibitors are of great interest because they help to maintain strand breaks generated by topoisomerases during replication. In this article, we discuss the characteristics of topoisomerase (DNA) I (TOP1) and its inhibitors, as well as the underlying DNA repair pathways and the use of TOP1 inhibitors in cancer therapy.

Biomolecules 2015, 5                                                                                                                           1653

  1. Type IB Topoisomerases and Inhibitors
    1.1. TOP1

DNA topoisomerases resolve topological constraints that may arise from DNA strand separation and are therefore important for transcription and replication [4]. There are six topoisomerases in humans, classified as Type IA, IB and IIA. Type IA topoisomerases TOP3a and TOP3b cleave one DNA strand to relax only negative supercoiling. In addition, TOP3a forms the BTR complex with BLM and RMI1/2, which plays a role in the dissolution of double-Holliday junctions [5]. Type IIA topoisomerases TOP2a and TOP2b generate double-strand breaks on one DNA molecule to allow the passing of other DNA strands [6]. Topoisomerases are attractive drug targets in cancer therapy. For example, the commonly used anticancer agents doxorubicin and etoposide (VP-16) are TOP2 inhibitors [7]. Type IB topoisomerases include the nuclear TOP1 and mitochondrial TOP1mt [4]. TOP1 initiates the DNA relaxation by nicking one DNA strand. It then forms a TOP1-DNA cleavage complex (TOP1cc) by covalently linked to the 3′-phosphate end via its tyrosine residue Y723 (3′-P-Y). Following the resolution of topological entanglements and the removal of TOP1, the 5′-hydroxyl end is realigned with the 3′-end for religation. Each nicking-closing cycle enables the relaxation of one DNA supercoiling (Figure 1).

Figure 1. A schematic representation of strand passages catalyzed by three types of topoisomerases (adapted from ref. [8]).


TOP1 is essential for embryonic development in mammals [9]. Although TOP1 plays an important role in the deconvolution of supercoils arising amid DNA replication, the precise steps involved with

Biomolecules 2015, 5                                                                                                                         1654

the recruitment of TOP1 to topological constraints remains to be revealed. It appears that in yeast TOP1 travels at a distance of 600 bp ahead of the replication fork [10] and remains associated with the GINS-MCM complex [11]. However, the yeast TOP1 is distinct from its human counterpart in that it has little effect on fork progression or the firing of replication origin [12]. In humans, TOP1 binds to the regions of the pre-replicative complex in cells during the M, early G1, and G1/S phases of the cell cycle to control the firing of replication origins [12]. This difference may explain why yeast cells are viable in the absence of TOP1. In addition, TOP1 also has functions in transcription that are independent of its role in resolving DNA topological entanglements. First, TOP1 is known to repress transcription by binding to TFIID [13]. Second, inhibition of TOP1 can cause the induction of c-Jun in leukemia cells, suggesting its additional role in the control of transcription [14]. Furthermore, TOP1 interacts with the splicing factor ASF/SF2 by which it promotes the maturation of RNA—through suppressing the formation of R-loops (RNA-DNA hybrids)—and prevents collision between transcription bubble and replication fork [15,16]. It appears that the levels of TOP1 have to be dynamically regulated. In B cells, TOP1 is reduced by activation-induced cytidine deaminase (AID) to facilitate class-switch recombination (CSR) and somatic hypermutation (SHM) [17,18]. Although TOP1mt is important for mitochondrial integrity and metabolism, mice lacking mitochondrial TOP1mt are viable and fertile but they are associated with increased negative supercoiling of mtDNA [19,20].

1.2. TOP1 Inhibitors

Stabilization of TOP1cc by topoisomerase poison is detrimental to cells due to the disruption of DNA uncoiling, increased strand breaks, and unstable RNA transcripts as well as incomplete DNA replication [21]. The TOP1 inhibitor camptothecin (CPT), first isolated from the Chinese tree Camptotheca acuminate, was clinically used for cancer treatment long before it was identified as a TOP1 inhibitor [22]. Due to side effects, CPT is no longer used clinically and it has been replaced by more effective and safer TOP1 inhibitors [23]. Currently, CPT derivatives topotecan (trade name: Hycamtin) and irinotecan (CPT-11, trade name: Camptosar) are routinely used to treat colorectal, ovarian and lung cancers, while a few other TOP1 inhibitors are being tested in clinical trials.

CPT is a 5-ring alkaloid that is active in its closed E-ring (lactone) form but it is inactive with an open E-ring (carboxylate) at physiological and alkaline pH [24]. Therefore, CPT is not effective for inhibiting TOP1mt due to a higher pH mitochondrial environment. The inactive form of CPT tends to bind to serum albumin, which might be a reason for its side effects. CPT is highly specific for TOP1 and the binding is of relatively low affinity and can be reversed after drug removal. These features make the action of CPT controllable [24], and in fact CPT is widely used in studies of replication-associated DNA damage response. There are a few CPT derivatives and non-CPT TOP1 inhibitors [4,8,24]. For example, CPT derivatives Diflomotecan and S39625 were designed to stabilize the E-ring. Irinotecan has the bis-piperidine side chain to increase its water solubility, but it also contributes to some side effects. Non-CPTs—such as indolocarbazoles, phenanthrolines (e.g., ARC-111) and indenoisoquinolines—refer to drugs that have no typical CPT E-ring structures but they can still specifically target TOP1 and bind irreversibly to TOP1cc. Some of the CPT derivatives (i.e., Gimatecan and Belotecan) and non-CPTs (i.e., NSC 725776 and NSC 724998) are presently tested in clinical trials [23].

Biomolecules 2015, 5                                                                                                                           1655

How does CPT trap TOP1cc? Analysis of the crystal structure and modeling suggest that CPT-TOP1-DNA forms a ternary complex to prevent the two DNA ends from religation [25–27]. Although it is still controversial on how CPT is intercalated into DNA, it seems that CPT traps TOP1cc with a thymine (T) at the -1 position and a guanine (G) at the +1 position on the scissile strand, and it is therefore sequence-specific [28]. Three amino acid residues of the TOP1 enzyme, R364, D533 and N722, combined with DNA bases, contribute to the stabilization of the ternary complex by forming hydrogen bonds and hydrophobic interactions. It is of note that several point mutations, including N722S, in Camptotheca acuminata TOP1 confer resistance to CPT [29]. Interestingly, the same amino acids also contribute to the inhibition of TOP1 by non-CPT drugs [24].

  1. Repair of TOP1 Poison-Induced DNA Lesions

As aforementioned, CPT-induced trapping of TOP1cc creates a single strand break with a free 5′-hydroxyl group, whereas the 3′-phosphate is connected to Y723 of TOP1 (3′-P-Y). At least two pathways contribute to the repair of DNA lesions created by TOP1 poison [30]. The tyrosyl-DNA-phosphodiesterase (TDP1) pathway starts with the ubiquitination and proteasome-mediated degradation of TOP1 in the CPT-TOP1-DNA complex to generate a 3′-P end linked to a short peptide [31]. TDP1 then cleaves the P-Y bond to release the 3′-P end; however, the 3′-P end cannot be directly ligated to the 5′-OH end because of the requirements of DNA ligases. The human polynucleotide kinase (PNKP) can process the DNA ends by functioning as both a 3′-phosphatase and a kinase to generate the required 3′-OH and 5′-P termini for direct ligation. The rest of the repair events can be best described by the single-strand break (SSB) repair pathway, which will be discussed below. Indeed, TDP1 and PNKP are tightly associated with the SSB repair machinery [32,33].

The endonuclease pathway requires multiple endonucleases to excise the DNA—usually at a few nucleotides away from the 3′-P-TOP1 end – on the scissile strand to release the DNA-TOP1 complex [30]. Initial studies were carried out to identify genes that functioned in CPT repair in the absence of TDP1 in yeast [34,35]. These studies led to the identification of RAD1-RAD10, SLX1-SLX4, MUS81-MMS4, MRE11-SAE2 as well as genes involved in recombination. The RAD1-RAD10 (human XPF/ERCC4-ERCC1) complex is a DNA structure-specific endonuclease that can act on 5′ overhang structures [36]. Interestingly, the cleavage site of XPF-ERCC1 is in the non-protruding DNA strand, about 3–4 nucleotides away from the 3′ end [36]. Therefore, trapped TOP1ccs can be removed by this endonuclease activity. Likewise, MUS81-MMS4 (human MUS81-EME1) can also cleave nicked duplex at the 5′ of the nick [37]. The SLX1-SLX4 endonuclease, although not tested on nicked duplexes, is able to process 3′ flap and other DNA structures [38,39]. In human cells, SLX4 also associates with XPF-ERCC1 and MUS81-EME1 endonucleases to process specific DNA intermediates [39,40]. Moreover, MRE11-RAD50 cleaves the 3′-P-Y bond and resects DNA to produce a 3′-OH end [41]. A direct role of SAE2 (human CtIP) in processing 3′-P-TOP1 is unknown, and its endonuclease activity appears to be limited to the 5′ flap or DNA “hairpin” structures [42,43]. Nonetheless, the endonuclease activity of CtIP is essential for processing CPT adducts [42]. In addition, like CtIP, the 5′ flap endonuclease RAD27 (human FEN1) seems to be unable to directly process 3′-P-TOP1 ends [44]. However, the gap endonuclease activity of FEN1 is important for processing stalled replication forks and CPT-induced adducts [45]. The role of FEN1 in SSB repair will be discussed further in the next section.

Biomolecules 2015, 5                                                                                                                           1656

During DNA replication, SSBs created by CPT are most likely converted to double-strand breaks (DSBs) by replication fork runoff. This conversion appears to be dependent on the proteolysis of TOP1 [46]. The repair of one-ended DSBs, as will be discussed in the next section, is largely dependent on homologous recombination (HR). However, low doses of CPT may also induce PARP1 and/or RAD51 dependent replication fork regression—generating no or few DSBs [47,48]. The regressed fork leads to the formation of a “chicken foot” DNA structure by newly synthesized strands [3,49,50]. The formation of regressed fork can be largely suppressed by ATR, EXO1, and DNA2 [51–53]. However, fork reversal can also be beneficial as it provides time for the repair of TOP1-induced DNA lesions by TDP1, thereby preventing DSB formation and the activation of error-prone non-homologous end-joining (NHEJ) [30].

  1. Pathways Involved in the Repair of CPT-Induced DNA Lesions

Normal cells use DNA damage response (DDR) pathways to maintain genomic stability [54]. As aforementioned, SSB and DSB repair mechanisms are the two major DDR pathways that repair TOP1-induced DNA lesions. Paradoxically, cancer cells exploit DDR pathways to accumulate necessary genomic alterations for promoting proliferation. Furthermore, altered DDR and apoptotic responses in cancer cells are the major obstacles to successful chemotherapy. Thus, the delineation of TOP1-related SSB and DSB repair mechanisms is of great importance for identifying drug targets that can selectively affect cancer cell survival.

3.1. Single-Strand Break (SSB) Repair

Trapping of TOP1cc results in a 3′-P-TOP1 end and a 5′-OH terminus. Because the two ends cannot be directly religated, the persisting SSB is likely to be detected by PARP1 in which activated PARP1 catalyzes the synthesis of poly(ADP-ribose) (PAR) chains for recruiting repair proteins [55]. This reaction can be rapidly reversed by PARG, which hydrolyzes the PAR chains. The PAR chains at the SSB sites are important for the recruitment of XRCC1 that functions as a loading dock for other SSB repair proteins including TDP1 and PNKP. TDP1 generates 3′-P and PNKP converts 3′-P to 3′-OH, and PNKP also converts 5′-OH to 5′-P, making ends compatible for religation with no base loss. The rejoining of the 3′-OH and 5′-P ends is mainly mediated by LIG3, in which XRCC1 mediates the recruitment of LIG3.

If the trapped TOP1cc intermediates are processed by endonucleases, the initial SSBs will be converted to 3′-OH and 5′-OH ends with a gap over a few nucleotides (in the case of XPF-ERCC1, the loss is in the range of 3–4 nt), leading to the activation of PARP1 and XRCC1 recruitment. Consequentially, Pol3 recruited by XRCC1 can catalyze the gap filling, and PCNA-Polö/E also plays a role in this process [55]. If the 5′-OH is not processed by PNKP, the 5′-flap resulted from gap filling is likely to be removed by FEN1, which explains why FEN1 deficiency also leads to an increased CPT sensitivity. The final ligation is catalyzed by LIG1 because of the presence of PCNA.

Biomolecules 2015, 5                                                                                                                           1657

3.2. Double-Strand Break (DSB) Repair

Successful DSB repair requires concerted actions of proteins involved in DNA damage signaling and repair [54]. To repair TOP1 poison-induced DNA lesions, ATR signaling is required due to the runoff of replication fork and the presence of long single-strand DNA (ssDNA) [56]. The full activation of ATR follows a “two-man” rule—the ssDNA-ATRIP-dependent recruitment of ATR kinase and the RAD17 clamp loader/9-1-1/TOPBP1 mediator loading at the ssDNA-dsDNA junction. ATR phosphorylates CHEK1 to harness cell cycle arrest. If one-ended DSB is formed, ATM will be activated through the action of the MRE11-RAD50-NBS1 (MRN) complex. ATM mainly phosphorylates CHEK2 to mediate cell cycle arrest. Both ATM and ATR are able to phosphorylate hundreds of proteins in response to DSB formation [57]. One remarkable substrate is the histone H2AX, which can be phosphorylated by both kinases to yield g-H2AX. It is conceived that the propagation of g-H2AX signaling along the chromatin facilitates MDC1 recruitment and BRCA1 signaling via the MDC1-RNF8-RNF168-RAP80 ubiquitin cascade—events that are essential for HR [58].

The repair of TOP1 poison-induced DNA lesions is in essence the repair of one-ended DSBs, which facilitates the restoration of replication forks to restart DNA replication. It is important to note that one-ended DSB repair occurs in the S phase and relies on HR rather than NHEJ [59]. The first step in HR is end resection to generate a 3′-overhang for homology searching. A TOP1 cleavage in the leading strand may require end resection by the MRN-CtIP-BRCA1 and BLM-EXO1-DNA2 complexes [60], whereas a cleavage in the lagging strand automatically forms a 3′-overhang. Rad51 then associates with the 3′-ssDNA to form a nucleofilament for strand invasion, which leads to the formation of a D-loop structure [61]. This process continues with DNA synthesis, branch migration and the resolution of Holliday junction structures to reconstitute a functional replication fork [62]. TOP1 poisons can also lead to the formation of two-ended DSB if two replication forks collide into each other at the site of SSB. The repair of this type of DSBs is not aimed for fork restoration and can be accomplished by the classical DSB repair mechanisms [61].

3.3. Genes Involved in CPT-Induced Damage Repair

A long list of genes, in which mutations confer sensitivity to CPT in yeast, chicken or mammalian cells, has been compiled [24,30,63]. With no surprise, many genes involved in SSB and DSB repair are on the list, such as PARP1, XRCC1, PNKP, TDP1 for SSB repair; MRN, ATM-CHK2, ATR-CHK1 for DSB signaling; BRCA1/2, XRCC2, XRCC3 for HR. Most recently, the hMSH5-FANCJ complex has also been implicated to play a role in CPT-induced DNA damage response and repair [64]. Mutations in the binding partners of these repair factors are also likely to sensitize cells to CPT treatment. For example, depletion of the MRN-binding partner hnRNPUL increases the sensitivity to CPT [65]; and deficiencies in ZRANB3 and SPIDR, binding partners of PCNA and RAD51, cause CPT hypersensitivity in cancer cells [66–68]. In addition, the two DNA helicases BLM and WRN have also been implicated in the repair of CPT-induced DNA lesions [69,70]. Early studies revealed that chicken BLM knockout cells and human BLM-deficient fibroblasts showed increased sensitivity to CPT [71,72]. On the contrary, mouse BLM knockout embryonic stem cells showed mild resistance to

Biomolecules 2015, 5                                                                                                                           1658

CPT [73]. This discrepancy is likely attributable to the complexity of CPT-induced DNA lesion repair as well as different treatment conditions and experimental systems.

Interstrand crosslinks (ICLs) resemble CPT-induced lesions in that they block both replication and transcription [74]. They may induce replication fork reversal and fork collapse, which require DNA incision for lesion processing and HR for repair. ICL repair is accomplished by the coordinated actions of 17 Fanconi anemia (FA) genes whose mutations contribute to FA in patients [75]. Depletion of FANCP/SLX4 or FANCQ/XPF causes cellular sensitivity to CPT because they form an endonuclease complex involved in the repair of trapped TOP1cc [38]. Likewise, depletion of FANCS/BRCA1, FANCD1/BRCA2, FANCN/PALB2 or FANCO/RAD51C sensitizes cells to CPT because of their involvement in HR [76]. Accordingly, depletion of the FA core complex except FANCM—involved in fork reversal—is not expected to increase CPT sensitivity because they are unable to recognize the trapped TOP1cc [76]. However, the roles of FANCI, D2, J and FAN1 in the process are elusive due to conflicting reports presumably reflecting different experimental systems [76–78]. For example, in a multicolor competition assay, loss of FANCI or FAN1 rendered cells sensitive to CPT treatment [77]. However, this observation could not be recapitulated in studies performed with FANCI-deficient lymphoblasts and FAN1-depleted HEK293 cells [76,79], indicating that the involvement of these two genes in CTP sensitivity might be cell type specific.

It is interesting to note that the MMS22L-TONSL complex plays a prominent role in mediating CPT sensitivity [80–83]. Depletion of this complex impairs RAD51 foci formation and triggers G2/M arrest, indicating that the MMS22L-TONSL complex participates in HR repair. Furthermore, this complex associates with MCM, FACT, ASF1 and histones. FACT and ASF1 are histone chaperones that function in H2A/H2B and H3/H4 chromatin assembly and disassembly, respectively [84]. They recycle parental histones from old DNA strands unwound by MCM and incorporate them into newly synthesized DNA strands. FACT and ASF1 also function in checkpoint signaling; therefore the involvement of MMS22L-TONSL in CPT response implies the existence of a close association between HR, DNA damage signaling and replication restart.

  1. TOP1 Inhibition in Cancer Treatment

The understanding of the function of TOP1 and the cellular effects of TOP1 inhibition has been a stepping-stone for the development of effective CPT derivatives in cancer therapy. Since TOP1 functions in normal and cancer cells, the use of low doses of TOP1 inhibitors are actively sought to treat cancers that heavily rely on the function of TOP1 for survival (e.g., highly malignant, rapid-dividing tumor cells). In fact, the FDA-approved CPT derivatives topotecan and irinotecan are currently used to treat ovarian and colorectal cancers, respectively [24].

Furthermore, the promising results from a Phase I trial have warranted further evaluation of the CPT derivative Diflomotecan in Phase II trials [85]. Other derivatives like Gimatecan, Lurtotecan and Exatecan are also being tested in clinical trials (Table 1). The non-CPT indolocarbazole BMS-250749 showed great anti-tumor activity against preclinical xenograft models [86], but no further evaluation beyond Phase I trials is presently available (Table 2). Another indolocarbazole compound Edotecarin has shown promising anti-tumor activity in xenograft models and it is now advanced to Phase II studies of patients with advanced solid tumors [87]. By contrast, Phenanthroline ARC-111 (topovale)

Biomolecules 2015, 5                                                                                                                             1659

was potently against human tumor xenografts and displayed anti-cancer activity in colon and Wilms’ tumors [88]; however, no result from Phase I clinical trials is available owing to profound bone marrow toxicity [89]. To date, indenoisoquinolines are the most promising non-CPT inhibitors in clinical trials. LMP400 (NSC 743400, indotecan) and LMP776 (NSC 725776, indimitecan) show significant anti-tumor activities in animal models and both are being evaluated in Phase I clinical trials for relapsed solid tumors and lymphomas [8,90].

Table 1. CPT derivatives in clinical trials [91].

Name                            Structure                     Clinical Trial            Malignancy        Reference

Biomolecules 2015, 5                                                                                                                           1660

Given the observation that CPT-mediated TOP1 inhibition provokes DNA repair activities, a synergistic effect is then anticipated on cancer cells by inhibition of TOP1 and downregulation of DNA repair activities. The rationale for this approach is to accelerate the accumulation of DNA breaks and trigger cellular apoptosis, probably through mitotic catastrophe [92]. Which DNA repair pathways can we exploit? Currently, the major interests are in SSB and DSB repair mechanisms. Indeed, PARP inhibitors can enhance the cytotoxicity of TOP1 inhibitors in cancer cell lines as well as in mouse models [93–96]. Phase I studies of combination therapy using PARP inhibitors veliparib or olaparib (FDA-approved) together with topotecan were carried out in patients with advanced solid tumors but showed some dose-dependent side effects [97,98]. TDP1 can be another potential target because it functions directly downstream of PARP1 in the repair of TOP1 poison-induced DNA lesions [99]. TDP1 inhibitors sensitize cells to CPT treatment in vitro [100,101], however in vivo evaluation is presently unavailable due to unsuitable properties of the compounds [102].

Table 2. Non-CPT derivatives in preclinical and clinical trials [91].

Name                       Structure               Clinical Trial            Malignancy             Reference

Phase II

(Edotecarin, Pfizer)

Stomach, breast
Anti-tumor activity
in preclinical
xenograft models
Anti-tumor activity

Preclinical                    in preclinical            [88,89,103]
xenograft models

(LMP400, LMP776)
Phase I                              Lymphomas             [8,90,103]

DSB repair can be targeted by either inhibition of DSB signaling or inhibition of HR. ATM and ATR inhibitors can largely increase the sensitivity to CPT in cancer cells [104,105]. This can be explained by the fact that abrogation of the cell cycle arrest will allow cells with unreplicated or unrepaired chromosomes to enter mitosis thereby triggering mitotic catastrophe and cell death. Similarly, CHEK1 and CHEK2 inhibitors are tested in Phase I studies in combination with irinotecan [106,107]. Inhibitors that can directly block HR proteins are very limited [108]. This is partially attributed to the fact that HR genes are often mutated in cancer cells, thus diminishing the enthusiasm for developing HR inhibitors. One diterpenoid compound, however, was found to be able to inhibit the function of BRCA1 and render cytotoxicity in human prostate cancer cells [109]. Several RAD51 inhibitors have also been

Biomolecules 2015, 5                                                                                                                           1661

identified but have not been tested in cell lines [110]. Inhibition of BRCA1 and RAD51 can be also achieved indirectly by harnessing corresponding kinases [106]. Clearly, defective hMRE11 sensitizes colon cancer cells to CPT treatment [111]. Although MRE11-deficeint tumor xenografts failed to display significant growth inhibition by irinotecan alone, combining thymidine with irinotecan caused a dramatic growth delay [112].

TOP1 inhibitors might be also useful for treating cancers with BRCA1/2 mutations. The successful use of PARP inhibitors in treating BRCA1/2-deficient tumors has ignited a broad interest in searching for synthetic lethality among DNA damage response and repair genes [113,114]. In the PARP-BRCA1/2 example, the accumulation of SSBs by PARP inhibition would lead to the formation of DSBs during replication. In HR-deficient cells, DSBs can only be repaired by illegitimate (toxic) NHEJ—joining one-ended DSBs from different locations—leading to cell death [115,116]. However, resistance to PARP inhibitors can arise in BRCA1-deficient tumors during treatment from either genetic reversion of BRCA1 mutations or the loss of NHEJ [117–122]. Therefore, it would be beneficial to explore the possibility of developing a similar synthetic lethal strategy to use TOP1 inhibitors in the treatment of BRCA1/2-deficient tumors.

Figure 2. An overview of the effects of TOP1 inhibition is provided. Inhibitors and key DNA repair factors are highlighted.

Biomolecules 2015, 5                                                                                                                         1662

  1. Conclusions

Trapping of TOP1 by inhibitors generates SSBs and DSBs that are repaired by their corresponding repair pathways (Figure 2). Therefore, developing effective TOP1 inhibitors not only provides powerful tools to study DNA replication and repair but also establishes a foundation to devise new synthetic lethal strategies for efficient cancer treatments. The accumulation of DNA strand breaks (SSBs and DSBs) by TOP1 inhibition in HR-deficient tumor cells is expected to enhance cytotoxicity. However, increased DNA repair activities in cancer cells can make TOP1 inhibitors less effective, so silencing of repair pathways in conjunction with the use of TOP1 inhibitors offers an attractive new means for cancer control. Since each tumor is unique, it would be advantageous to identify the individualities of DNA repair pathways or biomarkers reflecting the changes of DNA repair activities in tumor cells [92,123]. This will make it possible to achieve better and predictable prognosis through tailored therapeutic regimens. Given that TOP1 is essential for transcription and DNA replication, future design of novel TOP1 inhibitors and combinational therapy strategies should aim to increase therapeutic efficacy of the inhibitors, thus reducing side effects.


The work in the Her laboratory is supported by the NIH grant GM084353.

Author Contributions

Yang Xu and Chengtao Her wrote and revised the article.

Conflicts of Interest

The authors declare that they have no conflicts of interest with the contents of this article.

Please see the following file for the referencesReferences for top paper

From a 2015 Clinical Cancer Research paper:

Phase 1 clinical pharmacology study of F14512, a new polyamine-vectorized anti-cancer drug, in naturally occurring canine lymphoma

Dominique Tierny1, Francois Serres1, Zacharie Segaoula1, Ingrid Bemelmans1, Emmanuel Bouchaert1,

Aurelie Petain2, Viviane Brel3, Stephane Couffin4, Thierry Marchal5, Laurent Nguyen6, Xavier Thuru7,

Pierre Ferre2, Nicolas Guilbaud8, and Bruno Gomes9,*


Purpose: F14512 is a new topoisomerase II inhibitor containing a spermine moiety that facilitates selective uptake by tumor cells and increases topoisomerase II poisoning. F14512 is currently in Phase I/II clinical trial in patients with acute myeloid leukemia. The aim of this study was to investigate F14512 potential in a new clinical indication. Because of the many similarities between human and dog lymphomas, we sought to determine the tolerance, efficacy, PK/PD relationship of F14512 in this indication, and potential biomarkers that could be translated into human trials. Experimental design: Twenty-three dogs with stage III-IV naturally occurring lymphomas were enrolled in the Phase 1 dose-escalation trial which consisted of three cycles of F14512 intravenous injections. Endpoints included safety and therapeutic efficacy. Serial blood samples and tumor biopsies were obtained for PK/PD and biomarker studies. Results: Five dose levels were evaluated in order to determine the recommended dose. F14512 was well tolerated, with the expected dose-dependent hematological toxicity. F14512 induced an early decrease of tumoral lymph node cells, and a high response rate of 91% (21/23) with 10 complete responses, 11 partial responses, 1 stable disease and 1 progressive disease. Phosphorylation of histone H2AX was studied as a potential pharmacodynamic biomarker of F14512. Conclusions: This trial demonstrated that F14512 can be safely administered to dogs with lymphoma resulting in strong therapeutic efficacy. Additional evaluation of F14512 is needed to compare its efficacy with standards of care in dogs, and to translate biomarker and efficacy findings into clinical trials in humans.

AND From ASCO 2015 Annual Meeting

Survival impact of switching to different topoisomerase I or II inhibitors-based regimens (topo-I or topo-II) in extensive-disease small cell lung cancer (ED-SCLC): supplemental analysis from JCOG0509.


Background: The J0509 (phase III study for chemotherapy-naive ED-SCLC) demonstrated amrubicin plus cisplatin (AP) was inferior to irinotecan plus cisplatin (IP). However, median overall survival (OS) of both AP and IP (15 and 17 mo) was more favorable than those of previous trials (9-12 mo), probably because switching to different topo-I or topo-II in the second-line therapy, especially the use of topo-II in IP arm, was frequent. This analysis aimed to investigate whether observed survival benefit of IP arm can be explained by the treatment switching, and how post-protocol chemotherapy affected the result of J0509. Methods: Two analysis sets from J0509 were used: all randomized 283 pts and 250 pts who received post-protocol chemotherapy. One pt without initiation date of second-line therapy was excluded. A rank-preserving structural failure time (RPSFT) model was used to estimate “causal survival benefit” that would have been observed if all pts had been followed with the same type of regimen as randomized throughout the follow-up period. Additionally, to assess the survival impact of second-line use of topo-II, OS after initiating second-line therapy (OS2) was analyzed by multivariate Cox models. Results: %treatment switching in IP arm and AP arm was 65.2% (92/141) and 43.7% (62/142). By RPSFT model, estimated OS excluding the effect of the treatment switching was 2.7-fold longer in IP (topo-I) arm than AP (topo-II) arm. This causal survival benefit was stronger than the original report of J0509 (nearly 1.4-fold extension by Cox model), indicating that re-challenging topo-I in IP arm appeared beneficial. The multivariate Cox analysis for OS2 (n = 250) revealed second-line use of topo-II was detrimental (hazard ratio, 1.5; 95%CI, 1.1-2.1). Among sensitive relapsed pts in IP arm, OS2 was favorable in the following order: irinotecan-based regimen > the other topo-I > topo-II. Conclusions: IP remains the standard therapy. Re-challenging topo-I, especially irinotecan-based topo-I, seemed beneficial for IP-sensitive pts. This result should be confirmed in further investigations with large sample size. Clinical trial information: 000000720.





Below is actively recruiting clinical trials evaluating topoisomerase inhibitors. Shown are only a few trials for a complete list from please see this link:

A service of the U.S. National Institutes of Health

897 studies found for:    topoisomerase inhibitor | Open Studies

Include only open studies Exclude studies with Unknown status

Status Study
Recruiting A Study of Standard Treatment +/- Enoxaparin in Small Cell Lung Cancer

Condition: Small Cell Lung Cancer
Interventions: Drug: cisplatinum or carboplatin and e.g.etoposide.;   Drug: cisplatinum or carboplatin and e.g.etoposide+enoxaparin
Recruiting A Phase I Study of Indenoisoquinolines LMP400 and LMP776 in Adults With Relapsed Solid Tumors and Lymphomas

Conditions: Neoplasms;   Lymphoma
Interventions: Drug: LMP 400;   Drug: LMP 776
Recruiting A Dose-Ranging Study Evaluating the Efficacy, Safety, and Tolerability of GSK2140944 in the Treatment of Uncomplicated Urogenital Gonorrhea Caused by Neisseria Gonorrhoeae

Condition: Gonorrhea
Intervention: Drug: GSK2140944
Recruiting Selinexor in Combination With Irinotecan in Adenocarcinoma of Stomach and Distal Esophagus

Conditions: Esophageal Cancer;   Gastric Cancer
Interventions: Drug: Selinexor;   Drug: Irinotecan
Recruiting Multimodal Molecular Targeted Therapy to Treat Relapsed or Refractory High-risk Neuroblastoma

Condition: Neuroblastoma Recurrent
Interventions: Drug: Dasatinib;   Drug: Rapamycin;   Drug: Irinotecan;   Drug: Temozolomide
Unknown  Study of the Farnesyl Transferase Inhibitor, R115777, in Combination With Topotecan (NYU 99-32)

Condition: Cancer
Interventions: Drug: R115777 (farnesyl transferase inhibitor);   Drug: Topotecan
Recruiting Pegylated Irinotecan NKTR 102 in Treating Patients With Relapsed Small Cell Lung Cancer

Condition: Recurrent Small Cell Lung Carcinoma
Interventions: Other: Laboratory Biomarker Analysis;   Drug: Pegylated Irinotecan;   Other: Pharmacological Study
Recruiting Selinexor and Chemotherapy in Treating Patients With Relapsed or Refractory Acute Myeloid Leukemia

Conditions: Adult Acute Myeloid Leukemia With 11q23 (MLL) Abnormalities;   Adult Acute Myeloid Leukemia With Del(5q);   Adult Acute Myeloid Leukemia With Inv(16)(p13;q22);   Adult Acute Myeloid Leukemia With t(15;17)(q22;q12);   Adult Acute Myeloid Leukemia With t(16;16)(p13;q22);   Adult Acute Myeloid Leukemia With t(8;21)(q22;q22);   Recurrent Adult Acute Myeloid Leukemia;   Secondary Acute Myeloid Leukemia
Interventions: Drug: mitoxantrone hydrochloride;   Drug: etoposide;   Drug: cytarabine;   Drug: selinexor;   Other: laboratory biomarker analysis;   Other: pharmacological study
Recruiting WEE1 Inhibitor MK-1775 and Irinotecan Hydrochloride in Treating Younger Patients With Relapsed or Refractory Solid Tumors

Conditions: Childhood Solid Neoplasm;   Recurrent Childhood Medulloblastoma;   Recurrent Childhood Supratentorial Primitive Neuroectodermal Tumor;   Recurrent Neuroblastoma
Interventions: Drug: Irinotecan Hydrochloride;   Other: Laboratory Biomarker Analysis;   Other: Pharmacological Study;   Drug: WEE1 Inhibitor AZD1775
Recruiting PARP Inhibitor BMN-673 and Temozolomide or Irinotecan Hydrochloride in Treating Patients With Locally Advanced or Metastatic Solid Tumors

Conditions: Metastatic Cancer;   Unspecified Adult Solid Tumor
Interventions: Drug: PARP inhibitor BMN-673;   Drug: temozolomide;   Drug: irinotecan hydrochloride;   Other: pharmacological study;   Other: laboratory biomarker analysis
Recruiting A Phase II Multicenter, Randomized, Placebo Controlled, Double Blinded Clinical Study of KD018 as a Modulator of Irinotecan Chemotherapy in Patients With Metastatic Colorectal Cancer

Condition: Colorectal Neoplasms
Interventions: Drug: KD018;   Drug: Irinotecan;   Drug: Placebo
Recruiting The Efficacy of the 7 Days Tailored Therapy as 2nd Rescue Therapy for Eradication of H. Pylori Infection

Condition: Helicobacter Infection
Interventions: Procedure: H. pylori culture and antimicrobial susceptibility testing;   Drug: 14 days empirical bismuth quadruple therapy (Proton pump inhibitor);   Drug: Metronidazole;   Drug: Tetracycline;   Drug: tripotassium dicitrate bismuthate;   Drug: 7 days tailored therapy Proton Pump Inhibitor;   Drug: Moxifloxacin;   Drug: Amoxicillin
Recruiting G1T28 (CDK 4/6 Inhibitor) in Combination With Etoposide and Carboplatin in Extensive Stage Small Cell Lung Cancer (SCLC)

Condition: Small Cell Lung Cancer
Interventions: Drug: G1T28 + carboplatin/ etoposide;   Drug: Placebo + carboplatin/ etoposide
Recruiting Trial of Topotecan With VX-970, an ATR Kinase Inhibitor, in Small Cell Lung Cancer

Conditions: Carcinoma, Non-Small -Cell Lung;   Ovarian Neoplasms;   Small Cell Lung Carcinoma;   Uterine Cervical Neoplasms;   Carcinoma, Neuroendocrine
Interventions: Drug: Topotecan;   Drug: VX-970
Recruiting Prospective Analysis of UGT1A1 Promoter Polymorphism for Irinotecan Dose Escalation in Metastatic Colorectal Cancer Patients Treated With Bevacizumab Combined With FOLFIRI as the First-line Setting

Condition: Metastatic Colorectal Cancer
Interventions: Genetic: UGT1A1 genotyping (6,6);   Genetic: UGTIA1 genotyping (6,7);   Genetic: UGTIA1 genotyping (7,7);   Genetic: UGT1A1 non-genotyping;   Drug: bevacizumab (Avastin);   Drug: irinotecan;   Drug: Leucovorin;   Drug: 5-FU
Recruiting A Study of the Bruton’s Tyrosine Kinase Inhibitor, PCI-32765 (Ibrutinib), in Combination With Rituximab, Cyclophosphamide, Doxorubicin, Vincristine, and Prednisone in Patients With Newly Diagnosed Non-Germinal Center B-Cell Subtype of Diffuse Large B-Cell Lymphoma

Condition: Lymphoma
Interventions: Drug: Ibrutinib;   Drug: Placebo;   Drug: Rituximab;   Drug: Cyclophosphamide;   Drug: Doxorubicin;   Drug: Vincristine;   Drug: Prednisone (or equivalent)
Recruiting Irinotecan Combination Chemotherapy for Refractory or Relapsed Brain Tumor in Children and Adolescents

Condition: Brain Tumor
Intervention: Drug: Irinotecan combination chemotherapy
Recruiting A Study To Evaluate PF-04449913 With Chemotherapy In Patients With Acute Myeloid Leukemia or Myelodysplastic Syndrome

Condition: Acute Myeloid Leukemia
Interventions: Drug: PF-04449913;   Drug: Low dose ARA-C (LDAC);   Drug: Decitabine;   Drug: Daunorubicin;   Drug: Cytarabine
Recruiting Veliparib and Pegylated Liposomal Doxorubicin Hydrochloride in Treating Patients With Recurrent Ovarian Cancer, Fallopian Tube Cancer, or Primary Peritoneal Cancer or Metastatic Breast Cancer

Conditions: Estrogen Receptor Negative;   HER2/Neu Negative;   Male Breast Carcinoma;   Progesterone Receptor Negative;   Recurrent Breast Carcinoma;   Recurrent Fallopian Tube Carcinoma;   Recurrent Ovarian Carcinoma;   Recurrent Primary Peritoneal Carcinoma;   Stage IV Breast Cancer;   Triple-Negative Breast Carcinoma
Interventions: Other: Laboratory Biomarker Analysis;   Drug: Pegylated Liposomal Doxorubicin Hydrochloride;   Other: Pharmacological Study;   Drug: Veliparib
Recruiting A Study To Evaluate Ara-C and Idarubicin in Combination With the Selective Inhibitor Of Nuclear Export (SINE) Selinexor (KPT-330) in Patients With Relapsed Or Refractory AML

Condition: Acute Myeloid Leukemia (Relapsed/Refractory)
Interventions: Drug: Selinexor;   Drug: Idarubcin;   Drug: Cytarabine

Read Full Post »

Observations on Human Papilloma Virus and Cancer

Curator: Demet Sag, PhD, CRA, GCP 


What is Human Papilloma Virus?

 HPV 220px-HPV-16_genome_organization

Human papillomavirus

Taxonomy ID: 10566
Inherited blast name: viruses
Rank: species
Genetic code: Translation table 1 (Standard)
Host: vertebrates| human
Other names:

synonym: human papillomavirus HPV
synonym: Human Papilloma Virus

Lineage( full )

VirusesdsDNA viruses, no RNA stagePapillomaviridaeunclassified PapillomaviridaeHuman papillomavirus types

   Entrez records   
Database name Subtree links Direct links
Nucleotide 7,782 7,775
Protein 2,611 2,604
Structure 3 3
Genome 1 1
Popset 34 34
PubMed Central 4,742 4,742
Gene 21 21
SRA Experiments 43 43
Probe 12 12
Assembly 1 1
Bio Project 6 6
Bio Sample 53 53
PubChem BioAssay 5 5
Taxonomy 8 1
Human papillomavirus
Specialty Infectious diseasegynecologyHPV_

WHO_RHR_08.14_eng-Cervical cancer, human papillomavirus (HPV), and HPV vaccinesWHO= papilloma virus info

ICD10 B97.7
ICD9-CM 078.1 079.4
DiseasesDB 6032
eMedicine med/1037
MeSH D030361

ICTV homepage

WHO= papilloma virus info

WHO_RHR_08.14_eng-Cervical cancer, human papillomavirus (HPV), and HPV vaccines

Why is it related to Human Cancer?

 Since its first presumed diagnosis in women by an Italian Physician back in 1800s many developments took place to identify the real causative agents (PMID:19135222). Especially in 1970s the full discovery and relation between HPV and cancer established. Human papilloma virus (HPV)  is the second common cancer death in women, although HPV vaccines helped to decrease the morbidity rate there are complications due to vaccines.  Still there is an increase with cervical cancer estimated to be  490,000.

CDC also provided simple information for public on HPV since there is a misunderstanding that some people think it is like herpes or HIV viruses.  Yet, pathology is much different and changes based on age since younger women or girls can fight off but after age 30 predisposition of HPV as a cancer increases. (

Cervical cancer is responsible for 10–15% of cancer-related deaths in women worldwide1,2. The etiological role of infection with high-risk human papilloma viruses (HPV) in cervical carcinomas is well established.


  Relationship of mutational spectrum and rates with clinicopathological characteristics in cervical carcinoma presented 



Relationship of mutational spectrum and rates with clinicopathological characteristics in cervical carcinoma presented at

All panels are aligned with vertical tracks representing 115 individuals. The data is sorted in order by histology (middle panel) and total mutational rate (top panel). The relative frequencies of nucleotide mutations occurring at cytosines preceeded by thymines (Tp*C) or at cytosines followed by guanines (*CpG) sites are depicted in red and orange respectively, on the second panel. The bottom heatmap shows the distribution of mutations in significantly mutated genes (q<0.1) in squamous cell carcinomas and adenocarcinomas in the order listed in the following Table, TP53ERBB2 and KRAS were significant recurrence (q<0.1) among cancer driver genes reported in COSMIC.

Nature. Author manuscript; available in PMC 2014 Sep 12.  Published in final edited form as: Nature. 2014 Feb 20; 506(7488): 371–375.

Genes with Significantly Recurrent Somatic Mutations in Cervical Carcinomas

Gene Description Nonsilent mutations Relative frequency Patients Unique sites Silent mutations Indel + null q
FBXW7** F-box and WD repeat domain containing 7 12 15% 12 8 0 2 4.03E-12
PIK3CA phosphoinositide-3-kinase, catalytic, alpha polypeptide 11 14% 10 5 0 1 <9.08e-12
MAPK1** mitogen-activated protein kinase 1 6 8% 6 3 0 0 0.000671
HLA-B+ major histocompatibility complex, class I, B 7 9% 6 7 1 3 0.00169
STK11 serine/threonine kinase 11 3 4% 2 2 0 1 0.012
EP300+ E1A binding protein p300 13 16% 12 13 1 4 0.0354
NFE2L2+ nuclear factor (erythroid-derived 2)-like 2 3 4% 3 2 0 0 0.0597
PTEN phosphatase and tensin homolog (mutated in multiple advanced cancers 1) 5 6% 5 5 0 3 0.0693
ELF3* E74-like factor 3 (ets domain transcription factor, epithelial-specific) 3 13% 3 3 0 3 0.03
CBFB* core-binding factor, beta subunit 2 8% 2 2 0 1 0.0342

Indel: insertions or deletions;

Null: nonsense, frameshft or splice-site mutations;

q: q value, false discovery rate (Benjamini-Hochberg procedure).

**Genes with mutations observed in only squamous cell carcinomas

*Genes with mutations observed in only adenocarcinomas

+Genes with a majority of mutations occurring in squamous cell carcinomas.

Following figure (

  Novel recurrent somatic mutations in cervical carcinoma

Novel recurrent somatic mutations in cervical carcinoma

The locations of somatic mutations in novel significantly mutated genes in 115 cervical carcinoma, FBXW7, MAPK1HLA-BEP300NFE2L2 and ELF3 are shown in the context of protein domain models derived from UniProt and Pfam annotations. Numbers refer to amino acid residues. Each filled circle represents an individual mutated tumor sample: missense and silent mutations are represented by filled black and grey circles, respectively while nonsense, frameshift, and splice site mutations are represented by filled red circles and red text. Domains are depicted with various colors with an appropriate key located on the right hand of each domain model.

 Relationships between HPV integration, copy number amplifications and gene expression in cervical carcinoma

Relationships between HPV integration, copy number amplifications and gene expression in cervical carcinoma

Panel (a) shows comparative histograms of true and simulated genomic distances between HPV integration sites and the nearest copy number amplification (log segmean difference >0.5). Panel (b) shows boxplots of gene expression levels across 79 cervical tumors for 41 genes with chimeric human-HPV read pairs. The expression levels for tumors with HPV integration in the respective genes are highlighted in red circles. Panel (c) shows scatter plots comparing copy number alterations and gene expression levels across 79 tumors in selected integration site genes. The red circles represent data for the tumors with HPV integration events involving the respective genes.


Table. Diseases Associated With Specific HPV Types (e-Medicine)

Nongenital Cutaneous Disease HPV Type
Common warts (verrucae vulgaris) 1, 2, 4, 26, 27, 29, 41, 57, 65, 75-78
Plantar warts (myrmecias) 1, 2, 4, 60, 63
Flat warts (verrucae planae) 3, 10, 27, 28, 38, 41, 49
Butcher’s warts (common warts of people who handle meat, poultry, and fish) 1-4, 7, 10, 28
Mosaic warts 2, 27, 57
Ungual squamous cell carcinoma 16
Epidermodysplasia verruciformis (benign) 2, 3, 10, 12, 15, 19, 36, 46, 47, 50
Epidermodysplasia verruciformis (malignant or benign) 5, 8-10, 14, 17, 20-25, 37, 38
Nonwarty skin lesions 37, 38
Nongenital Mucosal Disease HPV Type
Respiratory papillomatosis 6, 11
Squamous cell carcinoma of the lung 6, 11, 16, 18
Laryngeal papilloma (recurrent respiratory papillomatosis)[17] 2, 6, 11, 16, 30, 40, 57
Laryngeal carcinoma 6, 11
Maxillary sinus papilloma 57
Squamous cell carcinoma of the sinuses 16, 18
Conjunctival papillomas 6, 11
Conjunctival carcinoma 16
Oral focal epithelial hyperplasia (Heck disease) 13, 32
Oral carcinoma 16, 18
Oral leukoplakia 16, 18
Squamous cell carcinoma of the esophagus 16, 18
Anogenital Disease HPV Type
Condylomata acuminata 1-6, 10, 11, 16, 18, 30, 31, 33, 35, 39-45, 51-59, 70, 83
Bowenoid papulosis 16, 18, 34, 39, 40, 42, 45
Bowen disease 16, 18, 31, 34
Giant condylomata (Buschke-Löwenstein tumors) 6, 11, 57, 72, 73
Unspecified intraepithelial neoplasia 30, 34, 39, 40, 53, 57, 59, 61, 62, 64, 66-69
Low-grade squamous intraepithelial lesions (LGSIL) 6, 11, 16, 18, 26, 27, 30, 31, 33-35, 40, 42-45, 51-58, 61, 62, 67-69, 71-74, 79, 81-84
High-grade squamous intraepithelial lesions (HGSIL) 6, 11, 16, 18, 31, 33, 35, 39, 42, 44, 45, 51, 52, 56, 58, 59, 61, 64, 66, 68, 82
Carcinoma of vulva 6, 11, 16, 18
Carcinoma of vagina 16
Carcinoma of cervix[18, 19] 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68, 70, 73, 82
Carcinoma of anus 16, 31, 32, 33
Carcinoma in situ of penis (erythroplasia of Queyrat) 16
Carcinoma of penis 16, 18


 epidemiology of HPV in the world

“Human papillomavirus (HPV) has become synonymous with cervical cancer, but its actual footprint is much bigger” said James Mitchell Crow. (PMID: 229324377  James Mitchell Crow. “HPV: The global burden”. Nature 488 S2–S3 (30 August 2012) doi:10.1038/488S2a Published online  29 August 2012).

Every year, over 27,000 women and men are affected by a cancer caused by HPV— that’s a new case every 20 minutes.

Persistent HPV infection can cause cervical and other cancers including:


Virus Diseases [C02]
   DNA Virus Infections [C02.256]

Papillomavirus Infections [C02.256.650]

Warts [C02.256.650.810]  +
Virus Diseases [C02]
   Tumor Virus Infections [C02.928]

Papillomavirus Infections [C02.928.725]



(PMID: 229324377)





In the lab few places propagating HPV. There are measures that need to be taken by the laboratory personnel. CDC as well as WHO published various articles to inform public.

Sensitivity and testing for Pap smear and HPV DNA testing in the detection of CIN2+

Test Sensitivity Specificity
Pap smear 53-55.4% 96.3-96.8%
High-risk HPV DNA testing 94.6-96.1% 90.7-94.1%
Pap smear + high-risk HPV testing 100% 92.5%

Cuzick J, Clavel C, Petry KU, Meijer CJ, Hoyer H, Ratnam S, Szarewski A, Birembaut P, Kulasingam S, Sasieni P, Iftner T. Overview of the European and North American studies on HPV testing in primary cervical cancer screening. Int J Cancer. 2006; 119(5):1095.

Mayrand MH, Duarte-Franco E, Rodrigues I, Walter SD, Hanley J, Ferenczy A, Ratnam S, Coutlée F, Franco EL, Canadian Cervical Cancer Screening Trial Study Group.

Human papillomavirus DNA versus Papanicolaou screening tests for cervical cancer N Engl J Med. 2007;357(16):1579.

Best Pract Res Clin Obstet Gynaecol. Author manuscript; available in PMC 2013 Apr 22. (

HPV Genotyping tests1

HPV genotyping test HPV types detected
Cervista® HPV 16/18 (Hologic, Inc;
Marlborough, MA)a
HR HPV types 16 and 18
Digene HPV Genotyping PS Test (Qiagen;
Hilden, Germany)
HR HPV types 16, 18, and 45
Roche LINEAR ARRAY HPV Genotyping
Test (Roche; Basel, Switzerland)
37 LR and HR HPV types
Innogenetics INNO-LiPA HPV Genotyping
Extra (Innogenetics; Gent, Belgium)
28 LR and HR HPV types
SPF10 Line Probe Assay HPV-typing System
(Roche; Basel,
Recognizes most genital
tract HPV types
Papillocheck1 (Greiner Bio-One;
Frickenhausen Germany)
18 HR and 6 LR HPV types
RealTime High Risk HPV Assay (Abbott
Laboratories;Abbott Park, IL)
HPV types 16 and 18
HPV Genotyping LQ Test (Qiagen Inc;
Valencia, CA)
18 HR HPV types
Seeplex HPV4A ACE (Seegene; Rockville,
HPV types 16 and 18
CLART HPV 2 (Genomica; Madrid, Spain) 35 LR and HR HPV types
GenoFlow HPV Array (DiagCor; North Point,
Hong Kong)
33 LR and HR HPV types
fHPV Typing (molGENTIX; Barcelona, Spain) 15 LR and HR HPV types

HPV, human papillomavirus; HR, high-risk; LR, low-risk.

aFDA-approved test.

1Schutzbank TE, Ginocchio CC. Assessment of clinical and analytical performance characteristics of an HPV genotyping test. Diagn Cytopathol. 2011 Apr 6. doi:10.1002/dc.21661.

Most papillomas are sufficiently distinct to be clinically recognizable. Bowenoid papulosis may be mistaken for lichen planus, psoriasis, seborrheic keratoses, or condylomata acuminata.

In additions to the conditions listed in the differential diagnosis, other problems to be considered include the following:

  • Acanthosis nigricans
  • Acrochordon
  • Actinic keratoses
  • Anogenital malignancy
  • Anogenital warts in children
  • Bowenoid papulosis
  • Carbon dioxide laser surgery for intraepithelial cervical neoplasms
  • Cervical polyp
  • Condyloma latum
  • Corns and calluses
  • Dermatitis papillaris
  • Endoscopic gynecologic surgery
  • Epidermodysplasia verruciformis
  • Fordyce spots
  • Hymenal remnants
  • Hypopigmentation
  • Keloid and hypertrophic scar
  • Keratoacanthoma
  • Laryngeal papillomatosis of neonates and infants
  • Malignant tumors of the mobile tongue
  • Micropapillomatosis labialis
  • Nevi
  • Pap test
  • Pityriasis versicolor
  • Psoriasis (plaque)
  • Recurrent respiratory papillomatosis
  • Seborrheic keratosis
  • Sinonasal papillomas, treatment
  • Skin tags (fibroepithelial polyps)
  • Verrucous carcinoma
  • Vestibular papillomatosis

Differential Diagnoses




1.       Immunomodulators

Class Summary

Immune response modifiers have immunomodulatory effects and are used for treatment of external anogenital warts (EGWs) or condylomata acuminata. Interferon alfa, beta, and gamma may be administered topically, systemically, and intralesionally. They stimulate production of cytokines and demonstrate strong antiviral activity.

View full drug information

Imiquimod (Aldara, Zyclara)

Imiquimod is an imidazoquinolinamine derivative that has no in vitro antiviral activity but does induce macrophages to secrete cytokines such as interleukin (IL)-2 and interferon alfa and gamma. Its mechanisms of action are unknown. Imiquimod has been studied extensively and is a new therapy relative to other EGW treatments. It may be more effective in women than in men.

Imiquimod is dispensed as an individual dose. Patients are advised to wash the affected area with mild soap and water upon awakening and to remove residual drug.

View full drug information

Interferon alfa-n3 (Alferon N)

Interferon alfa is a protein product either manufactured from a single-species recombinant DNA process or obtained from pooled units of donated human leukocytes that have been induced by incomplete infection with a murine virus.

The mechanisms by which interferon alfa exerts antiviral activity are not understood clearly. However, modulation of the host immune response may play an important role. This agent is indicated for intralesional treatment of refractory or recurring external condyloma acuminatum and is particularly useful for patients who have not responded satisfactorily to other treatment modalities (eg, podophyllin, surgical excision, laser therapy, or cryotherapy).

View full drug information

Interferon alfa-2b (Intron A)

This is a protein product manufactured by recombinant DNA technology. Its mechanism of antitumor activity is not clearly understood; however, direct antiproliferative effects against malignant cells and modulation of host immune response may play important roles. Its immunomodulatory effects include suppression of tumor cell proliferation, enhancement of macrophage phagocytic activity, and augmentation of lymphocyte cytotoxicity.

This agent is indicated for intralesional treatment of refractory or recurring external condyloma acuminatum and is particularly useful for patients who have not responded satisfactorily to other treatment modalities (eg, podophyllin, surgical excision, laser therapy, or cryotherapy).

2.       Keratolytic Agents

Class Summary

Antimitotic drugs arrest dividing cells in mitosis, resulting in the death of proliferating cells. They cause cornified epithelium to swell, soften, macerate, and then desquamate. Many of them are chemotherapeutic agents. The drugs listed below are used specifically for treatment of EGWs or condylomata acuminata.

Keratolytic agents are used to aid in removal of keratin in hyperkeratotic skin disorders, including corns, ichthyoses, common warts, flat warts, and other benign verrucae.

View full drug information

Podofilox (Condylox)

Podofilox is a topical antimitotic that can be synthesized chemically or purified from the plant families Coniferae and Berberidaceae (eg, species of Juniperus and Podophyllum). It is the active agent of podophyllin resin and is available as a 0.5% solution. Treatment results in necrosis of visible wart tissue; the exact mechanism of action is unknown. Treatment should be limited to no more than 10 cm2 of wart tissue, and no more than 0.5 mL/day of solution should be given. This is a patient-applied therapy.

View full drug information

Podophyllum resin (Podocon-25)

Podophyllin is derived from May apple (Podophyllum peltatum Linné) and contains the active agent podophyllotoxin, a cytotoxic substance that arrests mitosis in metaphase. American podophyllum contains one fourth the amount of podophyllotoxin that Indian podophyllum does. The potency of podophyllin varies considerably between batches. The exact mechanism of action is unknown.

Podophyllin is used as a topical treatment for benign growths, including external genital and perianal warts, papillomas, and fibroids. It results in necrosis when applied to anogenital warts. Only a trained medical professional can apply it, and it cannot be dispensed to a patient.

View full drug information

Trichloroacetic acid 85% (Tri-Chlor)

Trichloroacetic acid (TCA) is a highly corrosive desiccating agent that cauterizes skin, keratin, and other tissues and is used to burn lesions. Although it is caustic, it causes less local irritation and systemic toxicity than other agents in the same class. However, response often is incomplete, and recurrence is common.

Most clinicians use 25-50% TCA, although some use concentrations as high as 85% and then neutralize with either water or bicarbonate. Tissue sloughs and subsequently heals in 7-10 days. TCA therapy is less destructive than laser surgery, electrocautery, or cryotherapy.

View full drug information

Salicylic acid (Compound W, Dr. Scholl’s Clear Away Warts, Freezone)

By dissolving the intercellular cement substance, salicylic acid produces desquamation of the horny layer of skin without affecting the structure of viable epidermis. It is used for removal of nongenital cutaneous warts, particularly common or plantar warts. Before application, wash the affected area. The wart may be soaked in warm water for 5 minutes. Dry the area thoroughly.

3.       Antineoplastics, Antimetabolite

Class Summary

Antimetabolites interfere with nucleic acid synthesis and inhibit cell growth and proliferation. These are topical preparations that contain the fluorinated pyrimidine 5-fluorouracil (5-FU). Although these chemotherapeutic agents are not formally approved for use against warts, some studies have demonstrated a benefit against EGWs or condylomata acuminata.

View full drug information

Fluorouracil topical (Efudex, Carac, Fluoroplex)

Topical 5-FU interferes with DNA synthesis by blocking the methylation of deoxyuridylic acid and inhibits thymidylate synthetase, which subsequently reduces cell proliferation. Its primary indication is for topical treatment of actinic keratoses. Although it is not approved by the US Food and Drug Administration (FDA) for the treatment of warts, it has been used in adults, particularly for warts resistant to other forms of treatment. It is used for management of superficial basal cell carcinomas.

The solution contains either 2% or 5% 5-FU in propylene glycol, tris (hydroxymethyl) aminomethane, hydroxypropyl cellulose, paraben, and disodium edetate. The cream contains 5% 5-FU in white petrolatum, stearyl alcohol, propylene glycol, polysorbate 60, and paraben. When topical 5-FU is applied to the lesion, the area undergoes a sequence of erythema, vesiculation, desquamation, erosion, and reepithelialization.

4.       Topical Skin Products

Class Summary

Sinecatechins is another topical product that has gained FDA approval for genital warts.

View full drug information

Sinecatechins (Veregen)

Sinecatechins ointment is a botanical drug product for topical use that consists of extract from green tea leaves. It contains 15% sinecatechins and is available in 15- and 30-g tubes. Its mode of action is unknown, but it does elicit antioxidant activity in vitro. Sinecatechins ointment is indicated for topical treatment of external genital and perianal warts (condylomata acuminata) in immunocompetent patients.

5.       Vaccines, Inactivated, Viral

Class Summary

Three vaccines are available for the prevention of HPV-associated dysplasias and neoplasia, including cervical, vulvar, vaginal, and anal cancer; genital warts (condylomata acuminata); and precancerous genital lesions.

View full drug information

Human papillomavirus vaccine, nonavalent (Gardasil 9)

Recombinant vaccine that targets 9 HPV types (6, 11, 16, 18, 31, 33, 45, 52, and 58). It is indicated for females aged 9-26 years to prevent cervical, vulvar, vaginal, and anal cancer. It is also indicated to prevent genital warts and dysplastic lesions (eg, cervical, vulvar, vaginal, anal).

It is also indicated for boys aged 9-15 years for prevention of anal cancer, genital warts, and anal intraepithelial neoplasia. In addition to the approved indications, the CDC recommends vaccinating males aged 16 through 21 years not previously vaccinated. CDC recommendations also include men through age 26 years not previously vaccinated. Vaccination is also recommended by the CDC among men who have sex with men and among immunocompromised persons (including those with HIV infection) if not vaccinated previously through age 26 years.

View full drug information

Human papillomavirus vaccine, quadrivalent (Gardasil)

The quadrivalent HPV recombinant vaccine was the first vaccine indicated to prevent cervical cancer, genital warts (condylomata acuminata), and precancerous genital lesions (eg, cervical adenocarcinoma in situ; cervical intraepithelial neoplasia grades I-III; vulvar intraepithelial neoplasia grades II and III; and vaginal intraepithelial neoplasia grades II and III) due to HPV types 6, 11, 16, and 18. Its efficacy is mediated by humoral immune responses following immunization series.

The quadrivalent vaccine is FDA-approved for females aged 9-26 years and is under FDA priority review to evaluate efficacy in women aged 27-45 years. It is indicated for boys and men aged 11-26 years for prevention of condylomata acuminata caused by HPV types 6 and 11. It is also indicated in people aged 9-26 years for prevention of anal cancer and associated precancerous lesions.

View full drug information

Human papillomavirus vaccine, bivalent (Cervarix)

The bivalent HPV vaccine is a recombinant vaccine prepared from the L1 protein of HPV types 16 and 18. It is indicated in girls and women aged 10-25 years for the prevention of diseases caused by oncogenic HPV types 16 and 18 (eg, cervical cancer, cervical intraepithelial neoplasia grade II or higher, adenocarcinoma in situ, and cervical intraepithelial neoplasia grade I).


HPV Vaccines: Indications Approved and HPV Types by Specific Vaccines

Indicated to Prevent HPV 9-valent* HPV 4-valent HPV 2-valent
Girls and Women
Approved ages 9-26 y 9-26 y 9-25 y
Cervical cancer HPV types 16, 18, 31, 33, 45, 52, and 58 HPV types 16 and 18 HPV types 16 and 18
Vulvar cancer HPV types 16, 18, 31, 33, 45, 52, and 58 HPV types 16 and 18 Not approved
Vaginal cancer HPV types 16, 18, 31, 33, 45, 52, and 58 HPV types 16 and 18 Not approved
Anal cancer HPV types 16, 18, 31, 33, 45, 52, and 58 HPV types 16 and 18 Not approved
Genital warts (condyloma acuminata) HPV types 6 and 11 HPV types 6 and 11 Not approved
Cervical intraepithelial neoplasia (CIN) grade 2/3 and cervical adenocarcinoma in situ (AIS) HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58 HPV types 6, 11, 16, and 18 HPV types 16 and 18
Cervical intraepithelial neoplasia (CIN) grade 1 HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58 HPV types 6, 11, 16, and 18 HPV types 16 and 18
Vulvar intraepithelial neoplasia (VIN) grades 2 and 3 HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58 HPV types 6, 11, 16, and 18 Not approved
Vaginal intraepithelial neoplasia (VaIN) grades 2 and 3 HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58 HPV types 6, 11, 16, and 18 Not approved
Anal intraepithelial neoplasia (AIN) grades 1, 2, and 3 HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58 HPV types 6, 11, 16, and 18 Not approved
Boys and Men
Approved ages 9-15 y* 9-26 y Not approved
Anal cancer HPV types 16, 18, 31, 33, 45, 52, and 58 HPV types 16 and 18 Not approved
Genital warts (condyloma acuminata) HPV types 6 and 11 HPV types 6 and 11 Not approved
Anal intraepithelial neoplasia (AIN) grades 1, 2, and 3 HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58 HPV types 6, 11, 16, and 18 Not approved
*The CDC recommends vaccinating males 16-21 y not previously vaccinated, and through age 26 y among men who have sex with men and among immunocompromised persons (including those with HIV infection) if not vaccinated previously



Clinical Trials:


Two trials of clinically approved human papillomavirus (HPV) vaccines, Females United to Unilaterally Reduce Endo/Ectocervical Disease (FUTURE I/II) and the Papilloma Trial Against Cancer in Young Adults (PATRICIA), reported a 22% difference in vaccine efficacy (VE) against cervical intraepithelial neoplasia grade 2 or worse in HPV-naïve subcohorts; however, serological testing methods and the HPV DNA criteria used to define HPV-unexposed women differed between the studies.

The risk of newly detected human papillomavirus (HPV) infection and cervical abnormalities in relation to HPV type 16/18 antibody levels at enrollment in PATRICIA (Papilloma Trial Against Cancer in Young Adults; NCT00122681).

The control arm of PATRICIA (PApilloma TRIal against Cancer In young Adults,NCT00122681) was used to investigate the risk of progression from cervical HPV infection to cervical intraepithelial neoplasia (CIN) or clearance of infection, and associated determinants.


PMID: 25139208  PMCID: PMC4157699

Lang Kuhs KAPorras CSchiller JTRodriguez ACSchiffman MGonzalez PWacholder SGhosh ALi Y,Lowy DRKreimer ARPoncelet SSchussler JQuint Wvan Doorn LJSherman MESidawy MHerrero R,Hildesheim ASafaeian MCosta Rica Vaccine Trial Group. “Effect of different human papillomavirus serological and DNA criteria on vaccine efficacy estimates”. Am J Epidemiol. 2014 Sep 15;180(6):599-607. doi: 10.1093/aje/kwu168. Epub 2014 Aug 19.

PMID: 24610876-PMCID: PMC4111909

Castellsagué X1Naud P2Chow SN3Wheeler CM4Germar MJ5Lehtinen M6Paavonen J7Jaisamrarn U8,Garland SM9Salmerón J10Apter D11Kitchener H12Teixeira JC13Skinner SR14Limson G15Szarewski A16Romanowski B17Aoki FY18Schwarz TF19Poppe WA20Bosch FX21de Carvalho NS22Peters K23,Tjalma WA24Safaeian M25Raillard A26Descamps D27Struyf F27Dubin G28Rosillon D27Baril L27.”Risk of newly detected infections and cervical abnormalities in women seropositive for naturally acquired human papillomavirus type 16/18 antibodies: analysis of the control arm of PATRICIA”.  J Infect Dis. 2014 Aug 15;210(4):517-34. doi: 10.1093/infdis/jiu139. Epub 2014 Mar 8.  Comment in Naturally acquired immunity against human papillomavirus (HPV): why it matters in the HPV vaccine era. [J Infect Dis. 2014]

PMID: 25313727

Khode SR1Dwivedi RCRhys-Evans PKazi R.  Exploring the link between human papilloma virus and oral and oropharyngeal cancers. J Cancer Res Ther. 2014 Jul-Sep;10(3):492-8. doi: 10.4103/0973-1482.138213.


PMID: 24970795 PMCID: PMC4147298

Zaravinos A1. An updated overview of HPV-associated head and neck carcinomas.

Oncotarget. 2014 Jun 30;5(12):3956-69.

PMID: 24568473

Hamzi Abdul Raub S1Isa NMZailani HAOmar BAbdullah MFMohd Amin WANoor RMAyub MCAbidin ZKassim FVicknesh VZakaria ZKamaluddin MATan GCSyed Husain SN.  Distribution of HPV genotypes in cervical cancer in multi- ethnic Malaysia.  Asian Pac J Cancer Prev. 2014;15(2):651-6.


PMID: 24204068

de la Cruz MS1Young APRuffin MT 4thHuman papillomavirus (HPV) testing for normal cervical cytology in low-risk women aged 30-65 years by family physicians. J Am Board Fam Med. 2013 Nov-Dec;26(6):720-7. doi: 10.3122/jabfm.2013.06.120260.

PMID: 24127500

van der Zee RP1Richel Ode Vries HJPrins JM. The increasing incidence of anal cancer: can it be explained by trends in risk groups?  Neth J Med. 2013 Oct;71(8):401-11.


PMID: 23557172-PMCID: PMC3623745

van Poelgeest MI1Welters MJvan Esch EMStynenbosch LFKerpershoek Gvan Persijn van Meerten EL,van den Hende MLöwik MJBerends-van der Meer DMFathers LMValentijn AROostendorp JFleuren GJ,Melief CJKenter GGvan der Burg SH. HPV16 synthetic long peptide (HPV16-SLP) vaccination therapy of patients with advanced or recurrent HPV16-induced gynecological carcinoma, a phase II trial.  J Transl Med. 2013 Apr 4;11:88. doi: 10.1186/1479-5876-11-88.


PMID: 23441786

Prabhu SR1Wilson DF. Human papillomavirus and oral disease – emerging evidence: a review. Aust Dent J. 2013 Mar;58(1):2-10; quiz 125. doi: 10.1111/adj.12020. Epub 2013 Jan 31.


PMID: 24260180-PMCID:  PMC3834039

Jaisamrarn UCastellsagué XGarland SMNaud PPalmroth JDel Rosario-Raymundo MRWheeler CM,Salmerón JChow SNApter DTeixeira JCSkinner SRHedrick JSzarewski ARomanowski BAoki FY,Schwarz TFPoppe WABosch FXde Carvalho NSGermar MJPeters KPaavonen JBozonnat MC,Descamps DStruyf FDubin GORosillon DBaril LHPV PATRICIA Study GroupNatural history of progression of HPV infection to cervical lesion or clearance: analysis of the control arm of the large, randomised PATRICIA study.  PLoS One. 2013 Nov 19;8(11):e79260. doi: 10.1371/journal.pone.0079260. eCollection 2013.

Collaborators (124) Erratum in PLoS One. 2013;8(12). doi:10.1371/annotation/cea59317-929c-464a-b3f7-e095248f229a.


PMID: 22940493

Roset Bahmanyar EPaavonen JNaud PSalmerón JChow SNApter DKitchener HCastellsagué X,Teixeira JCSkinner SRJaisamrarn ULimson GAGarland SMSzarewski ARomanowski BAoki F,Schwarz TFPoppe WADe Carvalho NSHarper DMBosch FXRaillard ADescamps DStruyf FLehtinen MDubin GHPV PATRICIA Study Group.

Collaborators (151) Gynecol Oncol.

Prevalence and risk factors for cervical HPV infection and abnormalities in young adult women at enrolment in the multinational PATRICIA trial. 2012 Dec;127(3):440-50. doi: 10.1016/j.ygyno.2012.08.033. Epub 2012 Aug 30.

Comment in Optimizing cervical cancer prevention strategies in the United States. [Gynecol Oncol. 2012]


PMID:  22926339 PMCID:  PMC3494281

Pierce MC1Guan YQuinn MKZhang XZhang WHQiao YLCastle PRichards-Kortum RA pilot study of low-cost, high-resolution microendoscopy as a tool for identifying women with cervical precancer. Cancer Prev Res (Phila). 2012 Nov;5(11):1273-9. doi: 10.1158/1940-6207.CAPR-12-0221. Epub 2012 Aug 27.

Comment in Preventing cervical cancer globally. [Cancer Prev Res (Phila). 2012]

PMID: 22960955

Dim CC Towards improving cervical cancer screening in Nigeria: a review of the basics of cervical neoplasm and cytology.  Niger J Clin Pract. 2012 Jul-Sep;15(3):247-52. doi: 10.4103/1119-3077.100615. 1.


PMID: 22019724

Di Domenico F1Foppoli CCoccia RPerluigi MAntioxidants in cervical cancer: chemopreventive and chemotherapeutic effects of polyphenols. Biochim Biophys Acta. 2012 May;1822(5):737-47. doi: 10.1016/j.bbadis.2011.10.005. Epub 2011 Oct 12.


PMID: 22119058-PMCID: PMC3632360

Brown AJ1Trimble CLBest Pract Res Clin Obstet Gynaecol. New technologies for cervical cancer screening.  2012 Apr;26(2):233-42. doi: 10.1016/j.bpobgyn.2011.11.001. Epub 2011 Nov 25.

PMID: 22251005-PMCID:  PMC4164215

Jenkins M1Chiriva-Internati MMirandola LTonroy CTedjarati SSDavis ND’Cunha NTijani LHardwick F,Nguyen DKast WMCobos EPerspective for prophylaxis and treatment of cervical cancer: an immunological approach. Int Rev Immunol. 2012 Feb;31(1):3-21. doi: 10.3109/08830185.2011.637254.


PMID:  21796634-PMCID:  PMC3244688

Sylla BS1Wild CP. “A million africans a year dying from cancer by 2030: what can cancer research and control offer to the continent?Int J Cancer. 2012 Jan 15;130(2):245-50. doi: 10.1002/ijc.26333. Epub 2011 Aug 30.


PMID:  22075942-PMCID:  PMC3251847

Castañon A1Tristram AMesher DPowell NBeer HAshman SRieck GFielder HFiander ASasieni P. Effect of diindolylmethane supplementation on low-grade cervical cytological abnormalities: double-blind, randomised, controlled trial. Br J Cancer. 2012 Jan 3;106(1):45-52. doi: 10.1038/bjc.2011.496. Epub 2011 Nov 10.


PMID:  21527581

Einstein MH1Garcia FAMitchell ALDay SP.

Age-stratified performance of the Cervista HPV 16/18 genotyping test in women with ASC-US cytology. Cancer Epidemiol Biomarkers Prev. 2011 Jun;20(6):1185-9. doi: 10.1158/1055-9965.EPI-11-0116. Epub 2011 Apr 28.


PMID:  21768696

Kumaraswamy KL1Vidhya M. “Human papilloma virus and oral infections: an update”.   J Cancer Res Ther. 2011 Apr-Jun;7(2):120-7. doi: 10.4103/0973-1482.82915.


PMID: 21044009

PMCID: PMC3611662

Sell S1. Infection, stem cells and cancer signals. Curr Pharm Biotechnol. 2011 Feb 1;12(2):182-8.


PMID:  21121531

Juckett G1Hartman-Adams H. Human papillomavirus: clinical manifestations and prevention”.  Am Fam Physician. 2010 Nov 15;82(10):1209-13. Summary for patients in How to prevent cervical cancer. [Am Fam Physician. 2010]

Global Cancer Facts and Figures 207. Available from: ndfigures2007rev2p.pdf.

SEER data for 2000-2004. Available from:

Cancer Facts and Figures 2010. Available from:

Cancer Facts and Figures for African Americans 2009-2010. Available from: 0pdf.pdf.

Cancer Facts and Figures for Hispanic/Latinos 2009-2011. Available from: tinos20092011.pdf.


PMID: 229324377

James Mitchell Crow. “HPV: The global burden”. Nature 488 S2–S3 (30 August 2012) doi:10.1038/488S2a Published online  29 August 2012




Ojesina AI1Lichtenstein L2Freeman SS3Pedamallu CS4Imaz-Rosshandler I5Pugh TJ4Cherniack AD3,Ambrogio L3Cibulskis K3Bertelsen B6Romero-Cordoba S5Treviño V7Vazquez-Santillan K5Guadarrama AS5Wright AA8Rosenberg MW3Duke F9Kaplan B4Wang R10Nickerson E3Walline HM11Lawrence MS3Stewart C3Carter SL3McKenna A3Rodriguez-Sanchez IP12Espinosa-Castilla M5Woie K13Bjorge L14Wik E14Halle MK14Hoivik EA14Krakstad C14Gabiño NB5Gómez-Macías GS12Valdez-Chapa LD12,Garza-Rodríguez ML12Maytorena G15Vazquez J15Rodea C15Cravioto A15Cortes ML3Greulich H16,Crum CP17Neuberg DS18Hidalgo-Miranda A5Escareno CR19Akslen LA20Carey TE21Vintermyr OK20,Gabriel SB3Barrera-Saldaña HA12Melendez-Zajgla J5Getz G22Salvesen HB23Meyerson M24. “Landscape of genomic alterations in cervical carcinomas”.  Nature. 2014 Feb 20;506(7488):371-5. doi: 10.1038/nature12881. Epub 2013 Dec 25.

PMID:24256791, Scudellari M. “HPV: Sex, cancer and a virus”. Nature. 2013 Nov 21;503(7476):330-2. doi: 10.1038/503330a.

PMCID: PMC3740412.
 “Adey A1Burton JNKitzman JOHiatt JBLewis APMartin BKQiu RLee CShendure J. “The haplotype-resolved genome and epigenome of the aneuploid HeLa cancer cell line”.
Nature. 2013 Aug 8;500(7461):207-11. doi: 10.1038/nature12064.


PMID: 22932440

Humphries C. “Screening: Testing times.” Nature. 2012 Aug 30;488(7413):S8-9. doi: 10.1038/488S8a.

PMID: 22932437

Crow JM. “HPV: The global burden”. Nature. 2012 Aug 30;488(7413):S2-3. doi: 10.1038/488S2a.


Bowers EJ.”HPV vaccination: Clarifying the use of ‘prepubescent’”.Nature. 2011 Nov 9;479(7372):179. doi: 10.1038/479179c. Comment on The wrong message on vaccines. [Nature. 2011]

PMID: 19135222

zur Hausen H1. “Papillomaviruses in the causation of human cancers – a brief historical account”. Virology. 2009 Feb 20;384(2):260-5. doi: 10.1016/j.virol.2008.11.046. Epub 2009 Jan 8.



Herrero R, Quint W, Hildesheim A, Gonzalez P, Struijk L, Katki HA, Porras C, Schiffman M, Rodriguez AC, Solomon D, Jimenez S, Schiller JT, Lowy DR, van Doorn LJ, Wacholder S, Kreimer AR. CVT Vaccine Group. Reduced Prevalence of Oral Human Papillomavirus (HPV) 4 Years after Bivalent HPV Vaccination in a Randomized Clinical Trial in Costa Rica. PLoS One. 2013 Jul 17;8(7):e68329., Registry number NCT00128661.

PMID: 17699008

Hildesheim A, Herrero R, Wacholder S, Rodriguez AC, Solomon D, Bratti MC, Schiller JT, Gonzalez P, Dubin G, Porras C, Jimenez SE, Lowy DR. Costa Rican HPV Vaccine Trial Group. Effect of human papillomavirus 16/18 L1 viruslike particle vaccine among young women with preexisting infection: a randomized trial. JAMA. 2007;298(7):743–753. TRIAL REGISTRATION: Identifier: NCT00128661.

PMCID: PMC282145

McCance D, Kopan R, Fuchs E, et al. Human papillomavirus type 16 alters human epithelial cell differentiation in vitro. Proc. Natl. Acad. Sci. 1988;85:7169–7173.

PMCID: PMC3137403

Castellsague X, Munoz N, Pitisuttithum P, Ferris D, Monsonego J, Ault K, et al. End-of-study safety, immunogenicity, and efficacy of quadrivalent HPV (types 6, 11, 16, 18) recombinant vaccine in adult women 24–45 years of age. Br J Cancer. 2011;105:28–37.


CDC. FDA Licensure of Bivalent Human Papillomavirus Vaccine (HPV2, Cervarix) for Use in Females and Updated HPV Vaccination Recommendations from the Advisory Committee on Immunization Practices (ACIP) MMWR. 2010;59(20):626–629.


CDC. Recommendations on the Use of Quadrivalent Human Papillomavirus Vaccine in Males-Advisory Committee on Immunization Practices (ACIP) MMWR. 2011;60(50):1705–1708.

CDC. [accessed October 28, 2013];Biosafety in Microbiological and Biomedical Laboratories (BMBL) (5th Edition).


Chow LT, Duffy AA, Wang HK, Broker TR. A highly efficient system to produce infectious human papillomavirus. Cell Cycle. 2009;8:1319–1323.

Clinical Trials Publications:

Kreimer AR, Rodriguez AC, Hildesheim A, Herrero R, Porras C, Schiffman M, González P, Solomon D, Jiménez S, Schiller JT, Lowy DR, Quint W, Sherman ME, Schussler J, Wacholder S; CVT Vaccine Group. Proof-of-principle evaluation of the efficacy of fewer than three doses of a bivalent HPV16/18 vaccine. J Natl Cancer Inst. 2011 Oct 5;103(19):1444-51. doi: 10.1093/jnci/djr319. Epub 2011 Sep 9.

Kemp TJ, Hildesheim A, Safaeian M, Dauner JG, Pan Y, Porras C, Schiller JT, Lowy DR, Herrero R, Pinto LA. HPV16/18 L1 VLP vaccine induces cross-neutralizing antibodies that may mediate cross-protection. Vaccine. 2011 Mar 3;29(11):2011-4. doi: 10.1016/j.vaccine.2011.01.001. Epub 2011 Jan 15.
Additional publications automatically indexed to this study by Identifier (NCT Number):

Kreimer AR, Struyf F, Del Rosario-Raymundo MR, Hildesheim A, Skinner SR, Wacholder S, Garland SM, Herrero R, David MP, Wheeler CM; Costa Rica Vaccine Trial and PATRICIA study groups. Efficacy of fewer than three doses of an HPV-16/18 AS04-adjuvanted vaccine: combined analysis of data from the Costa Rica Vaccine and PATRICIA trials. Lancet Oncol. 2015 Jul;16(7):775-86. doi: 10.1016/S1470-2045(15)00047-9. Epub 2015 Jun 9.

Gonzalez P, Hildesheim A, Herrero R, Katki H, Wacholder S, Porras C, Safaeian M, Jimenez S, Darragh TM, Cortes B, Befano B, Schiffman M, Carvajal L, Palefsky J, Schiller J, Ocampo R, Schussler J, Lowy D, Guillen D, Stoler MH, Quint W, Morales J, Avila C, Rodriguez AC, Kreimer AR; Costa Rica HPV Vaccine Trial (CVT) Group. Rationale and design of a long term follow-up study of women who did and did not receive HPV 16/18 vaccination in Guanacaste, Costa Rica. Vaccine. 2015 Apr 27;33(18):2141-51. doi: 10.1016/j.vaccine.2015.03.015. Epub 2015 Mar 18.

Lang Kuhs KA, Porras C, Schiller JT, Rodriguez AC, Schiffman M, Gonzalez P, Wacholder S, Ghosh A, Li Y, Lowy DR, Kreimer AR, Poncelet S, Schussler J, Quint W, van Doorn LJ, Sherman ME, Sidawy M, Herrero R, Hildesheim A, Safaeian M; Costa Rica Vaccine Trial Group. Effect of different human papillomavirus serological and DNA criteria on vaccine efficacy estimates. Am J Epidemiol. 2014 Sep 15;180(6):599-607. doi: 10.1093/aje/kwu168. Epub 2014 Aug 19.

Hildesheim A, Wacholder S, Catteau G, Struyf F, Dubin G, Herrero R; CVT Group. Efficacy of the HPV-16/18 vaccine: final according to protocol results from the blinded phase of the randomized Costa Rica HPV-16/18 vaccine trial. Vaccine. 2014 Sep 3;32(39):5087-97. doi: 10.1016/j.vaccine.2014.06.038. Epub 2014 Jul 10.

Lang Kuhs KA, Gonzalez P, Rodriguez AC, van Doorn LJ, Schiffman M, Struijk L, Chen S, Quint W, Lowy DR, Porras C, DelVecchio C, Jimenez S, Safaeian M, Schiller JT, Wacholder S, Herrero R, Hildesheim A, Kreimer AR; Costa Rica Vaccine Trial Group. Reduced prevalence of vulvar HPV16/18 infection among women who received the HPV16/18 bivalent vaccine: a nested analysis within the Costa Rica Vaccine Trial. J Infect Dis. 2014 Dec 15;210(12):1890-9. doi: 10.1093/infdis/jiu357. Epub 2014 Jun 23.

Lang Kuhs KA, Gonzalez P, Struijk L, Castro F, Hildesheim A, van Doorn LJ, Rodriguez AC, Schiffman M, Quint W, Lowy DR, Porras C, Delvecchio C, Katki HA, Jimenez S, Safaeian M, Schiller J, Solomon D, Wacholder S, Herrero R, Kreimer AR; Costa Rica Vaccine Trial Group. Prevalence of and risk factors for oral human papillomavirus among young women in Costa Rica. J Infect Dis. 2013 Nov 15;208(10):1643-52. doi: 10.1093/infdis/jit369. Epub 2013 Sep 6.

Herrero R, Quint W, Hildesheim A, Gonzalez P, Struijk L, Katki HA, Porras C, Schiffman M, Rodriguez AC, Solomon D, Jimenez S, Schiller JT, Lowy DR, van Doorn LJ, Wacholder S, Kreimer AR; CVT Vaccine Group. Reduced prevalence of oral human papillomavirus (HPV) 4 years after bivalent HPV vaccination in a randomized clinical trial in Costa Rica. PLoS One. 2013 Jul 17;8(7):e68329. doi: 10.1371/journal.pone.0068329. Print 2013.

Clarke M, Schiffman M, Wacholder S, Rodriguez AC, Hildesheim A, Quint W; Costa Rican Vaccine Trial Group. A prospective study of absolute risk and determinants of human papillomavirus incidence among young women in Costa Rica. BMC Infect Dis. 2013 Jul 8;13:308. doi: 10.1186/1471-2334-13-308.

Castro FA, Quint W, Gonzalez P, Katki HA, Herrero R, van Doorn LJ, Schiffman M, Struijk L, Rodriguez AC, DelVecchio C, Lowy DR, Porras C, Jimenez S, Schiller J, Solomon D, Wacholder S, Hildesheim A, Kreimer AR; Costa Rica Vaccine Trial Group. Prevalence of and risk factors for anal human papillomavirus infection among young healthy women in Costa Rica. J Infect Dis. 2012 Oct 1;206(7):1103-10. Epub 2012 Jul 30.

Kreimer AR, González P, Katki HA, Porras C, Schiffman M, Rodriguez AC, Solomon D, Jiménez S, Schiller JT, Lowy DR, van Doorn LJ, Struijk L, Quint W, Chen S, Wacholder S, Hildesheim A, Herrero R; CVT Vaccine Group. Efficacy of a bivalent HPV 16/18 vaccine against anal HPV 16/18 infection among young women: a nested analysis within the Costa Rica Vaccine Trial. Lancet Oncol. 2011 Sep;12(9):862-70. doi: 10.1016/S1470-2045(11)70213-3. Epub 2011 Aug 22. Erratum in: Lancet Oncol. 2011 Nov;12(12):1096.

Wacholder S, Chen BE, Wilcox A, Macones G, Gonzalez P, Befano B, Hildesheim A, Rodríguez AC, Solomon D, Herrero R, Schiffman M; CVT group. Risk of miscarriage with bivalent vaccine against human papillomavirus (HPV) types 16 and 18: pooled analysis of two randomised controlled trials. BMJ. 2010 Mar 2;340:c712. doi: 10.1136/bmj.c712.

Dessy FJ, Giannini SL, Bougelet CA, Kemp TJ, David MP, Poncelet SM, Pinto LA, Wettendorff MA. Correlation between direct ELISA, single epitope-based inhibition ELISA and pseudovirion-based neutralization assay for measuring anti-HPV-16 and anti-HPV-18 antibody response after vaccination with the AS04-adjuvanted HPV-16/18 cervical cancer vaccine. Hum Vaccin. 2008 Nov-Dec;4(6):425-34. Epub 2008 Nov 11.

Hildesheim A, Herrero R, Wacholder S, Rodriguez AC, Solomon D, Bratti MC, Schiller JT, Gonzalez P, Dubin G, Porras C, Jimenez SE, Lowy DR; Costa Rican HPV Vaccine Trial Group. Effect of human papillomavirus 16/18 L1 viruslike particle vaccine among young women with preexisting infection: a randomized trial. JAMA. 2007 Aug 15;298(7):743-53.

Related Articles at Leaders in Pharmaceutical Intelligence on HPV:

Head and Neck Cancer Studies Suggest Alternative Markers More Prognostically Useful than HPV DNA Testing

Reporter: Aviva Lev-Ari, PhD, RN Head and Neck Cancer Studies Suggest Alternative Markers More Prognostically Useful than HPV DNA Testing September 18, 2012 By a GenomeWeb staff reporter NEW YORK (GenomeWeb News) – The presence or absence of human papillomavirus DNA on its own in an individual’s head or neck cancer does not provide enough information […]

Read Full Post »

What is Human Papilloma Virus?   Human papillomavirus Taxonomy ID: 10566 Inherited blast name: viruses Rank: species Genetic code: Translation table 1 (Standard) Host: vertebrates| human Other names: synonym: human papillomavirus HPV synonym: Human Papilloma Virus Lineage( full ) Viruses; dsDNA viruses, no RNA stage; Papillomaviridae; unclassified Papillomaviridae; Human papillomavirus types    Entrez records    Database name Subtree links Direct links Nucleotide 7,782 7,775 Protein 2,611 2,604 Structure 3 […]

Read Full Post »

Viruses and Cancer: A Walk on the Memory Lane Curator: Demet Sag, PhD, CRA, GCP   One of the other mechanism where cancer and microorganisms establish a close relationship is viruses. They are vicious sometimes as they adept fast even we don’t call them a real organism since they require a living cell to survive. […]

Read Full Post »

Bacillus Calmette–Guérin (BCG) for Superficial Bladder Cancer Curator: Demet Sag, PhD, CRA, GCP Bladder cancer is arising from the epithelial lining, specifically the urothelium of the urinary bladder with a five year survival rate. The common one is transitional cell carcinoma (90%) and the remaining 10% of bladder cancer are squamous cell carcinoma, adenocarcinoma, sarcoma, small cell carcinoma.  This is a […]

Read Full Post »

Helicobacter pylorum Larry H. Bernstein, MD, FCAP, Curator LPBI The Nobel Prize in Physiology or Medicine 2005 Photo: C. Northcott Barry J. Marshall Photo: U. Montan J. Robin Warren  Affiliation at the time of the award: NHMRC Helicobacter pylori Research Laboratory, QEII Medical Centre, Nedlands, Australia, University of Western Australia, Perth, Australia The Nobel Prize […]

Read Full Post »

 Papilloma viruses for cervical cancer Larry H. Bernstein, MD, FCAP, Curator LPBI Practice Bulletin No. 131: Screening for Cervical Cancer Obstetrics & Gynecology: November 2012 doi: http://10.1097/AOG.0b013e318277c92a The incidence of cervical cancer in the United States has decreased more than 50% in the past 30 years because of widespread screening with cervical cytology. In 1975, […]

Read Full Post »

Curation of Recently Halted Oncology Trials Due to Serious Adverse Events – 2015 Curator: Stephen J. Williams, Ph.D. The following is reports of oncology clinical trials in 2015 which have been halted for Serious Adverse Events (SAE), in most instances of an idiopathic nature. For comparison I have listed (as of this writing) the oncology […]

Read Full Post »

Papilloma viruses for cervical cancer

Larry H. Bernstein, MD, FCAP, Curator


Practice Bulletin No. 131: Screening for Cervical Cancer

Obstetrics & Gynecology:

The incidence of cervical cancer in the United States has decreased more than 50% in the past 30 years because of widespread screening with cervical cytology. In 1975, the rate was 14.8 per 100,000 women. By 2008, it had been reduced to 6.6 per 100,000 women. Mortality from the disease has undergone a similar decrease from 5.55 per 100,000 women in 1975 to 2.38 per 100,000 women in 2008 (1). The American Cancer Society (ACS) estimates that there will be 12,170 new cases of cervical cancer in the United States in 2012, with 4,220 deaths from the disease (2). Cervical cancer is much more common worldwide, particularly in countries without screening programs, with an estimated 530,000 new cases of the disease and 275,000 resultant deaths each year (3, 4). When cervical cancer screening programs have been introduced into communities, marked reductions in cervical cancer incidence have followed (5, 6).

New technologies for cervical cancer screening continue to evolve as do recommendations for managing the results. In addition, there are different risk-benefit considerations for women at different ages, as reflected in age-specific screening recommendations. The ACS, the American Society for Colposcopy and Cervical Pathology (ASCCP), and the American Society for Clinical Pathology (ASCP) have recently updated their joint guidelines for cervical cancer screening (7), and an update to the U.S. Preventive Services Task Force recommendations also has been issued (8). The purpose of this document is to provide a review of the best available evidence regarding screening for cervical cancer.

Study Backs Co-Testing for Cervical Cancer

A positive co-test result was more sensitive than either a positive HPV-only test or a positive Pap-only test.

Charles Bankhead

Cervical cancer screening with a test for human papillomavirus (HPV) resulted in a 50% higher rate of false-negative results versus Pap testing and three times greater versus co-testing, a large retrospective study showed.

Data encompassing more than 250,000 women showed a false-negative rate of 18.6% compared with 12.2% for Pap testing. With a false-negative rate of 5.5%, screening women with the HPV test and Pap test missed the fewest cancers.

The results support clinical guidelines that recommend co-testing, according to authors of a report in Cancer Cytopathology. The results differ dramatically, however, from those of previous studies that have consistently shown greater diagnostic accuracy for the HPV test compared with the Pap test.

“The reason that women are screened is that they want to be protected from cervical cancer,” study author R. Marshall Austin, MD, PhD, of Magee-Women’s Hospital and the University of Pittsburgh, told MedPage Today. “The previous trials have generally focused on cervical intraepithelial neoplasia 2 or 3, so-called precancer. The difference is that most of what we call precancer will actually never develop into cancer.

“The unique thing about this study, and what makes it so important, is that we looked at over 500 invasive cervical cancers, which are what women want to be protected against, and looked at the effectiveness of the methods of testing.”

A year ago, the FDA approved Roche’s cobas assay for HPV DNA as a first-line test for cervical cancer screening, following a unanimous vote for approval by an FDA advisory committee.

The approval was based primarily on a pivotal trial involving 47,200 women at high risk for cervical cancer. The primary endpoint was the proportion of patients who developed grade ≥3 cervical intraepithelial neoplasia (≥CIN3). The results showed a greater than 50% reduction in the incidence of ≥CIN3 with the DNA test versus Pap testing.

Austin and colleagues retrospectively analyzed clinical records for 256,648 average-risk women, ages 30 to 65, all of whom underwent co-testing as a screen for cervical cancer and subsequently had a cervical biopsy within a year of co-testing. The primary objective was to determine the sensitivity of the three screening methods for detection of biopsy-proven ≥CIN3 and invasive cancer.

The results showed that 74.7% of the women had a positive HPV test, 73.8% had an abnormal Pap test (atypical squamous cells of undetermined significance or worse), 89.2% had a positive co-test, and 1.6% had ≥CIN3.

Biopsy results showed that co-testing had the highest sensitivity for ≥CIN3 (98.8% versus 94% for HPV test only and 91.3% for Pap testing alone, P<0.0001). The Pap test had greater specificity versus HPV testing alone or co-testing (26.3% versus 25.6% versus 10.9%, P<0.0001).

Investigators identified 526 patients who developed biopsy-proven invasive cervical cancer. Of those patients, 98 tested negative by HPV assay only, 64 by Pap test only, and 29 by co-testing.

Given the average risk of the patient population included in the study, the results are broadly applicable to women in the age range studied, regardless of baseline risk for cervical cancer, Austin said.

The results are clearly at odds with previously reported comparative data showing superiority for the HPV assay versus Pap testing as a standalone screening test, but the reasons for the inconsistency aren’t clear, said Debbie Saslow, PhD, of the American Cancer Society (ACS) in Atlanta.

The data also show that co-testing is better than either test alone, which supports current ACS recommendations for cervical cancer screening.

“The current approach, according to the American Cancer Society and 25 other organizations that worked with us on our last guideline, co-testing is the preferred strategy,” Saslow told MedPage Today. “This paper completely backs that up. Even though a Pap alone is acceptable, clearly, co-testing is the best way to go.”

Noting that only half of women in the U.S. do not under go co-testing despite clinical guidelines recommending it for more than a decade, Saslow asked, “What’s taking so long?”

Earlier this year, several organizations released joint “interim guidance” regarding cervical cancer screening. Described as an aid to clinical decision-making until existing guidelines are updated, the interim guidance characterized the HPV-DNA test as an acceptable alternative to Pap testing as a primary screening test.

Acknowledging that the guidance focused on use of the HPV assay as a single test, interim guidance lead author Warner Huh, MD, of the University of Alabama at Birmingham, noted that “Every single study worldwide that has looked at this issue shows the same result: HPV testing outperforms Pap testing.”

In their article, Austin and colleagues argued that the HPV assay should be evaluated in comparison with the Pap test but as an alternative to co-testing.

“HPV-only primary screening for cervical cancer presents many challenges for clinicians,” the authors said. “Questions arise regarding its effectiveness, its long-term risk, and when it is the best option for a particular patient.

“Clinicians had similar questions when co-testing was first recommended for women 30 and older in 2006,” they added. “Since then the adoption of co-testing has steadily increased, with approximately 50% of physicians co-testing women 30 and older, but it is still not done at the recommended level.”

The study had some limitations. The authors could not confirm that the cervical biopsy results were from women who did not have an intervening screening test or treatment with a different provider during the study period.

Also, the authors were unable to draw conclusions based on the overall population of women who were screened for cervical cancer because the dataset consisted of screening results of women who underwent biopsies.


Read Full Post »

Can IntraTumoral Heterogeneity Be Thought of as a Mechanism of Resistance?

Curator/Reporter: Stephen J. Williams, Ph.D.

Therapeutic resistance remains one of the most challenging problems for the oncologist, despite the increase of new therapeutics in the oncologist’s toolkit. As new targeted therapies are developed, and new novel targets are investigated as potential therapies, especially cytostatic therapies which it has become evident our understanding of chemoresistance is expanding beyond mechanisms to circumvent a drug’s pharmacologic mechanism of action (i.e. increased DNA repair and cisplatin) or pharmacokinetic changes (i.e. increased efflux by acquisition of a MDR phenotype).

In a talk at the 2015 AACR National Meeting, Dr. Charles Swanton discusses the development of tumor heterogeneity in the light of developing, or acquired, drug resistance. Chemoresistance is either categorized as acquired resistance (where resistance develops upon continued exposure to drug) or inherent resistance (related to a tumor being refractory or unresponsive to drug). Dr Swanton discusses findings where development of this heterogeneity (discussed here in a posting on Issues in Personalized Medicine in Cancer: Intratumor Heterogeneity and Branched Evolution Revealed by Multiregion Sequencing) and here (Notes On Tumor Heterogeneity: Targets and Mechanisms, from the 2015 AACR Meeting in Philadelphia PA) on recent findings on Branched Chain Heterogeneity) is resulting in clones resistant to the initial drug treatment.

To recount a bit of background I list the overall points of the one of previous posts on tumor heterogeneity (and an interview with Dr. Charles Swanton) are as follows:

Multiple biopsies of primary tumor and metastases are required to determine the full mutational landscape of a patient’s tumor

The intratumor heterogeneity will have an impact on the personalized therapy strategy for the clinician

Metastases arising from primary tumor clones will have a greater genomic instability and mutational spectrum than the tumor from which it originates

Tumors and their metastases do NOT evolve in a linear path but have a branched evolution and would complicate biomarker development and the prognostic and resistance outlook for the patient


The following is a curation of various talks and abstracts from the 2015 AACR National Meeting in Philadelphia on effects of clonal evolution and intratumoral heterogeneity of a tumor with respect to development of chemoresistance. As this theory of heterogeneity and clonal evolution is particularly new I attempted to present all works (although apologize for the length upfront) to forgo bias and so the reader may extract any information pertinent to their clinical efforts and research. However I will give a brief highlight summary below:


From the 2015 AACR National Meeting in Philadelphia






Presentation Title: Polyclonal and heterogeneous resistance to targeted therapy in leukemia
Presentation Time: Monday, Apr 20, 2015, 10:40 AM -10:55 AM
Location: Room 201, Pennsylvania Convention Center
Author Block: Catherine C. Smith, Amy Paguirigan, Chen-Shan Chin, Michael Brown, Wendy Parker, Mark J. Levis, Alexander E. Perl, Kevin Travers, Corynn Kasap, Jerald P. Radich, Susan Branford, Neil P. Shah. University of California, San Francisco, CA, Fred Hutchinson Cancer Research Center, Seattle, WA, Pacific Biosciences, Menlo Park, CA, Royal Adelaide Hospital, Adelaide, Australia, Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD, Abramson Cancer Center of the University of Pennsylvania, Philadelphia, PA, University of California, San Francisco, CA
Abstract Body: Genomic studies in solid tumors have revealed significant branching intratumoral clonal genetic heterogeneity. Such complexity is not surprising in solid tumors, where sequencing studies have revealed thousands of mutations per tumor genome. However, in leukemia, the genetic landscape is considerably less complex. Chronic myeloid leukemia (CML) is the human malignancy most definitively linked to a single genetic lesion, the BCR-ABL gene fusion. Genome wide sequencing of acute myeloid leukemia (AML) has revealed that AML is the most genetically straightforward of all extensively sequenced adult cancers to date, with an average of 13 coding mutations and 3 or less clones identified per tumor.
In CML, tyrosine kinase inhibitors (TKIs) of BCR-ABL have resulted in high rates of remission. However, despite excellent initial response rates with TKI monotherapy, patients still relapse, including virtually all patients with Philadelphia-positive acute lymphoblastic leukemia and blast crisis CML. Studies of clinical resistance highlight BCR-ABL as the sole genetic driver in CML as secondary kinase domain (KD) mutations that prevent drug binding are the predominant mechanism of relapse on BCR-ABL TKIs.
In AML, a more diverse panel of disease-defining genetic mutations has been uncovered. However, in individual patients, a single oncogene can still drive disease. This is the case in FLT3 mutant AML, in which the investigational FLT3 TKI quizartinib achieved an initial response rate of ~50% in relapsed/refractory AML patients with activating FLT3 internal tandem duplication (ITD) mutations, though most patients eventually relapsed. Confirming the importance of FLT3 in disease maintenance, we showed that 8 of 8 patients who relapsed on quizartinib did so due to acquired drug-resistant FLT3 KD mutations.
Studies in CML have revealed that sequential TKI therapy is associated with additional complexity where multiple mutations can coexist separately in an individual patient (“polyclonality”) or in tandem on a single allele (“compound mutations”). In AML, we observed polyclonal FLT3-ITD KD mutations in 2 of 8 patients examined in our initial study of quizartinib resistance.
In light of the polyclonal KD mutations observed in CML and AML at the time of TKI relapse, we undertook next generation sequencing studies to determine the true genetic complexity in CML and AML patients at the time of relapse on targeted therapy. We used Pacific Biosciences RS Single Molecule Real Time (SMRT) third generation sequencing technology to sequence the entire ABL KD or the entire FLT3 juxtamembrane and KD on a single strand of DNA. Using this method, we assessed a total of 103 samples from 79 CML patients on ABL TKI therapy and 36 paired pre-treatment and relapse samples from 18 FLT3-ITD+ AML patients who responded to investigational FLT3 TKI therapy.
In CML, using SMRT sequencing, we detected all mutations previously detected by direct sequencing. Of samples in which multiple mutations were detectable by direct sequencing, 85% had compound mutant alleles detectable in a variety of combinations. Compound mutant alleles were comprised of both dominant and minor mutations, some which were not detectable by direct sequencing. In the most complex case, 12 individual mutant alleles comprised of 7 different mutations were identified in a single sample.
For 12 CML patients, we interrogated longitudinal samples (2-4 time points per patient) and observed complex clonal relationships with highly dynamic shifts in mutant allele populations over time. We detected compound mutations arising from ancestral single mutant clones as well as parallel evolution of de novo polyclonal and compound mutations largely in keeping with what would be expected to cause resistance to the second generation TKI therapy received by that patient.
We used a phospho-flow cytometric technique to assesses the phosphorylation status of the BCR-ABL substrate CRKL in as a method to test the ex vivo biochemical responsiveness of individual mutant cell populations to TKI therapy and assess functional cellular heterogeneity in a given patient at a given timepoint. Using this technique, we observed co-existing cell populations with differential ex vivo response to TKI in 2 cases with detectable polyclonal mutations. In a third case, we identified co-existence of an MLL-AF9 containing cell population that retained the ability to modulate p-CRKL in response to BCR-ABL TKIs along with a BCR-ABL containing only population that showed biochemical resistance to all TKIs, suggesting the co-existence of BCR-ABL independent and dependent resistance in a single patient.
In AML, using SMRT sequencing, we identified acquired quizartinib resistant KD mutations on the FLT3-ITD (ITD+) allele of 9 of 9 patients who relapsed after response to quizartinib and 4 of 9 patients who relapsed after response to the investigational FLT3 inhibitor, PLX3397. In 4 cases of quizartinib resistance and 3 cases of PLX3397 resistance, polyclonal mutations were observed, including 7 different KD mutations in one patient with PLX3397 resistance. In 7 quizartinib-resistant cases and 3 PLX3397-resistant cases, mutations occurred at the activation loop residue D835. When we examined non-ITD containing (ITD-) alleles, we surprisingly uncovered concurrent drug-resistant FLT3 KD mutations on ITD- alleles in 7 patients who developed quizartinib resistance and 4 patients with PLX3397 resistance. One additional PLX3397-resistant patient developed a D835Y mutation only in ITD- alleles at the time of resistance, suggesting selection for a non-ITD containing clone. All of the individual substitutions found on ITD- alleles were the same substitutions identified on ITD+ alleles for each individual patient.
Given that the same individual mutations found on ITD- alleles were also found on ITD+ alleles, we sought to determine whether these mutations were found in the same cell or were indicative of polyclonal blast populations in each patient. To answer this question, we performed single cell sorting of viably frozen blasts from 3 quizartinib-resistant patients with D835 mutations identified at the time of relapse and genotyped single cells for the presence or absence of ITD and D835 mutations. This analysis revealed striking genetic heterogeneity. In 2/3 cases, polyclonal D835 mutations were found in both ITD+ and ITD- cells. In all cases, FLT3-ITD and D835 mutations were found in both heterozygous and homozygous combinations. Most surprisingly, in all 3 patients, approximately 30-40% of FLT3-ITD+ cells had no identified quizartinib resistance-causing FLT3 KD mutation to account for resistance, suggesting the presence of non-FLT3 dependent resistance in all patients.
To determine that ITD+ cells lacking FLT3 KD mutations observed in patients relapsed on quizartinib are indeed consistent with leukemic blasts functionally resistant to quizartinib and do not instead represent a population of differentiated or non-proliferating cells, we utilized relapse blasts from another patient who initially achieved clearance of bone marrow blasts on quizartinib and developed a D835Y mutation at relapse. We performed a colony assay in the presence of 20nM quizartinib. As expected, this dose of quizartinib was unable to suppress the colony-forming ability of blasts from this relapsed patient when compared to DMSO treatment. Genotyping of individual colonies grown from this relapse sample in the presence of 20nM quizartinib again showed remarkable genetic heterogeneity, including ITD+ and ITD- colonies with D835Y mutations in homozygous and heterozygous combinations as well as ITD+ colonies without D835Y mutations, again suggesting the presence of blasts with non-FLT3 dependent resistance. Additionally, 4 colonies with no FLT3 mutations at all were identified in this sample, suggesting the presence of a quizartinib-resistant non-FLT3 mutant blast population. To see if we could identify specific mechanisms of off-target resistance, we performed targeted exome sequencing 33-AML relevant genes from relapse and pre-treatment DNA from all four patients and detected no new mutations in any genes other than FLT3 acquired at the time of disease relapse. Clonal genetic heterogeneity is not surprising in solid tumors, where multiple driver mutations frequently occur, but in CML and FLT3-ITD+ AML, where disease has been shown to be exquisitely dependent on oncogenic driver mutations, our studies suggest a surprising amount of clonal diversity. Our findings show that clinical TKI resistance in these diseases is amazingly intricate on the single allele level and frequently consists of both polyclonal and compound mutations that give rise to an complicated pool of TKI-resistant alleles that can change dynamically over time. In addition, we demonstrate that cell populations with off-target resistance can co-exist with other TKI-resistant populations, underscoring the emerging complexity of clinical TKI resistance. Such complexity argues strongly that monotherapy strategies in advanced CML and AML may be ultimately doomed to fail due to heterogeneous cell intrinsic resistance mechanisms. Ultimately, combination strategies that can address both on and off target resistance will be required to effect durable therapeutic responses.
Session Title: Tumor Heterogeneity and Evolution
Session Type: Educational Session
Session Start/End Time: Saturday, Apr 18, 2015, 1:00 PM – 3:00 PM
Location: Terrace Ballroom II-III (400 Level), Pennsylvania Convention Center
CME: CME-Designated
CME/CE Hours: 2
Session Description: One of the major challenges for both the measurement and management of cancer is its heterogeneity. Recent studies have revealed both extensive inter- and intra-tumor heterogeneity at the genotypic and phenotypic levels. Leaders in the field will discuss this challenge, its origins, dynamics and clinical importance. They will also review how we can best measure and deal with tumor heterogeneity, particularly intra-tumor heterogeneity.
Saturday, Apr 18, 2015, 1:00 PM – 3:00 PM
Carlo C. Maley. UCSF Helen Diller Family Comp. Cancer Center, San Francisco, CA
Universal biomarkers: How to handle tumor heterogeneity
Saturday, Apr 18, 2015, 1:00 PM – 1:25 PM
Carlo C. Maley. UCSF Helen Diller Family Comp. Cancer Center, San Francisco, CA
Saturday, Apr 18, 2015, 1:25 PM – 1:30 PM
Heterogeneity of resistance to cancer therapy
Saturday, Apr 18, 2015, 1:30 PM – 1:55 PM
Ivana Bozic. HARVARD UNIV., Cambridge, MA
Saturday, Apr 18, 2015, 1:55 PM – 2:00 PM
Determinants of phenotypic intra-tumor heterogeneity: integrative approach
Saturday, Apr 18, 2015, 2:00 PM – 2:25 PM
Andriy Marusyk, Michalina Janiszewska, Doris Tabassum. Dana-Farber Cancer Institute, Boston, MA, Dana-Farber Cancer Institute, Boston, MA
Saturday, Apr 18, 2015, 2:25 PM – 2:30 PM
Cancer clonal complexity and evolution at the macro- and microheterogeneity scale
Saturday, Apr 18, 2015, 2:30 PM – 2:55 PM
Marco Gerlinger. Institute of Cancer Research, London, United Kingdom
Saturday, Apr 18, 2015, 2:55 PM – 3:00 PM

From Ivana Bozic:

A spatial model predicts that dispersal and cell turnover limit intratumour heterogeneity.

Waclaw B, Bozic I, Pittman ME, Hruban RH, Vogelstein B, Nowak MA.

Nature. 2015 Sep 10;525(7568):261-4. doi: 10.1038/nature14971. Epub 2015 Aug 26.



Similar articles

Select item 253494242.

Timing and heterogeneity of mutations associated with drug resistance in metastatic cancers.

Bozic I, Nowak MA.

Proc Natl Acad Sci U S A. 2014 Nov 11;111(45):15964-8. doi: 10.1073/pnas.1412075111. Epub 2014 Oct 27.



Free PMC Article

Similar articles

Select item 238053823.

Evolutionary dynamics of cancer in response to targeted combination therapy.

Bozic I, Reiter JG, Allen B, Antal T, Chatterjee K, Shah P, Moon YS, Yaqubie A, Kelly N, Le DT, Lipson EJ, Chapman PB, Diaz LA Jr, Vogelstein B, Nowak MA.

Elife. 2013 Jun 25;2:e00747. doi: 10.7554/eLife.00747.



Free PMC Article

Similar articles

Select item 227228434.

The molecular evolution of acquired resistance to targeted EGFR blockade in colorectal cancers.

Diaz LA Jr, Williams RT, Wu J, Kinde I, Hecht JR, Berlin J, Allen B, Bozic I, Reiter JG, Nowak MA, Kinzler KW, Oliner KS, Vogelstein B.

Nature. 2012 Jun 28;486(7404):537-40. doi: 10.1038/nature11219.



Free PMC Article

Similar articles


Session Title: Mechanisms of Cancer Therapy Resistance
Session Type: Educational Session
Session Start/End Time: Saturday, Apr 18, 2015, 1:00 PM – 3:00 PM
Location: Room 204, Pennsylvania Convention Center
CME: CME-Designated
CME/CE Hours: 2
Session Description: Despite dramatic advances in the treatment of cancer, therapy resistance remains the most significant hurdle in improving the outcome of cancer patients. In this session, we will discuss many different aspects of therapy resistance, including a summary of our current understanding of therapy resistant tumor cell populations as well as analyses of the challenges associated with intratumoral heterogeneity and adaptive responses to targeted therapies.
Saturday, Apr 18, 2015, 1:00 PM – 3:00 PM
Charles Swanton. Cancer Research UK London Research Institute, London, United Kingdom
Tumor heterogeneity and drug resistance
Saturday, Apr 18, 2015, 1:00 PM – 1:30 PM
Charles Swanton. Cancer Research UK London Research Institute, London, United Kingdom

Saturday, Apr 18, 2015, 1:30 PM – 1:40 PM
Discussion Discussion, Discussion

Principles of resistance to targeted therapy
Saturday, Apr 18, 2015, 1:40 PM – 2:10 PM
Levi A. Garraway. Dana-Farber Cancer Institute, Boston, MA

Saturday, Apr 18, 2015, 2:10 PM – 2:20 PM
Discussion Discussion, Discussion

Adaptive re-wiring of signaling pathways driving drug resistance to targeted therapies
Saturday, Apr 18, 2015, 2:20 PM – 2:50 PM
Taru E. Muranen. Harvard Medical School, Boston, MA

Saturday, Apr 18, 2015, 2:50 PM – 3:00 PM
Discussion Discussion, Discussion

Presentation Abstract  




Abstract Number: 737
Presentation Title: Clonal evolution of the HER2 L755S mutation as a mechanism of acquired HER-targeted therapy resistance
Presentation Time: Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Location: Section 30
Poster Board Number: 29
Author Block: Xiaowei Xu1, Agostina Nardone1, Huizhong Hu1, Lanfang Qin1, Sarmistha Nanda1, Laura Heiser2, Nicholas Wang2, Kyle Covington1, Edward Chen1, Alexander Renwick1, Tamika Mitchell1, Marty Shea1, Tao Wang1, Carmine De Angelis1, Alejandro Contreras1, Carolina Gutierrez1, Suzanne Fuqua1, Gary Chamness1, Chad Shaw1, Marilyn Li1, David Wheeler1, Susan Hilsenbeck1, Mothaffar Fahed Rimawi1, Joe Gray2, C.Kent Osborne1, Rachel Schiff1. 1Baylor College of Medicine, Houston, TX; 2Oregon Health & Science University, Portland, OR
Abstract Body: Background: Targeting HER2 with lapatinib (L), trastuzumab (T), or the LT combination, is effective in HER2+ breast cancer (BC), but acquired resistance commonly occurs. In our 12-week neoadjuvant
trial (TBCRC006) of LT without chemotherapy in HER2+ BC, the overall pathologic complete response (pCR) rate was 27%. To investigate resistance mechanisms, we developed 10 HER2+ BC cell line
models resistant (R) to one or both drugs (LR/TR/LTR). To discover potential predictive markers/therapeutic targets to circumvent resistance, we completed genomic profiling of the cell lines and a
subset of pre-treatment specimens from TBCRC006.
Methods: Parental (P) and LR/TR/LTR lines of 10 cell line models were profiled with whole exome/RNA sequencing. Mutations detected in R lines but not in P lines of the same model were identified. Mutation-specific Q-PCR was designed for sensitive quantification. Resistant cell and xenograft tumor growth were measured in response to drugs. Whole exome sequencing (>100X) and Ampliseq of 17 baseline tumor/normal pairs from TBCRC006 were performed.
Results: We found and validated the HER2 L755S mutation in the BT474/ATCC-LTR line and BT474/AZ-LR line (in ~30% of DNA/RNA), in which the HER pathway was reactivated for resistance. Overexpression of this mutation was previously shown to induce LR in HER2-negative BC cell lines, and resistant growth of BT474/AZ-LR line is significantly inhibited by HER2-L755S-specific siRNA knock-down, suggesting its role as an acquired L/LT resistance driver in HER2+ BC. Sequencing of BT474/AZ-LR single cell clones found the mutation in ~30% of HER2 copies in every cell. Using mutation-specific Q-PCR, we found statistically higher HER2 L755S levels in two BT474 parentals compared to P lines of SKBR3, AU565, and UACC812. These data suggest that HER2 L755S resistant subclones preexist in the BT474 parentals and were selected by L treatment to become the major clone in the two R lines. The HER1/2 irreversible tyrosine kinase inhibitor (TKI) afatinib (Afa) robustly inhibited growth of BT474/AZ-LR and BT474/ATCC-LTR cells (IC50: Afa 0.02µM vs. L 3 µM) and BT474/AZ-LR xenografts. Whole exome sequencing/Ampliseq of TBCRC006 found the HER2 L755S mutation in 1/17 primaries. This patient did not achieve pCR. The variant was present in 2% of DNA on both platforms, indicating a subclonal event of the resistance mutation.
Conclusion: Acquired L/LT resistance in the two BT474 R lines is due to selection of HER2 L755S subclones present in parental cells. The higher HER2 L755S
levels in BT474 parentals compared with other parentals, and detection of its subclonal presence in a pre-treatment HER2+ BC patient, suggest that sensitive mutation detection methods will be needed to identify patients with potentially actionable HER family mutations in primary tumor. Treating this patient group
with an irreversible TKI like Afa may prevent resistance and improve clinical outcome of this subset of HER2+ BC.
Presentation Number: SY07-04
Presentation Title: The evolutionary landscape of CLL: Therapeutic implications
Presentation Time: Sunday, Apr 19, 2015, 2:25 PM – 2:45 PM
Location: Grand Ballroom (300 Level), Pennsylvania Convention Center
Author Block: Catherine J. Wu. Dana-Farber Cancer Institute, Boston, MA
Abstract Body: Clonal evolution is a key feature of cancer progression and relapse. Recent studies across cancers have demonstrated the extensive degree of intratumoral heterogeneity present within individual cancers. We hypothesized that evolutionary dynamics contribute to the variations in disease tempo and response to therapy that are highly characteristic of chronic lymphocytic leukemia (CLL). We have recently investigated this phenomenon by developing a pipeline that estimates the fraction of cancer cells harboring each somatic mutation within a tumor through integration of whole-exome sequence (WES) and local copy number data (Landau et al., Cell 2013). By applying this analysis approach to 149 CLL cases, we discovered earlier and later cancer drivers, uncovered patterns of clonal evolution in CLL and linked the presence of subclones harboring driver mutations with adverse clinical outcome. Thus, our study, generated from a heterogeneous sample cohort, strongly supports the concept that CLL clonal evolution arises from mass extinction and therapeutic bottlenecks which lead to the emergence of highly fit (and treatment resistant) subclones. We further hypothesized that epigenetic heterogeneity also shapes CLL clonal evolution through interrelation with genetic heterogeneity. Indeed, in recent work, we have uncovered stochastic methylation disorder as the primary cause of methylation changes in CLL and cancer in general, and that this phenomena impacts gene transcription, genetic evolution and clinical outcome. Thus, integrated studies of genetic and epigenetic heterogeneity in CLL have revealed the complex and diverse evolutionary trajectories of these cancer cells.
Immunotherapy is exquisitely suited for specifically and simultaneously targeting multiple lesions. We have developed an approach that leverages whole-exome sequencing to systematically identify personal tumor mutations with immunogenic potential, which can be incorporated as antigen targets in multi-epitope personalized therapeutic vaccines. We are pioneering this approach in an ongoing trial in melanoma and will now expand this concept to address diverse malignancies. Our expectation is that the choice of tumor neoantigens for a vaccine bypasses thymic tolerance and thus generates highly specific and potent high-affinity T cell responses to eliminate tumors in any cancer, including both ‘trunk’ and ‘branch’ lesions.


Abstract Number: LB-056
Presentation Title: TP53 and RB1 alterations promote reprogramming and antiandrogen resistance in advanced prostate cancer
Presentation Time: Sunday, Apr 19, 2015, 4:50 PM – 5:05 PM
Location: Room 122, Pennsylvania Convention Center
Author Block: Ping Mu, Zhen Cao, Elizabeth Hoover, John Wongvipat, Chun-Hao Huang, Wouter Karthaus, Wassim Abida, Elisa De Stanchina, Charles Sawyers. Memorial Sloan Kettering Cancer Center, New York, NY
Abstract Body: Castration-resistant prostate cancer (CRPC) is one of the most difficult cancers to treat with conventional methods and is responsible for nearly all prostate cancer deaths in the US. The Sawyers laboratory first showed that the primary mechanism of resistance to antiandrogen therapy is elevated androgen receptor (AR) expression. Research based on this finding has led to the development of next-generation antiandrogen: enzalutamide. Despite the exciting clinical success of enzalutamide, about 60% of patients exhibit various degrees of resistance to this agent. Highly variable responses to enzalutamide limit the clinical benefit of this novel antiandrogen, underscoring the importance of understanding the mechanisms of enzalutamide resistance. Most recently, an unbiased SU2C-Prostate Cancer Dream Team metastatic CRPC sequencing project led by Dr. Sawyers and Dr. Chinnaiyan revealed that mutations in the TP53 locus are the most significantly enriched alteration in CRPC tumors when compared to primary prostate cancers. Moreover, deletions and decreased expressions of the TP53 and RB1 loci (co-occurrence and individual occurrence) are more commonly associated with CRPC than with primary tumors. These results established that alteration of the TP53 and RB1 pathways are associated with the development of antiandrogen resistance.
By knockdowning TP53 or/and RB1 in the castration resistant LNCaP/AR model, we demonstrate that the disruption of either TP53 or RB1 alone confers significant resistance to enzalutamide both in vitro and in vivo. Strikingly, the co-inactivation of these pathways confers the most dramatic resistance. Since up-regulation of either AR or AR target genes is not observed in the resistant tumors, loss of TP53 and RB1 function confers enzalutamide resistance likely through an AR independent mechanism. In the clinic, resistance to enzalutamide is increasingly being associated with a transition to a poorly differentiated or neuroendocrine-like histology. Interestingly, we observed significant up-regulations of the basal cell marker Ck5 and the neuroendocrine-like cell marker Synaptophysin in the TP53 and RB1 inactivated cells, as well as down-regulation of the luminal cell marker Ck8. The differences between these markers became even greater after enzalutamide treatment. By using the p53-stabilizing drug Nutlin, level of p53 is rescued and consequently the the decrease of AR protein caused by RB1 and TP53 knockdown is reversed. These results strongly suggest that interference of TP53 and RB1 pathways confers antiandrogen resistance by “priming” prostate cancer cells to reprogramming or transdifferentiation, likely neuroendocrine-like differentiation, in response to treatment. Futher experiments will be performed to assess the molecular mechanism of TP53/RB1 alterations in mediating cell programming and conferring antiandrogen resistance.


Abstract Number: LB-146
Presentation Title: TGF-β-induced tumor heterogeneity and drug resistance of cancer stem cells
Presentation Time: Monday, Apr 20, 2015, 1:00 PM – 5:00 PM
Location: Section 41
Author Block: Naoki Oshimori1, Daniel Oristian1, Elaine Fuchs2. 1Rockefeller University, New York, NY; 2HHMI/Rockefeller University, New York, NY
Abstract Body: Among the most common and life-threatening cancers world-wide, squamous cell carcinoma (SCC) exhibit high rates of tumor recurrence following anti-cancer therapy. Subsets of cancer stem cells (CSCs) often escape anti-cancer therapeutics and promote recurrence. However, its sources and mechanisms that generate tumor heterogeneity and therapy-resistant cell population are largely unknown. Tumor microenvironment may drive intratumor heterogeneity by transmitting signaling factors, oxygen and metabolites to tumor cells depending on their proximity to the local sources. While the hypothesis is attractive, experimental evidence is lacking, and non-genetic mechanisms that drive functional heterogeneity remain largely unknown. As a potential non-genetic factor, we focused on TGF-β because of its multiple roles in tumor progression.
Here we devise a functional reporter system to monitor, track and modify TGF-β signaling in mouse skin SCC in vivo. Using this approach, we found that perivascular TGF-β in the tumor microenvironment generates heterogeneity in TGF-β signaling in neighboring CSCs. This heterogeneity is functionally important: small subsets of TGF-β-responding CSCs proliferate more slowly than their non-responding counterparts. They also exhibit invasive morphology and a malignant differentiation program compared to their non-responding neighbors. By lineage tracing, we show that although TGF-β-responding CSCs clonally expand more slowly they gain a growth advantage in a remarkable ability to escape cisplatin-induced apoptosis. We show that indeed it is their progenies that make a substantial contribution in tumor recurrence. Surprisingly, the slower proliferating state of this subset of CSCs within the cancer correlated with but did not confer the survival advantage to anti-cancer drugs. Using transcriptomic, biochemical and genetic analyses, we unravel a novel mechanism by which heterogeneity in the tumor microenvironment allows a subset of CSCs to respond to TGF-β, and evade anti-cancer drugs.
Our findings also show that TGF-β established ability to suppress proliferation and promote invasion and metastasis do not happen sequentially, but rather simultaneously. This new work build upon the roles of this factor in tumor progression, and sets an important paradigm for a non-genetic factor that produces tumor heterogeneity.
Abstract Number: LB-129
Presentation Title: Identifying tumor subpopulations and the functional consequences of intratumor heterogeneity using single-cell profiling of breast cancer patient-derived xenografts
Presentation Time: Monday, Apr 20, 2015, 1:00 PM – 5:00 PM
Location: Section 41
Author Block: Paul Savage1, Sadiq M. Saleh1, Ernesto Iacucci1, Timothe Revil1, Yu-Chang Wang1, Nicholas Bertos1, Anie Monast1, Hong Zhao1, Margarita Souleimanova1, Keith Szulwach2, Chandana Batchu2, Atilla Omeroglu1, Morag Park1, Ioannis Ragoussis1. 1McGill University, Montreal, QC, Canada; 2Fluidigm Corporation, South San Francisco, CA
Abstract Body: Human breast tumors have been shown to exhibit extensive inter- and intra-tumor heterogeneity. While recent advances in genomic technologies have allowed us to deconvolute this heterogeneity, few studies have addressed the functional consequences of diversity within tumor populations. Here, we identified an index case for which we have derived a patient-derived xenograft (PDX) as a renewable tissue source to identify subpopulations and perform functional assays. On pathology, the tumor was an invasive ductal carcinoma which was hormone receptor-negative, HER2-positive (IHC 2+, FISH average HER2/CEP17 2.4), though the FISH signal was noted to be heterogeneous. On gene expression profiling of bulk samples, the primary tumor and PDX were classified as basal-like. We performed single cell RNA and exome sequencing of the PDX to identify population structure. Using a single sample predictor of breast cancer subtype, we have identified single basal-like, HER2-enriched and normal-like cells co-existing within the PDX tumor. Genes differentially expressed between these subpopulations are involved in proliferation and differentiation. Functional studies distinguishing these subpopulations are ongoing. Microfluidic whole genome amplification followed by whole exome capture of 81 single cells showed high and homogeneous target enrichment with >75% of reads mapping uniquely on target. Variant calling using GATK and Samtools revealed founder mutations in key genes as BRCA1 and TP53, as well as subclonal mutations that are being investigated further. Loss of heterozygocity was observed in 16 TCGA cancer driver genes and novel mutations in 7 cancer driver genes. These findings may be important in understanding the functional consequences of intra-tumor heterogeneity with respect to clinically important phenotypes such as invasion, metastasis and drug-resistance.
Abstract Number: 2847
Presentation Title: High complexity barcoding to study clonal dynamics in response to cancer therapy
Presentation Time: Monday, Apr 20, 2015, 4:35 PM – 4:50 PM
Location: Room 118, Pennsylvania Convention Center
Author Block: Hyo-eun C. Bhang1, David A. Ruddy1, Viveksagar Krishnamurthy Radhakrishna1, Rui Zhao2, Iris Kao1, Daniel Rakiec1, Pamela Shaw1, Marissa Balak1, Justina X. Caushi1, Elizabeth Ackley1, Nicholas Keen1, Michael R. Schlabach1, Michael Palmer1, William R. Sellers1, Franziska Michor2, Vesselina G. Cooke1, Joshua M. Korn1, Frank Stegmeier1. 1Novartis Institutes for BioMedical Research, Cambridge, MA; 2Dana-Farber Cancer Institute, Boston, MA
Abstract Body: Targeted therapies, such as erlotinib and imatinib, lead to dramatic clinical responses, but the emergence of resistance presents a significant challenge. Recent studies have revealed intratumoral heterogeneity as a potential source for the emergence of therapeutic resistance. However, it is still unclear if relapse/resistance is driven predominantly by pre-existing or de novo acquired alterations. To address this question, we developed a high-complexity barcode library, ClonTracer, which contains over 27 million unique DNA barcodes and thus enables the high resolution tracking of cancer cells under drug treatment. Using this library in two clinically relevant resistance models, we demonstrate that the majority of resistant clones pre-exist as rare subpopulations that become selected in response to therapeutic challenge. Furthermore, our data provide direct evidence that both genetic and non-genetic resistance mechanisms pre-exist in cancer cell populations. The ClonTracer barcoding strategy, together with mathematical modeling, enabled us to quantitatively dissect the frequency of drug-resistant subpopulations and evaluate the impact of combination treatments on the clonal complexity of these cancer models. Hence, monitoring of clonal diversity in drug-resistant cell populations by the ClonTracer barcoding strategy described here may provide a valuable tool to optimize therapeutic regimens towards the goal of curative cancer therapies.
Abstract Number: 3590
Presentation Title: Resistance mechanisms to ALK inhibitors
Presentation Time: Tuesday, Apr 21, 2015, 8:00 AM -12:00 PM
Location: Section 31
Poster Board Number: 13
Author Block: Ryohei Katayama1, Noriko Yanagitani1, Sumie Koike1, Takuya Sakashita1, Satoru Kitazono1, Makoto Nishio1, Yasushi Okuno2, Jeffrey A. Engelman3, Alice T. Shaw3, Naoya Fujita1. 1Japanese Foundation for Cancer Research, Tokyo, Japan; 2Graduate School of Medicine, Kyoto University, Kyoto, Japan; 3Massachusetts General Hospital Cancer Center, Boston, MA
Abstract Body: Purpose: ALK-rearranged non-small cell lung cancer (NSCLC) was first reported in 2007. Approximately 3-5% of NSCLCs harbor an ALK gene rearrangement. The first-generation ALK tyrosine kinase inhibitor (TKI) crizotinib is a standard therapy for patients with advanced ALK-rearranged NSCLC. Several next-generation ALK-TKIs have entered the clinic and have shown promising antitumor activity in crizotinib-resistant patients. As patients still relapse even on these next-generation ALK-TKIs, we examined mechanisms of resistance to one next-generation ALK-TKI – alectinib – and potential strategies to overcome this resistance.
Experimental Procedure: We established a cell line model of alectinib resistance, and analyzed resistant tumor specimens from patients who had relapsed on alectinib. Cell lines were also established under an IRB-approved protocol when there was sufficient fresh tumor tissue. We established Ba/F3 cells expressing EML4-ALK and performed ENU mutagenesis to compare potential crizotinib or alectinib-resistance mutations. In addition, we developed Ba/F3 models harboring ALK resistance mutations and evaluated the potency of multiple next-generation ALK-TKIs including 3rd generation ALK inhibitor in these models and in vivo. To elucidate structure-activity-relationships of ALK resistance mutations, we performed computational thermodynamic simulation with MP-CAFEE.
Results: We identified multiple resistance mutations, including ALK I1171N, I1171S, and V1180L, from the ENU mutagenesis screen and the cell line model. In addition we found secondary mutations at the I1171 residue from the Japanese patients who developed resistance to alectinib or crizotinib. Both ALK mutations (V1180L and I1171 mutations) conferred resistance to alectinib as well as to crizotinib, but were sensitive to ceritinib and other next-generation ALK-TKIs. Based on thermodynamics simulation, each resistance mutation is predicted to lead to distinct structural alterations that decrease the binding affinity of ALK-TKIs for ALK.
Conclusions: We have identified multiple alectinib-resistance mutations from the cell line model, patient derived cell lines, and tumor tissues, and ENU mutagenesis. ALK secondary mutations arising after alectinib exposure are sensitive to other next generation ALK-TKIs. These findings suggest a potential role for sequential therapy with multiple next-generation ALK-TKIs in patients with advanced, ALK-rearranged cancers.
Session Title: Mechanisms of Resistance: From Signaling Pathways to Stem Cells
Session Type: Major Symposium
Session Start/End Time: Tuesday, Apr 21, 2015, 10:30 AM -12:30 PM
Location: Terrace Ballroom II-III (400 Level), Pennsylvania Convention Center
CME: CME-Designated
CME/CE Hours: 2
Session Description: Even the most effective cancer therapies are limited due to the development of one or more resistance mechanisms. Acquired resistance to targeted therapies can, in some cases, be attributed to the selective propagation of a small population of intrinsically resistant cells. However, there is also evidence that cancer drugs themselves can drive resistance by triggering the biochemical- or genetic-reprogramming of cells within the tumor or its microenvironment. Therefore, understanding drug resistance at the molecular and biological levels may enable the selection of specific drug combinations to counteract these adaptive responses. This symposium will explore some of the recent advances addressing the molecular basis of cancer cell drug resistance. We will address how tumor cell signaling pathways become rewired to facilitate tumor cell survival in the face of some of our most promising cancer drugs. Another topic to be discussed involves how drugs select for or induce the reprogramming of tumor cells toward a stem-like, drug resistant fate. By targeting the molecular driver(s) of rewired signaling pathways and/or cancer stemness it may be possible to select drug combinations that prevent the reprogramming of tumors and thereby delay or eliminate the onset of drug resistance.
Tuesday, Apr 21, 2015, 10:30 AM -12:30 PM
David A. Cheresh. UCSD Moores Cancer Center, La Jolla, CA
Tuesday, Apr 21, 2015, 10:30 AM -10:40 AM
Resistance to tyrosine kinase inhibitors: Heterogeneity and therapeutic strategies.
Tuesday, Apr 21, 2015, 10:40 AM -10:55 AM
Jeffrey A. Engelman. Massachusetts General Hospital, Boston, MA
Tuesday, Apr 21, 2015, 10:55 AM -11:00 AM
NG04: Clinical acquired resistance to RAF inhibitor combinations in BRAF mutant colorectal cancer through MAPK pathway alterations
Tuesday, Apr 21, 2015, 11:00 AM -11:15 AM
Ryan B. Corcoran, Leanne G. Ahronian, Eliezer Van Allen, Erin M. Coffee, Nikhil Wagle, Eunice L. Kwak, Jason E. Faris, A. John Iafrate, Levi A. Garraway, Jeffrey A. Engelman. Massachusetts General Hospital Cancer Center, Boston, MA, Dana-Farber Cancer Institute, Boston, MA
Tuesday, Apr 21, 2015, 11:15 AM -11:20 AM
SY27-02: Tumour heterogeneity and therapy resistance in melanoma
Tuesday, Apr 21, 2015, 11:20 AM -11:35 AM
Claudia Wellbrock. Univ. of Manchester, Manchester, United Kingdom

Presentation Number: SY27-02
Presentation Title: Tumour heterogeneity and therapy resistance in melanoma
Presentation Time: Tuesday, Apr 21, 2015, 11:20 AM -11:35 AM
Location: Terrace Ballroom II-III (400 Level), Pennsylvania Convention Center
Author Block: Claudia Wellbrock. Univ. of Manchester, Manchester, United Kingdom
Abstract Body: Solid tumors are structurally very complex; they consist of heterogeneous cancer cell populations, other non-cancerous cell types and a distinct extracellular matrix. Interactions of cancer cells with non-cancerous cells is well investigated, and our recent work in melanoma has demonstrated that the cellular environment that surrounds cancer cells has a major impact on the way a patient responds to MAP-kinase pathway targeting therapy.
We have shown that intra-tumor signaling within a heterogeneous tumor can have a major impact on the efficacy of BRAF and MEK inhibitors. With the increasing evidence of genetic and phenotypic heterogeneity within tumors, intra-tumor signaling between individual cancer-cell subpopulations is therefore a crucial factor that needs to be considered in future therapy approaches. Our work has identified the ‘melanocyte-lineage survival oncogene’ MITF as an important player in phenotypic heterogeneity (MITFhigh and MITFlow cells) in melanoma, and MITF expression levels are crucial for the response to MAP-kinase pathway targeted therapy. We found that ‘MITF heterogeneity’ can be caused by cell-autonomous mechanisms or by the microenvironment, including the immune-microenvironment.
We have identified various mechanisms underlying MITF action in resistance to BRAF and MEK inhibitors in melanoma. In MITFhigh expressing cells, MITF confers cell-autonomous resistance to MAP-kinase pathway targeted therapy. Moreover, it appears that in melanomas heterogeneous for MITF expression (MITFhigh and MITFlow cells), individual subpopulations of resistant and sensitive cells communicate and MITF can contribute to overall tumor-resistance through intra-tumor signaling. Finally, we have identified a novel approach of interfering with MITF action, which profoundly sensitizes melanoma to MAP-kinase pathway targeted therapy.
Tuesday, Apr 21, 2015, 11:35 AM -11:40 AM
SY27-03: Breast cancer stem cell state transitions mediate therapeutic resistance
Tuesday, Apr 21, 2015, 11:40 AM -11:55 AM
Max S. Wicha. University of Michigan, Comprehensive Cancer Center, Ann Arbor, MI
Tuesday, Apr 21, 2015, 11:55 AM -12:00 PM
SY27-04: Induction of cancer stemness and drug resistance by EGFR blockade
Tuesday, Apr 21, 2015, 12:00 PM -12:15 PM
David A. Cheresh. UCSD Moores Cancer Center, La Jolla, CA


Cellular Reprogramming in Carcinogenesis: Implications for Tumor Heterogeneity, Prognosis, and Therapy
Session Type: Major Symposium
Session Start/End Time: Tuesday, Apr 21, 2015, 10:30 AM -12:30 PM
Location: Room 103, Pennsylvania Convention Center
CME: CME-Designated
CME/CE Hours: 2
Session Description: Cancers, both solid and liquid, consist of phenotypically heterogeneous cell types that make up the full cellular complement of disease. Deep sequencing of bulk cancers also frequently reveals a genetic intratumoral heterogeneity that reflects clonal evolution in space and in time and under the influence of treatment. How the distinct phenotypic and genotypic cells contribute to individual cancer growth and progression is incompletely understood. In this symposium, we will discuss issues of cancer heterogeneity and effects on growth and treatment resistance, with emphasis on cancer cell functional properties and influences of the microenvironment, interclonal genomic heterogeneity, and lineage relationships between cancer cells with stem cell and differentiated properties. Understanding these complex cellular relationships within cancers will have critical implications for devising more effective treatments.
Tuesday, Apr 21, 2015, 10:30 AM -12:30 PM
Peter B. Dirks. Univ. of Toronto Hospital for Sick Children, Toronto, ON, Canada

Tuesday, Apr 21, 2015, 10:30 AM -10:40 AM

Origins, evolution and selection in childhood leukaemia
Tuesday, Apr 21, 2015, 10:40 AM -11:00 AM
Tariq Enver. Cancer Research UK, London, United Kingdom

Tuesday, Apr 21, 2015, 11:00 AM -11:05 AM

Cytokine-controlled stem cell plasticity inintestinal tumorigenesis
Tuesday, Apr 21, 2015, 11:05 AM -11:25 AM
Florian Greten. Georg-Speyer-Haus, Frankfurt, Germany

Tuesday, Apr 21, 2015, 11:25 AM -11:30 AM

SY23-03: Intratumoural heterogeneity in human serous ovarian carcinoma
Tuesday, Apr 21, 2015, 11:30 AM -11:50 AM
John P. Stingl. Cancer Research UK Cambridge Research Inst., Cambridge, United Kingdom

Tuesday, Apr 21, 2015, 11:50 AM -11:55 AM

Functional and genomic heterogeneity in brain tumors
Tuesday, Apr 21, 2015, 11:55 AM -12:15 PM


Proc Natl Acad Sci U S A. 2015 Jan 20;112(3):851-6. doi: 10.1073/pnas.1320611111. Epub 2015 Jan 5.

Single cell-derived clonal analysis of human glioblastoma links functional and genomic heterogeneity.

Meyer M1, Reimand J2, Lan X3, Head R1, Zhu X1, Kushida M1, Bayani J4, Pressey JC5, Lionel AC6, Clarke ID7, Cusimano M8, Squire JA9, Scherer SW6, Bernstein M10, Woodin MA5, Bader GD11, Dirks PB12.

Author information


Glioblastoma (GBM) is a cancer comprised of morphologically, genetically, and phenotypically diverse cells. However, an understanding of the functional significance of intratumoral heterogeneity is lacking. We devised a method to isolate and functionally profile tumorigenic clones from patient glioblastoma samples. Individual clones demonstrated unique proliferation and differentiation abilities. Importantly, naïve patient tumors included clones that were temozolomide resistant, indicating that resistance to conventional GBM therapy can preexist in untreated tumors at a clonal level. Further, candidate therapies for resistant clones were detected with clone-specific drug screening. Genomic analyses revealed genes and pathways that associate with specific functional behavior of single clones. Our results suggest that functional clonal profiling used to identify tumorigenic and drug-resistant tumor clones will lead to the discovery of new GBM clone-specific treatment strategies.



739: Tumor cell plasticity with transition to a mesenchymal phenotype is a mechanism of chemoresistance that is reversed by Notch pathway inhibition in lung adenocarcinoma
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Khaled A. Hassan. University Of Michigan, Ann Arbor, MI

745: Oncostatin M receptor activation leads to molecular targeted therapy resistance in non-small cell lung cancer
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Kazuhiko Shien1, Vassiliki A. Papadimitrakopoulou1, Dennis Ruder1, Nana E. Hanson1, Neda Kalhor1, J. Jack Lee1, Waun Ki Hong1, Ximing Tang1, Roy S. Herbst2, Luc Girard3, John D. Minna3, Jonathan M. Kurie1, Ignacio I. Wistuba1, Julie G. Izzo1. 1University of Texas MD Anderson Cancer Center, Houston, TX; 2Yale Cancer Center, Yale School of Medicine, New Haven, CT; 3Hamon Center for Therapeutic Oncology Research, University of Texas Southwestern Medical Center, Dallas, TX

746: Activation of EGFR bypass signaling through TGFα overexpression induces acquired resistance to alectinib in ALK-translocated lung cancer cells
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Tetsuo Tani, Hiroyuki Yasuda, Junko Hamamoto, Aoi Kuroda, Daisuke Arai, Kota Ishioka, Keiko Ohgino, Ichiro Kawada, Katsuhiko Naoki, Hayashi Yuichiro, Tomoko Betsuyaku, Kenzo Soejima. Keio University, Tokyo, Japan

752: Elucidating the mechanisms of acquired resistance in lung adenocarcinomas
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Sandra Ortiz-Cuarán1, Lynnette Fernandez-Cuesta1, Christine M. Lovly2, Marc Bos1, Matthias Scheffler3, Sebastian Michels3, Kerstin Albus4, Lydia Meyer4, Katharina König4, Ilona Dahmen1, Christian Mueller1, Luca Ozretić4, Lars Tharun4, Philipp Schaub1, Alexandra Florin4, Berit Pinther1, Nike Bahlmann1, Sascha Ansén3, Martin Peifer1, Lukas C. Heukamp4, Reinhard Buettner4, Martin L. Sos1, Jürgen Wolf3, William Pao2, Roman K. Thomas1. 1University of Cologne, Cologne, Germany; 2Department of Medicine, Vanderbilt University, Nashville, TN; 3Department of Internal Medicine, Center for Integrated Oncology Köln-Bonn, University Hospital Cologne, Cologne, Germany; 4Institute of Pathology, University Hospital Cologne, Cologne, Germany

760: On the evolution of erlotinib-resistant NSCLC subpopulations
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Michael E. Ramirez1, Robert J. Steininger, III1, Lani F. Wu2, Steven J. Altschuler2. 1UT Southwestern, Dallas, TX; 2UCSF, San Francisco, CA
763: Implications of resistance patterns with NSCLC targeted agents
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
David J. Stewart, Paul Wheatley-Price, Rob MacRae, Jason Pantarotto. University of Ottawa, Ottawa, ON, Canada


768: A kinome-wide siRNA screen identifies modifiers of sensitivity to the EGFR T790M-targeted tyrosine kinase inhibitor (TKI), AZD9291, in EGFR mutant lung adenocarcinoma
Sunday, Apr 19, 2015, 1:00 PM – 5:00 PM
Eiki Ichihara1, Joshua A. Bauer2, Pengcheng Lu3, Fei Ye3, Darren Cross4, William Pao1, Christine M. Lovly1. 1Vanderbilt University School of Medicine, Nashville, TN; 2Vanderbilt Institute of Chemical Biology High-Throughput Screening Facility, Nashville, TN; 3Vanderbilt University Medical Center, Nashville, TN; 4AstraZeneca Oncology Innovative Medicines, United Kingdom

LB-055: Clinical acquired resistance to RAF inhibitor combinations in BRAF-mutant colorectal cancer through MAPK pathway alterations
Sunday, Apr 19, 2015, 4:35 PM – 4:50 PM
Leanne G. Ahronian1, Erin M. Sennott1, Eliezer M. Van Allen2, Nikhil Wagle2, Eunice L. Kwak1, Jason E. Faris1, Jason T. Godfrey1, Koki Nishimura1, Kerry D. Lynch3, Craig H. Mermel1, Elizabeth L. Lockerman1, Anuj Kalsy1, Joseph M. Gurski, Jr.1, Samira Bahl4, Kristin Anderka4, Lisa M. Green4, Niall J. Lennon4, Tiffany G. Huynh3, Mari Mino-Kenudson3, Gad Getz1, Dora Dias-Santagata3, A. John Iafrate3, Jeffrey A. Engelman1, Levi A. Garraway2, Ryan B. Corcoran1. 1Massachusetts General Hospital Cancer Center, Boston, MA; 2Dana Farber Cancer Institute, Boston, MA; 3Massachusetts General Hospital Department of Pathology, Boston, MA; 4Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, MA


Other Articles on this Site Related to Tumor Heterogeneity Include

Notes On Tumor Heterogeneity: Targets and Mechanisms, from the 2015 AACR Meeting in Philadelphia PA

Issues in Personalized Medicine: Discussions of Intratumor Heterogeneity from the Oncology Pharma forum on LinkedIn

Issues in Personalized Medicine in Cancer: Intratumor Heterogeneity and Branched Evolution Revealed by Multiregion Sequencing

CANCER COMPLEXITY: Heterogeneity in Tumor Progression and Drug Response – 2015 Annual Symposium @Koch Institute for Integrative Cancer Research at MIT – W34, 6/12/2015 9:00 AM EDT – 4:30 PM EDT

In vitro Models of Tumor Microenvironment for New Cancer Target and Drug Discovery, 11/17 – 11/19/2014, Hyatt Boston Harbor

What can we expect of tumor therapeutic response?


Read Full Post »

Viruses and Cancer: A Walk on the Memory Lane

Curator: Demet Sag, PhD, CRA, GCP


One of the other mechanism where cancer and microorganisms establish a close relationship is viruses. They are vicious sometimes as they adept fast even we don’t call them a real organism since they require a living cell to survive. Vaccination against these viruses or using them as a tool to deliver genes to cure certain human diseases also become very attractive. They come various shapes, sizes, and content.

At first the discoveries of human viral cancers was done by tedious viral technology but later for the last four human cancer viruses molecular biology techniques used.

It was in 2011 Francis Peyton Rous’s landmark experiments on an avian cancer virus the connection between viruses and cancer is established yet we discover new ones. Currently we believe that about 10-215% cancers originated from viruses.

They were very interesting due to their dual actions through infections or non-infectious cancer causes with their effects on immune system, innate immunity, and tumor suppressor proteins.

Since their discoveries it was also identified that 20 % or one in five cancer cases born as a result of viral infections. Therefore, in the world now two of them have widely used vaccines, hepatitis B virus (HPV) and human papilloma virus (HPV). On the other hand, one may wonder what their efficacy is.

Of course these discoveries came with the highest recognitions:

Nobel Prizes awarded for the discoveries of viruses in timeline.

The origin of cancer viruses and cancer sometimes bring a misconception. For a virus tumors are dead end since they can’t replicate and invade the organisms unlike many thought that viruses infect the host to increase their replication. Thus, most of time only in very rare occasions they transmit to another human so the big fat truth is most if the human tumor viruses are asymptomatic. Even if they can be very mildly symptomatic, they don’t make neoplasia.

On the other hand, the question is why and how the viruses make oncogenes and why they initiate tumorogenecity begs the question. Of course, there is an evolution but also they have a common functional targets in the human genome. Like viruses human genome has various replicating sequences or inversions. When these viruses expressing oncoproteins they mainly target the RB1 and p53.  In addition, these tumor targets attack telomerase reverse transcriptase (TERT), cytoplasmic PI3K–AKT–mTOR, nuclear factor-κB (NF-κB), β-catenin (also known as CTNNB1) and interferon signaling pathways.

Thus immunity and inflammation reactions present different pathways against the virulent action and initiation of tumor forming for cancer.

1966  Nobel Prize awarded to Rous

Tumorigenic retroviruses have been central to cancer biology, leading to the development of focus formation assays, discovery of reverse transcription, identification of more than 20 cellular oncogenes, and ultimately Nobel Prize recognition for Rous 57 years after his initial experiments. Then these discoveries led to discoveries of oncogenes and tumor suppressor genes.


1975 Nobel Prize awarded to Temin, Baltimore, and Dulbeco


1976 Nobel Prize awarded to Blumberg

HBV, discovered shortly after EBV in the mid-1960s and leading to a Nobel Prize for Baruch Blumberg in 1976, has only recently been successfully propagated in culture and was first linked by serology to acute hepatitis rather than to cancer25,26. The role of HBV in hepatocellular carcinoma was established more than a decade later by Beasley et al.27 through longitudinal studies of Taiwanese insurance company cohorts.


1989 Nobel Prize awarded to Bishop and Varmus


2008 Nobel Prize awarded to Harald zur Hausen, François Barré-Sinoussi and Luc Montagnier.


Nobel Prizes awarded in 2008 for the discovery by Harald zur Hausen of high-risk HPV strains that cause cervical cancer and the discovery of HIV, an agent that does not initiate cancer but indirectly ‘sets the stage’ for malignancy through immuno suppression, by François Barré-Sinoussi and Luc Montagnier.

Furthermore, human cancer viruses span the entire range of virology and include:

  • complex exogenous retroviruses
    • such as HTLV-I,
  • positive-stranded RNA viruses
    • such as hepatitis C virus (HCV),
  • DNA viruses with retroviral features
    • such as HBV
  • both large double-stranded DNA viruses :
    • such as EBV and
    • Kaposi’s sarcoma herpesvirus

(KSHV; also known as human herpesvirus 8 (HHV8))

  • small double-stranded DNA viruses
    • HPV and
    • Merkel cell polyomavirus (MCV)).



The human cancer viruses:

Virus Genome Notable cancers Year first
Epstein–Barr virus (EBV; also
known as human herpesvirus 4
Double-stranded DNA herpesvirus Most Burkitt’s lymphoma and nasopharyngeal
carcinoma, most lymphoproliferative disorders,
some Hodgkin’s disease, some non-Hodgkin’s
lymphoma and some gastrointestinal lymphoma


Epstein MA, Achong BG, Barr YM. Virus particles in cultured lymphoblasts from Burkitt’s lymphoma. Lancet. 1964;15:702–703.

Hepatitis B virus (HBV) Single-stranded and
double-stranded DNA
Some hepatocellular carcinoma 1965


Blumberg BS, Alter HJ, Visnich S. A “new” antigen in leukemia sera. JAMA. 1965;191:541–546.

Human T-lymphotropic virus-I
Positive-strand, single-stranded RNA
Adult T cell leukaemia 1980


Poiesz BJ, et al. Detection and isolation of type C retrovirus particles from fresh and cultured lymphocytes of a patient with cutaneous T-cell lymphoma. Proc. Natl Acad. Sci. USA. 1980;77:7415–7419.

High-risk human papillomaviruses
(HPV) 16 and HPV 18 (some other
α-HPV types are also carcinogens)
Double-stranded DNA
Most cervical cancer and penile cancers and some
other anogenital and head and neck cancers


Durst M, Gissmann L, Ikenberg H, zur Hausen H. A papillomavirus DNA from a cervical carcinoma and its prevalence in cancer biopsy samples from different geographic regions. Proc. Natl Acad. Sci. USA. 1983;80:3812–3815.


Boshart M, et al. A new type of papillomavirus DNA, its presence in genital cancer biopsies and in cell lines derived from cervical cancer. EMBO J. 1984;3:1151–1157.

Hepatitis C virus (HCV) Positive-strand, single-stranded
RNA flavivirus
Some hepatocellular carcinoma and some


Choo QL, et al. Isolation of a cDNA clone derived from a blood-borne non-A, non-B viral hepatitis genome. Science. 1989;244:359–362.

Kaposi’s sarcoma herpesvirus
(KSHV; also known as human
herpesvirus 8 (HHV8))
Double-stranded DNA herpesvirus Kaposi’s sarcoma, primary effusion lymphoma and
some multicentric Castleman’s disease


Chang Y, et al. Identification of herpesvirus-like DNA sequences in AIDS-associated Kaposi’s sarcoma. Science. 1994;265:1865–1869.

Merkel cell polyomavirus (MCV) Double-stranded DNA polyomavirus Most Merkel cell carcinoma 2008


Feng H, Shuda M, Chang Y, Moore PS. Clonal integration of a polyomavirus in human Merkel cell carcinoma. Science. 2008;319:1096–1100.

Common cellular targets for unrelated tumour virus oncoproteins

An incomplete but diverse list of animal and human tumour virus proteins that target RB1, p53, interferon and PI3K–mTOR signalling pathways. Most of these viral proteins are evolutionarily distinct from each other and have unique mechanisms for regulating or ablating these signalling pathways. Convergent evolution of tumour viruses to target these (and other cellular signalling pathways (not shown), including interleukin-6 (IL-6)–signal transducer and activator of transcription 3 signalling, telomerase and nuclear factor-κB (NF-κB) signalling pathways) reveals commonalities among the cancer viruses in tumour supressor and oncoprotein targeting. CBP, cAMP-response element binding protein; CDKI, cyclin-dependent kinase inhibitor; EBV, Epstein–Barr virus; HCV, hepatitis C virus; HPV, human papillomavirus; HTLV, human T-lymphotropic virus; IFNR, interferon receptor; IRF, interferon regulatory factor; KSHV, Kaposi’s sarcoma herpesvirus; LMP, latent membrane protein; miRNA, microRNA. Nat Rev Cancer. Author manuscript; available in PMC 2013 Jul 22.

Two views for the origins of viral oncoproteins

a | The tumour virus proteins target RB1 and p53 to drive a quiescent G0 cell into S phase of the cell cycle, allowing viral access to the nucleotide pools and replication machinery that are needed for replication and transmission100. Viral tumourigenesis is a by-product of the molecular parasitism by viruses to promote their own replication. Cells respond to virus infection by activating RB1 and p53 to inhibit virus replication as part of the innate immune response86. To survive, tumour viruses have evolved the means for inactivating these and other immune signalling pathways that place the cell at risk for cancerous transformation. This view holds that many tumour suppressor proteins have dual functions in preventing cancer formation and virus infection. b | An illustration of the overlap between intracellular innate immune and tumour suppressor signalling. Under typical circumstances, viruses do not cause cancers except in the settings of immunosuppression and/or complementing host cell mutations. Non-tumorigenic viruses, which constitute the overwhelming majority of viruses, target many of the same innate immune and tumour suppressor pathways as tumour viruses but do so in ways that do not place the host at risk for carcinogenesis. Apart from p53, RB1 and p300, additional proteins are likely to have both tumour suppressor and innate immune functions.

The molecular evolution of a human tumour virus

Merkel cell polyomavirus (MCV), which has tumour-specific truncation mutations, illustrates common features among the human tumour viruses involving immunity, virus replication and tumour suppressor targeting. Although MCV is a common infection, loss of immune surveillance through ageing, AIDS or transplantation and subsequent treatment with immunosuppressive drugs may lead to resurgent MCV replication in skin cells161. If a rare integration mutation into the host cell genome occurs34, the MCV T antigen can activate independent DNA replication from the integrated viral origin that will cause DNA strand breaks in the proto-tumour cell157. A second mutation that truncates the T antigen, eliminating its viral replication functions but sparing its RB1 tumour suppressor targeting domains, is required for the survival of the nascent Merkel tumour cell. Exposure to sunlight (possibly ultraviolet (UV) irradiation) and other environmental mutagens may enhance the sequential mutation events that turn this asymptomatic viral infection into a cancer virus.


Antibody panning cDNA from a tumour is used to express proteins in bacteria and transferred to replicate filters. Antibody screening of the filters can then be used to identify colonies expressing the specific cDNA encoding an antigen.
Bayesian reasoning A scientific approach developed from Bayes theorem, combining features of the Logical Positivist and Kuhnian schools of science philosophy, and describing how the probability of a hypothesis (in this case, virus A causes cancer B) changes with new evidence. In simple terms, it can be described as the repeated application of the scientific method to falsify a hypothesis such that the hypothesis has a high probability of being either true or false.
Digital transcriptome subtraction DTS. Method to discover new viruses by exhaustively sequencing cDNA libraries and aligning known human sequences by computer leaving a smaller candidate pool of potential viral sequences for analysis36.
Endogenous retrovirus ERV. Retrovirus that has inserted into the metazoan germline genome over evolutionary timescales and is now transmitted to offspring as a genetic element through Mendelian inheritance. Approximately 8% of the human genome is estimated to be derived from retroviral precursors.
High-risk papillomaviruses More than 160 different genotypes or strains of HPV have been described but only a few genotypes belonging to a high-risk carcinogenic clade of the α-HPV genus are responsible for invasive HPV-related anogenital cancers211.
Longitudinal study Virus infection is measured initially in a cohort of patients who are then followed over time to determine cancer occurrence.
Prodromal phase An early set of nonspecific symptoms that occur before the onset of specific disease symptoms.
Representational difference analysis A PCR-based subtractive hybridization technique that can subtract common human sequences from a tumour genomic library using a control human tissue genomic library35.
Serology The measurement of antibodies against viruses in blood or bodily fluids. This usually does not distinguish ongoing infections from past viral infections.

Read Full Post »

Science Teaching in Math and Technology

Larry H. Bernstein, MD, FCAP, Curator


2015 Best High Schools for STEM Rankings Methodology

U.S. News looked at 500 public high schools to identify the best in math and science education.

By Robert Morse May 11, 2015

U.S. News & World Report’s Best High Schools for STEM rankings methodology is based on the key principle that students at the Best High Schools for STEM must participate in and pass a robust curriculum of college-level math and science courses. STEM stands for science, technology, engineering and math.

To be included in the U.S. News Best High Schools for STEM rankings, a public high school first had to be listed as a gold medal winner in the 2015 U.S. News Best High Schools rankings. That meant that the top 500 nationally ranked high schools were eligible for the STEM rankings.

Those eligible schools were next judged nationally on their level of math and science participation and success, using Advanced Placement STEM test data for 2013 graduates as the benchmark to conduct the analysis. The U.S. News Best High Schools for STEM rankings methodology does not rely on any data from the U.S. Department of Education.

AP is a College Board program that offers college-level courses at high schools across the country. College Board defines STEM Math as AP courses in Calculus AB, Calculus BC, Computer Science A and Statistics; and STEM Science as AP courses in Biology, Chemistry, Environmental Science, Physics B, Physics C: Electricity and Magnetism and Physics C: Mechanics.

Math and science success at the high school level was assessed by computing a STEM Achievement Index for each school that ranked in the top 500 of the 2015 Best High Schools. The index was based on the percentage of all the AP test-takers in a school’s 2013 graduating class who took and passed college-level AP STEM Math and AP STEM Science tests. The higher a high school scored on the STEM Achievement Index, the better it placed in the Best High Schools for STEM rankings.

The maximum STEM Achievement Index value is 100. No public high school evaluated achieved that top score. The highest index was 98.3.

The first step in the rankings process was to compute the STEM Math Achievement Index. It was derived from two variables. The first was the percentage of AP test-takers in the 2013 graduating class who took at least one AP STEM Math course during high school, which was weighted 25 percent. The second was the percentage of those AP STEM Math test-takers who passed at least one AP STEM Math test during high school, receiving an AP score of 3 or higher. This variable was weighted 75 percent.

The next step was to calculate a STEM Science Achievement Index. Much like the math index, it was derived from the percentage of AP test-takers in the 2013 graduating class who took at least one AP STEM Science course during high school – weighted 25 percent – and the percentage of those AP STEM Science test-takers who passed at least one AP STEM Science test during high school, receiving an AP score of 3 or higher – weighted 75 percent.

This means that the methodology weights students taking AP math and science STEM courses at the high school level at 25 percent and passing those same AP STEM courses at 75 percent. In other words, passing both AP math and science tests was three times as important in the rankings as simply taking AP math and science courses.

The final step in the rankings process was to calculate the overall STEM Achievement Index, a combination of the STEM Math Achievement Index and the STEM Science Achievement Index. Each index was weighted at 50 percent, and then added together to create a composite value that is the STEM Achievement Index score.

The STEM rankings were based on sorting the unrounded – to many decimal places – STEM Achievement Index in descending order, with the top-ranked schools having the highest index values. The STEM Achievement Index was then rounded to the nearest 10th for online publication.

The top 250 high schools that achieved a value of greater than or equal to 66.8 in their STEM Achievement Index scored high enough to be numerically ranked. That high index cutoff point was used since it meant that all the high schools in the STEM rankings had, on average, more than two-thirds of the AP test-takers in their 2013 graduating class take and pass at least one AP STEM Math and one AP STEM Science test.

AP® and Advanced Placement® are registered trademarks of the College Board. Used with permission.

Top 50 Science Teacher Blogs

Bringing the subject of science to life for students is the challenge shared by the teachers who author these 50 amazing and insightful science education blogs. Sharing narratives set within and beyond the classroom walls, these next generation educators embrace technology but are never so dazzled by it that they lose sight of their common goal.

Physics Teacher Frank Noschese discusses science education topics like whether Khan Academy is effective at teaching physics, applying Angry Birds in physics lessons, and the idea of pseudoteaching.

Teaching High School Psychology
Teaching High School Psychology is a joint collaboration that explores the deeper lesson of the Stanford Prison Experiment, Gamification and its implications as a behavioral motivator, and opportunities for teaching Operant Conditioning with TV’s Big Bang Theory.

Little Miss Hypothesis
Inspiring Kindergarten scientists and giving a too-often neglected subject its due is the aim of Little Miss Hypothesis, where Mrs. Coe chronicles activities with growing brains, harvesting Spirit Garden salads and the development of science centers in a classroom that is home to Bluebonnet the Betta fish and the crab shack’s resident hermits.

Science for Kids
Sue Cahalane shares her passion for teaching science to elementary students in grades PK-4 on Science for Kids with ideas for classroom experiments, tutorials for science lessons, updates on science education news, and photos of students engaged in science activities.

Science Education on the Edge
Chris Ludwig, a high school science teacher from Colorado, writes about improving assessment and instruction in science and education technology.

Teach Science for All
Kirk Robbins shares helpful resources and tools for science teachers including reports, useful websites, and online tools.

Teaching | chemistry
Ellena Bethea, a high school chemistry teacher, writes about grading practices, online tools, and lab activities.

Adventures with the Lower Level
Tracie Schroeder shares her experiences teaching science, teaching methods, and thoughts on learning.

Physics in Flux
Dan Fullerton provides a resource for teachers with details of his successes and failures, technology guides, and physics book reviews.

Think Thank Thunk
On his blog, Think Thank Thunk, Shawn Cornally celebrates the Merlin within every teacher, the need for repackaging education, the debate surrounding Standards Based Grading and the dread of being dull as he chronicles his plight as an educator.

Marine biology teacher Sean Nash gets inspired by WordFoto and invites educators to appreciate and aim for “Whoa” moments on his blog, nashworld.

Science Teacher
A science teacher and former pediatrican finds an exemplary model in Dr. Seuss, challenges technophiles to understand deeply, and explains why he has made a tradition of culminating each school year with a field trip to watch horseshoe crabs in the throes of romance.

Teach Science
At Teach Science, Ed Hitchcock muses on the DNA shared by Socrates and explains why science’s greatest appeal is the unexpected.

Quantum Progress
At Quantum Progress, 9th grade Atlanta physics teacher John Burk relives a childhood tradition at Physics Teacher Camp, promotes blogging as a tool for professional development, and ponders why physics buildings never win campus beauty contests.

Pedagogue Padawan
At Pedagogue Padawan, Geoff Schmit shares innovative ideas for using Sudoku to teach Circuit Analysis, Angry Birds as a lesson in holography, and wikispaces as a tool for science projects.

Re:thinking blends personal reflection with a challenge to rethink school culture and policy as 9th grade teacher Ben Wildeboer finds teachable moments in events like the Japanese quake and explains the importance of “hard fun” for students.

Journey in Technology
At Journey in Technology a Dallas Physics teacher discusses implementing Khan Academy, discovering community and deep connections at Educon, and transforming the pseudoteaching of “cookbook” lab projects into real learning in the classroom.

Always Formative
Jason Buell is a middle school science teacher from California who writes about standards-based grading, education conferences, education books and more.

Stretching Forward
At Stretching Forward, Earth science teacher Janelle Wilson shares experiences from the Space Academy for Educators, discusses class blogging, and shares thoughts on engaging students and parents in science.

Tearing Down Walls
Derrick Willard teaches AP Environmental Science and discusses using social media and online tools to extend lessons outside the walls of the physical classroom.

Teaching Computer Science
Alfred Thompson is a former high school computer science teacher who currently works at Microsoft and writes about computer science education and resources.

A+ Computer Science Blog
High school computer science teacher Stacey Armstrong discusses why computer science is cool, game programming, career options in computer science, and computer science resources.

Teaching CS in Dallas
Kathleen Weaver writes about teaching robotics, Android development, and computer science education topics on her blog.

In Need of a Base Case
This blog discusses the need for change in computer science education, computer science project ideas, and the value of learning computer science.

Hélène Martin
Hélène Martin muses on the power of childhood playthings to fuel tech career ambitions and describes how lost airport luggage is a reminder to look for ways to leverage computing to solve real-life problems in this blog from the perspective of a computer science teacher.

Garth’s CS Education Blog
A computer science and programming teacher at a private school writes about teaching fun and important concepts and preparing students for computer science careers.

The Blog of Phyz
The Blog of Phyz is California teacher Dean Baird’s platform for debunking “Magnet Boys” and magic wristbands, and touting a 75 cent investment guaranteed to wow even the most cynical student.

Mr. Gonzalez’s Classroom
An Olympic Odyssey customarily culminates the academic year for middle school teacher Alfonso Gonzalez, who explores the challenge of giving terms like “on-task” and “structured learning” 21st century relevance on his blog, Mr. Gonzalez’s Classroom.

Free/Libre Open Source Science Education
Pseudoteaching and trends like the “reverse lecture” are hot topics on Free/Libre Open Source Science Education, where Steve Dickie shares his own innovative methods, including cartooning with GoAnimate and creating his own textbooks.

The Science Classroom
Oklahoma physics teacher Jody Bowie reports on the thrill of seeing students connect classroom lessons in everyday life, explains why everyone needs a whetstone to hone their thinking and divulges his identification with the Wizard in Wicked on The Science Classroom blog.

Jacobs Physics
On his blog Greg Jacobs calls course evaluations brutal but vital and bucks a few trends by advocating daily work and disparaging summer assignments in favor of starting each year “from the ground up”.

New Physics Modeler
Bryan Battaglia explains the appeal of professional conferences, the career changing power of blogging, and reflects that teachers gain as many lessons by year’s end as their students.

Just Call Me Ms Frizzle
Becky offers a distinctive first-year teacher perspective on Just Call Me Ms Frizzle, contrasting the low of leaving the room in frustration with the high of a Friday classroom on its best behavior, along with the challenge of teaching a non-traditional class.

And Yet it Moves
On his blog, And Yet it Moves, Ben Chun explains why problem-solving skills trump smarts, tackles the debate over doing away with honors classes, and challenges the AP curriculum.

Reflections of a Science Teacher
Sandra McCarron dismisses the notion of a rubric for thinking, believes that a successful classroom starts out with a vision and ponders the merits of science fairs that have been sacrificed to make way for education reforms on her blog, Reflections of a Science Teacher.

Hurricane Maine
A veteran teacher discusses ideas in education and technology, interesting articles, and how to make school more like play rather than work.

The Physics of Learning
Doug Smith authors this physics education blog that discusses topics like whether to use iPads in the classroom, the myths of merit pay, and scientific literacy.

Room 611
Mr. Young teaches Earth science and other subjects in Canada and provides insights into class by outlining what is covered in class almost every school day.

Using Blogs in Science Education
Stacey Baker is a high school biology teacher and writes about how to use classroom blogs to help students learn science.

Physics! Blog!
Physics! Blog! shares results of The No Homework Experiment and discusses standards based grading, the goal of testing, and teaching students how to learn from mistakes.

Ideas for Teaching Computer Technology to Kids
A blog sharing ideas and resources for teaching computer technology including robots for computer science education, programming resources, and computer science teaching tools.
A physics teacher shares interesting science articles like Nobel prize winning sentences, things from movies that cannot exist, and cool science videos.

Teach. Brian. Teach.
Brian discusses what makes for a good science conversation, reflects on teaching, shares observations of students, and explains why it is important to point out when students are having fun doing science.

The Skeptical Teacher
A high school physics teacher discusses science education and promotes critical thinking on his blog.

Physics & Physical Science Demos, Labs, & Projects for High School Teachers
A physics teacher provides a resource for science teachers to share ideas for labs and demonstrations and commentary on what works.

The Art of Teaching Science
Jack Hassard is a professor of science education and explores issues in teaching science, shares resources for science teachers, and discusses why teaching science is important.

SuperFly Physics
At SuperFly Physics, Andy Rundquist shares ideas for teaching physics, fun science experiments, and interesting physics problems.

Newton’s Minions
A physics blog sharing student work, anecdotes from the classroom, thoughts on student assessment, and ideas for teaching complex physics lessons.

Mr. Barlow’s Blog
Mr. Barlow is a high school science teacher and podcaster from Melbourne, Australia who shares interesting science studies, cool science news, and optical illusions at his blog.

What is JASON?

We are a non-profit organization that connects students, in the classroom and out, to real science and exploration to inspire and motivate them to study and pursue careers in Science, Technology, Engineering and Math (STEM).

We embed exciting STEM professionals and cutting-edge research into award-winning, standards-aligned in and out-of-school curricula. Live webcasts connect students with inspirational STEM role models. Student materials include reading selections with read-to-me functionality, inquiry-based labs, videos, and online games. For teachers and informal educators, we provide lesson plans, assessments, and comprehensive professional development programs.

Ten Websites for Science Teachers

Originally Published: February 7, 2012 | Updated: October 10, 2014

We all know that the web is full of excellent web resources for science teachers and students. However, unless you live on the web, finding the best websites can become quite a challenge. This isn’t a “Top Ten” list — instead, it is a list of websites that I either use on a regular basis or just find interesting. From teaching resources for the nature of science and authentic field journals to wacky videos about numbers, I am sure that you will find something in the following list the works for you!

1) Understanding Science

UC Berkeley’s Understanding Science website is a “must use” for all science teachers. It is a great resource for learning more about the process of science. The resource goes much deeper than the standard “PHEOC” model of the scientific method by emphasizing peer review, the testing of ideas, a science flowchart and “what is science?” checklist. Understanding Science also provides a variety of teaching resources including case studies of scientific discoveries and lesson plans for every grade level.

2) Field Research Journals

The Field Book Project from the National Museum of Natural History and the Smithsonian Institution Archives intends to create a “one stop” archive for field research journals and other documentation. You can find plenty of examples from actual field research journals for your classes.

3) Evolution

Berkeley’s Understanding Evolution website is the precursor to their Understanding Science efforts. The Understanding Evolution website provides a plethora of resources, news items and lessons for teaching about evolution. Lessons provide appropriate “building blocks” to help students at any grade level work towards a deeper understanding of evolution. The Evo 101 tutorial provides a great overview of the science behind evolution and the multiple lines of evidence that support the theory.

4) PhET Simulations

PhET from the University of Colorado provides dozens of fantastic simulations for physics, chemistry and biology. The website also includes a collection of teacher contributed activities, lab experiences, homework assignments and conceptual questions that can be used with the simulations.

5) Earth Exploration

The Earth Exploration Toolbook provides a series of activities, tools and case studies for using data sets with your students.

6) EdHead Interactives

Edheads is an organization that provides engaging web simulations and activities for kids. Current activities focus on simulated surgical procedures, cell phone design (with market research), simple and compound machines, and weather prediction.

7) Plant Mentors

Do you teach about plants? Check out Planting Science to connect your middle or high school students to science mentors and a collaborative inquiry project. From the project:

Planting Science is a learning and research resource, bringing together students, plant scientists, and teachers from across the nation. Students engage in hands-on plant investigations, working with peers and scientist mentors to build collaborations and to improve their understanding of science.

8) Periodic Table of Videos

Check out The Periodic Table of Videos for a wide array of videos about the elements and other chemistry topics.

9) More Videos!

Students can read and watch video about 21 Smithsonian scientistsincluding a volcano watcher, fossil hunter, art scientist, germinator and zoo vet.

10) Even More Videos!

How many videos were watched on YouTube in 2010? If you said 22 billion, you are sort of correct… Those 22 billion views only represent the number of times education videos were watched! In addition to this list of science and math YouTube channels, here are two of my favorites:

  • SciShow is all about teaching scientific concepts in an accessible and easy-to-understand manner. This channel includes a variety of short (3 minute) and long (10 minute) videos. New videos are released weekly.
  • Former BBC journalist Brady Haran is crazy about math and science. If you love numbers, you will love his Numberphile channel, dedicated to exploring the stories behind numbers.
  • And let’s close with a particularly good SciShow on Climate Change:

Best High Schools

 School for the Talented and Gifted


DALLAS, TX 75203

Dallas Independent School District

GOLD Medal


Near National Avg

253 Students

17 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#2 BASIS Scottsdale

11440 NORTH 136TH ST


BASIS Schools Inc.

GOLD Medal



698 Students

N/A Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#3 Thomas Jefferson High School for Science and Technology



Fairfax County Public Schools

GOLD Medal


Near National Avg

1,846 Students

111 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#4 Gwinnett School of Mathematics, Science and Technology



Gwinnett County Public Schools

GOLD Medal


Near National Avg

851 Students

48 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)


School of Science and Engineering Magnet


DALLAS, TX 75203

Dallas Independent School District

GOLD Medal




Near National Avg

386 Students

24 Teachers



Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#6 Carnegie Vanguard High School

1501 TAFT


Houston Independent School District

GOLD Medal


Near National Avg

590 Students

34 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#7 Academic Magnet High School



Charleston County School District

GOLD Medal


Near National Avg

610 Students

44 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#8 University High School



Tolleson Union High School District

GOLD Medal


Larger than National Avg

460 Students

14 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#9 Lamar Academy

1009 NORTH 10TH ST


Mcallen Independent School District

GOLD Medal


Smaller than National Avg

106 Students

19 Teachers


Above National Avg

100% Tested (IB)

100% Passed (IB)

#10 Gilbert Classical Academy High School



Gilbert Unified District

GOLD Medal


Smaller than National Avg

220 Students

20 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#11 The High School of American Studies at Lehman College


BRONX, NY 10468

New York City Public Schools

GOLD Medal


Near National Avg

393 Students

25 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#12 American Indian Public High School



Oakland Unf

GOLD Medal


Larger than National Avg

243 Students

13 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#13 International Studies Charter High School

2480 SW 8TH ST

MIAMI, FL 33135

Miami-Dade County Public Schools

GOLD Medal


Near National Avg

359 Students

27 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#14 High School for Dual Language and Asian Studies


NEW YORK, NY 10002

New York City Public Schools

GOLD Medal


Near National Avg

392 Students

25 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#15 Northside College Preparatory High School



Chicago Public Schools

GOLD Medal


Near National Avg

1,069 Students

74 Teachers


Above National Avg

100% Tested (AP®)

100% Passed (AP®)

#16 Oxford Academy



Anaheim Union High

GOLD Medal


Larger than National Avg

1,152 Students

38 Teachers


Above National Avg

100% Tested (AP®)

99% Passed (AP®)

#17 University High School


TUCSON, AZ 85711

Tucson Unified School District

GOLD Medal


Larger than National Avg

934 Students

44 Teachers


Above National Avg

100% Tested (AP®)

99% Passed (AP®)

#18 Pacific Collegiate School



Santa Cruz County Office Of Education

GOLD Medal


Larger than National Avg

515 Students

28 Teachers


Above National Avg

100% Tested (AP®)

99% Passed (AP®)

#19 Biotechnology High School



Monmouth County Vocational School District

GOLD Medal


Near National Avg

311 Students

23 Teachers


Above National Avg

100% Tested (IB)

99% Passed (IB)

#20 High Technology High School



Monmouth County Vocational School District

GOLD Medal


Near National Avg

280 Students

22 Teachers


Above National Avg

99% Tested (AP®)

99% Passed (AP®)

The United States has developed as a global leader, in large part, through the genius and hard work of its scientists, engineers, and innovators. In a world that’s becoming increasingly complex, where success is driven not only by what you know, but by what youcan do with what you know, it’s more important than ever for our youth to be equipped with the knowledge and skills to solve tough problems, gather and evaluate evidence, and make sense of information. These are the types of skills that students learn by studying science, technology, engineering, and math—subjects collectively known as STEM.

Yet today, few American students pursue expertise in STEM fields—and we have an inadequate pipeline of teachers skilled in those subjects. That’s why President Obama has set a priority of increasing the number of students and teachers who are proficient in these vital fields.


The need

All young people should be prepared to think deeply and to think well so that they have the chance to become the innovators, educators, researchers, and leaders who can solve the most pressing challenges facing our nation and our world, both today and tomorrow. But, right now, not enough of our youth have access to quality STEM learning opportunities and too few students see these disciplines as springboards for their careers.expand/collapse

The goals

President Obama has articulated a clear priority for STEM education: within a decade, American students must “move from the middle to the top of the pack in science and math.” The Obama Administration also is working toward the goal of fairness between places, where an equitable distribution of quality STEM learning opportunities and talented teachers can ensure that all students have the chance to study and be inspired by science, technology, engineering, and math—and have the chance to reach their full potential.expand/collapse

The plan

The Committee on STEM Education (CoSTEM), comprised of 13 agencies—including all of the mission-science agencies and the Department of Education—are facilitating a cohesive national strategy, with new and repurposed funds, to increase the impact of federal investments in five areas: 1.) improving STEM instruction in preschool through 12th grade; 2.) increasing and sustaining public and youth engagement with STEM; 3.) improving the STEM experience for undergraduate students; 4.) better serving groups historically underrepresented in STEM fields; and 5.) designing graduate education for tomorrow’s STEM workforce.expand/collapse

Supporting Teachers and Students in STEM

At the Department of Education, we share the President’s commitment to supporting and improving STEM education. Ensuring that all students have access to high-quality learning opportunities in STEM subjects is a priority, demonstrated by the fact that dozens of federal programs have made teaching and learning in science, technology, engineering, and math a critical component of competitiveness for grant funding. Just this year, for the very first time, the Department announced that its Ready-to-Learn Television grant competition would include a priority to promote the development of television and digital media focused on science.

The Department’s Race to the Top-District program supports educators in providing students with more personalized learning—in which the pace of and approach to instruction are uniquely tailored to meet students’ individual needs and interests—often supported by innovative technologies. STEM teachers across the country also are receiving resources, support, training, and development through programs like Investing in Innovation (i3), the Teacher Incentive Fund, the Math and Science Partnershipsprogram, Teachers for a Competitive Tomorrow, and the Teacher Quality Partnerships program.

Because we know that learning happens everywhere—both inside and outside of formal school settings—the Department’s 21st Century Community Learning Centers program is collaborating with NASA, the National Park Service, and the Institute of Museum and Library Services to bring high-quality STEM content and experiences to students from low-income, high-need schools. This initiative has made a commitment to Native-American students, providing about 350 young people at 11 sites across six states with out-of-school STEM courses focused on science and the environment.

And in higher education, the Hispanic-Serving Institutions-STEM program is helping to increase the number of Hispanic students attaining degrees in STEM subjects.

This sampling of programs represents some of the ways in which federal resources are helping to assist educators in implementing effective approaches for improving STEM teaching and learning; facilitating the dissemination and adoption of effective STEM instructional practices nationwide; and promoting STEM education experiences that prioritize hands-on learning to increase student engagement and achievement.

Learn more

How Do I Find…?

Information About…



A new ranking of how well the United States’ schools are preparing students for science and engineering careers shows that although there’s a small number of high performers, most states are doing a poor job of educating students in these subjects.

According to the ranking of schools teaching kindergarten through 12th grade, Massachusetts leads the pack with a score of 4.82 on a scale of 1 to 5, while Mississippi trails behind as “worst in the United States” with a 1.11 score. Twenty-one states in total, including California, earned what the ranking classified as “below average” or “far below average” scores, and only 10 states earned scores above the national average of 2.82.

Read Full Post »

« Newer Posts - Older Posts »