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M.D. Anderson Cancer Center Breast Cancer SPORE
Overall Abstract The M.D. Anderson Cancer Center SPORE in Breast Cancer includes three main areas: research, comprised of the five main projects; research support, comprised of the three cores; and research development, comprised of the Developmental Research Program and Career Development Program. Each of these is outlined below: Project 1 With the widespread use of screening, detection of smaller, early stage breast cancer is increasingly more common. Optimal care of patients with early, localized breast cancer remains controversial and many women are receiving unnecessarily aggressive therapy and others receiving insufficient treatment because of uncertainties that remain in current prognostication. A large number of women recur or develop distant disease following therapy, with rates in Stage I of approximately 2% per year and in Stage II of 4% per year. The clinical management of early disease in ethnic minorities is even less clear, although strong evidence exists for worse disease outcomes among African-American women even among stage-matched patients. The concerns about optimal management are more complex in different ethnic groups where it is clear that prognosis as well as expression of molecular markers are related to ethnicity. Little is known about the biologic (genetic and epigenetic events in tumors) and epidemiologic factors that determine progression of early-stage disease. Transcriptional profiling has been demonstrated to be able to classify breast cancers into distinct subsets. In several small studies, these molecular profiles have been able to predict patient prognosis or responsiveness to therapy. However, these studies have been limited by the lack of analysis of different ethnic groups, insufficient numbers of patients to bypass the multiple parameter problem, lack of comparison of different stages and lack of follow-up of sufficient patient numbers to determine relationships with different patient management. Thus, with exception of HER2 levels or ER/PR statu, genomic profiles or molecular markers are not routinely used in patient management. The assessment of the ability of molecular profiling to predict optimal patient management has been limited by an inability to apply global technologies to retrospective samples with adequate follow up where frequently only paraffin blocks are available. Our hypothesis is that tumor genotype, defined by measures of DNA copy number and allelic imbalance, will add predictive power to current predictive methods such as tumor and clinicoepidemiologic characteristics. Further, we hypothesize that the genetic composition of tumors will vary by ethnicity and that these variations will explain differences in patient outcome in different ethnic groups. As a corollary, we predict that a high-resolution global analysis of genetic defects in patient tumors will reveal patterns of genetic change able to predict tumor behavior and response to therapy and that these will vary between different ethnic groups. Specific Aims are to (1) Determine if Stage I and II breast cancers are intrinsically different diseases or represent a continuum of the same disease by assessing: Whether changes in DNA copy number as single loci and/or co-occurring allele-specific imbalances differ by stage and correlate with known prognostic histopathologic and clinical variables. Whether epidemiologic factors (i.e., family history, age, smoking history, reproductive factors) correlate with the pattern of copy number aberrations and allelic imbalance in early stage breast tumors. Whether ethnicity affects the pattern of copy number aberrations, loci involved or degree of allelic imbalance in stage matched patients. (2) Determine if changes in DNA copy number at specific loci and/or co-occurring allele specific imbalances independently predict recurrence (local or distant) after treatment with tamoxifen and/or anthracycline-based chemotherapy. (3) Develop and test new statistical approaches to determine 'robust' sample size for future training and test sets to generate high level evidence for use of genomic or other high dimensional data obtained from tumors in prognostication and treatment planning. (4) Construct statistical models to determine if tumor DNA copy number, known prognostic markers and/or epidemiologic factors combined are better predictors of recurrence and treatment response than the standard histopathologic and clinical parameters. Project 2 Molecular genetic anomalies in breast cancer often result in excessive growth and proliferation. Normal cell cycle control mechanisms become disrupted as a consequence of oncogenic stimuli or loss of tumor suppressor function. Over-expression of several cyclins has been observed in human breast cancer, and their over-expression is, in general, associated with worse clinical outcomes. We have found in 395 breast cancer patients that cyclin E is over-expressed and present in low molecular weight (LMW) isoforms in breast cancer cells and tumor tissues, and that such alteration of cyclin E is a very strong predictor of poor outcome in breast cancer patients. Cyclin E over-expression had a 100% predictive value for the development of metastatic disease in patients with Stage I breast cancer. The mechanism by which these LMW forms give tumor cells a growth advantage in breast cancer is not known yet and is the subject of our studies. Preclinical and some clinical data suggest that the LMW forms of cyclin E may serve as a molecular predictor of response to endocrine therapy. Treatment of breast cancer with several chemotherapeutic drugs results in the induction of p21. Since the LMW forms of cyclin E are resistant to p21, it follows that one of the biological consequences of over-expression of LMW forms of cyclin E resistance to the growth inhibitory effects of drugs whose mechanism of growth inhibition is through induction of p21 and/or p27. We hypothesize that cyclin E is deregulated in breast cancer and its deregulation can be targeted both prognostically and therapeutically. The main goal of this SPORE project is to prospectively validate our previous retrospective findings, to identify the mechanisms by which cyclin E and its low molecular forms influence clinical behavior of breast cancer and resistance to endocrine and cytotoxic therapy and to identify cyclin E/CDK2 inhibitors that selectively target activity of the low molecular weight forms of cyclin E and study their effects on growth and progression of breast cancer. Specific Aims are to (1) Establish cyclin E as a prognostic marker and a predictor of clinical response to neoadjuvant chemotherapy and adjuvant hormonal therapy in patients with stage II-III breast cancer. (2) Determine the mechanism of altered sensitivity to chemotherapy and hormonal therapy in cyclin E over-expressing breast cancer cells. (3) Identify cyclin E/CDK2 inhibitors which selectively target activity of the low molecular weight forms of cyclin E Project 3 Metastatic tumors and stroma interactions are known to be critical for the survival of tumors, including breast cancer. There are data supporting the role of IFN$ as a natural anti-angiogenic factor and potent immune modulator in suppression of human tumor development. In this proposal, we direct our attention to the unique biological characteristics of mesenchymal stem cells (MSC), define their involvement in tumor development, and determine if modified MSC can be used as cellular delivery vehicles to control metastatic tumor development. We have preliminary data that indicate stromal cells found in solid tumors can originate from bone marrow MSC. We demonstrate that MSC are able to reach a metastatic lesion after intravenous administration, interact with neoplastic clones and proliferate locally in response to tumor microenvironmental signals. We will determine if MSC expressing anti-tumor molecules are able to home to growing tumors and suppress tumor growth in vivo. In our initial work, we chose IFN$ as a potent tumor inhibitor as this molecule has been shown to suppress the growth of a wide range of tumors. We hypothesize that MSC will home to and selectively proliferate in the stroma of breast cancer xenografts and their metastases and that MSC engineered to express therapeutic gene products such as IFN$ will suppress tumor growth in vitro and in vivo. The proposed studies are designed to understand the factors responsible for MSC interactions in the tumor microenvironment and will optimize the cellular delivery of therapeutic genes into the stroma of breast tumors. Specific Aims are to (1) Determine the mechanisms of MSC interactions in the tumor microenvironment. (2) Investigate the use of gene-modified MSC as a cellular delivery vehicle for control of tumor growth in established metastatic and implanted xenograft models. (3) Determine the antitumor effect and safety of gene-modified mesenchymal stem cells in patients with advanced metastatic breast cancer Project 4 The HER2 (neu or erbB-2) gene is amplified and overexpressed in 20% to 30% of breast cancers. Gene amplification or over-expression is associated with adverse prognosis and a more malignant clinical course. The HER2/ErbB2-targeting antibody, trastuzumab (Herceptin), that specifically binds to the extra-cellular domain of ErbB2, has remarkable therapeutic efficacy in certain patients with HER2/ErbB2-overexpressing breast cancer. As the only FDA approved therapeutic antibody for metastatic breast cancer, trastuzumab has demonstrated durable responses as a single agent and striking therapeutic efficacy in combination with other chemotherapeutics The overall response rate with trastuzumab, however, is limited (11% to 35%) and the cause of trastuzumab resistance is poorly understood. Trastuzumab resistance-conferring factors may serve as molecular targets for overcoming trastuzumab resistance. Unfortunately, we have limited information on the mechanisms of trastuzumab resistance of breast cancer cells. Currently, there is no clinically verified factor that can be used to predict trastuzumab resistance. PTEN (phosphatase and tensin homologue, also named MMAC1/TEP) is a dual phosphatase that mainly dephosphorylates position D3 of membrane phosphatidylinositol-3,4,5 trisphosphate (PI3,4,5P3), which is the site for recruiting the plecstrin-homology domain of Akt to the cell membrane. Since phosphatidylinositol 3-Kinase (PI3K) catalyzes the production of PI3,4,5P3, PTEN antagonizes this PI3K function and negatively regulates Akt activities. Loss of PTEN function due to PTEN mutations, PTEN haploinsufficiency from LOH at the PTEN locus, and epigenetic down-modulation of PTEN have been reported in nearly 50% of breast cancers and in many other cancer types. Restoration of PTEN expression in PTEN-null cells leads to inhibition of Akt activities and tumor suppression. Thus, PTEN is an important tumor suppressor gene. We have found that PTEN activation contributes to trastuzumab's anti-tumor activity. Reducing PTEN expression in ErbB2-overexpressing BT474 breast cancer cells by antisense oligonucleotides (AS-PTEN) conferred trastuzumab resistance in vitro and in vivo. More importantly, among a small cohort of patients having ErbB2-overexpressing breast cancers treated with trastuzumab and paclitaxel, those with PTEN-deficient cancers had significantly poorer responses to trastuzumab-based therapy than those with normal PTEN (p<0.01). We also found that PI3K inhibitors LY294002 and worthmannin rescued PTEN loss-induced trastuzumab resistance in vitro and in vivo. Thus, we hypothesize that loss of PTEN in an ErbB2-overexpressing breast tumor is a sensitive and specific predictor for trastuzumab resistance, and novel combination therapies targeting the PI3K/PTEN pathways can be developed to overcome the PTEN loss-mediated trastuzumab resistance. Specific Aims are to (1) Determine if PTEN deficiency can predict trastuzumab resistance in large-scale retrospective studies. (2) Establish animal models bearing PTEN-deficient, ErbB2-over-expressing mammary tumors. (3) Test therapeutics for overcoming PTEN deficiency-mediated trastuzumab resistance in vitro and in vivo. (4) Determine if combination therapy of trastuzumab and PI3K inhibitors can overcome PTEN loss-mediated trastuzumab resistance in patients. Project 5 Bik is an effective apoptosis-inducing gene that suppresses mammary tumor development in a gene therapy setting on an orthotopic animal model. Bik, known as nbk, is a pro-apoptotic gene that contains only one of the Bcl-2 homology regions, the BH3 domain, and has recently been recognized as an essential initiator of apoptosis. The 18-kDa Bik interacts with E1B 19K and forms heterodimers with various anti-apoptotic proteins, such as Bcl-2 and Bcl-XL and thus inhibits their anti-apoptotic function. We have shown that the Bik gene complexed with a nonviral gene delivery system and delivered intravenously significantly inhibited the growth and metastasis of human breast cancer cells implanted in nude mice and prolonged the life span of the treated animals. In addition, we found that a Bik mutant, BikDD, in which threonine 33 and/or serine 35 were changed to aspartic acid to mimic the phosphorylation at these two residues, enhanced its binding affinity with the anti-apoptotic protein Bcl-XL and Bcl-2, and was more potent than wild-type Bik in inducing apoptosis and inhibiting proliferation of human breast cancer cells. BikDD also suppressed tumorigenicity and tumor-take rate in a mouse ex vivo model. Moreover, Bik mutant (BikDD)-liposome complexes inhibited tumor growth and prolonged life span more effectively than the wild-type Bik-liposome complex in an in vivo orthotopic breast cancer animal model (4). All these results suggest that BikDD is a potential therapeutic gene therapy for breast cancer and we have initiated an animal toxicity study of BikDD/liposome to identify the starting dose for human clinical trials and have submitted a preliminary package to communicate with the FDA. In addition, we have recently constructed a breast cancer-specific expression vector (BCSV) that has been tested in human breast cancer cell lines and animal models of mammary tumor. This BCSV contains a modified TopII promoter, which allows the therapeutic gene to be preferentially expressed in breast cancer cells over the normal cells/tissues. This BCSV will be used to drive the BikDD gene and test its therapeutic efficacy in an orthotopic animal model. We predict this BCSV-BikDD construct will be more specific in suppressing breast cancer cell growth and may have minimal toxicity compared with a non-specific promoter such as the cytomegalovirus (CMV) promoter. In the current application, we propose a phase I clinical trial for BikDD/liposome gene therapy administered by intravenous injection and continuation of plans to develop a preclinical study of combination therapy of BikDD/liposome with other breast cancer-targeting treatments to identify more effective therapies for breast cancer. Specific Aims are (1) Determination of anti-tumor activity of BikDD/liposome via non-invasive imaging systems in mammary tumor-bearing animal models. (2) Development of a targeted gene therapy for breast cancer. (3) Development of combination therapy with BikDD/liposome and chemotherapy. (4) Development of a phase I clinical trial using BikDD/liposome formulation.
Core A We believe that the projects described above represent innovative, well thought-out, hypothesis-driven and highly feasible multidisciplinary translational research. In order to accomplish this research, however, we must also have scrupulous administration, open communication, and meticulous fiscal oversight. Core A, the Administrative Core, will provide leadership and general administration for all activities related to the Breast SPORE. This core will be responsible for all Project and Core resources of the SPORE, and will ensure compliance with all NCI regulations and requirements. Core A will convene all necessary meetings, including the Executive Committee meetings, the Internal and External Advisory Committee meetings, monthly investigators' meetings, quarterly research meetings, lectures, and symposia. Core A will also generate and submit all reports related to SPORE activities, and will ensure compliance with all general, governmental, and specific NCI regulations and requirements, including timely communication and consultation with the NCI project officer and other NCI staff. Another important function of Core A is oversight of all fiscal and budgetary issues. In addition, Core A will coordinate a yearly breast cancer conference. Core A will work closely with Cores B and C to ensure that they have adequate resources to respond to the needs of the SPORE investigators. An important function of the Administrative Core is to coordinate quality control and quality assurance. Areas of particular concern for the Administrative Core include data management, confidentiality assurance issues, and obtaining proper consent. The Administrative Core will be responsible for monitoring for misconduct and fraud. All personnel will be instructed on the absolute necessity of avoiding even the appearance or suspicion of such behavior and the utmost importance of reporting any observation of suspected infractions to the Administrative Core. Evidence of misconduct or fraud will be reported to the NCI staff immediately.
Core B Expert assistance with biostatistical and data management issues is essential to the design and conduct of translational research studies. This support involves issues of study design, including appropriate sample size for primary objectives and power for detecting alternative hypotheses, as well as identifying unmet needs and developing new methodology for biomarker-integrated translational studies and for monitoring the conduct and possible early termination of such studies. Once data are collected, they must be efficiently entered, managed and stored with the highest integrity and attention to data quality, safety and confidentiality. These are the goals of the Biostatistics and Data Management Core. This core will serve as a comprehensive, multi-lateral resource for data acquisition and management, designing clinical and basic science experiments, developing innovative statistical methodology, statistical analysis, and publishing translational research generated by the Breast SPORE. Core B will incorporate sound experimental design principles within each project to increase the clarity and enhance interpretability of study results. This core will provide each SPORE project with tailored analyses, accompanied by novel statistical development as necessary, to reveal apparent and hidden relationships among data. This core will also provide expertise in the design of an integrated data management system to facilitate communication among all projects and cores. This process includes prospective data collection, data quality control, data security, and assurance of patient confidentiality. In addition, Core B will collaborate with all project investigators to facilitate the timely publication of research accomplished through the SPORE. Data quality is of the highest priority for the Breast SPORE application, and painstaking measures have been taken to ensure that all data are correctly, safely, and confidentially stored.
Core C Effective tissue procurement and utilization is vital to the effective functioning of large-scale translational research projects. This is especially the case in research involving samples from early breast cancers, which tend to be very small. Prompt handling, careful orientation, sampling and preservation of these precious specimens are of the utmost importance for proper preservation of architectural and cellular tissue components, and for allowing reproducible and accurate analysis of biomarkers. M.D. Anderson Cancer Center's Breast Cancer Tissue Bank was first established in 1992 in order to meet these important goals, and has flourished since that time. It is IRB-approved and has fully functioning guidelines in place for the collection of written informed consent, blood and tissue procurement, and maintenance of patient confidentiality. Since 1992, tissue has been collected from almost 5000 patients, and over 4034 snap-frozen tissue samples are archived. There are also 10455 samples of non-malignant tissue, and more than 10,000 paraffin-embedded archival samples. This archive is an extraordinarily valuable resource for current and proposed research studies. Core C will subsume the Breast Cancer Tissue Bank and will continue its important mission of obtaining, preserving, organizing, and distributing tissue and surrogate materials to M. D. Anderson investigators. The serum bank started in 2002 contains serum and blood cells from over 450 individuals and it includes patients and healthy subjects. Plans are underway to collect samples from more than 1,200 individuals, including all stages of breast cancer as well as, high-risk and low-risk subjects without known cancer. Core C will work with each SPORE project to ensure maximal, efficient, and highly coordinated procurement, use, and storage of human tissue samples. The Core will establish and maintain a repository of tissue samples (including tumor tissue, premalignant tissue, adjacent non-malignant tissue, peripheral blood lymphocytes, and surrogate tissues) and tumor cell lines for laboratory use, with an effective coding system for all laboratory specimens to ensure patient confidentiality and prevent experimental bias. Continuous communication between the surgeons, research nurses, and pathologists, as well as standardized operating procedures, will allow optimal tissue collection and the accurate processing, analysis, and storage of each sample.
Gabriel N. Hortobagyi, MD, FACP - SPORE Principal Investigator
Mien-Chie Hung, PhD - SPORE Co-Principal Investigator
Constance Albarracin, MD, PhD
Michael Andreeff, MD, PhD
Banu Arun, MD
Benjamin Nebiyou Bekele, PhD
Isabel Bedrosian, MD
Donald Berry, PhD
Melissa L. Bondy, PhD
Malcolm Brenner, PhD, FCRP
Thomas A. Buchholz, MD
Kwai Wa Cheng, BSc, MPhil, PhD
Kevin R. Coombes, MD
Kim Do, PhD
Francisco Esteva, MD, PhD
Juri G. Gelovani, MD, PhD
Joe W. Gray, PhD
Kenneth R. Hess, PhD
Edward Horwitz, MD, PhD
Kelly K. Hunt, MD
Khandan Keyomarsi, PhD
Ralf Krahe, PhD
Rakesh Kumar, PhD
Lei Li, PhD
Frank Marini, PhD
Laurent Meijer, PhD
Gordon B. Mills, MD, PhD
Aysegul A. Sahin, MD
Elizabeth Sphall, MD
Patricia Thompson-Carino, PhD
Susan Tucker, PhD
David Yang, PhD
Dihua Yu, MD, PhD |
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