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Dana Farber/Harvard Cancer Center Ovarian Cancer SPORE
OVERALL ABSTRACT This Specialized Program of Research Excellence (SPORE) in Ovarian Cancer is submitted from the Dana-Farber/Harvard Cancer Center (DF/HCC) and combines 5 translational projects, 3 supportive cores, and 2 synergistic programs. Project 1 will examine gene-environmental interactions affecting modifiable exposures, such as anti-inflammatory drug or genital talc use, in combined data from the Nurses’ Health Cohort and a large case-control study. Project 2 seeks to identify histology-specific molecular genetic changes in large and well-characterized collections of early-staged tumors using innovative DNA-based technologies coupled with bioinformatics. Project 3 will identify biomarkers of occult ovarian cancers or precursor lesions through proteomic analysis of pre- and post-operative blood and urine specimens from women attending DF/HCC and Cancer Genetics Network Clinics for prophylactic salpingo-oophorectomy. Project 4 will identify genetic pathways associated with paclitaxel and carboplatin drug resistance, building upon exciting cell line work that points towards acquired pathways and transcriptional profiling that seeks to identify intrinsic mechanisms. Project 5 proposes Phase I and II trials to assess the value of two distinct vaccine approaches—one using recombinant vaccinia viruses and another involving dendritic cell and ovarian cancer cell fusions. Projects 1 and 3 satisfy SPORE requirements related to prevention or screening while Projects 2, 4, and 5 address biologic foundations that may translate into interventional strategies. Administrative, Biostatistical, and Tissue Cores are collaboratively structured and will support the aims of current and future projects. A Developmental Research Program will reinforce scientific and technological foundations for the DF/HCC Ovarian Cancer SPORE and maintain the balance and diversity of its projects, while a complementary Career Development Program will increase the number of researchers committed to ovarian cancer and maximize their potential to be leaders. The overall mission of the DF/HCC SPORE is to address the full range of prevention, early detection, and treatment of ovarian cancer. PROJECT 1 The primary prevention of ovarian cancer through the identification of modifiable exposures or practical chemopreventive agents has been an elusive goal of epidemiologic studies of the disease, partly because the risk factors identified are relatively weak or inconsistent among studies. This project proposes two strategies to address these limitations. To explain why certain risk factors are modest in effect, it may be necessary to identify gene-environment interactions, modification between risk factors, or variation by histologic type of ovarian cancer. To address the issue of consistency, researchers with separate data sets should work more closely together during data analysis to be certain that common definitions of exposures and confounders are used. Consistency between case-control and cohort studies would provide especially compelling evidence about risk factors. For this proposal, researchers working with existing data and biologic specimens from two large studies, the Nurses’ Health Study Cohort and a New England based case-control study of ovarian cancer, will join forces to address the following hypotheses. First, by free radical damage generated from chronic inflammation, genital exposure to talc increases risk for ovarian cancer; and the association may be modified by variants of genes in detoxification pathways. Second, by a variety of mechanisms, anti-inflammatory drugs may reduce the risk for ovarian cancer; and the association may be modified by variants of genes in metabolic pathways. Third, by enhancing steroid production, caffeine consumption may affect ovarian cancer risk differentially by menopausal status; and the association may be further modified by cholesterol consumption, smoking, hormonal use, or variants of genes in caffeine metabolism pathways. Finally, by their ability to scavenge reactive oxygen species or affect growth pathways, carotenoids may reduce risk for ovarian cancer; but a systematic investigation of factors contributing to heterogeneity between studies is necessary as well as an examination of the effect of other types of antioxidants such as the flavonoids. Histologic variation will be assessed by conducting analyses for all epithelial types combined and the following categories: serous invasive, serous borderline, mucinous, endometrioid and clear cell, and other/undifferentiated types. The common lifestyle exposures to be assessed in this project, as opposed to stronger risk factors like pregnancy history, may be more amenable to change. Clarifying their role would permit stronger public health advice and potentially decrease ovarian cancer occurrence. PROJECT 2 Understanding the cascade of genetic changes that leads to the formation of cancer promises a revolution in the prevention, early detection, and treatment of cancer by approaches that are innovative and specific. In addition, molecular profiling of tumors is likely to have far greater prognostic ability than traditional staging and grading. Advances in understanding early molecular genetic changes for ovarian cancer have been slow likely due to: a relative paucity of early-staged cancers for study of this disease, infrequent availability of fresh tissue for early-stage lesions, and histologic diversity of the disease which may confound efforts to identify common pathways. For this study, we will utilize innovative high throughput technologies for DNA and RNA analyses of early stage ovarian cancers obtained from two sources: 272 well-annotated formalin-fixed specimens from the Gynecologic Oncology Group protocol 157 and 100 specimens collected from Partners Hospital with snap frozen tissue available. These specimens will be evaluated using the NCI cDNA array platform for comparative genomic hybridization analysis to detect DNA copy number abnormalities and by single nucleotide polymorphisms (SNP)-array-based loss of heterozygosity analysis on the 10K SNP array platform to detect allelic loss profiles. Candidate genes will be first selected by performing expression profiling with the cDNA array platform and then further validated by fluorescent in situ hybridization (FISH), quantitative PCR, immunohistochemistry and enzyme linked immunosorbant assay (ELISA). All analyses will be histology-specific and use bioinformatics techniques suitable for assay based data to identify genetic changes correlated with clinical outcomes. Our hypothesis is that the identification of histology-specific molecular genetic changes will suggest potential targets for prevention, early detection, or treatment strategies, and that the molecular genetic profiles will provide important predictors. PROJECT 3 The aim of this study is to identify novel ovarian cancer biomarkers using high throughput mass spectrometry and other proteomic techniques on serum, plasma, and urine samples from high risk women undergoing risk reducing salpingo oophorectomy (RRSO). An often used approach to identifying cancer biomarkers is to obtain samples from subjects already clinically identified as having the target cancer but prior to any treatment intervention, and compare with samples from subjects without the disease. Putative markers which separate well the cases from the controls are then proposed as candidates for further testing, especially markers which separate early stage cases from controls. High throughput mass spectroscopy coupled with non-linear statistical analyses has recently demonstrated that patterns of peaks in the spectra can separate all cases from most controls. However, two issues arise with using pre-operative samples. The first is that clinically identified early stage disease is likely to be bulky, symptomatic disease, and the markers identified may be indicators only of bulky disease late in the carcinogenesis process. The second issue is that clinically identified early stage disease is not the target disease for an early detection program. In fact, an early detection program aims to identify asymptomatic subjects in early stage disease that would have been clinically identified in late stage disease. Subjects planning on RRSO form an ideal cohort for identification of biomarkers which are sensitive to low volume, asymptomatic, early stage disease. Usually individuals who undergo RRSO are at high risk of ovarian cancer due to known BRCA mutations or a strong family history of ovarian and breast cancer. Occult ovarian cancer has been identified in approximately 10% of ovaries following RRSO. Biospecimens will be obtained from a large cohort of subjects undergoing RRSO prior to and following surgery. A comprehensive pathology review will identify the subjects with occult ovarian cancer (cases) and subjects without ovarian cancer (controls). High throughput mass spectrometry followed by non-linear statistical classification methods will be utilized to identify patterns of peaks which separate cases as much as possible from non-cases. An alternative methodology, 2D DIGE (2 dimensional digital gel electrophoresis) will also be applied to identify potential serum/plasma or urine biomarkers. Following identification of the most promising peak/spot pattern, proteins and peptides corresponding to the peaks/spots will be identified through LC-MS/MS. Monoclonal antibodies will be developed for the six most important proteins/peptides in the pattern, immunoassays developed from the antibodies, and finally tested against the remaining aliquots of biospecimens to verify and enhance pattern identification through further application of non-linear classification methods. PROJECT 4 Taxol and Carboplatin are drugs of major clinical importance in the treatment of ovarian carcinoma; but the majority of patients will eventually develop resistance to these drugs. Molecular mechanisms in the development of drug resistance might involve genetic properties of tumors acquired during chemotherapy as well as intrinsic genetic properties of the tumors that contribute to resistance. The types of studies that may be useful for defining mechanism of drug resistance include in-vitro studies on cell lines and transcriptional profiling studies in human specimens. We have developed extensive preliminary in-vitro data on transcripts linked to Taxol resistance. Other cell line data have identified genes in the Fanconi Anemia (FA)/ BRCA pathway and their methylation as being potentially related to platinum resistance stemming from observations of the cisplatin hypersensitivity of cells from Fanconi-anemia patients. This project seeks to build upon preliminary work by the investigators addressing acquired mechanisms of drug resistance as well as exploring robust transcriptional models addressing intrinsic mechanisms of drug resistance by the following specific aims. First, evaluate the expression of a refined list of about 50 transcripts linked to Taxol resistance in cell lines using either quantitative PCR comparing primary and recurrent paired tumors or immuno-histochemistry in archived paired primary and recurrent tumor specimens. Second, evaluate the role of FA/BRCA gene family in initial platinum sensitivity and evolving platinum resistance by methylation and functional studies of FA genes and pathway in matched sets of germ line, primary ovarian tumor, and recurrent tumor DNA. Third, evaluate gene expression profiles in micro-dissected epithelial cells from primary ovarian cancer tumor specimens that distinguish women who had clinical remission for at least one year versus those who relapsed within six months of completing therapy. This project is designed to identify genes and/or pathways that are associated with intrinsic or acquired mechanisms of Taxol and Carboplatin resistance with the ultimate goal of identifying potential therapeutic targets for future drug development. PROJECT 5 We have developed a vaccine for patients with cancer by fusing their tumor cells with dendritic cells (DC). Fusion cell vaccines have been shown to be effective in the treatment of metastatic mouse tumor models and in reversing immunologic unresponsiveness to the human MUC1 carcinoma antigen in MUC1-transgenic mice. A Phase I clinical trial of the fusion cell vaccine has demonstrated the induction of immunologic and clinical responses. Our overall hypothesis is that ovarian cancer cells express antigens that can be exploited as targets for the induction of anti-tumor immunity. Certain ovarian carcinoma-associated antigens, such as the MUC1 glycoprotein, represent known targets. Given the genetic instability associated with progression of human tumors, there are conceivably many epitopes unique to ovarian cancer cells that, while unknown, represent additional targets for vaccine therapy. Our objective is to develop a fusion cell vaccine that induces immunity against multiple antigens expressed by ovarian cancer cells, is associated with substantial clinical efficacy and is non-toxic. Our hypotheses are that we can generate an effective fusion cell vaccine for the treatment of ovarian cancer by: 1) using mature autologous DC; 2) combining the vaccine with recombinant human IL-12 (rhIL-12); and 3) vaccinating patients with metastatic disease earlier in their treatment course. Effectiveness of the fusion cell vaccine will be assessed in vitro and in a Phase I/II trial that monitors anti-tumor activity and induction of immunity in patients with advanced ovarian cancer. The findings from these studies will be used to design a Phase II study to compare the efficacy of the approach and a vaccinia virus-based vaccine that targets the MUC1 antigen (rV-MUC1+rV-TRICOM). The development of vaccines that are non-toxic and effective against advanced ovarian cancer could provide alternative treatment options and potentially improve prognosis for patients with this disease. The specific aims are: 1) To assess fusions of ovarian carcinoma cells with immature and mature autologous DC by evaluating cytokine production (IL-12, IL-10) and potency of the fusions in generating tumor specific immunity in vitro; 2) To conduct a Phase I/II trial of the ovarian carcinoma-DC fusion vaccine generated with mature DC and administered with rhIL-12; 3) To assess the induction of immunity against ovarian cancer cells and MUC1 in patients treated with the fusion cell vaccine alone and with rhIL-12; and 4) To perform a Phase II trial that will define the immunologic efficacy of the fusion cell vaccine and the rV-MUC1+rV-TRICOM vaccine in the treatment of patients with advanced ovarian cancer who have achieved a complete remission following primary chemotherapy. Core 1 The Administration, Communication, Evaluation, and Planning (i.e., Administration) Core will manage the DF/HCC Ovarian SPORE across several DF/HCC institutions. Its importance to the Ovarian SPORE is to free project leaders to concentrate on their research by centralizing as many of the administrative tasks as possible. Specific aims of this Core are to: 1) monitor research progress and human subjects compliance; 2) provide fiscal oversight; 3) integrate the Ovarian SPORE into the DF/HCC structure; 4) create and manage a “virtual” specimen bank; 5) promote communication and collaboration at many levels and forums including the National SPORE Workshop; and 6) plan SPORE priorities and facilitate the search for new research opportunities. Aim 1 will be accomplished through structured interactions among the SPORE PI, Co-PI, Executive Committee, Advisory Committees and the DF/HCC Executive Committee as well as central documentation of human subjects’ protocols and certifications of investigator training. Fiscal oversight will be maintained by experienced grant managers with access to discretionary institutional support. The participation of DF/HCC Executive Committee members as internal advisors and SPORE investigators who are also key members of DF/HCC committees will facilitate the Cancer Center integration promised under Aim 3.Creation of a secure, virtually-unified specimen bank from the seven repositories that currently exist will be achieved through conversion of the current Access database systems to Oracle based systems that have confidential data and specimen details in separate files. Communication and collaboration will be fostered by a dedicated BWH Ovarian SPORE website in combination with a number of meetings, conferences, and newsletters (including one for patients). Key meetings will be organized before the National SPORE Workshop to select projects for presentation and after to review workshop highlights. Planning SPORE priorities and searching for new research opportunities are focuses of the Developmental Research and Career Development Programs, whose administrative aspects will be incorporated into the Administration Core and managed by a leader with responsibility for both. Finally, overall, the mission of the DF/HCC Ovarian Administration Core is to insure efficient and ethical management of resources, foster collaboration and communication, and adjust SPORE priorities. Core 2 The Biostatistics Core facility provides the statistical and computational support for all ovarian cancer SPORE investigators. The Core will support consultation and collaboration on all aspects of study design, database development and quality control, and the analysis and interpretation of data. The specific aims of this core are to:
Provide a scientific computing facility suitable to meet the statistical analysis of ovarian cancer SPORE investigators, including technical assistance in moving data between various computers and operating system environments. Core 3 The mission of the DF/HCC Ovarian SPORE Tissue Bank and Pathology Core is to maximize the research use of clinical specimens available from DF/HCC patients in a confidential and ethical manner, provide state of the art research pathology services and expert morphologic interpretation of tissue slides and to maximize the cost effectiveness and quality of these research pathology services to the SPORE investigators and qualified investigators at other institutions. This mission will be achieved through the following specific aims:
Existing banks at DFCI already provide a variety of high quality clinical specimens including snap frozen and frozen OCT-embedded ovarian and normal tissue suitable for retrieval of DNA and RNA, fixed paraffin-embedded ovarian and normal tissue, and frozen blood components, body fluids and urine. To maximize their suitability and availability for research, collection and storage methods will be made uniform and the specimens catalogued. Collection of clinical specimens occurring at MGH and DFCI will be expanded to other DF/HCC institutions under rigorous human subject’s guidelines. To achieve the critical goal to create a “virtually-unified” bank, data collection methods will be standardized and a secure, centralized database created. A governance system will be established to permit a fair, timely evaluation of requests for research materials with determination of level of linkage to clinical data permitted by the informed consent agreements. The DF/HCC Tissue Core will be lead by pathologists and clinical researchers who have a track record of collaborative research documented by nearly 70 joint publications. Dana Farber/Harvard Cancer Center Ovarian Cancer SPORE Investigators Daniel W. Cramer, MD Michael Seiden, MD David Avigan, MD Debra A. Bell, MD Ross Berkowitz, MD Michael Birrer, MD, PhD John Blessing, PhD Stephen A. Cannistra, MD lan D’Andrea, MD Dianne Finkelstein, PhD Judy Garber, MD Frank G. Haluska, MD Susan Hankinson, ScD Harvard School of Public Health Jonathan L. Hecht, MD David Hunter, MD Donald Kufe, MD Sam Mok, PhD Cynthia Morton, MD Steven J. Skates, PhD Eugen Sobel , MD Linda Titus-Ernstoff, PhD William R. Welch, MD |
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