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Study Finds that Both Women and their Primary Care Physicians Confusion over Ovarian Cancer Symptoms May Lead to Misdiagnosis

Reporter: Stephen J. Williams, Ph.D.

This post discusses the recently released “The Every Woman Study” conducted by the World Ovarian Cancer Coalition.  For full PDF of the study please click here: WOCC-Every-Woman-Study-Summary-Report-Oct-16

The findings are summarized nicely in the NPR article from Joanne Silberner below but just want to list a few takeaways from the study

  1.  Ovarian Cancer, while not the most common cancer in women, is still one of the most deadly malignancies.  A major reason for this is the inability to catch the disease in its early, and most treatable stages.  Much work is being done on early detection (a few posts on this area from this online journal are given at the end of this post for reference)
  2. The symptoms of ovarian cancer closely mimic symptoms of gastrointestinal distress and disorders and many times these symptoms are overlooked by women as benign, temporary issues and may be mis-self diagnosed.  In addition, if mistaken for common gastrointestinal discomfort or gynecologic discomfort (cramping)  women may self-medicate with over the counter agents which mask the symptoms of ovarian cancer
  3. certain lessons can be learned from the experiences in other countries regarding access to healthcare and diagnosis. For instance

Looking at the key findings of the study it becomes clear that countries have significant potential to
learn from each other:
• Women in Germany had the shortest time to diagnosis, but much less access to
specialist clinicians that are key to successful treatment.
• Women in the UK have almost universal access to specialists but the lowest
proportion of women diagnosed within a month of visiting a doctor.
• Women in Japan had one of the shortest times to diagnosis, but very little access to
genetic testing, and were least likely to get the emotional support they needed.
• Women in the USA were most likely to wait more than three months before
consulting a doctor about symptoms, but most likely to receive genetic testing.
• Women with ovarian cancer in Hungary were most aware of ovarian cancer before
their diagnosis, but were much less likely to be offered surgery to treat their disease.

 

In summary it appears there are three key areas needing to be addressed with regard to improving early reporting of symptoms of ovarian cancer

  1. information and awareness of symptoms by BOTH women and their physicians
  2. family risk assessment programs are very important to make women aware of their risks and needs for screening
  3. access to specialist treatment is important in the early diagnosis and treatment of this disease

 

Learn the Symptoms

Symptoms (from the Sandy Rollman Ovarian Cancer Foundation)

Historically ovarian cancer was called the “silent killer” because symptoms were not thought to develop until the chance of cure was poor. However, recent studies have shown this term is untrue and that the following symptoms are much more likely to occur in women with ovarian cancer than women in the general population. These symptoms include:

  • Bloating
  • Pelvic or abdominal pain
  • Difficulty eating or feeling full quickly
  • Urinary symptoms (urgency or frequency)

Women with ovarian cancer report that symptoms are persistent and represent a change from normal for their bodies. The frequency and/or number of such symptoms are key factors in the diagnosis of ovarian cancer. Several studies show that even early stage ovarian cancer can produce these symptoms.

Women who have these symptoms almost daily for more than a few weeks should see their doctor, preferably a gynecologist. Prompt medical evaluation may lead to detection at the earliest possible stage of the disease. Early stage diagnosis is associated with an improved prognosis.

Several other symptoms have been commonly reported by women with ovarian cancer. These symptoms include fatigue, indigestion, back pain, pain with intercourse, constipation and menstrual irregularities. However, these other symptoms are not as useful in identifying ovarian cancer because they are also found in equal frequency in women in the general population who do not have ovarian cancer.

 

In addition there are serum biomarker tests which have shown useful in the screening for ovarian cancer however these tests have their caveats and not generally suggested for whole population screening due to number of false postitives which may occur (these tests will be discussed in further posts)

Serum biomarker tests include:

 Taken From NPR at https://www.npr.org/sections/goatsandsoda/2018/10/21/658798956/report-women-everywhere-dont-know-enough-about-ovarian-cancer

Report: Women Everywhere Don’t Know Enough About Ovarian Cancer

Colored scanning electron micrograph of dividing ovarian cancer cells.

Steve Gschmeissner/Science Source

new study of women with ovarian cancer shows that ignorance about the condition is common among patients in all 44 countries surveyed. And that ignorance has a cost. The disease is more treatable, even potentially curable, in its early stages.

The women’s answers also suggested their doctors were ignorant. Many of them reported that diagnosis took a long time and that they weren’t referred to proper specialists.

The study was based on an online survey of 1,531 women who had been diagnosed with the cancer and was conducted by the World Ovarian Cancer Coalition, a nonprofit support group between March and May of this year.

Ovarian cancer is the eighth leading cause of cancer in women, according to the World Health Organization. Nearly 300,000 women will develop it this year. The World Ovarian Cancer Coalition estimates that one in six will die within three months of diagnosis and fewer than half will be alive in five years.

Prior to their diagnosis, two-thirds of the women surveyed either had never heard of ovarian cancer or were familiar with the name but didn’t know anything about the disease.

 

Other articles related to Ovarian Cancer on this online Open Access Journal Include:

Model mimicking clinical profile of patients with ovarian cancer @ Yale School of Medicine

New Findings in Endometrial Cancer: Mutations, Molecular Types and Immune Responses Evoked by Mutation-prone Endometrial, Ovarian Cancer Subtypes

Good and Bad News Reported for Ovarian Cancer Therapy

Efficacy of Ovariectomy in Presence of BRCA1 vs BRCA2 and the Risk for Ovarian Cancer

Testing for Multiple Genetic Mutations via NGS for Patients: Very Strong Family History of Breast & Ovarian Cancer, Diagnosed at Young Ages, & Negative on BRCA Test

Ultrasound-based Screening for Ovarian Cancer

Warning signs may lead to better early detection of ovarian cancer

Epigenetics, Environment and Cancer: Articles of Note @PharmaceuticalIntelligence.com

Early Diagnosis [Early Detection Research Networks]

 

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Lesson 9 Cell Signaling:  Curations and Articles of reference as supplemental information for lecture section on WNTs: #TUBiol3373

Stephen J. Wiilliams, Ph.D: Curator

UPDATED 4/23/2019

This has an updated lesson on WNT signaling.  Please click on the following and look at the slides labeled under lesson 10

cell motility 9b lesson_2018_sjw

Remember our lessons on the importance of signal termination.  The CANONICAL WNT signaling (that is the β-catenin dependent signaling)

is terminated by the APC-driven degradation complex.  This leads to the signal messenger  β-catenin being degraded by the proteosome.  Other examples of growth factor signaling that is terminated by a proteosome-directed include the Hedgehog signaling system, which is involved in growth and differentiation as well as WNTs and is implicated in various cancers.

A good article on the Hedgehog signaling pathway is found here:

The Voice of a Pathologist, Cancer Expert: Scientific Interpretation of Images: Cancer Signaling Pathways and Tumor Progression

All images in use for this article are under copyrights with Shutterstock.com

Cancer is expressed through a series of transformations equally involving metabolic enzymes and glucose, fat, and protein metabolism, and gene transcription, as a result of altered gene regulatory and transcription pathways, and also as a result of changes in cell-cell interactions.  These are embodied in the following series of graphics.

Figure 1: Sonic_hedgehog_pathwaySonic_hedgehog_pathway

The Voice of Dr. Larry

The figure shows a modification of nuclear translocation by Sonic hedgehog pathway. The hedgehog proteins have since been implicated in the development of internal organs, midline neurological structures, and the hematopoietic system in humans. The Hh signaling pathway consists of three main components: the receptor patched 1 (PTCH1), the seven transmembrane G-protein coupled receptor smoothened (SMO), and the intracellular glioma-associated oncogene homolog (GLI) family of transcription factors.5The GLI family is composed of three members, including GLI1 (gene activating), GLI2 (gene activating and repressive), and GLI3 (gene repressive).6 In the absence of an activating signal from either Shh, Ihh or Dhh, PTCH1 exerts an inhibitory effect on the signal transducer SMO, preventing any downstream signaling from occurring.7 When Hh ligands bind and activate PTCH1, the inhibition on SMO is released, allowing the translocation of SMO into the cytoplasm and its subsequent activation of the GLI family of transcription factors.

 

And from the review of  Elaine Y. C. HsiaYirui Gui, and Xiaoyan Zheng   Regulation of Hedgehog Signaling by Ubiquitination  Front Biol (Beijing). 2015 Jun; 10(3): 203–220.

the authors state:

Finally, termination of Hh signaling is also important for controlling the duration of pathway activity. Hh induced ubiquitination and degradation of Ci/Gli is the most well-established mechanism for limiting signal duration, and inhibiting this process can lead to cell patterning disruption and excessive cell proliferation (). In addition to Ci/Gli, a growing body of evidence suggests that ubiquitination also plays critical roles in regulating other Hh signaling components including Ptc, Smo, and Sufu. Thus, ubiquitination serves as a general mechanism in the dynamic regulation of the Hh pathway.

Overview of Hedgehog signaling showing the signal termination by ubiquitnation and subsequent degradation of the Gli transcriptional factors. obtained from Oncotarget 5(10):2881-911 · May 2014. GSK-3B as a Therapeutic Intervention in Cancer

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Note that in absence of Hedgehog ligands Ptch inhibits Smo accumulation and activation but upon binding of Hedgehog ligands (by an autocrine or paracrine fashion) Ptch is now unable to inhibit Smo (evidence exists that Ptch is now targeted for degradation) and Smo can now inhibit Sufu-dependent and GSK-3B dependent induced degradation of Gli factors Gli1 and Gli2.  Also note the Gli1 and Gli2 are transcriptional activators while Gli3 is a transcriptional repressor.

UPDATED 4/16/2019

Please click on the following links for the Powerpoint presentation for lesson 9.  In addition click on the mp4 links to download the movies so you can view them in Powerpoint slide 22:

cell motility 9 lesson_SJW 2019

movie file 1:

Tumorigenic but noninvasive MCF-7 cells motility on an extracellular matrix derived from normal (3DCntrol) or tumor associated (TA) fibroblasts.  Note that TA ECM is “soft” and not organized and tumor cells appear to move randomly if  much at all.

Movie 2:

 

Note that these tumorigenic and invasive MDA-MB-231 breast cancer cells move in organized patterns on organized ECM derived from Tumor Associated (TA) fibroblasts than from the ‘soft’ or unorganized ECM derived from normal  (3DCntrl) fibroblasts

 

The following contain curations of scientific articles from the site https://pharmaceuticalintelligence.com  intended as additional reference material  to supplement material presented in the lecture.

Wnts are a family of lipid-modified secreted glycoproteins which are involved in:

Normal physiological processes including

A. Development:

– Osteogenesis and adipogenesis (Loss of wnt/β‐catenin signaling causes cell fate shift of preosteoblasts from osteoblasts to adipocytes)

  – embryogenesis including body axis patterning, cell fate specification, cell proliferation and cell migration

B. tissue regeneration in adult tissue

read: Wnt signaling in the intestinal epithelium: from endoderm to cancer

And in pathologic processes such as oncogenesis (refer to Wnt/β-catenin Signaling [7.10]) and to your Powerpoint presentation

 

The curation Wnt/β-catenin Signaling is a comprehensive review of canonical and noncanonical Wnt signaling pathways

 

To review:

 

 

 

 

 

 

 

 

 

 

 

Activating the canonical Wnt pathway frees B-catenin from the degradation complex, resulting in B-catenin translocating to the nucleus and resultant transcription of B-catenin/TCF/LEF target genes.

Fig. 1 Canonical Wnt/FZD signaling pathway. (A) In the absence of Wnt signaling, soluble β-catenin is phosphorylated by a degradation complex consisting of the kinases GSK3β and CK1α and the scaffolding proteins APC and Axin1. Phosphorylated β-catenin is targeted for proteasomal degradation after ubiquitination by the SCF protein complex. In the nucleus and in the absence of β-catenin, TCF/LEF transcription factor activity is repressed by TLE-1; (B) activation of the canonical Wnt/FZD signaling leads to phosphorylation of Dvl/Dsh, which in turn recruits Axin1 and GSK3β adjacent to the plasma membrane, thus preventing the formation of the degradation complex. As a result, β-catenin accumulates in the cytoplasm and translocates into the nucleus, where it promotes the expression of target genes via interaction with TCF/LEF transcription factors and other proteins such as CBP, Bcl9, and Pygo.

NOTE: In the canonical signaling, the Wnt signal is transmitted via the Frizzled/LRP5/6 activated receptor to INACTIVATE the degradation complex thus allowing free B-catenin to act as the ultimate transducer of the signal.

Remember, as we discussed, the most frequent cancer-related mutations of WNT pathway constituents is in APC.

This shows how important the degradation complex is in controlling canonical WNT signaling.

Other cell signaling systems are controlled by protein degradation:

A.  The Forkhead family of transcription factors

Read: Regulation of FoxO protein stability via ubiquitination and proteasome degradation

B. Tumor necrosis factor α/NF κB signaling

Read: NF-κB, the first quarter-century: remarkable progress and outstanding questions

1.            Question: In cell involving G-proteins, the signal can be terminated by desensitization mechanisms.  How is both the canonical and noncanonical Wnt signal eventually terminated/desensitized?

We also discussed the noncanonical Wnt signaling pathway (independent of B-catenin induced transcriptional activity).  Note that the canonical and noncanonical involve different transducers of the signal.

Noncanonical WNT Signaling

Note: In noncanonical signaling the transducer is a G-protein and second messenger system is IP3/DAG/Ca++ and/or kinases such as MAPK, JNK.

Depending on the different combinations of WNT ligands and the receptors, WNT signaling activates several different intracellular pathways  (i.e. canonical versus noncanonical)

 

In addition different Wnt ligands are expressed at different times (temporally) and different cell types in development and in the process of oncogenesis. 

The following paper on Wnt signaling in ovarian oncogenesis shows how certain Wnt ligands are expressed in normal epithelial cells but the Wnt expression pattern changes upon transformation and ovarian oncogenesis. In addition, differential expression of canonical versus noncanonical WNT ligands occur during the process of oncogenesis (for example below the authors describe the noncanonical WNT5a is expressed in normal ovarian  epithelia yet WNT5a expression in ovarian cancer is lower than the underlying normal epithelium. However the canonical WNT10a, overexpressed in ovarian cancer cells, serves as an oncogene, promoting oncogenesis and tumor growth.

Wnt5a Suppresses Epithelial Ovarian Cancer by Promoting Cellular Senescence

Benjamin G. Bitler,1 Jasmine P. Nicodemus,1 Hua Li,1 Qi Cai,2 Hong Wu,3 Xiang Hua,4 Tianyu Li,5 Michael J. Birrer,6Andrew K. Godwin,7 Paul Cairns,8 and Rugang Zhang1,*

A.           Abstract

Epithelial ovarian cancer (EOC) remains the most lethal gynecological malignancy in the US. Thus, there is an urgent need to develop novel therapeutics for this disease. Cellular senescence is an important tumor suppression mechanism that has recently been suggested as a novel mechanism to target for developing cancer therapeutics. Wnt5a is a non-canonical Wnt ligand that plays a context-dependent role in human cancers. Here, we investigate the role of Wnt5a in regulating senescence of EOC cells. We demonstrate that Wnt5a is expressed at significantly lower levels in human EOC cell lines and in primary human EOCs (n = 130) compared with either normal ovarian surface epithelium (n = 31; p = 0.039) or fallopian tube epithelium (n = 28; p < 0.001). Notably, a lower level of Wnt5a expression correlates with tumor stage (p = 0.003) and predicts shorter overall survival in EOC patients (p = 0.003). Significantly, restoration of Wnt5a expression inhibits the proliferation of human EOC cells both in vitro and in vivo in an orthotopic EOC mouse model. Mechanistically, Wnt5a antagonizes canonical Wnt/β-catenin signaling and induces cellular senescence by activating the histone repressor A (HIRA)/promyelocytic leukemia (PML) senescence pathway. In summary, we show that loss of Wnt5a predicts poor outcome in EOC patients and Wnt5a suppresses the growth of EOC cells by triggering cellular senescence. We suggest that strategies to drive senescence in EOC cells by reconstituting Wnt5a signaling may offer an effective new strategy for EOC therapy.

Oncol Lett. 2017 Dec;14(6):6611-6617. doi: 10.3892/ol.2017.7062. Epub 2017 Sep 26.

Clinical significance and biological role of Wnt10a in ovarian cancer. 

Li P1Liu W1Xu Q1Wang C1.

Ovarian cancer is one of the five most malignant types of cancer in females, and the only currently effective therapy is surgical resection combined with chemotherapy. Wnt family member 10A (Wnt10a) has previously been identified to serve an oncogenic function in several tumor types, and was revealed to have clinical significance in renal cell carcinoma; however, there is still only limited information regarding the function of Wnt10a in the carcinogenesis of ovarian cancer. The present study identified increased expression levels of Wnt10a in two cell lines, SKOV3 and A2780, using reverse transcription-polymerase chain reaction. Functional analysis indicated that the viability rate and migratory ability of SKOV3 cells was significantly inhibited following Wnt10a knockdown using short interfering RNA (siRNA) technology. The viability rate of SKOV3 cells decreased by ~60% compared with the control and the migratory ability was only ~30% of that in the control. Furthermore, the expression levels of β-catenin, transcription factor 4, lymphoid enhancer binding factor 1 and cyclin D1 were significantly downregulated in SKOV3 cells treated with Wnt10a-siRNA3 or LGK-974, a specific inhibitor of the canonical Wnt signaling pathway. However, there were no synergistic effects observed between Wnt10a siRNA3 and LGK-974, which indicated that Wnt10a activated the Wnt/β-catenin signaling pathway in SKOV3 cells. In addition, using quantitative PCR, Wnt10a was overexpressed in the tumor tissue samples obtained from 86 patients with ovarian cancer when compared with matching paratumoral tissues. Clinicopathological association analysis revealed that Wnt10a was significantly associated with high-grade (grade III, P=0.031) and late-stage (T4, P=0.008) ovarian cancer. Furthermore, the estimated 5-year survival rate was 18.4% for patients with low Wnt10a expression levels (n=38), whereas for patients with high Wnt10a expression (n=48) the rate was 6.3%. The results of the present study suggested that Wnt10a serves an oncogenic role during the carcinogenesis and progression of ovarian cancer via the Wnt/β-catenin signaling pathway.

Targeting the Wnt Pathway includes curations of articles related to the clinical development of Wnt signaling inhibitors as a therapeutic target in various cancers including hepatocellular carcinoma, colon, breast and potentially ovarian cancer.

 

2.         Question: Given that different Wnt ligands and receptors activate different signaling pathways, AND  WNT ligands  can be deferentially and temporally expressed  in various tumor types and the process of oncogenesis, how would you approach a personalized therapy targeting the WNT signaling pathway?

3.         Question: What are the potential mechanisms of either intrinsic or acquired resistance to Wnt ligand antagonists being developed?

 

Other related articles published in this Open Access Online Scientific Journal include the following:

Targeting the Wnt Pathway [7.11]

Wnt/β-catenin Signaling [7.10]

Cancer Signaling Pathways and Tumor Progression: Images of Biological Processes in the Voice of a Pathologist Cancer Expert

e-Scientific Publishing: The Competitive Advantage of a Powerhouse for Curation of Scientific Findings and Methodology Development for e-Scientific Publishing – LPBI Group, A Case in Point 

Electronic Scientific AGORA: Comment Exchanges by Global Scientists on Articles published in the Open Access Journal @pharmaceuticalintelligence.com – Four Case Studies

 

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Ultrasound-based Screening for Ovarian Cancer

Author: Dror Nir, PhD

Occasionally, I check for news on ovarian cancer screening. I do that for sentimental reasons; I started the HistoScanning project aiming to develop an effective ultrasound-based screening solution for this cancer.

As awareness for ovarian cancer is highest in the USA, I checked for the latest news on the NCI web-site. I found that to-date: “There is no standard or routine screening test for ovarian cancer. Screening for ovarian cancer has not been proven to decrease the death rate from the disease.

Screening for ovarian cancer is under study and there are screening clinical trials taking place in many parts of the country. Information about ongoing clinical trials is available from the NCI Web site.”

I also found that:

Estimated new cases and deaths from ovarian cancer in the United States in 2013:

  • New cases: 22,240
  • Deaths: 14,030

To get an idea on the significance of these numbers, lets compare them to the numbers related to breast cancer:

Estimated new cases and deaths from breast cancer in the United States in 2013:

  • New cases: 232,340 (female); 2,240 (male)
  • Deaths: 39,620 (female); 410 (male)

Death rate of ovarian cancer patients is almost 4 times higher than the rate in breast cancer patients!

Therefore, I decided to raise awareness to the results achieved for ovarian HistoScanning in a double-blind multicenter European study that was published in European Radiology three years ago. The gynecologists who recruited patients to this study used standard ultrasound machines of GE-Medical. I would like as well to disclose that I am one of the authors of this paper:

A new computer-aided diagnostic tool for non-invasive characterisation of malignant ovarian masses: results of a multicentre validation study, Olivier Lucidarme et.al., European Radiology, August 2010, Volume 20, Issue 8, pp 1822-1830

Abstract

Objectives

To prospectively assess an innovative computer-aided diagnostic technology that quantifies characteristic features of backscattered ultrasound and theoretically allows transvaginal sonography (TVS) to discriminate benign from malignant adnexal masses.

Methods

Women (n = 264) scheduled for surgical removal of at least one ovary in five centres were included. Preoperative three-dimensional (3D)-TVS was performed and the voxel data were analysed by the new technology. The findings at 3D-TVS, serum CA125 levels and the TVS-based diagnosis were compared with histology. Cancer was deemed present when invasive or borderline cancerous processes were observed histologically.

Results

Among 375 removed ovaries, 141 cancers (83 adenocarcinomas, 24 borderline, 16 cases of carcinomatosis, nine of metastases and nine others) and 234 non-cancerous ovaries (107 normal, 127 benign tumours) were histologically diagnosed. The new computer-aided technology correctly identified 138/141 malignant lesions and 206/234 non-malignant tissues (98% sensitivity, 88% specificity). There were no false-negative results among the 47 FIGO stage I/II ovarian lesions. Standard TVS and CA125 had sensitivities/specificities of 94%/66% and 89%/75%, respectively. Combining standard TVS and the new technology in parallel significantly improved TVS specificity from 66% to 92% (p < 0.0001).

table 3

table 4

An example of an ovary considered to be normal with TVS.

An example of an ovary considered to be normal with TVS.

The same TVS false-negative ovary with OVHS-detected foci of malignancy. The presence of an adenocarcinoma was confirmed histologically.

The same TVS
false-negative ovary with OVHS-detected foci of malignancy. The presence of an
adenocarcinoma was confirmed histologically.

Conclusions

Computer-aided quantification of backscattered ultrasound is  highly sensitive for the diagnosis of malignant ovarian masses.

 Personal note:

Based on this study a promising offer for ultrasound-based screening method for ovarian cancer was published in:  Int J Gynecol Cancer. 2011 Jan;21(1):35-43. doi: 10.1097/IGC.0b013e3182000528.: Mathematical models to discriminate between benign and malignant adnexal masses: potential diagnostic improvement using ovarian HistoScanning. Vaes EManchanda RNir RNir DBleiberg HAutier PMenon URobert A.

Regrettably, the results of these studies were never transformed into routine clinical products due to financial reasons.

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

Beta-Blockers help in better survival in ovarian cancer

Ovarian Cancer and fluorescence-guided surgery: A report

Role of Primary Cilia in Ovarian Cancer

Squeezing Ovarian Cancer Cells to Predict Metastatic Potential: Cell Stiffness as Possible Biomarker

BRCA1 a tumour suppressor in breast and ovarian cancer – functions in transcription, ubiquitination and DNA repair

Warning signs may lead to better early detection of ovarian cancer

 

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Author, Editor: Tilda Barliya PhD

Surgery is being commonly used to diagnose, treat and even help  prevent cancer. In which the surgeon will cut into the body to remove the cancer along with some surrounding healthy tissue to ensure that all of the cancer is removed. However distinguishing cancer cells from healthy ones during surgery can prove difficult, if not impossible. Sometimes lesions are detected only postoperatively, leading to more surgery down the line. Currently, surgeons rely on vision and touch to detect tumors during surgery but in many cases there is still no good way to determine a tumor’s margins.

In recent years, major progression has been made in imaging-guided surgery and doctors believe that  use of fluorescent dye could boost survival rates by guiding them to tiny clusters of malignant cells.

The first fluorescence-guided surgery in ovarian cancer patients have yielded great results and are summarized herein.

Dr. Phillip Low, a Ralph C. Corely Distinguished Professor of Chemistry from Purdue University has invented a fluorescent imaging agent to a modified form of the vitamin folic acid, which acts as a “homing device” to seek out and attach to ovarian cancer cells (1)

” Of all gynecologic malignancies, epithelial ovarian cancer (EOC) is the most frequent cause of death, both in the United States and in Europe. The relative absence of a clear, distinctive clinical presentation in early stages, combined with the lack of a screening tool, often results in the disease being diagnosed only at more advanced stages. The overall 5-year survival rate is 45%, and for stages III and IV it is only 20–25%.” Cytoreduction surgery followed by chemotherapy is considereed the most effective treatment.  Radiologic approaches such as X-ray, CT, MRI and ultrasound have been considered for use in assisting surgical procedures, but these are not tumor specific and generally are not useful for intraoperative applications. Therefore, a better tumor-specific detection strategy may drastically improve the patient survival.

The overexpression of folate receptor-α (FR-α) in 90–95% of epithelial ovarian cancers prompted the investigation of intraoperative tumor-specific fluorescence imaging in ovarian cancer surgery using an FR-α–targeted fluorescent agent.  Moreover, the absence of FR-α on healthy cells leads to high tumor-to-normal ratios.

Intraoperative tumor-specific fluorescence imaging in ovarian cancer by folate receptor-α targeting (http://spie.org/newsroom/technical-articles-archive/4003-shifting-the-paradigm-in-surgical-vision-with-fluorescence-molecular-imaging)

As a ligand of FR-α, folate has already been conjugated to DTPA for SPECT/CT imaging and to several PET tracers. It has also been linked to fluorescein for use in imaging metastatic disease in murine tumor models, although this was never tested in humans.

In this article, the authors have conjugated the folate to fluorescein isothiocyanate (FITC) for the use in surgery together with a real-time multispectral intraoperative fluorescence imaging system.

The authors have conducted the first clinical trial using the fluorescence-guided surgery in ovarian cancer patient. Described herein:

Tumor-specific fluorescent agent:

Targeting of the FR-α in ovarian cancer in patients, the imaging agent was produced at clinical grade according to GMP conditions by Endocyte Inc. Folate hapten (vitamin B9) was conjugated with fluorescein isothiocyanate (FITC), yielding folate-FITC (See Fitgure). Folate-FITC has an excitation wavelength of 495 nm and emits light at 520 nm. The conjugate has a very high sensitivity and  clusters of cancer cells as small as one-tenth of a millimeter can be detected, as opposed to the earlier average minimal cluster size of 3 millimeters in diameter based on current methods of visual and tactile detection.

Folate-FITC was dissolved in 10 ml sterile normal saline and injected at a dose of 0.3 mg per kg body weight over a period of 10 min and was injected 2 hrs prior to the surgery.

Patients:

10 patients with different stages of the over cancer were recruited, The mean age of all patients was 61.2 ± 11.4 (mean ± s.d.). Four patients were diagnosed with a malignant epithelial ovarian tumor (two serous carcinomas, one undifferentiated carcinoma and one mucinous carcinoma) and one patient with a serous borderline tumor. Five patients were diagnosed with a benign ovarian tumor, as confirmed by histopathology: two fibrothecomas, one cellular fibroma, one cystic teratoma and one benign multicystic ischemic ovary.

Multispectral fluorescence camera system:

The camera system (developed by the Technical University Munich/Helmholtz Center) consists of a charge-coupled digital (EM-CCD) camera (Andor Technology) for sensitive fluorescence detection and two separate cameras for detection of intrinsic fluorescence and color (PCO AG). The system is controlled by a synchronized multi-CPU computer system (Dell Computer) for simultaneous processing of raw data and image registration and rendering. The system allows color imaging and simultaneous sensitive fluorescence detection in the visible light spectrum, as appropriate for FITC imaging.  Surgery and imaging procedure are described in detail in the article (1). Shortly, a live imaging during surgery enabled the surgeon to locate the tumor and remove it, biopsy was taken for further histopathology.

Results:

Fluorescence was detectable intraoperatively in all patients with a malignant tumor and FR-αexpression but was absent in the patient with a malignant tumor but no FR-α expression and in those with benign tumors (Table 1)

Table 1: Demographics an  individual data for patients

Study no. Age (years) Histopathology FIGO stage In vivo fluorescence IHC FR-α expression FM FITC

n = 10 patients. ++, strong; +, moderate; 0, weak; −, absent; FIGO, International Federation of Gynecology and Obstetrics; IHC FR-α, immunohistochemistry folate-receptor alpha; FM FITC, fluorescence microscopy for folate-FITC; n.a., not applicable.

Malignant tumor
1 72 Serous ovarian carcinoma III ++ ++ ++
7 76 Serous ovarian carcinoma III + + +
9 64 Undifferentiated carcinoma III
10 61 Mucineus ovarian carcinoma III + + +
Borderline tumor
5 48 Serous borderline tumor I 0 + 0/+
Benign tumor
2 59 Fibrothecoma n.a.
3 74 Fibrothecoma n.a.
4 53 Mature cystic teratoma n.a.
6 64 Benign multicystic ischemic ovary n.a.
8 41 Fibroma n.a.

Healthy tissue did not show any fluorescence signal either in vivo, ex vivo or on histopathological validation. In two separate still images of patients with ovarian cancer, the mean tumor-to-background ratio (as compared to healthy peritoneal surface) for ten demarcated fluorescent tumor deposits in each still image was 3.1 (± 0.8 s.d.). In the patient with a high-grade serous carcinoma and extensive peritoneal disseminated disease (stage III, FR-α positive), widespread tumor-specific fluorescence (white spots) was present throughout the abdominal cavity, as confirmed by ex vivo histopathology. Real-time image-guided excision of fluorescent tumor deposits of size <1 mm was feasible.

A video of the surgery is presented herein:

http://www.purdue.edu/newsroom/research/2011/110918LowSurgery.html

Detection of Tumor Deposits:

Five surgeons independently identified tumor deposits on three separate color images (shown on a representative image in (Left) and on their corresponding fluorescence image of precisely the same area (Right).

The number of tumor deposits detected by surgeons when guided by tumor-specific fluorescence (median 34, range 8–81) was significantly higher than with visual observation alone (median 7, range 4–22, P < 0.001).

Summary:

In this limited series, the authored showed that the use of intraoperative tumor-specific fluorescence imaging of the systemically administered FR-α–targeted agent folate-FITC offers specific and sensitive real-time identification of tumor tissue during surgery in patients with ovarian cancer and the presence of FR-α–positive tumors. Nevertheless, one patient presented with a malignant tumor that did not express FR-α, and consequently, no fluorescence was detected.

  • A major advantage over current imaging modalities is that an intraoperative fluorescence imaging system offers a large field of view for inspection and staging. This, in turn, may permit future patient-tailored surgical interventions and may decrease the number of needless extensive surgical procedures and the associated morbidity.
  • The second major advantage of intraoperative imaging as compared to current standard techniques is that it may guide the surgeon in debulking efforts, thus contributing to more efficient cytoreduction and ultimately improving the effect of adjuvant chemotherapy in patients with reduced tumor load
  • Improving the detection of cancer deposits to submillimeter size might ultimately improve survival rates, but whether this is the case needs to established by additional clinical studies.

Advantages:

  • In ovarian cancer, the FR-α appears to constitute a good target because it is overexpressed in 90–95% of malignant tumors, especially serous carcinomas.
  • Targeting ligand, folate, is attractive as it is nontoxic, inexpensive and relatively easily conjugated to a fluorescent dye to create a tumor-specific fluorescent contrast agent.

Disadvantages:

  • Overexpression of FR-α varies strongly between different solid tumors originating from different organs, a characteristic that reduces the general applicability of folate-FITC in cancer.
  • Many organs have autofluorescence in the excitation and emission parameters of the FITC dye.

Development of new fluorescent agents in the near-infrared spectrum will allow for identification of more deeply seated tumors, based on the stronger penetration properties of near-infrared dyes with an excitation wavelength >700 nm compared to FITC.

This is the first in-human proof-of-principle and the potential benefit of intraoperative tumor-specific fluorescence imaging in staging and debulking surgery for ovarian cancer using the systemically administered targeted fluorescent agent folate-FITC. Larger international multicenter studies using standardized, uniformly calibrated multispectral fluorescence camera systems combined with folate-FITC are needed to confirm our data and further elucidate the diagnostic (accuracy, sensitivity and specificity) and therapeutic value of the reported approach in larger series of ovarian cancer patients.

Note:  Other similar approaches have been explored for  brain tumors (3a, 3b) in human clinical trials using 5-aminolevulinic acid (5-ALA). We will not address this trial in this discussion.

Ref:

1. Gooitzen M van Dam, George Themelis, Lucia M A Crane, Niels J Harlaar, Rick G Pleijhuis, Wendy Kelder, Athanasios Sarantopoulos, Johannes S de Jong, Henriette J G Arts, Ate G J van der Zee, Joost Bart, Philip S Low & Vasilis Ntziachristos. Intraoperative tumor-specific fluorescence imaging in ovarian cancer by folate receptor-αtargeting: first in-human results. Nature Medicine 17, 1315–1319 (2011). http://www.nature.com.rproxy.tau.ac.il/nm/journal/v17/n10/full/nm.2472.html

http://www.nature.com.rproxy.tau.ac.il/nm/journal/v17/n10/pdf/nm.2472.pdf

http://www.purdue.edu/newsroom/research/2011/110918LowSurgery.html

Video: http://www.youtube.com/watch?v=cPlRP0qrxts

http://www.guardian.co.uk/science/2011/sep/18/ovarian-cancer-fluorescence-detection

2. Lung cancer: http://emoryhealthmagazine.emory.edu/issues/2012/winter/briefs/a-yellow-brick-path/index.html

3a. Stummer W, Pichlmeier U, Meinel T, Wiestler OD, Zanella F, Reulen HJ; ALA-Glioma Study group. Fluorescence-guided surgery with 5-aminolevulinic acid for resection of malignant glioma: a randomised controlled multicentre phase III trial. Lancet Oncol  2006 May;7(5):392-401.

3b. Clinical trial set up: http://clinicaltrials.gov/show/NCT01502280

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