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Posts Tagged ‘Non-small cell lung cancer’


Argos Announces Start of Phase II AGS-003 Trial in NSCLC

Reported from source http://www.oncotherapynetwork.com/lung-cancer-targets/argos-announces-start-phase-ii-ags-003-trial-nsclc?GUID=D63BFB74-A7FD-4892-846F-A7D1FFE0F131&rememberme=1&ts=29032016 by Stephen J. Williams, Ph.D.

News | March 28, 2016 | Lung Cancer Targets
By Bryant Furlow
The Cancer Research Network of Nebraska has initiated a phase II clinical trial of the autologous dendritic cell immunotherapy AGS-003 with standard platinum-doublet chemotherapy, for non-small cell lung cancer (NSCLC), Argos Therapeutics, Inc. has announced.
AGS-003 is produced using RNA from a patient’s tumor sample, and dendritic cells. It is designed to provoke memory T-cell immune responses specifically targeting an individual patient’s tumor neoantigens, which arise from tumor-specific gene mutations.

“The standard of treatment of NSCLC has been chemotherapy after surgery, but now we can offer this exciting new option of individualized immunotherapy,” said co-principal investigator Stephen Lemon, MD, Oncology Associates in Omaha.

The nonrandomized, open-label, phase II safety study will enroll 20 patients newly diagnosed with stage III NSCLC, administering AGS-003 either concurrently or sequentially with standard carboplatin and paclitaxel chemotherapy regimens, with or without radiotherapy. The primary study endpoint is the effect of AGS-003 on the toxicity associated with standard chemotherapy. Secondary endpoints include memory T-cell activation among patients who complete induction therapy and are administered five or more doses of AGS-003.

AGS-003 is also under study in the phase III ADAPT clinical trial for patients with metastatic renal cell carcinoma (mRCC). Argos is an immuno-oncology firm developing and commercializing “truly individualized” anticancer immunotherapies.

– See more at: http://www.oncotherapynetwork.com/lung-cancer-targets/argos-announces-start-phase-ii-ags-003-trial-nsclc?GUID=D63BFB74-A7FD-4892-846F-A7D1FFE0F131&rememberme=1&ts=29032016#sthash.I9FPkdTf.dpuf

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Prognostic biomarker for NSCLC and Cancer Metastasis

Larry H. Bernstein, MD, FCAP, Curator

LPBI

 

Membranous CD24 expression as detected by the monoclonal antibody SWA11 is a prognostic marker in non-small cell lung cancer patients

Michael MajoresAnne SchindlerAngela FuchsJohannes SteinLukas HeukampPeter Altevogt and Glen Kristiansen

BMC Clinical Pathology201515:19   http://dx.doi.org:/10.1186/s12907-015-0019-z

Background    Lung cancer is one of the most common malignant neoplasms worldwide and has a high mortality rate. To enable individualized therapy regimens, a better understanding of the molecular tumor biology has still to be elucidated. The expression of the cell surface protein CD24 has already been claimed to be associated with shorter patient survival in non-small cell lung cancer (NSCLC), however, the prognostic value and applicability of CD24 immunostaining in paraffin embedded tissue specimens has been questioned due to the recent acknowledgement of restricted epitope specificity of the commonly used antibody SN3b.   Methods    A cohort of 137 primary NSCLC cases was immunostained with a novel CD24 antibody (clone SWA11), which specifically recognizes the CD24 protein core and the resulting expression data were compared with expression profiles based on the monoclonal antibody SN3b. Furthermore, expression data were correlated to clinico-pathological parameters. Univariate and multivariate survival analyses were conducted with Kaplan Meier estimates and Cox regression, respectively. Results    CD24 positivity was found in 34 % resp. 21 % (SN3b) of NSCLC with a membranous and/or cytoplasmic staining pattern. Kaplan-Meier analyses revealed that membranous, but not cytoplasmic CD24 expression (clone SWA11) was associated with lympho-nodular spread and shorter overall survival times (both p < 0.05). CD24 expression established by SN3b antibodies did not reveal significant clinicopathological correlations with overall survival, neither for cytoplasmic nor membranous CD24 staining.  Conclusions    Membranous CD24 immunoreactivity, as detected with antibody clone SWA11 may serve as a prognostic factor for lymphonodular spread and poorer overall survival. Furthermore, these results corroborate the importance of a careful distinction between membranous and cytoplasmic localisation, if CD24 is to be considered as a potential prognostic biomarker.

 

Lung cancer is a major cause of carcinoma related death, being responsible for 17.8 % of all cancer deaths and accounting for more than a million deaths worldwide per year [1]. Despite intense studies to improve therapy options, its prognosis has remained poor with a 5-year overall survival rate of less than 15 % [2].

In the past decade, the largest subgroup of lung cancer, i.e. non-small cell lung cancer (NSCLC), has been subjected to exerted research for a better understanding of the underlying molecular biology of lung cancer. More than ten years ago, CD24 has already been suggested as a novel and promising biomarker for carcinoma progression in NSCLC [3] and several groups have confirmed this finding on protein and transcript level [2, 4]. CD24 is a highly glycosylated protein, that binds to the cell surface through a GPI (glycosyl-phosphatidylinositol)-anchor and functions as a cell adhesion molecule and is involved in cell-cell-interaction via its P-selectin binding site [5]. CD24 has been found to be expressed by pre-B-lymphocytes [5]. It is assumed that CD24-positive cells can attach more easily to platelets and activated endothelial cells [6, 7]. Notably, CD24 has also been observed in many human carcinomas, such as ovarian cancer, renal cell cancer, breast cancer and NSCLC [3, 812]. In epithelial ovarian cancer high scores of cytoplasmic CD24 were highly predictive of shorter patient survival times (mean 97.8 vs. 36.5 months), whereas membranous CD24 expression seemed to have no influence on survival times. Interestingly, CD24 positivity (membranous or cytoplasmic) of prostate cancer samples was significantly associated to younger patient age and higher pT stages and a higher 3-year prostate-specific antigen (PSA) relapse rate compared with CD24-negative tumours.

In patients with gallbladder carcinoma, tumors with up-regulation of CD24 revealed lymph node metastasis and lymphovascular invasion more frequently. Moreover, up-regulation of CD24 tended to show deeper invasion depth and higher TNM stage [13]. Together, these findings support CD24 as a prognostic marker for carcinoma progression and poorer survival.

Despite these intriguing findings, major concerns regarding a lack of epitope specificity of the commonly used monoclonal antibody SN3b have been raised [14]. Recent findings indicate that the mAb (monoclonal antibody) SN3b does not bind to the protein core itself, but binds to a glycan structure that decorates the CD24 molecule. On the one hand, this motif is not present on all forms of CD24 and—on the other hand—it can be present in other epitopes irrespective of CD24 [14]. These limitations underline the need for more specific CD24 antibodies, such as the mAb SWA11 antibody that has been suggested to be more specific as it binds to the protein core [14].

As CD24 is a promising biomarker for the risk assessment of disease progression, the goal of the present study was to investigate CD24 expression in NSCLC using the novel, more specific monoclonal antibody (mAb) SWA11. Special emphasis was put on the comparison of SN3b- and SWA11-mediated CD24 detection regarding a) the subcellular distribution of CD24 expression (i.e. membranous versus cytoplasmic expression) and b) its correlation with various clinicopathological features including patient survival times.

Table 1

Clinicopathological characteristics of the NSCLC cohort

  AC SCC
N (%) N (%)
Tumour stage (pT)
1 29 (21.2 %) 5 (3.6)
2 51 (37.2 %) 23 (16.8 %)
3 6 (4.4 %) 6 (4.4 %)
4 1 (0.7 %) 0 (0 %)
Nodal Status (pN) 0 37 (27.0 %) 15 (10.9 %)
1 15 (10.9 %) 9 (6.6 %)
2 14 (10.2 %) 3 (2.2 %)
3 1 (0.7 %) 0 (0.0 %)
Grading (G) 1 5 (3.6 %) 0 (0.0 %)
2 41 (29.9 %) 16 (11.6 %)
3 44 (32.1 %) 17 (12.4 %)
Mean age at surgery 64,2 64,56
(median age) (65) (67)
Sex (m:w) 68:34 30:5
Median OS (months) 52 24
(SD; 95 % CI [months]) (±23.7; 5.5– 98.5) (± 12.8;0.0– 49.0)

 

Immunohistochemical detection of CD24 expression using clone SWA11 and SN3b

Using the mAb SWA11, 47 of 137 (34.3 %) NSCLC revealed CD24 expression (either cytoplasmic or membranous) (Table 2). CD24 expression was observed more frequently in adenocarcinomas (AC) than in squamous cell carcinomas (SCC). In AC cytoplasmic expression was observed more frequently than membranous expression. In SCC, both cyptoplasmic and membranous expression was rare. Normal lung parenchyma (i.e. alveolar surface cells) showed no expression of CD24. Bronchial epithelium showed a strong membranous and cytoplasmic staining of the brush border (Fig. 1).

Table 2

Cytoplasmic and membranous expression of CD24

SWA11 (mAb clone) SN3b (mAB clone)
  AC SCC   AC SCC
Cytoplasmic N (%) N (%) Cytoplasmic N (%) N (%)
0 45 (32.6 %) 19 (13.8 %) 0 76 (55.1 %) 31 (22.5 %)
1 22 (15.9 %) 8 (5.8 %) 1 12 (8.7 %) 1 (0.7 %)
2 17 (12.3 %) 4 (2.9 %) 2 7 (5.1 %) 2 (1.4 %)
3 18 (13.0 %) 4 (2.9 %) 3 1 (0.7 %) 0 (0 %)
AC SCC AC SCC
Membranous N (%) N (%) Membranous N (%) N (%)
0 68 (49.3 %) 21 (15.2 %) 0 64 (46.4 %) 30 (21.7 %)
1 21 (15.2 %) 5 (3.6 %) 1 10 (7.2 %) 2 (1.4 %)
2 8 (5.8 %) 4 (2.9 %) 2 12 (8.7 %) 2 1.4 %)
3 5 (3.6 %) 5 (3.6 %) 3 10 (7.2 %) 0 (0 %)

Staining intensities are determined as follows:

0: negative or equivocal, 1: weak, 2: moderate and 3: strong CD24 staining

 

https://static-content.springer.com/image/art%3A10.1186%2Fs12907-015-0019-z/MediaObjects/12907_2015_19_Fig1_HTML.gif

Fig 1

The immunohistochemical characterization reveals membranous and/or cytoplasmic CD24 (mAb SWA11) expression. Strong cytoplasmic CD24 expression is found in a proportion of both AC (a) and SCC (b, d) specimens. Membranous CD24 expression can be pronounced with only scant or even absent cytoplasmic staining as shown in the AC (c). Also, both membranous and cytoplasmic CD24 detection can be found in some instances (d), the insert is showing the corresponding squamous carcinoma in-situ with membranous staining. Simultaneous membranous and cytoplasmic CD24 expression is also found in AC specimens (e, f). In normal tissue, alveolar epithelial cells do not express CD24 (g), whereas CD24 staining is found at the apical cell membrane of bronchial respiratory epithelia (h)

Using the mAb SN3b, 29 of 137 (21.2 %) NSCLC revealed CD24 expression (either cytoplasmic or membranous) (Table 2). As above, CD24 expression was observed more frequently in adenocarcinomas (AC) than in squamous cell carcinomas (SCC). However, in contrast to mAb SWA11 cytoplasmic expression was observed less frequently than membranous expression in AC. In SCC, both cytoplasmic and membranous expression was rare. Normal lung parenchyma (i.e. alveolar surface cells) showed a distinct membranous immunoreactivity. Bronchial epithelium revealed both membranous and cytoplasmic staining of CD24.

Correlation between SWA11 and SN3b: As SWA11 and SN3b detect different epitopes, we evaluated the correlation of the immunohistochemical staining patterns. Of 132 NSCLC specimens with matched expression data, only 9 specimens (6.8 %) revealed a concordant CD24 expression. Of these cases, 4 cases revealed a concordant cytoplasmic staining and another 5 cases revealed a concordant membranous CD24 expression. Statistically, no significant correlation between the two mAb could be observed (cc = −0.63, p = 0.470; Fisher’s exact test p = 0.665). The correlation of cytoplasmic and membranous expression (for each antibody) was as follows: cc = 0.475 (p < 0.05) for SWA11 (n = 108) and cc = 0.140 (p = 0.11) for SN3b (n = 103).

Survival analyses

Recent studies indicate that CD24 expression is associated with tumor progression and poorer survival rates. Therefore, we performed follow up analyses with a special emphasis on 1) the prognostic value of mAb SWA11 in dependence on subcellular staining characteristics and 2) the prognostic values of different clinicopathological parameters:

Prognostic value of CD24 in Kaplan Meier Analyses

Only membranous CD24 (SWA11) staining revealed significantly poorer survival rates (median overall survival 21 vs. 52 months; p = 0.005) as illustrated in Fig. 2. In contrast, cytoplasmic CD24 (SWA11) staining did not affect the survival rates (median OS 34 vs. 35 months; p = 0.884) (Table 3). When stratifying the cohort into SCC (n = 35) and AC (n = 102) in Kaplan Meier analyses, membranous CD24 (SWA11) expression did not affect patients’ survival, neither in SCC (p = 0.243) nor AC (p = 0.135) (Table 3), probably due to the small number of observations (Fisher exact test: p > 0.05). After stratification for AC subtypes, membranous CD24 expression (SWA11) showed a tendency towards an association with poorer survival in acinar subtype AC, but failed significance (p = 0.328).
https://static-content.springer.com/image/art%3A10.1186%2Fs12907-015-0019-z/MediaObjects/12907_2015_19_Fig2_HTML.gif

Fig 2

Survival analysis. Kaplan-Meier curves according to SWA11 expression. Cases with moderate to strong expression were bundled in a ‘high expression’ and cases with negative or weak expression in a ‘low expression’ group. Membranous expression of CD24 detected by SWA11 proved to be an independent marker for shorter survival times in NSCLC (p = 0.005)

Table 3

Univariate survival analysis

SWA11 No. of cases Mean survival time Median survival time p-value
(months +/− s.e.) (months +/− s.e.)
Mem CD24
Negative 76 84.833 +/− 10.395 52.000 +/− 27.030 0.005
Positive 16 27.925 +/− 6.379 21.000 +/− 4.000
Cyto CD24
Negative 66 75.209 +/− 10.577 35.000 +/− 12.422 0.884
Positive 26 60.540 +/− 11.551 34.000 +/− 12.196
Total CD24
Negative 64 76.972 +/− 10.841 35.000 +/− 13.726 0.633
Positive 28 57.535 +/− 10.895 34.000 +/− 9.303
SCC
Mem CD24 negative 16 52.063 +/− 14.668 16.000 +/− 16.000 0.243
Mem CD24 positive 7 21.571 +/− 7.201 24.000 +/− 23.568
AC
Mem CD24 negative 59 88.953 +/− 11.631 56.000 +/− 22.885 0.135
Mem CD24 positive 8 39.167 +/− 11.674 21.000 +/− 8.485
pN0 31 103.641 +/− 14.940 93.000 +/− 28.224 0.012
pN1+ 30 54.911 +/− 10.646 26.000 +/− 0.983

 

…..

Univariate survival analysis according to the Cox regression model (mAb SWA11)

  Beta HR (hazard ratio) 95 % CI of HR P-value
SWA11 mem all 0.856 2.353 1.268–4.364 0.007
pN 0.963 2.620 1.389–4.943 0.003
pT 0.844 2.325 1.279–4.224 0.006
Tumour type 0.975 2.651 1.999–3.517 0.000

Table 5

Multivariate survival analysis according to the Cox regression model (mAb SWA11)

  Beta HR (hazard ratio) 95 % CI of HR P-value
SWA11 mem all 0.944 2.571 1.211–5.458 0.014
pN 0.737 2.091 1.087–4.021 0.027
pT 0.587 1.799 0.755–4.283 0.185

 

…..

In the present study, we have analyzed immunohistochemical staining characteristics and the prognostic value of CD24 expression in NSCLC with a special emphasis on the comparison of the CD24 antibodies SWA11 and SN3b. The most important result of our study is that the prognostic relevance of CD24 is critically dependent on the careful consideration of sub-cellular compartments and the epitope specificity of the antibody used.

Overall, about one third of the NSCLC cohort revealed a significant CD24 expression (either cytoplasmic or membranous). These results are in line with the findings of other studies. In another NSCLC cohort, CD24 (SN3b) expression was found in 33 % of the samples (87 of 267 cases) [2]. Consistent with those results, we have found similar rates of high CD24 expression levels (35 % of the cases) for SWA11. Originally, we would have expected lower rates than those found by Lee et al, as they used the antibody SN3b, that also recognizes yet unidentified other glycoproteins next to CD24. Furthermore, they used whole mount sections instead of tissue microarrays. A possible explanation for rather equal detection rates would be the fact that it has been demonstrated that the epitope recognized by SN3b is indeed present in CD24, but is not found in all glycoforms of CD24 [14]. In contrast to the commonly used mAb SN3b, mAb SWA11 binds to the protein core of CD24 and does not depict other glycan moieties next to CD24. The protein core of CD24 is linear, consisting of the amino acid sequence leucine-proline-alanine (LAP) next to a glycosyl-phosphatidylinositol anchor [15].

CD24 expression has been associated with disease progression and cancer-related death in the majority of malignant tumors [2, 3, 16, 17], although a caveat to these data is that most of these studies are based on the supposedly less specific CD24 clone SN3b. Lee et al demonstrated a significant association between CD24-high expression (SN3b) and shorter patient survival times. Furthermore, Lee and colleagues and ourselves in former studies referred the results to cytoplasmic CD24 expression [2, 3].

Switching Off Cancers’ Ability to Spread

http://www.technologynetworks.com/rnai/news.aspx?ID=189704

A key molecule in breast and lung cancer cells can help switch off the cancers’ ability to spread around the body.

The findings by researchers at Imperial College London, published in the journal EMBO Reports, may help scientists develop treatments that prevent cancer travelling around the body – or produce some kind of test that allows doctors to gauge how likely a cancer is to spread. During tumour growth, cancer cells can break off and travel in the bloodstream or lymph system to other parts of the body, in a process called metastasis.

Patients whose cancers spread tend to have a worse prognosis, explains Professor Justin Stebbing, senior author of the study from the Department of Surgery and Cancer at Imperial: “The ability of a cancer to spread around the body has a large impact on a patient’s survival. However, at the moment we are still in the dark about why some cancers spread around the body – while others stay in one place. This study has given important insights into this process.”

The researchers were looking at breast and lung cancer cells and they found that a protein called MARK4 enables the cells to break free and move around to other parts of the body, such as the brain and liver. Although scientist are still unsure how it does this, one theory is it affects the cell’s internal scaffolding, enabling it to move more easily around the body. The team found that a molecule called miR-515-5p helps to silence, or switch off, the gene that produces MARK4.

In the study, the team used human breast cancer and lung cancer cells to show that the miR-515-5p molecule silences the gene MARK4. They then confirmed this in mouse models, which showed that increasing the amount of miR-515-5p prevents the spread of cancer cells. The findings also revealed that the silencer molecule was found in lower levels in human tumours that had spread around the body. The team then also established that patients with breast and lung cancers whose tumours had low amounts of these silencer molecules – or high amounts of MARK4 – had lower survival rates.

Researchers are now investigating whether either the MARK4 gene or the silencer molecule could be targeted with drugs. They are also investigating whether these molecules could be used to develop a test to indicate whether a patient’s cancer is likely to spread. Professor Stebbing said: “In our work we have shown that this silencer molecule is important in the spread of cancer. This is very early stage research, so we now need more studies to find out more about this molecule, and if it is present in other types of cancer.”

Dr Olivier Pardo, lead author of the paper, also from the Department of Surgery and Cancer at Imperial, added: “Our work also identified that MARK4 enables breast and lung cancer cells to both divide and invade other parts of the body. These findings could have profound implications for treating breast and lung cancers, two of the biggest cancer killers worldwide.” The study was supported by the NIHR Imperial Biomedical Research Centre, the Medical Research Council, Action Against Cancer and the Cancer Treatment and Research Trust.

 

‘Silencer molecules’ switch off cancer’s ability to spread around body

by Kate Wighton

main image

Scientists have revealed that a key molecule in breast and lung cancer cells can help switch off the cancers’ ability to spread around the body

The findings by researchers at Imperial College London, published in the journal EMBO Reports, may help scientists develop treatments that prevent cancer travelling around the body – or produce some kind of test that allows doctors to gauge how likely a cancer is to spread.

During tumour growth, cancer cells can break off and travel in the bloodstream or lymph system to other parts of the body, in a process called metastasis.

Patients whose cancers spread tend to have a worse prognosis, explains Professor Justin Stebbing, senior author of the study from the Department of Surgery and Cancer at Imperial: “The ability of a cancer to spread around the body has a large impact on a patient’s survival. However, at the moment we are still in the dark about why some cancers spread around the body – while others stay in one place. This study has given important insights into this process.”

The researchers were looking at breast and lung cancer cells and they found that a protein called MARK4 enables the cells to break free and move around to other parts of the body, such as the brain and liver. Although scientist are still unsure how it does this, one theory is it affects the cell’s internal scaffolding, enabling it to move more easily around the body.

 

miR‐515‐5p controls cancer cell migration through MARK4 regulation

Olivier E Pardo, Leandro Castellano, Catriona E Munro, Yili Hu, Francesco Mauri,Jonathan Krell, Romain Lara, Filipa G Pinho, Thameenah Choudhury, Adam EFrampton, Loredana Pellegrino, Dmitry Pshezhetskiy, Yulan Wang, JonathanWaxman, Michael J Seckl, Justin Stebbing    

EMBO reports http://embor.embopress.org/content/early/2016/02/10/embr.201540970     http://dx.doi.org:/
Here, we show that miR‐515‐5p inhibits cancer cell migration and metastasis. RNA‐seq analyses of both oestrogen receptor receptor‐positive and receptor‐negative breast cancer cells overexpressing miR‐515‐5p reveal down‐regulation of NRAS, FZD4, CDC42BPA, PIK3C2B and MARK4 mRNAs. We demonstrate that miR‐515‐5p inhibits MARK4 directly 3′ UTR interaction and that MARK4 knock‐down mimics the effect of miR‐515‐5p on breast and lung cancer cell migration. MARK4 overexpression rescues the inhibitory effects of miR‐515‐5p, suggesting miR‐515‐5p mediates this process through MARK4 down‐regulation. Furthermore, miR‐515‐5p expression is reduced in metastases compared to primary tumours derived from both in vivo xenografts and samples from patients with breast cancer. Conversely, miR‐515‐5p overexpression prevents tumour cell dissemination in a mouse metastatic model. Moreover, high miR‐515‐5p and low MARK4 expression correlate with increased breast and lung cancer patients’ survival, respectively. Taken together, these data demonstrate the importance of miR‐515‐5p/MARK4 regulation in cell migration and metastasis across two common cancers.
Embedded Image

miR‐515‐5p inhibits cancer progression, cell migration and metastasis through its direct target MARK4, a regulator of the cytoskeleton and cell motility. Moreover, reduced miR‐515‐5p and increased MARK4 levels in metastatic lung and breast cancer correlate with poor patient prognosis.

  • MARK4 down‐regulation promotes microtubule polymerisation.

  • Increased cell spreading downstream of miR‐515‐5p overexpression or MARK4 silencing hinders cell motility and invasiveness.

  • miR‐515‐5p overexpression or MARK4 silencing prevent organ colonisation by circulating tumour cells.

  • MARK4 inhibitors may represent novel therapeutic agents to control cancer dissemination.breasat cancer

 

Liquid Biopsy for NSCLC

http://www.technologynetworks.com/Diagnostics/news.aspx?ID=190276

‘Liquid biopsy’ blood test accurately detects key genetic mutations in most common form of lung cancer, study finds.

A simple blood test can rapidly and accurately detect mutations in two key genes in non-small cell lung tumors, researchers at Dana-Farber Cancer Institute and other institutions report in a new study – demonstrating the test’s potential as a clinical tool for identifying patients who can benefit from drugs targeting those mutations.

The test, known as a liquid biopsy, proved so reliable in the study that Dana-Farber/Brigham and Women’s Cancer Center (DF/BWCC) expects to offer it soon to all patients with non-small cell lung cancer (NSCLC), either at the time of first diagnosis or of relapse following previous treatment.

NSCLC is the most common form of lung cancer, diagnosed in more than 200,000 people in the United States each year, according to the American Cancer Society. An estimated 30 percent of NSCLC patients have mutations in either of the genes included in the study, and can often be treated with targeted therapies. The study is being published online today by the journal JAMA Oncology.

The liquid biopsy tested in the study – technically known as rapid plasma genotyping – involves taking a test tube-full of blood, which contains free-floating DNA from cancer cells, and analyzing that DNA for mutations or other abnormalities. (When tumor cells die, their DNA spills into the bloodstream, where it’s known as cell-free DNA.) The technique, which provides a “snapshot” of key genetic irregularities in a tumor, is a common tool in research for probing the molecular make-up of different kinds of cancers.

“We see plasma genotyping as having enormous potential as a clinical test, or assay – a rapid, noninvasive way of screening a cancer for common genetic fingerprints, while avoiding the challenges of traditional invasive biopsies,” said the senior author of the study, Geoffrey Oxnard, MD, thoracic oncologist and lung cancer researcher at Dana-Farber and Brigham and Women’s Hospital. “Our study was the first to demonstrate prospectively that a liquid biopsy technique can be a practical tool for making treatment decisions in cancer patients. The trial was such a success that we are transitioning the assay into a clinical test for lung cancer patients at DF/BWCC.”

The study involved 180 patients with NSCLC, 120 of whom were newly diagnosed, and 60 of whom had become resistant to a previous treatment, allowing the disease to recur. Participants’ cell-free DNA was tested for mutations in the EGFR and KRAS genes, and for a separate mutation in EGFR that allows tumor cells to become resistant to front-line targeted drugs. The test was performed with a technique known as droplet digital polymerase chain reaction (ddPCR), which counts the individual letters of the genetic code in cell-free DNA to determine if specific mutations are present. Each participant also underwent a conventional tissue biopsy to test for the same mutations. The results of the liquid biopsies were then compared to those of the tissue biopsies.

The data showed that liquid biopsies returned results much more quickly. The median turnaround time for liquid biopsies was three days, compared to 12 days for tissue biopsies in newly diagnosed patients and 27 days in drug-resistant patients.

Liquid biopsy was also found to be highly accurate. In newly diagnosed patients, the “predictive value” of plasma ddPCR was 100 percent for the primary EGFR mutation and the KRAS mutation – meaning that a patient who tested positive for either mutation was certain to have that mutation in his or her tumor. For patients with the EGFR resistance mutation, the predictive value of the ddPCR test was 79 percent, suggesting the blood test was able to find additional cases with the mutation that were missed using standard biopsies.

“In some patients with the EGFR resistance mutation, ddPCR detected mutations missed by standard tissue biopsy,” Oxnard remarked. “A resistant tumor is inherently made up of multiple subsets of cells, some of which carry different patterns of genetic mutations. A single biopsy is only analyzing a single part of the tumor, and may miss a mutation present elsewhere in the body. A liquid biopsy, in contrast, may better reflect the distribution of mutations in the tumor as a whole.”

When ddPCR failed to detect these mutations, the cause was less clear-cut, Oxnard says. It could indicate that the tumor cells don’t carry the mutations or, alternatively, that the tumor isn’t shedding its DNA into the bloodstream. This discrepancy between the test results and the presence of mutations was less common in patients whose cancer had metastasized to multiple sites in the body, researchers found.

The ddPCR-based test, or assay, was piloted and optimized for patients at the Translational Resarch lab of the Belfer Center for Applied Cancer Science at Dana-Farber. It was then validated for clinical use at Dana-Farber’s Lowe Center for Thoracic Oncology.

An advantage of this form of liquid biopsy is that it can help doctors quickly determine whether a patient is responding to therapy. Fifty participants in the study had repeat testing done after starting treatment for their cancer. “Those whose blood tests showed a disappearance of the mutations within two weeks were more likely to stay on the treatment than patients who didn’t see such a reduction,” said the study’s lead author, Adrian Sacher, MD, of Dana-Farber and Brigham and Women’s Hospital.

And because tumors are constantly evolving and acquiring additional mutations, repeated liquid biopsies can provide early detection of a new mutation – such as the EGFR resistance mutation – that can potentially be treated with targeted agents.

“The study data are compelling,” said DF/BWCC pathologist Lynette Sholl, MD, explaining the center’s decision to begin offering ddPCR-based liquid biopsy to all lung cancer patients. “We validated the authors’ findings by cross-comparing results from liquid and tissue biopsies in 34 NSCLC patients. To work as a real-world clinical test, liquid biopsy needs to provide reliable, accurate data and be logistically practical. That’s what we’ve seen with the ddPCR-based blood test.

“The test has great utility both for patients newly diagnosed with NSCLC and for those with a recurrence of the disease,” she continued. “It’s fast, it’s quantitative (it indicates the amount of mutant DNA in a sample), and it can be readily employed at a cancer treatment center.”

The co-authors of the study are Cloud Paweletz, PhD, Allison O’Connell, BSc, and Nora Feeney, BSc, of the Belfer Center for Applied Cancer Science at Dana-Farber; Ryan S. Alden BSc, and Stacy L. Mach BA, of Dana-Farber; Suzanne E. Dahlberg, PhD, of Dana-Farber and Harvard T.H. Chan School of Public Health; and Pasi A. Jänne, MD, PhD, of Dana-Farber, the Belfer Center, and Brigham and Women’s Hospital.

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Multiple Lung Cancer Genomic Projects Suggest New Targets, Research Directions for Non-Small Cell Lung Cancer

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

lung cancer

(photo credit: cancer.gov)

A report Lung Cancer Genome Surveys Find Many Potential Drug Targets, in the NCI Bulletin,

http://www.cancer.gov/ncicancerbulletin/091812/page2

summarizes the clinical importance of five new lung cancer genome sequencing projects. These studies have identified genetic and epigenetic alterations in hundreds of lung tumors, of which some alterations could be taken advantage of using currently approved medications.

The reports, all published this month, included genomic information on more than 400 lung tumors. In addition to confirming genetic alterations previously tied to lung cancer, the studies identified other changes that may play a role in the disease.

Collectively, the studies covered the main forms of the disease—lung adenocarcinomas, squamous cell cancers of the lung, and small cell lung cancers.

“All of these studies say that lung cancers are genomically complex and genomically diverse,” said Dr. Matthew Meyerson of Harvard Medical School and the Dana-Farber Cancer Institute, who co-led several of the studies, including a large-scale analysis of squamous cell lung cancer by The Cancer Genome Atlas (TCGA) Research Network.

Some genes, Dr. Meyerson noted, were inactivated through different mechanisms in different tumors. He cautioned that little is known about alterations in DNA sequences that do not encode genes, which is most of the human genome.

Four of the papers are summarized below, with the first described in detail, as the Nature paper used a multi-‘omics strategy to evaluate expression, mutation, and signaling pathway activation in a large cohort of lung tumors. A literature informatics analysis is given for one of the papers.  Please note that links on GENE names usually refer to the GeneCard entry.

Paper 1. Comprehensive genomic characterization of squamous cell lung cancers[1]

The Cancer Genome Atlas Research Network Project just reported, in the journal Nature, the results of their comprehensive profiling of 230 resected lung adenocarcinomas. The multi-center teams employed analyses of

  • microRNA
  • Whole Exome Sequencing including
    • Exome mutation analysis
    • Gene copy number
    • Splicing alteration
  • Methylation
  • Proteomic analysis

Summary:

Some very interesting overall findings came out of this analysis including:

  • High rates of somatic mutations including activating mutations in common oncogenes
  • Newly described loss of function MGA mutations
  • Sex differences in EGFR and RBM10 mutations
  • driver roles for NF1, MET, ERBB2 and RITI identified in certain tumors
  • differential mutational pattern based on smoking history
  • splicing alterations driven by somatic genomic changes
  • MAPK and PI3K pathway activation identified by proteomics not explained by mutational analysis = UNEXPLAINED MECHANISM of PATHWAY ACTIVATION

however, given the plethora of data, and in light of a similar study results recently released, there appears to be a great need for additional mining of this CGAP dataset. Therefore I attempted to curate some of the findings along with some other recent news relevant to the surprising findings with relation to biomarker analysis.

Makeup of tumor samples

230 lung adenocarcinomas specimens were categorized by:

Subtype

33% acinar

25% solid

14% micro-papillary

9% papillary

8% unclassified

5% lepidic

4% invasive mucinous
Gender

Smoking status

81% of patients reported past of present smoking

The authors note that TCGA samples were combined with previous data for analysis purpose.

A detailed description of Methodology and the location of deposited data are given at the following addresses:

Publication TCGA Web Page: https://tcga-data.nci.nih.gov/docs/publications/luad_2014/

Sequence files: https://cghub.ucsc.edu

Results:

Gender and Smoking Habits Show different mutational patterns

 

WES mutational analysis

  1. a) smoking status

– there was a strong correlations of cytosine to adenine nucleotide transversions with past or present smoking. In fact smoking history separated into transversion high (past and previous smokers) and transversion low (never smokers) groups, corroborating previous results.

mutations in groups              Transversion High                   Transversion Low

TP53, KRAS, STK11,                 EGFR, RB1, PI3CA

     KEAP1, SMARCA4 RBM10

 

  1. b) Gender

Although gender differences in mutational profiles have been reported, the study found minimal number of significantly mutated genes correlated with gender. Notably:

  • EGFR mutations enriched in female cohort
  • RBM10 loss of function mutations enriched in male cohort

Although the study did not analyze the gender differences with smoking patterns, it was noted that RBM10 mutations among males were more prevalent in the transversion high group.

Whole exome Sequencing and copy number analysis reveal Unique, Candidate Driver Genes

Whole exome sequencing revealed that 62% of tumors contained mutations (either point or indel) in known cancer driver genes such as:

KRAS, EGFR, BRMF, ERBB2

However, authors looked at the WES data from the oncogene-negative tumors and found unique mutations not seen in the tumors containing canonical oncogenic mutations.

Unique potential driver mutations were found in

TP53, KEAP1, NF1, and RIT1

The genomics and expression data were backed up by a proteomics analysis of three pathways:

  1. MAPK pathway
  2. mTOR
  3. PI3K pathway

…. showing significant activation of all three pathways HOWEVER the analysis suggested that activation of signaling pathways COULD NOT be deduced from DNA sequencing alone. Phospho-proteomic analysis was required to determine the full extent of pathway modification.

For example, many tumors lacked an obvious mutation which could explain mTOR or MAPK activation.

 

Altered cell signaling pathways included:

  • Increased MAPK signaling due to activating KRAS
  • Higher mTOR due to inactivating STK11 leading to increased proliferation, translation

Pathway analysis of mutations revealed alterations in multiple cellular pathways including:

  • Reduced oxidative stress response
  • Nucleosome remodeling
  • RNA splicing
  • Cell cycle progression
  • Histone methylation

Summary:

Authors noted some interesting conclusions including:

  1. MET and ERBB2 amplification and mutations in NF1 and RIT1 may be unique driver events in lung adenocarcinoma
  2. Possible new drug development could be targeted to the RTK/RAS/RAF pathway
  3. MYC pathway as another important target
  4. Cluster analysis using multimodal omics approach identifies tumors based on single-gene driver events while other tumor have multiple driver mutational events (TUMOR HETEROGENEITY)

Paper 2. A Genomics-Based Classification of Human Lung Tumors[2]

The paper can be found at

http://stm.sciencemag.org/content/5/209/209ra153

by The Clinical Lung Cancer Genome Project (CLCGP) and Network Genomic Medicine (NGM),*,

Paper Summary

This sequencing project revealed discrepancies between histologic and genomic classification of lung tumors.

Methodology

– mutational analysis by whole exome sequencing of 1255 lung tumors of histologically

defined subtypes

– immunohistochemistry performed to verify reclassification of subtypes based on sequencing data

Results

  • 55% of all cases had at least one oncogenic alteration amenable to current personalized treatment approaches
  • Marked differences existed between cluster analysis within and between preclassified histo-subtypes
  • Reassignment based on genomic data eliminated large cell carcinomas
  • Prospective classification of 5145 lung cancers allowed for genomic classification in 75% of patients
  • Identification of EGFR and ALK mutations led to improved outcomes

Conclusions:

It is feasible to successfully classify and diagnose lung tumors based on whole exome sequencing data.

Paper 3. Genomic Landscape of Non-Small Cell Lung Cancer in Smokers and Never-Smokers[3]

A link to the paper can be found here with Graphic Summary: http://www.cell.com/cell/abstract/S0092-8674%2812%2901022-7?cc=y?cc=y

Methodology

  • Whole genome sequencing and transcriptome sequencing of cancerous and adjacent normal tissues from 17 patients with NSCLC
  • Integrated RNASeq with WES for analysis of
    • Variant analysis
    • Clonality by variant allele frequency anlaysis
    • Fusion genes
  • Bioinformatic analysis

Results

  • 3,726 point mutations and more than 90 indels in the coding sequence
  • Smokers with lung cancer show 10× the number of point mutations than never-smokers
  • Novel lung cancer genes, including DACH1, CFTR, RELN, ABCB5, and HGF were identified
  • Tumor samples from males showed high frequency of MYCBP2 MYCBP2 involved in transcriptional regulation of MYC.
  • Variant allele frequency analysis revealed 10/17 tumors were at least biclonal while 7/17 tumors were monoclonal revealing majority of tumors displayed tumor heterogeneity
  • Novel pathway alterations in lung cancer include cell-cycle and JAK-STAT pathways
  • 14 fusion proteins found, including ROS1-ALK fusion. ROS1-ALK fusions have been frequently found in lung cancer and is indicative of poor prognosis[4].
  • Novel metabolic enzyme fusions
  • Alterations were identified in 54 genes for which targeted drugs are available.           Drug-gable mutant targets include: AURKC, BRAF, HGF, EGFR, ERBB4, FGFR1, MET, JAK2, JAK3, HDAC2, HDAC6, HDAC9, BIRC6, ITGB1, ITGB3, MMP2, PRKCB, PIK3CG, TERT, KRAS, MMP14

Table. Validated Gene-Fusions Obtained from Ref-Seq Data

Note: Gene columns contain links for GeneCard while Gene function links are to the    gene’s GO (Gene Ontology) function.

GeneA (5′) GeneB (3′) GeneA function (link to Gene Ontology) GeneB function (link to Gene Ontology) known function (refs)
GRIP1 TNIP1 glutamate receptor IP transcriptional repressor
SGMS1 STK10 sphingolipid synthesis ser/thr kinase
RASSF3 TTYH2 GTP-binding protein chloride anion channel
KDELR2 ROS1, GOPC ER retention seq. binding proto-oncogenic tyr kinase
ACSL4 DCAF6 fatty acid synthesis ?
MARCH8 PRKG1 ubiquitin ligase cGMP dependent protein kinase
APAF1 UNC13B, TLN1 caspase activation cytoskeletal
EML4 ALK microtubule protein tyrosine kinase
EDR3,PHC3 LOC441601 polycomb pr/DNA binding ?
DKFZp761L1918,RHPN2 ANKRD27 Rhophilin (GTP binding pr ankyrin like
VANGL1 HAO2 tetraspanin family oxidase
CACNA2D3 FLNB VOC Ca++ channel filamin (actin binding)

Author’s Note:

There has been a recent literature on the importance of the EML4-ALK fusion protein in lung cancer. EML4-ALK positive lung tumors were found to be les chemo sensitive to cytotoxic therapy[5] and these tumor cells may exhibit an epitope rendering these tumors amenable to immunotherapy[6]. In addition, inhibition of the PI3K pathway has sensitized EMl4-ALK fusion positive tumors to ALK-targeted therapy[7]. EML4-ALK fusion positive tumors show dependence on the HSP90 chaperone, suggesting this cohort of patients might benefit from the new HSP90 inhibitors recently being developed[8].

Table. Significantly mutated genes (point mutations, insertions/deletions) with associated function.

Gene Function
TP53 tumor suppressor
KRAS oncogene
ZFHX4 zinc finger DNA binding
DACH1 transcription factor
EGFR epidermal growth factor receptor
EPHA3 receptor tyrosine kinase
ENSG00000205044
RELN cell matrix protein
ABCB5 ABC Drug Transporter

Table. Literature Analysis of pathways containing significantly altered genes in NSCLC reveal putative targets and risk factors, linkage between other tumor types, and research areas for further investigation.

Note: Significantly mutated genes, obtained from WES, were subjected to pathway analysis (KEGG Pathway Analysis) in order to see which pathways contained signicantly altered gene networks. This pathway term was then used for PubMed literature search together with terms “lung cancer”, “gene”, and “NOT review” to determine frequency of literature coverage for each pathway in lung cancer. Links are to the PubMEd search results.

KEGG pathway Name # of PUBMed entries containing Pathway Name, Gene ANDLung Cancer
Cell cycle 1237
Cell adhesion molecules (CAMs) 372
Glioma 294
Melanoma 219
Colorectal cancer 207
Calcium signaling pathway 175
Prostate cancer 166
MAPK signaling pathway 162
Pancreatic cancer 88
Bladder cancer 74
Renal cell carcinoma 68
Focal adhesion 63
Regulation of actin cytoskeleton 34
Thyroid cancer 32
Salivary secretion 19
Jak-STAT signaling pathway 16
Natural killer cell mediated cytotoxicity 11
Gap junction 11
Endometrial cancer 11
Long-term depression 9
Axon guidance 8
Cytokine-cytokine receptor interaction 8
Chronic myeloid leukemia 7
ErbB signaling pathway 7
Arginine and proline metabolism 6
Maturity onset diabetes of the young 6
Neuroactive ligand-receptor interaction 4
Aldosterone-regulated sodium reabsorption 2
Systemic lupus erythematosus 2
Olfactory transduction 1
Huntington’s disease 1
Chemokine signaling pathway 1
Cardiac muscle contraction 1
Amyotrophic lateral sclerosis (ALS) 1

A few interesting genetic risk factors and possible additional targets for NSCLC were deduced from analysis of the above table of literature including HIF1-α, mIR-31, UBQLN1, ACE, mIR-193a, SRSF1. In addition, glioma, melanoma, colorectal, and prostate and lung cancer share many validated mutations, and possibly similar tumor driver mutations.

KEGGinliteroanalysislungcancer

 please click on graph for larger view

Paper 4. Mapping the Hallmarks of Lung Adenocarcinoma with Massively Parallel Sequencing[9]

For full paper and graphical summary please follow the link: http://www.cell.com/cell/abstract/S0092-8674%2812%2901061-6

Highlights

  • Exome and genome characterization of somatic alterations in 183 lung adenocarcinomas
  • 12 somatic mutations/megabase
  • U2AF1, RBM10, and ARID1A are among newly identified recurrently mutated genes
  • Structural variants include activating in-frame fusion of EGFR
  • Epigenetic and RNA deregulation proposed as a potential lung adenocarcinoma hallmark

Summary

Lung adenocarcinoma, the most common subtype of non-small cell lung cancer, is responsible for more than 500,000 deaths per year worldwide. Here, we report exome and genome sequences of 183 lung adenocarcinoma tumor/normal DNA pairs. These analyses revealed a mean exonic somatic mutation rate of 12.0 events/megabase and identified the majority of genes previously reported as significantly mutated in lung adenocarcinoma. In addition, we identified statistically recurrent somatic mutations in the splicing factor gene U2AF1 and truncating mutations affecting RBM10 and ARID1A. Analysis of nucleotide context-specific mutation signatures grouped the sample set into distinct clusters that correlated with smoking history and alterations of reported lung adenocarcinoma genes. Whole-genome sequence analysis revealed frequent structural rearrangements, including in-frame exonic alterations within EGFR and SIK2 kinases. The candidate genes identified in this study are attractive targets for biological characterization and therapeutic targeting of lung adenocarcinoma.

Paper 5. Integrative genome analyses identify key somatic driver mutations of small-cell lung cancer[10]

Highlights

  • Whole exome and transcriptome (RNASeq) sequencing 29 small-cell lung carcinomas
  • High mutation rate 7.4 protein-changing mutations/million base pairs
  • Inactivating mutations in TP53 and RB1
  • Functional mutations in CREBBP, EP300, MLL, PTEN, SLIT2, EPHA7, FGFR1 (determined by literature and database mining)
  • The mutational spectrum seen in human data also present in a Tp53-/- Rb1-/- mouse lung tumor model

 

Curator Graphical Summary of Interesting Findings From the Above Studies

DGRAPHICSUMMARYNSLCSEQPOST

The above figure (please click on figure) represents themes and findings resulting from the aforementioned studies including

questions which will be addressed in Future Posts on this site.

References:

  1. Comprehensive genomic characterization of squamous cell lung cancers. Nature 2012, 489(7417):519-525.
  2. A genomics-based classification of human lung tumors. Science translational medicine 2013, 5(209):209ra153.
  3. Govindan R, Ding L, Griffith M, Subramanian J, Dees ND, Kanchi KL, Maher CA, Fulton R, Fulton L, Wallis J et al: Genomic landscape of non-small cell lung cancer in smokers and never-smokers. Cell 2012, 150(6):1121-1134.
  4. Takeuchi K, Soda M, Togashi Y, Suzuki R, Sakata S, Hatano S, Asaka R, Hamanaka W, Ninomiya H, Uehara H et al: RET, ROS1 and ALK fusions in lung cancer. Nature medicine 2012, 18(3):378-381.
  5. Morodomi Y, Takenoyama M, Inamasu E, Toyozawa R, Kojo M, Toyokawa G, Shiraishi Y, Takenaka T, Hirai F, Yamaguchi M et al: Non-small cell lung cancer patients with EML4-ALK fusion gene are insensitive to cytotoxic chemotherapy. Anticancer research 2014, 34(7):3825-3830.
  6. Yoshimura M, Tada Y, Ofuzi K, Yamamoto M, Nakatsura T: Identification of a novel HLA-A 02:01-restricted cytotoxic T lymphocyte epitope derived from the EML4-ALK fusion gene. Oncology reports 2014, 32(1):33-39.
  7. Yang L, Li G, Zhao L, Pan F, Qiang J, Han S: Blocking the PI3K pathway enhances the efficacy of ALK-targeted therapy in EML4-ALK-positive nonsmall-cell lung cancer. Tumour biology : the journal of the International Society for Oncodevelopmental Biology and Medicine 2014.
  8. Workman P, van Montfort R: EML4-ALK fusions: propelling cancer but creating exploitable chaperone dependence. Cancer discovery 2014, 4(6):642-645.
  9. Imielinski M, Berger AH, Hammerman PS, Hernandez B, Pugh TJ, Hodis E, Cho J, Suh J, Capelletti M, Sivachenko A et al: Mapping the hallmarks of lung adenocarcinoma with massively parallel sequencing. Cell 2012, 150(6):1107-1120.
  10. Peifer M, Fernandez-Cuesta L, Sos ML, George J, Seidel D, Kasper LH, Plenker D, Leenders F, Sun R, Zander T et al: Integrative genome analyses identify key somatic driver mutations of small-cell lung cancer. Nature genetics 2012, 44(10):1104-1110.

Other posts on this site which refer to Lung Cancer and Cancer Genome Sequencing include:

Multi-drug, Multi-arm, Biomarker-driven Clinical Trial for patients with Squamous Cell Carcinoma called the Lung Cancer Master Protocol, or Lung-MAP launched by NCI, Foundation Medicine, and Five Pharma Firms

US Personalized Cancer Genome Sequencing Market Outlook 2018 –

Comprehensive Genomic Characterization of Squamous Cell Lung Cancers

International Cancer Genome Consortium Website has 71 Committed Cancer Genome Projects Ongoing

Non-small Cell Lung Cancer drugs – where does the Future lie?

Lung cancer breathalyzer trialed in the UK

Diagnosing Lung Cancer in Exhaled Breath using Gold Nanoparticles

Multi-drug, Multi-arm, Biomarker-driven Clinical Trial for patients with Squamous Cell Carcinoma called the Lung Cancer Master Protocol, or Lung-MAP launched by NCI, Foundation Medicine, and Five Pharma Firms

Read Full Post »


FDA approves EGFR mutation detection test for NSCLC drug, Tarceva

Author/Reporter: Ritu Saxena, Ph.D.

The cobas EGFR Mutation Test, Roche Molecular Diagnostics, identifies mutations in epidermal growth factor receptor (EGFR) exons 18, 19, 20 and 21 of patients. The FDA has approved the companion diagnostic for the cancer drug Tarceva (erlotinib). It would select non-small cell lung cancer (NSCLC) patients for treatment with EGFR inhibitors. This is the first FDA-approved companion diagnostic that detects EGFR gene mutations, which are present in approximately 10-30% of non-small cell lung cancers (NSCLC). The test is being approved with an expanded use for Tarceva as a first-line treatment for patients with NSCLC that has metastasized and who have certain mutations in the EGFR gene.

Lung cancer, the leading cause of cancer death among both men and women leads to death of more people than colon, breast, and prostate cancers combined. The American Cancer Society’s most recent estimates for lung cancer in the United States for 2012 reveal that about 226,160 new cases of lung cancer will be diagnosed (116,470 in men and 109,690 in women), and there will be an estimated 160,340 deaths from lung cancer (87,750 in men and 72,590 among women), accounting for about 28% of all cancer deaths. NSCLC is the most common type of lung cancer and usually grows and spreads more slowly than small cell lung cancer. Activating EGFR mutations occur in 10–30% NSCLC cases, and lead to hyperdependence of tumors on EGFR signaling and increased sensitivity of EGFR to inhibition by erlotinib. Genentech/OSI Pharmaceuticals/Roche/Chugai Pharmaceutical’s erlotinib (Tarceva) is a small molecule quinazoline and directly and reversibly inhibits the EGFR tyrosine kinase.

Tarceva has been indicated for first-line treatment of cancer with EGFR mutations including NSCLC. The approval is Tarceva’s fourth indication and the third use for lung cancer. The FDA approved Tarceva on April 16, 2010, for maintenance treatment of patients with locally advanced or metastatic NSCLC whose disease has not progressed after four cycles of platinum-based first-line chemotherapy. Tarceva was originally approved in November 2004 for the treatment of patients with locally advanced or metastatic NSCLC after failure of at least one prior chemotherapy regimen.

In a recent multicenter, open label, randomized, phase III clinical trial (EURTAC trial; NCT0044625; http://clinicaltrials.gov/ct2/show/NCT00446225 ), Tarceva was investigated in patients with advanced NSCLC with mutations in the tyrosine kinase (TK) domain of the EGFR. The EURTAC trial was initiated in February 2007 and completed in December 2012 and enrolled around 174 patients. Patients were divided into two experimental arms. Patients in arm 1 were administered Tarceva (150 mg/day) while patients in arm 2 underwent chemotherapy as platinum-based doublets. The chemotherapeutic drugs were administered as Cisplatin (75 mg/m2) / Docetaxel (75 mg/m2); Cisplatin (75 mg/m2) / Gemcitabine (1250 mg/m2; day 1 and 8); Docetaxel (75 mg/m2) /carboplatin (AUC=6); Gemcitabine (1000 mg/m2; day 1 and 8) / Carboplatin (AUC=5). Results revealed that Erlotinib is better tolerated in Chinese population (grade 3-4 toxicities 17%) then in European patients (grade 3-4 toxicities 45%). Erlotinib scored significantly better than chemotherapy in terms of progression-free survival (PFS) with 9.7 versus 5.2 months, respectively (HR 0.37, 95% CI 0.25-0.54). Thus, the results of the trial strengthen the rationale for routine baseline tissue-based assessment of EGFR mutations in patients with NSCLC and for treatment of mutation-positive patients with EGFR tyrosine-kinase inhibitors. (Gridelli C and Rossi A, J Thorac Dis. 2012 Apr 1;4(2):219-20; http://www.ncbi.nlm.nih.gov/pubmed/22833832 )

In conclusion, FDA approval of cobas EGFR Mutation Test is a recent example of how genotyping patients in clinical trials could lead to crucial information regarding personalizing the diagnostic and therapeutic approaches.

Reference:

News brief

Clinical lab products http://www.clpmag.com/all-news/24074-fda-approves-first-companion-diagnostic-to-detect-gene-mutation-linked-with-a-type-of-lung-cancer

Clinical trial http://clinicaltrials.gov/ct2/show/NCT00446225

Research articles

Melosky B. EURTAC first line therapy for non small cell lung carcinoma in epidermal growth factor receptor mutation positive patients: A choice between two TKIs. J Thorac Dis. 2012 Apr 1;4(2):221-2; http://www.ncbi.nlm.nih.gov/pubmed/22833833

Gridelli C and Rossi AJ. EURTAC first-line phase III randomized study in advanced non-small cell lung cancer: Erlotinib works also in European population. Thorac Dis. 2012 Apr 1;4(2):219-20; http://www.ncbi.nlm.nih.gov/pubmed/22833832

Related reading

Nguyen KS and Neal JW. First-line treatment of EGFR-mutant non-small-cell lung cancer: the role of erlotinib and other tyrosine kinase inhibitors. Biologics. 2012;6:337-45; http://www.ncbi.nlm.nih.gov/pubmed/23055691

https://pharmaceuticalintelligence.com/2012/11/06/non-small-cell-lung-cancer-drugs-where-does-the-future-lie/ Curator: Ritu Saxena, Ph.D.

https://pharmaceuticalintelligence.com/2013/03/03/personalized-medicine-in-nsclc/ Curator: Larry H. Bernstein, M.D.

https://pharmaceuticalintelligence.com/2012/11/08/lung-cancer-nsclc-drug-administration-and-nanotechnology/ Author: Tilda Barliya, Ph.D.

https://pharmaceuticalintelligence.com/2012/09/18/personalized-rx-decisions-in-nsclc-treatments-symposium-in-thoracic-oncology/ Reporter: Aviva Lev-Ari, Ph.D., R.N.

https://pharmaceuticalintelligence.com/2013/05/15/diagnosis-of-cardiovascular-disease-treatment-and-prevention-current-predicted-cost-of-care-and-the-promise-of-individualized-medicine-using-clinical-decision-support-systems/ Author/Curator: Larry H. Bernstein, M.D.

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Author and Curator: Ritu Saxena, Ph.D

Although cancer stem cells constitute only a small percentage of the tumor burden, their self-renewal capacity and possible link with recurrence of cancer post treatment makes them a sought after therapeutic target in cancer. The post on cancer stem cells published on the 22nd of March, 2013, describes the identity of CSCs, their functional characteristics, possible cell of origin and biomarkers. This post focuses on the therapeutic potential of CSCs, their resistance to conventional anti-tumor therapies and current therapeutic targets including biomarkers, signaling pathways and niches.

CSCs Are Resistant to conventional anticancer therapies including chemotherapy, radiotherapy and surgery that are used either alone or in combination. However, these strategies have failed several times to eradicate CSCs resulting in metastasis and relapse, hence, a fatal disease outcome.

The properties of CSCs that contribute to or lead to chemoresistance include:

Quiescent Phenotype

Chemotherapeutic agents target fast-growing cells; however, some CSCs that remain in the dormant or quiescent stage are spared from lethal damage. Later, when the dormant CSCs enter cell cycle, tumor proliferation is stimulated.

Antiapoptosis

Antiapoptotic proteins such as BCL-2 and some self-renewal pathways such as transforming growth factor β, Wnt/ β -catenin or BMI-1 are activated in CSCs. Consequently, DNA damage repair capability of CSCs is enhanced after genotoxic stress or activation of autocrine loops through the production of growth factors like epidermal growth factor (Moserle L, Cancer Lett, 1 Feb 2010;288(1):1-9).

Expression of Drug Efflux Pumps

CSCs express some proteins that have typically been known to contribute to multidrug resistance. The proteins are drug efflux pumps ABCC1, ABCG2 or MDR1. Multidrug resistance-associated proteins (ABCC subfamily) are members of the ATP-binding cassette (ABC) superfamily of transport proteins and act as cellular efflux transporters for a wide variety of substrates, in particular glutathione, glucuronide and sulfate conjugates of diverse compounds.

Radiotherapy is mainly used in breast cancer and glioblastoma multiforme. In glioblastoma multiforme, the properties of CSCs that contribute to radiotherapy resistance is the presence of CD133 marker. CD133+ CSCs preferentially activate DNA damage repair pathway and significantly induced checkpoint kinases that leads to reduced apoptosis in CSCs compared to the CD133- tumor cells (Bao S, Nature, 7 Dec 2006;444(7120):756-60).

Radiotherapy resistance in breast cancer is due to reduced levels of reactive oxygen species in CSCs. In addition, radiation resistance of progenitor cells in an immortalized breast cancer cell line was mediated by the Wnt/β catenin pathway proteins (Diehn M, et al, Nature, 9 Apr 2009;458(7239):780-3; Chen MS, et al, J Cell Sci, 1 Feb 2007;120(Pt 3):468-77).

As mentioned in the previous post on CSCs, CSC targeting therapy could either eliminate CSCs by either killing them after differentiating them from other tumor population, and/or by disrupting their niche. Efficient eradication of CSCs may require the combined ablation of CSCs themselves and their niches. Thus, identification of appropriate and specific markers of CSCs is crucial for targeting them and preventing tumor relapse. Table 1 (adapted from a review article on CSCs by Zhao et al) describes the currently used biomarkers for CSC-targeted therapy (Zhao L, et al, Eur Surg Res, 2012;49(1):8-15).

Table 1

Specific Target Cancer type Marker properties and therapy
Targeting cell markers
CD24+CD44+ESA+ Pancreatic cancer Pancreatic CSCs, elevated during tumorigenesis
CD44+CD24–ESA+ Breast cancer Breast CSCs
EpCAM high CD44+CD166+ Colorectal cancer
CD34+CD38– AML broad use as a target for chemotherapy
CD133+ Prostate cancer and breast cancer 5-transmembrane domain cell surface glycoprotein,also a marker for neuron epithelial, hematopoietic and endothelialprogenitor cells
Stro1+CD105+CD44+ Bone sarcoma
Nodal/activin Knockdown or pharmacological inhibition of its receptorAlk4/7 abrogated self-renewal capacity and in vivo tumorigenicity of CSCs.
Targeting signaling pathways
Hedgehog signaling Upregulated in several cancer types inhibitors: GDC-0449,PF04449913, BMS-833923, IPI-926 and TAK-441
Wnt/β-catenin signaling CML, squamous cell carcinoma Be required for CSC self-renewal and tumor growthinhibitors: PRI-724, WIF-1 and telomerase
Notch signaling Several cancer types An important regulator in normal development, adult stem cell maintenance,and tumorigenesis in multiple organs,inhibitors: RO4929097, BMS-906024, IPI-926 and MK0752
PI3K/Akt/PTEN/mTOR, Several cancer types The pathway is deregulated in many tumors and used to preferentially target CSCsinhibitors: temsirolimus, everolimus FDA-approved therapy for renal cell carcinoma
Targeting CSC Niche
Angiogenesis Niche Colon cancer, breast cancer, NSCLC Inhibitor: bevacizumab results in a disruption of the CSC niche, depleted vasculature and a dramatic reduction in the number of CSCs.
Hypoxia (HIF pathway) Ovarian cancer, lung cancer, cervical cancer Inhibitors: topotecan and digoxin have been approved for ovarian, lung and cervical cancer
Targeting Micro RNA
miR-200 family Inhibits EMT and cancer cell migration by direct targeting of E-cadherin transcriptional repressors ZEB1 and ZEB2
Let-7 family Regulates BT-IC stem cell-like properties by silencing more than one target
miR-124 Related to neuronal differentiation, targets laminin γ1 and integrin β1.
miR-21 Suppresses the self-renewal of embryonic stem cells

The challenge is to develop an effective treatment regimen that prevents survival, self-renewal and differentiation of CSCs and also disturbs their niche without damaging normal stem cells. In order to evaluate the efficiency of CSC-targeting therapies, in vitro models and mouse xenotransplantation models have been used for preclinical studies. Some potential CSC targeting agents in preclinical stages include notch inhibitors for glioblastoma stem cells and telomerase peptide vaccination after chemoradiotherapy of non-small cell lung cancer stem cells Stem Cells (Hovinga KE, et al, Jun 2010;28(6):1019-29; Serrano D, Mol Cancer, 9 Aug 2011;10:96). In addition, several phase II and phase III trials are currently underway to test CSC-targeting drugs focusing on efficacy and safety of treatment.

Reference:

Bao S, Nature, 7 Dec 2006;444(7120):756-60).

Diehn M, et al, Nature, 9 Apr 2009;458(7239):780-3

Chen MS, et al, J Cell Sci, 1 Feb 2007;120(Pt 3):468-77

Zhao L, et al, Eur Surg Res, 2012;49(1):8-15

Hovinga KE, et al, Jun 2010;28(6):1019-29

Serrano D, Mol Cancer, 9 Aug 2011;10:96

Pharmaceutical Intelligence posts:

https://pharmaceuticalintelligence.com/2013/03/22/in-focus-identity-of-cancer-stem-cells/ Author and curator: Ritu Saxena, PhD

https://pharmaceuticalintelligence.com/2012/08/15/to-die-or-not-to-die-time-and-order-of-combination-drugs-for-triple-negative-breast-cancer-cells-a-systems-level-analysis/ Authors: Anamika Sarkar, PhD and Ritu Saxena, PhD

https://pharmaceuticalintelligence.com/2013/03/07/the-importance-of-cancer-prevention-programs-new-perceptions-for-fighting-cancer/ Author: Ziv Raviv, PhD

https://pharmaceuticalintelligence.com/2013/03/03/treatment-for-metastatic-her2-breast-cancer/ Reporter: Larry H Bernstein, MD

https://pharmaceuticalintelligence.com/2013/03/02/recurrence-risk-for-breast-cancer/ Larry H Bernstein, MD

https://pharmaceuticalintelligence.com/2013/02/14/prostate-cancer-androgen-driven-pathomechanism-in-early-onset-forms-of-the-disease/ Curator: Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2013/01/15/exploring-the-role-of-vitamin-c-in-cancer-therapy/ Curator: Ritu Saxena, PhD

https://pharmaceuticalintelligence.com/2013/01/12/harnessing-personalized-medicine-for-cancer-management-prospects-of-prevention-and-cure-opinions-of-cancer-scientific-leaders-httppharmaceuticalintelligence-com/ Curator: Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2013/01/10/the-molecular-pathology-of-breast-cancer-progression/ Author and reporter: Tilda Barliya PhD

https://pharmaceuticalintelligence.com/2012/11/30/histone-deacetylase-inhibitors-induce-epithelial-to-mesenchymal-transition-in-prostate-cancer-cells/ Reporter and Curator: Stephen J. Williams, PhD

https://pharmaceuticalintelligence.com/2012/10/22/blood-vessel-generating-stem-cells-discovered/ Reporter: Ritu Saxena, PhD

https://pharmaceuticalintelligence.com/2012/10/17/stomach-cancer-subtypes-methylation-based-identified-by-singapore-led-team/ Reporter: Aviva Lev-Ari, PhD, RN

https://pharmaceuticalintelligence.com/2012/09/17/natural-agents-for-prostate-cancer-bone-metastasis-treatment/ Reporter: Ritu Saxena, PhD

https://pharmaceuticalintelligence.com/2012/08/28/cardiovascular-outcomes-function-of-circulating-endothelial-progenitor-cells-cepcs-exploring-pharmaco-therapy-targeted-at-endogenous-augmentation-of-cepcs/ Aviva Lev-Ari, PhD, RN

 

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State of the art in oncologic imaging of lungs.

Author-Writer: Dror Nir, PhD

 This is the second post in a series in which I will address the state of the art in oncologic imaging based on a review paper; Advances in oncologic imaging that provides updates on the latest approaches to imaging of 5 common cancers: breast, lung, prostate, colorectal cancers, and lymphoma. This paper is published at CA Cancer J Clin 2012. © 2012 American Cancer Society.

The paper gives a fair description of the use of imaging in interventional oncology based on literature review of more than 200 peer-reviewed publications.

In this post I summaries the chapter on lung cancer imaging.

Lung Cancer Imaging

“Lung cancer remains the most common cause of death from cancer worldwide, having resulted in 1.38 million deaths (18.2% of all cancer deaths) in 2008.48 It also represents the leading cause of death in smokers and the leading cause of cancer mortality in men and women in the United States. In 2012, it was estimated that 226,160 new cases of lung cancer would be diagnosed (accounting for about 14% of cancer diagnoses) and that lung cancer would cause 160,340 deaths (about 29% of cancer deaths in men and 26% of cancer deaths in women) in the United States.1 The 1-year relative survival rate for the disease increased from 35% to 43% from 1975 through 1979 to 2003 through 2006.49 The 5-year survival rate is 53% for disease that is localized when first detected, but only 15% of lung cancers are diagnosed at this early stage.”

For cancer with such poor survival rates removal of the primary lesion by surgery at an early-stage disease is the best option. The current perception in regards to lung cancr is that patients may have subclinical disease for years before presentation. It is also known that early lung cancer lesions; adenocarcinoma in situ (AIS) and minimally invasive adenocarcinoma (MIA) are slow-growing, doubling time which can exceed 2 years.52 But, since at present, no lung cancer early-detection biomarker is clinically available, the diagnosis of this disease is primarily based on symptoms, and detection often occurs after curative intervention and when it’s already too late – see: Update on biomarkers for the detection of lung cancer and also Diagnosing lung cancer in exhaled breath using gold nanoparticles. Until biomarker is found, the burden of screening for this disease is on imaging.

“AIS and MIA generally appear as a single peripheral ground-glass nodule on CT. A small solid component may be present if areas of alveolar collapse or fibroblastic proliferation are present,5051 but any solid component should raise concern for a more invasive lesion (Fig. 8). Growth over time on imaging can often be difficult to assess due to the long doubling time of these AIS and MIA, which can exceed 2 years.52 However, indicators other than growth, such as air bronchograms, increasing density, and pleural retraction within a ground-glass nodule are suggestive of AIS or MIA.

CT image shows a ground glass nodule, which is the typical appearance of AIS, in the right upper lobe.

CT image shows a ground glass nodule, which is the typical appearance of AIS, in the right upper lobe.

 

CT (A) demonstrated extensive consolidation with air bronchograms in the left upper lobe, which at surgical resection were found to represent adenocarcinoma of mixed subtype with predominate (70%) mucinous bronchioloalveolar subtype. PET imaging in the same patient (B) demonstrated uptake in the lingula higher than expected for bronchioloalveolar carcinoma and probably due to secondary inflammation/infection. CT (C) obtained 3 years after images (A) and (B) demonstrated biopsy-proven recurrent soft-tissue mass near surgical site. Fused FDG/PET images (D) demonstrate no uptake in the area. This finding is consistent with the decreased uptake usually seen in tumors of bronchioloalveolar histology (new terminology of MIA).

CT (A) demonstrated extensive consolidation with air bronchograms in the left upper lobe, which at surgical resection were found to represent adenocarcinoma of mixed subtype with predominate (70%) mucinous bronchioloalveolar subtype. PET imaging in the same patient (B) demonstrated uptake in the lingula higher than expected for bronchioloalveolar carcinoma and probably due to secondary inflammation/infection. CT (C) obtained 3 years after images (A) and (B) demonstrated biopsy-proven recurrent soft-tissue mass near surgical site. Fused FDG/PET images (D) demonstrate no uptake in the area. This finding is consistent with the decreased uptake usually seen in tumors of bronchioloalveolar histology (new terminology of MIA).

In August 2011 the results of the “National Lung Screening Trial “ which was funded by the National Cancer Institute (NCI) were published in NEJM; Reduced Lung-Cancer Mortality with Low-Dose Computed Tomographic Screening. This randomized study results showed that with low-dose CT screening of high-risk persons, there was a significant reduction of 20% in the mortality rate from lung cancer as compared to chest radiographs screening.

Based on these results one can find the following information regarding Lung Cancer Screening on the NCI web-site:

Three screening tests have been studied to see if they decrease the risk of dying from lung cancer.

The following screening tests have been studied to see if they decrease the risk of dying from lung cancer:

  • Chest x-ray: An x-ray of the organs and bones inside the chest. An x-ray is a type of energy beam that can go through the body and onto film, making a picture of areas inside the body.
  • Sputum cytology: Sputum cytology is a procedure in which a sample of sputum (mucus that is coughed up from the lungs) is viewed under a microscope to check for cancer cells.
  • Low-dose spiral CT scan (LDCT scan): A procedure that uses low-dose radiation to make a series of very detailed pictures of areas inside the body. It uses an x-ray machine that scans the body in a spiral path. The pictures are made by a computer linked to the x-ray machine. This procedure is also called a low-dose helical CT scan.

Screening with low-dose spiral CT scans has been shown to decrease the risk of dying from lung cancer in heavy smokers.

A lung cancer screening trial studied people aged 55 years to 74 years who had smoked at least 1 pack of cigarettes per day for 30 years or more. Heavy smokers who had quit smoking within the past 15 years were also studied. The trial used chest x-rays or low-dose spiral CT scans (LDCT) scans to check for signs of lung cancer.

LDCT scans were better than chest x-rays at finding early-stage lung cancer. Screening with LDCT also decreased the risk of dying from lung cancer in current and former heavy smokers.

Guide is available for patients and doctors to learn more about the benefits and harms of low-dose helical CT screening for lung cancer.

Screening with chest x-rays or sputum cytology does not decrease the risk of dying from lung cancer.

Chest x-ray and sputum cytology are two screening tests that have been used to check for signs of lung cancer. Screening with chest x-ray, sputum cytology, or both of these tests does not decrease the risk of dying from lung cancer.

The authors of Advances in oncologic imaging found out that for pre-treatment staging and post treatment follow-up of lung cancer patients mainly involves CT (preferably contrast enhanced, FDG PET and PET/CT. “Integrated PET/CT has been found to be more accurate than PET alone, CT alone, or visual correlation of PET and CT for staging NSCLC (Non-small-cell lung carcinoma).59 “

The standard treatment of choice for localized disease remains surgical resection with or without chemo-radiation therapy (stage dependant). “The current recommendations for routine follow-up after complete resection of NSCLC are as follows: for 2 years following surgery a contrast-enhanced chest CT scan every 4 to 6 months and then yearly non-contrast chest CT scans.62 Detection of recurrence on CT is the primary goal in the initial years, and therefore, optimally, a contrast-enhanced scan should be obtained to evaluate the mediastinum. In subsequent years, when identifying an early second primary lung cancer becomes of more clinical importance, a non-contrast CT chest scan suffices to evaluate the lung parenchyma.

CT (A) of 78-year-old male who was status post–left lobe lobectomy and left upper lobe wedge resection shows recurrent nodule at the surgical resection site. Fused PET/CT (B) demonstrates increased [18F]FDG uptake in the corresponding nodule at the surgical resection site consistent with recurrent tumor.

CT (A) of 78-year-old male who was status post–left lobe lobectomy and left upper lobe wedge resection shows recurrent nodule at the surgical resection site. Fused PET/CT (B) demonstrates increased [18F]FDG uptake in the corresponding nodule at the surgical resection site consistent with recurrent tumor.

In patients undergoing chemotherapies: “ [18F]FDG PET response correlates with histologic response.63 [18F]FDG PET scan data can provide an early readout of response to chemotherapy in patients with advanced-stage lung cancer.64

In patients treated by recently developed “Targeted Therapies” such as Radiofrequency ablation (RFA) the authors found out that PET/CT is the preferred imaging modality for post treatment follow-up.

“ Most patients treated with pulmonary ablation will have had a pre-procedure CT or a fusion PET/CT scan, which allows more precise anatomic localization of abnormalities seen on PET. Generally, either CT or PET/CT is performed within a few weeks of the procedure to provide a new baseline to which future images can be compared to assess for changes in size, degree of enhancement or [18F]FDG avidity.67

CT (A) demonstrates new left upper lobe mass representing new primary NSCLC in a patient who had a status post–right pneumonectomy for a prior NSCLC. CT (B) obtained in the same patient 2 weeks after radiofrequency ablation (RFA) demonstrates the postablation density in the left upper lobe. Fused PET/CT (C) obtained 4 months after RFA demonstrates mild [18F]FDG uptake at RFA site in the left upper lobe consistent with posttreatment inflammation. Fused PET/CT (D) obtained 7 months after RFA demonstrates new focal [18F]FDG uptake at post-RFA-opacity consistent with recurrent tumor.

CT (A) demonstrates new left upper lobe mass representing new primary NSCLC in a patient who had a status post–right pneumonectomy for a prior NSCLC. CT (B) obtained in the same patient 2 weeks after radiofrequency ablation (RFA) demonstrates the postablation density in the left upper lobe. Fused PET/CT (C) obtained 4 months after RFA demonstrates mild [18F]FDG uptake at RFA site in the left upper lobe consistent with posttreatment inflammation. Fused PET/CT (D) obtained 7 months after RFA demonstrates new focal [18F]FDG uptake at post-RFA-opacity consistent with recurrent tumor.

Prostate Cancer Imaging

To be followed…

Other research papers related to the management of Lung cancer were published on this Scientific Web site:

Diagnosing lung cancer in exhaled breath using gold nanoparticles

Lung Cancer (NSCLC), drug administration and nanotechnology

Non-small Cell Lung Cancer drugs – where does the Future lie?

Comprehensive Genomic Characterization of Squamous Cell Lung Cancers

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Author: Tilda Barliya PhD

Dr. Saxena has greatly introduced us to lung cancer , the associated drug treatments and their market share in the post titled ” NSCLC and where the future lie?”. Since lung cancer is the most leading cause of death in both man and women, and have gained lots of attention I am interested in elaborating on NSCLC and explore the potential use of nanotechnology in this matter.

As previously mentioned, there are 3 common types of lung cancer:

  • Adenocarcinomas are often found in an outer area of the lung. (Most common)
  • Squamous cell carcinomas are usually found in the center of the lung next to an air tube (bronchus).
  • Large cell carcinomas can occur in any part of the lung. They tend to grow and spread faster than the other two types. (Least common).

Figure 1. The Signs and symptoms of lung cancer anatomy.

Image

Since each type develops in different areas/part of the lung, it is hypothesized that they might need different routs of administration. The possible routes of administration are:

  • IV (systemic)————->through the blood
  • Inhaled aerosols (more localized)———–>through the airways

In order to understand what does “different routs of administration” refers to, we need to dig into the anatomy of the lung, i.e, airways and blood circulation as well as understand the lung-blood barriers components that may affect drug absorption.

The Blood Circulation

Two different circulatory systems, the bronchial and the pulmonary, supply the lungs with blood (Staub, 1991). The bronchial circulation is a part of the systemic circulation and is under high pressure. It receives about 1% of the cardiac output and supplies the airways (from the trachea to the terminal bronchioles), pulmonary blood vessels and lymph nodes with oxygenated blood and nutrients and conditions the inspired air (Staub, 1991). In addition, it may be important to the distribution of systemically administered drugs to the airways and to the absorption of inhaled drugs from the airways (Chediak et al., 1990). The pulmonary circulation comprise an extensive low pressure vascular bed, which receives the entire cardiac output. It perfuses the alveolar capillaries to secure efficient gas exchange and supplies nutrients to the alveolar walls. Anastomoses between bronchial and pulmonary arterial circulations have been found in the walls of medium-sized bronchi and bronchioles (Chediak et al., 1990; Kröll et al.,1987)

Image

Advantages:

  • Fast: 15–30 seconds to 1-2 hours
  • suitable for drugs not absorbed by the digestive system
  • IV can deliver continuous medication

Disadvantages:

  • Patients are not typically able to self-administer
  • It is the most dangerous route of administration because it bypasses most of the body’s natural defenses, exposing the user to health problems, known as chemo side affects.
  • Finally dose at the organ site is much lower than the administrated dose

Most of the conventional chemotherapy are mainly administrated IV (Docetaxel, Paxlitaxel, Gemcitiabine, Avastin etc).

The Airways

The human respiratory system can be divided in two functional regions: the conducting airways and the respiratory region. The conducting airways, which are composed of the nasal cavity and associated sinuses, the pharynx, larynx, trachea, bronchi, and bronchioles, filter and condition the inspired air. From trachea to the periphery of the airway tree, the airways repeatedly branch dichotomously into two daughter branches with smaller diameters and shorter length than the parent branch (Weibel, 1991). For each new generation of airways, the number of branches is doubled and the crosssectional area is exponentially increased. The conducting region of the airways generally constitutes generation 0 (trachea) to 16 (terminal bronchioles). The respiratory region, where gas exchange takes place, generally constitutes generation 17-23 and is composed of respiratory bronchioles, the alveolar ducts, and the alveolar sacs.

The air-blood barrier of the gas exchange area is composed of the alveolar epithelial cells (surface area 140 m2) on one side and the capillary bed (surface area 130 m2) on the other side of a thin basement membrane (Simionescu, 1991; Stone et al., 1992). The extensive surface area of the air-blood barrier in combination with its extreme thinness (0.1-0.5 μm) permit rapid gas exchange by passive diffusion (Plopper, 1996).

Image

The lung is a very attractive target for drug delivery. It provides direct access to disease in the treatment of respiratory diseases, while providing an enormous surface area and a relatively low enzymatic, controlled environment for systemic absorption of medications. (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1884307/)

Advantages:

  • Can be self medicated
  • Easy to use
  • Reduced side effects associated with systemic delivery

Disadvantages:

  • Slower route of action
  • Potential problem of deposition to the deeper alveolar (higher generations, like G 8-10)
  • Immuno-defense system
  • Difficulty in measuring the exact dose inside the lung
  • inhaled aerosol is entrapped in the mucus in the conducting airways

Need to be reminded that in addition, a drug’s efficacy may be affected by where in the respiratory tract it is deposited, its delivered dose and the disease it may be trying to treat.

Major components of the lung – barriers to drug absorption
As one of the primary interfaces between the organism and the environment, the respiratory system is constantly exposed to airborne particles, potential pathogens, and toxic gases in the inspired air (Plopper, 1996). As a result a sophisticated respiratory host defense system, present from the nostrils to the alveoli, has evolved to clear offending agents (Twigg, 1998).

The system comprises of:

  • mechanical (i.e. air filtration,cough, sneezing, and mucociliary clearance),
  • chemical (antioxidants, antiproteases and surfactant lipids),
  • immunological defense mechanisms and is tightly regulated to minimize inflammatory reactions that could impair the vital gas-exchange

**Intratracheal inhalation is another  administration option but will be left out of the discussion for now

From a drug delivery perspective, the components of the host defense system comprise barriers that must be overcome to ensure efficient drug deposition and absorption from the respiratory tract.

Generally, lung physiological investigations show that the airway and alveolar epithelia, not the interstitium and the endothelium, constitute the main barrier that restricts the movement of drugs and solutes from the airway lumen into the cells or the blood circulation.

Aerosols are defined as An aerosol is a suspensions of fine solid particles or liquid droplets in a gas.The major aspect affect the efficacy of aerosols as a drug delivery system is Drug Deposition.

Aerosol Drug deposition is affected by:

  • particle properties (e.g. size, shape, density, and charge),
  • respiratory tract morphology,
  • the breathing pattern (e.g. airflow rate and tidal volume)

These parameters determine not only the quantity of particles that are deposited but also in what region of the respiratory tract the particles are deposited.

Particle properties

As the cross-sectional area of the airways increases, the airflow rate rapidly decreases, and consequently the residence time of the particles in the lung increases from the large conducting airways towards the lung periphery. The most important mechanisms of particle deposition in the respiratory tract are (1) inertial impaction, (2) sedimentation, and (3) diffusion.

  • Inertial impaction – Inertial impaction occurs predominantly in the extrathoracic airways and in the tracheobronchial tree, where the airflow velocity is high and rapid changes in airflow direction occurs. Generally, particles with a diameter larger than 10 μm are most likely deposited in the extrathoracic region, whereas 2- to 10-μm particles are deposited in the tracheobronchial tree by inertial impaction. A long residence time of the inspired air favors particle deposition by sedimentation and diffusion.
  • Sedimentation – Sedimentation is of greatest importance in the small airways and alveoli and is most pronounced for particles with a diameter of 0.5-2 μm, Ultrafine particles (<0.5 μm in diameter) are deposited mainly by diffusional transport in the small airways and lung parenchyma where there is a maximal residence time of the inspired air.

Most therapeutic aerosols are almost always heterodisperse, consisting of a wide range of particle sizes and described by the log-normal distribution with the log of the particle diameters plotted against particle number, surface area or volume (mass) on a linear or probability scale and expressed as absolute values or cumulative percentage (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1884307/)

Optimal drug delivery to the lungs depends on an interaction between;

  • the inhaler device,
  • the drug formulation properties,
  • the inhalation maneuver

The devices currently available for pulmonary drug administration of pharmaceutical aerosols in clinical therapy include nebulizers, pressurized metered dose inhalers (pMDIs), and dry powder inhalers (DPIs).

However, much effort is put into the development of new inhaler devices and formulations to optimize the pulmonary delivery system for local or systemic drug targeting.

One of the major problems in aerosol delivery is

One disadvantage of the aerosol inhalation is, however, that a substantial portion of the aerosolized drug is not delivered to the lungs (i.e. delivered to the nose, mouth, skin, exhaled). only 10–15% of the emitted dose in the lungs.

In general the aerosol exposure techniques have a low dosing effectiveness, which often requires longer exposure times to administer the target dose and renders investigations of rapid kinetic events difficult. In addition, aerosol exposure requires an advanced equipment for exposure and ml-quantities of test formulation to fill up the device.

Airway geometry and humidity

Progressive branching and narrowing of the airways encourage impaction of particles. The larger the particle size, the greater the velocity of incoming air, the greater the bend angle of bifurcations and the smaller the airway radius, the greater the probability of deposition by impaction. The lung has a relative humidity of approximately 99.5%. The addition and removal of water can significantly affect the particle size of a hygroscopic aerosol and thus deposition. Drug particles are known to be hygroscopic and grow or shrink in size in high humidity, such as in the lung. A hygroscopic aerosol that is delivered at relatively low temperature and humidity into one of high humidity and temperature would be expected to increase in size when inhaled into the lung. The rate of growth is a function of the initial diameter of the particle, with the potential for the diameter of fine particles <1 µm to increase five-fold compared with two-to-three-fold for particles >2 µm. he increase in particle size above the initial size should affect the amount of drug deposited and particularly, the distribution of the aerosolized drug within the lung,

Lung Clearance Mechanism

Once deposited in the lungs, inhaled drugs are either cleared from the lungs, absorbed into the systemic circulation or degraded via drug metabolism. Drug particles deposited in the conducting airways are primarily removed through mucociliary clearance and, to a lesser extent, are absorbed through the airway . epithelium into the blood or lymphatic system. a low-viscosity periciliary or sol layer covered by a high-viscosity gel layer. Insoluble particles are trapped in the gel layer and are moved toward the pharynx (and ultimately to the gastrointestinal tract) by the upward movement of mucus generated by the metachronous beating of cilia. In the normal lung, the rate of mucus movement varies with the airway region and is determined by the number of ciliated cells and their beat frequency. Movement is faster in the trachea than in the small airways and is affected by factors influencing ciliary functioning and the quantity and quality of mucus.

Drugs deposited in the alveolar region may be phagocytosed and cleared by alveolar macrophages or absorbed into the pulmonary circulation. Alveolar macrophages are the predominant phagocytic cell for the lung defence against inhaled microorganisms, particles and other toxic agents. There are approximately five to seven alveolar macrophages per alveolus in the lungs of healthy nonsmokers. Macrophages phagocytose insoluble particles that are deposited in the alveolar region and are either cleared by the lymphatic system or moved into the ciliated airways along currents in alveolar fluid and then cleared via the mucociliary escalator.

Very little is known about how the drug-metabolizing activities of the lung affect the concentration and therapeutic efficacy of inhaled drugs. All metabolizing enzymes found in the liver are found to a lesser extent in the lung. Therefore assuming, drug deposition could have been calculated it would be hard to impossible to evaluate it’s metabolism.

In summary:

As the end organ for the treatment of local diseases or as the route of administration for systemic therapies, the lung is a very attractive target for drug delivery. It provides direct access the site of disease for the treatment of respiratory diseases without the inefficiencies and unwanted effects of systemic drug delivery. It provides an enormous surface area and a relatively low enzymatic, controlled environment for systemic absorption of medications. But it is not without barriers. Airway geometry, humidity, clearance mechanisms and presence of lung disease influence the deposition of aerosols and therefore influence the therapeutic effectiveness of inhaled medications. A drug’s efficacy may be affected by the site of deposition in the respiratory tract and the delivered dose to that site. To provide an efficient and effective inhalant therapy, these factors must be considered. Aerosol particle size characteristics can play an important role in avoiding the physiological barriers of the lung, as well as targeting the drug to the appropriate lung region.

Drug formulations and chemo drug delivery will be further discussed in a another post.

Ref:

1. N R Labiris and M B Dolovich. “Pulmonary drug delivery. Part I: Physiological factors affecting therapeutic effectiveness of aerosolized medications”. Br J Clin Pharmacol. 2003 December; 56(6): 588–599. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1884307/.

2. Tronde A. “Pulmonary drug absorption”. Acta Universities Upsalninesis Uppsala 2002. uu.diva-portal.org/smash/get/diva2:161887/FULLTEXT01

3. Naushad Khan Ghilzai. Pulmonary drug delivery. http://www.drugdel.com/Pulm_review.pdf.

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