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Organization of the Organ Systems Branch Current SPORE Programs Information for the Public PART Program Information for Applicants IntraSPORE Communications
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UCSF Bay Area Breast Cancer Translational Research Program
SPORE Director: Joe Gray, PhD Bay Area Breast Cancer SPORE investigators pursue translational research on development of novel, molecularly targeted therapeutic agents and on identification of molecular markers that predict disease recurrence or response to therapy. All projects are based on and contribute to our knowledge of the basic genetic and biological events that enable cancer genesis and progression. The suite of therapeutic and marker studies now under study address all aspects of breast cancer progression. This research is carried out in 5 Projects and 4 Cores. New ideas and investigators are bought into the SPORE through Developmental Project and Career Development awards. Project 1 is developing immunoliposomes for targeted delivery of a suite of therapeutic agents. Project 2 focuses on identification of markers that improve prediction of response to RTK pathway inhibitors. Project 3 is exploiting a new in vitro model of early cancer development to identify markers of risk and targets for chemoprevention. Project 4 is developing therapeutic agents that inhibit proliferation and induce apoptosis by forcing telomerase misfunction. Project 5 is a population-based effort to identify predictors of recurrence in women with DCIS. Core 1 provides administration and organizational support for all SPORE investigators. Project 6 is exploring the development of therapeutic antibodies to treat ERBB2 negative tumors. Core 2 collects and manages tissue resources and outcomes information on breast cancer patients for SPORE investigators. Core 3 supports preclinical evaluations of novel therapeutic agents in xenograft models. Core 4 supports the involvement of breast cancer advocates in SPORE research. Two current Development Projects are (a) the role of type 1 collagen remodeling during breast carcinogenesis and (b) the role of PI3-kinase isoforms in mediating breast cancer motility and invasion. The current career development awardee is developing an approach to breast cancer treatment using inhibitors of integrin signaling. The SPORE is located organizationally in the Cancer Center Breast Oncology Program and takes full advantage of 13 Cancer Center Cores.
Project 1. Targeted Drug Delivery Via Immunoliposome Technology We have developed immunoliposome technology for targeted intracellular drug delivery. Immunoliposomes (ILs) were designed to provide maximal efficacy via a receptor-targeted and internalizing drug carrier that is stable, long circulating, non-immunogenic, and versatile. We have previously constructed anti-HER2 ILs, which showed efficient binding and internalization in HER2-overexpressing cells in vitro and in vivo; immunoliposomes loaded with doxorubicin (anti-HER2 ILs-dox) demonstrated marked antitumor efficacy in multiple HER2-overexpressing models. Anti-HER2 ILs-dox has been systematically optimized with respect to each component (MAb fragment, liposomal drug, conjugation method) and developed for clinical trials. The modular organization of immunoliposome constructs makes possible a combinatorial strategy for the generation of new therapeutics: monoclonal antibody (MAb) fragments, derived from available MAbs or newly selected from phage antibody libraries, can be coupled to an appropriate liposomal drug, chosen from a repertoire of liposomal drugs. We will therefore apply this platform technology to develop new agents, and will prioritize these for highest clinical potential and rapid development. Our specific aims are to (1) complete advanced preclinical studies and clinical translation of anti-HER2 ILs-dox. We will collaborate with NCI and a new pharmaceutical partner to complete GMP manufacturing, formal toxicology, and IND filing. In parallel, we will develop assays of binding, internalization, and drug delivery that can be applied to clinical studies for mechanistic evaluation, which will be performed in collaboration with the newly awarded Molecular Assessment Program at UCSF. (2) Develop anti-HER2 ILs for targeted delivery of additional anticancer drugs. Anti-HER2 immunoliposomes can be used to deliver potent small molecules in addition to doxorubicin, including other FDA-approved chemotherapeutic drugs as well as novel compounds that we have recently encapsulated in liposomes. (3) Develop immunoliposomes targeted to other breast cancer-associated antigens, anti-EGFR ILs. We have constructed anti-EGFR immunoliposomes, using available MAbs and newly selected scFvs, and will evaluate them for EGFR-targeted drug delivery. (4) Develop immunoliposomes targeted to tumor-associated endothelial antigens, anti-VEGFR2 ILs. We have constructed anti-VEGFR2 immunoliposomes and will evaluate them for VEGFR2-targeted drug delivery in endothelial cells. (5) Implement clinical translation of new construct(s). Immunoliposome agents that appear the most promising based on Aims 2-4 will be moved to a development track for clinical testing, using our established production protocols and model for collaborative drug development. Project 2. Molecular Determinants of Response to RTK Pathway Inhibitors Deregulation of receptor tyrosine kinase (RTK) signaling is now recognized as important in the genesis and progression of human epithelial cancers, including breast cancer. This knowledge has stimulated the development of a broad range of RTK pathway inhibitors as anticancer agents. Iressa (ZD-1839) and Herceptin are two examples that are now in clinical use. Unfortunately, responses to these and other RTK inhibitors vary dramatically, even between patients that over express the target receptor. We postulate that this is due, in part, to the considerable variation in RTK pathway function that occurs downstream of the receptor. We believe that aberrations that will interfere with response to upstream RTK inhibitors and/or that predict unexpected response can be detected by analysis of a collection of 60 breast cancer cell lines that carry the same spectrum of RTK pathway aberrations as found in primary breast tumors. Use of cell lines to predict inhibitor response failure is attractive because of the savings in cost and time and because the information can be obtained before early phase clinical trials begin so that it can be used to guide trial design. Thus, the central goal of this project is to test this hypothesis. Should this approach prove out, it would have broad utility for development of predictive strategies for the wealth of RTK inhibitors and other targeted therapeutics that are now under development and would substantially increase the speed and decrease the cost of agent trials. This will be accomplished in three aims. (1.) Biological responses and molecular profiles will be measured before and during treatment with Iressa and Herceptin for 60 breast cancer cell lines and analyzed to identify aberrations that interfere with Herceptin- or Iressa-induced changes in mitotic index, motility, growth rate, and/or apoptotic rate. (2.) Molecular predictors of response developed in Aim 1 will be tested in vitro by modulating the expression of RTK pathway genes predicted to influence response to Herceptin and/or Iressa and in vivo by examining biological and clinical responses of human breast cancer xenografts. 3. Predictors of response will be tested in 200 patients treated with Herceptin as a single agent and, if successful, in ~1400 samples from patients enrolled in the NSABP B-31 trial in which samples are being collected from 2700 patients treated with chemotherapy +/-Herceptin. Predictors of response to Iressa will be tested in 40 patients treated with Iressa as a single agent. Project 3. A New Model System to Identify Markers for Risk and Targets for Chemoprevention We have identified a rare subpopulation of human mammary epithelial cells (vHMEC) obtained from women with no known breast disease. These cells express a phenotype that allows them to enter a period of growth that is extremely vulnerable to mutagenesis and is typified by loss of specific cell cycle control and accumulation of a tremendous number of chromosomal abnormalities. Should these cells arise in vivo , they could be the origin of the initial lesions of human breast cancer. These observations identify novel opportunities, providing potential markers for assessing susceptibility to neoplastic transformation in individuals as well as potential targets for prevention and therapy. Multiple markers clearly identify the different cellular states and may allow the identification of these cells in vivo . Remarkably, the earliest lesions in breast cancer, hyperplasias, demonstrate abnormally controlled proliferation but relatively few chromosomal structural abnormalities, a phenotype similar to early-passage post-selection HMEC. The more progressed lesions in breast cancer, DCIS (ductal carcinoma in situ ), additionally demonstrate the types of chromosomal aberrations observed in late-passage HMEC. We hypothesize that the above-described properties of HMEC in vitro are critically relevant to their transformation processes in vivo . We hypothesize that this in vitro model system will not only yield insights to the earliest steps in carcinogenesis in breast cancer, but will also allow us to identify and test new agents for prevention based on the molecular features of the HMECs as they transform. We hypothesize that the frequency of these cells in high risk women will be a possible marker for targeted prevention studies and postulate that agents that selectively kill post-selection HMEC should decrease the incidence of breast cancer. We will use this newly acquired data to take our SPORE project in a new direction and devise a model system for studying risk assessment and chemoprevention of breast cancer in vivo . The general goals of this new effort are to use this system to identify molecular characteristics of post-selection HMEC as endpoints to monitor the potential for malignant transformation activity and then use these to evaluate candidate chemopreventive agents. We will achieve this by (1) determining the frequency, distribution and defined molecular characteristics of vHMEC in vivo in women at low risk for developing breast cancer, (2) determining if the frequency, distribution and defined molecular characteristics of vHMECs in tissues from individuals at high risk for breast cancer are different as compared to tissue from low risk individuals, and (3) identifying agents which will selectively kill vHMECs in vitro and providing information to test their utility as preventive agents in vivo. Project 4. Breast Cancer Therapeutic Agents that Force Telomerase Misfunction This Breast Cancer SPORE Project focuses on a new strategy: to turn the action of an active telomerase against the breast cancer cells. In the course of the previous funding cycle, we have successfully demonstrated that a low threshold of expression of mutant-template telomerase RNA (MT-hTer) genes in human breast cancer cells is sufficient for a potent killing and growth inhibitory effect on these cells. This answers the main question we had sought to address: can telomerase be a molecular target for anti-cancer therapy. For the new five-year funding period the goal of the project is therefore the exploitation of these novel, surprisingly potent and dominant effects, which are based on "toxic" telomeres, as a potential anti-cancer therapeutic strategy: The specific aims for the next 5 years are: ( 1) Test the effects of delivering MT-hTer genes by receptor (HER2)-targeted immunoliposomes to HER2-overexpressing human breast cancer cells. Test cells a) in culture, b) grown as tumors in the nude mouse xenograft system. Using the same delivery and nude mouse xenograft system, test the effects on tumor growth of delivering telomerase RNA-specific oligonucleotides, designed to induce MT-hTer-mimetic effects ("AbTel" oligonucleotides). (2): Test MT-hTers for the ability to prevent the growth of breast epithelial tumor cells taken directly from surgically removed human tumors or collected by ductal lavage. Similarly, test any "AbTel" oligonucleotides identified in Specific Aim #1. Determine whether individual cancers with different genetic properties differ with respect to the "toxic telomerase" effects, with the goal of eventual tailoring of therapy to individual tumors. (3): Determine which MT-hTer mutants and which gene delivery vectors have the most potent effects on cell growth and apoptosis rates. (4): Determine whether combining MT-hTer with a) anti-wild type telomerase RNA ribozyme, b) small interfering RNA directed against WT-hTER or telomerase hTERT mRNA, of c) anti-sense nucleic acids, increases the potency of the MT-hTer effects on cell growth and apoptosis rates. Project 5: Predictors of Recurrence in Women with DCIS We propose to define the risk of disease recurrence or progression by clinical and histopathologic characteristics and molecular and genetic markers in a population-based study of women with ductal carcinoma in situ (DCIS) of the breast. Three study cohorts will be evaluated: (1) a population-based cohort of 1,744 women diagnosed with DCIS from 1983-96, identified by the Northern California Surveillance, Epidemiology, and End Results Program, who underwent lumpectomy alone for treatment, (2) women diagnosed with DCIS whose tissue has been collected at UCSF for the Breast Oncology Program database (N= 457), and (3) a subset of the 1,310 women initially diagnosed with DCIS who underwent lumpectomy and radiation therapy in the NSABPB-17 and NSABPB-24 trials. We will determine the risk of recurrence or progression of DCIS and/or invasive cancer in women treated by lumpectomy alone according to: (1) nuclear grade, necrosis type, margin width, and established molecular markers (p53, erbB-2, ER, PgR, and Ki67); (2) candidate molecular markers of cell cycle dysregulation (Cyclin D1, p16 methylation), tumor invasiveness (e.g., VEGF, factor VIII related antigen, Cox-2), and stromal-breast epithelium interactions (e.g., uPA and related proteins, E-cadherin, b1 integrin); and (3) a subset of chromosomal alterations detected by array-based comparative genomic hybridization. In addition, to validate our findings, we will evaluate women that underwent lumpectomy and radiation therapy to assess that the same measures that predict recurrence among women treated by lumpectomy alone also predict recurrence among women who received adjuvant radiotherapy (Group 3). The results of these studies will be used to develop a risk assessment tool based on measures that have been found to be independently associated with recurrence in a multivariate model. The risk assessment tool will estimate the risk of recurrence for an individual woman as a function of her clinical (e.g., age at diagnosis, menopause status) and histopathology (e.g., nuclear grade, margin width) information, established molecular markers (e.g., ER status, Ki67), and candidate markers. Project 6: Identification and Targeting of Novel Breast Cancer Antigens for Antibody Based Breast Cancer Therapy (Formerly Developmental Project 1) A major goal of cancer research has been to identify antigens on the surface of tumor cells which are qualitatively or quantitatively different from antigens on normal cells. The presence of such antigens can be targeted by monoclonal antibodies, an important new class of therapeutic agents. Herceptin, the first such drug approved for beast cancer therapy, targets the growth factor receptor ErbB2. The goal of this project is to select antibodies directed against novel antigens that are overexpressed on the surface of breast cancer cells that have growth inhibitory or apoptosis inducing effects. We will concentrate on generating therapeutics for the approximately 70% of tumors that do not express high levels of ErbB2. Antibodies will be generated from human antibody gene repertoires displayed on the surface of filamentous bacteriophage (antibody phage display). This will yield antibodies which are entirely human in sequence and optimal for treatment of human disease. The approach relies on probing the surface of breast tumor cells with phage antibody libraries to identify differentially expressed cell surface antigens. The enriched phage antibody pool will then be screened for tumor cell specificity by flow cytometry using the collection of well-characterized breast cancer cell lines developed by the interSPORE cell line consortium and maintained in Project 2. Further characterization will be performed using the native 25kDa single chain Fv antibody fragment (scFv) which is displayed on the phage surface. ScFv antibodies binding antigens that are overexpressed in breast tumors will be evaluated for their ability to inhibit tumor cell growth or induce apoptosis in vitro in a high throughout assay allowing the screening of hundreds to thousands of antibodies. Inhibiting antibodies will be assessed to: (1) identify the antigen bound by scFv; and (2) construct human IgG from scFv genes and evaluate anti-tumor activity in an appropriate xenograft model. Such studies are described in detail in an expanded proposal in the appendix. After 5 years, we anticipate identification of multiple lead candidate antibodies, one of which will be taken into clinical trials for breast cancer therapy. We also anticipate that most scFv will trigger receptor-mediated endocytosis. Some of these scFv may be transferred to Project 1 for development of immunoliposomes. In addition, tumor antigens recognized by the antibodies may prove useful as vaccine candidates or as targets for small molecule drug discovery.
Developmental Project 3: Role of Type 1 Collagen During Epithelial Carcinogenesis We hypothesize that type I collagen remodeling is a rate-limiting step for neoplastic progression in general, and not limited to skin carcinogenesis. To test this hypothesis, we propose to intercross MMTV- neu transgenic mice predisposed to mammary carcinoma development with Collagen r/r mice to determine the functional significance of Type I collagen remodeling during mammary carcinogenesis. Collagen r/r mice provides us with a unique opportunity to define the functional role of collagen remodeling during tumor development and to provide proof-of-principle that eliminating productive access to a critical MMP substrate is an efficacious intervention for neoplastic progression. Our hope is that understanding the importance of this equilibrium will demonstrate that altering the productive access of critical extracellular proteinases to their major targets represents a viable approach for development of novel anti-cancer therapeutics.
Developmental Project 4: The Role of Individual PI3-kinase Isoforms in Mediating Breast Cancer Motility and Invasion A number of genes in the phosphoinositide 3-kinase (PI3-kinase) signaling pathway have been shown to lie in regions of increased copy number in a number of different tumor types. For example, PIK3CA, the p110 a catalytic subunit of type I PI3-kinases is amplified in ovarian, cervical, head and neck and lung cancers. Although the current evidence seems to implicate PIK3CA as a predominant target of tumor-derived genetic alterations, other components of the PI3-kinase are also affected. For example, in a panel of primary breast tumors and breast cancer cell lines, p110 b is increased in copy number in 14% cases, whereas p110 d copy number is decreased in 21% cases. The different roles of individual PI3-kinase catalytic subunits is only beginning to be understood, through a number of strategies including targeted gene knockouts (for example, the phenotypes of p110 a -/- mice are quite distinct from those of p110 g -/- mice) and inhibitory antibody studies [Vanhaesebroeck, 1999 #2700]. In this developing project, we propose to examine the role of different type 1 PI3-kinase catalytic subunit isoforms in breast cancer cells, by inducible expressing each subunit ( a , b , g and d , together with regulatory subunits if required) in selected breast cancer cell lines. Migration and motility will be carefully studied, as recent data suggests that these biological effects are controlled by specific PI3-kinase isoforms. Additional phenotypes associated with PI3-kinase activity, such as invasion, proliferation and survival will also be examined.
Career Development Project 1: Cell Surface Receptor Mediated Therapy for Breast Cancer Recent developments of biologic agents aimed at targeted surface receptors have shown great promise. b 1 integrin belongs to the family of heterodimeric transmembrane integrin receptors and plays a critical role in cell-ECM interactions influencing cell polarity, adhesion, and migration. Aberrant b 1 integrin signaling has been associated with malignant progression and metastatic potential in breast cancer models. Conversely, normalization of b 1 integrin expression in human mammary tumor cells, using monoclonal antibody therapy (AIIB2) has been associated with restoration of a non-malignant phenotype. Our long-range goal is to determine how b 1 integrins influence cancer progression, and ultimately to develop strategies to abrogate the effects of aberrant signaling. Our initial studies will focus on aspects of clinical-translational biology that will lead to a better understanding of how b 1 integrin biology may be exploited for clinical treatment in the future. We will investigate the prognostic value of b 1 integrin in a series of human breast cancer specimens to identify a subgroup of patients who may benefit from this therapy. In parallel, we propose to investigate the potential efficacy of anti- b 1 integrin monoclonal antibody (AIIB2) as a novel therapy in breast cancer in vivo. We will test the ability of AIIB2 to abrogate cancer growth in nude mice models of primary, localized breast cancer, and bone metastasis. In addition, we will evaluate the potential for AIIB2 to be used in combination with radiation therapy, as a tumor cell sensitizer using a physiologically relevant 3-dimensional breast cancer model.
Administrative Core The Administrative Core is the backbone of the Bay Area Breast Cancer Translational Research Program. It supports all of the projects, cores, and investigators of the SPORE by providing assistance with communications, grants administration, and budgetary oversight. The Core facilitates and stimulates interaction between SPORE investigators by organizing SPORE meetings including the weekly seminar series, the annual retreat, the annual External Advisory Board meeting, and other ad hoc research and clinical meetings. The Core is responsible for coordinating the Career Development and Developmental Project Program of the SPORE and maintains the SPORE web site. The Administrative Core consists of the SPORE Director, the Executive Committee, the Program Manager, and an Administrative Assistant.
Tissue-Outcomes Core The Tissue and Outcomes Core is responsible for identifying women diagnosed with breast cancer at the University of California, San Francisco (UCSF, including San Francisco General Hospital - SFGH) and the California Pacific Medical Center (CPMC), collecting fresh tissue (at CPMC) and paraffin blocks (at all sites), collecting and entering clinical, epidemiologic, pathology and follow-up information into a database (for all sites), and distributing breast tissue with associated clinical information to SPORE investigators. Tissue is collected prospectively at the time of surgery (following consent) , and banked as fresh-frozen cassettes or formalin-fixed blocks. Fresh tissue for culture is also collected. The Core also identifies archival formalin blocks for studies , and coordinates with other tumor banks to obtain additional material for investigators. All tissue is reviewed by Core pathologists for histopathologic features and to confirm the presence of tumor. Requests for tissue and clinical data are approved by a Tissue Utilization Tissue & Data Utilization Committee, which reviews requests for project feasibility and priorities. The Core extracts DNA and RNA for studies , and coordinates preparation of tissue microarray blocks. In addition to baseline data collected on women with newly diagnosed breast cancer, the Core obtains informed consent for tissue use and follow-up information to determine disease status on all women in the database by mailing women a survey every 18-months. Annual linkage is done with the Northern California Surveillance, Epidemiology, and End Results (SEER) program to determine vital status and disease specific mortality. Baseline and follow-up information from over 2,301 women with breast cancer (1,819 invasive and 482 DCIS cases) diagnosed at UCSF or CPMC with a median follow-up time of five years are currently in a relational database. The overall goal for the next five years is to maintain and expand the database ( by including patients at SFGH , starting in 2002 , and at additional outside sites) so that it can continue to serve as a resource to conduct high quality, clinically significant translational research and to acquire breast cancer outcomes for study populations.
Pre-Clinical Animal Core The Preclinical Animal Core is the primary vehicle by which breast cancer investigators generate pre-clinical data in vivo and mechanistically validate hypotheses generated in vitro . Pre-clinical animal tumor model systems are essential to evaluate the anti-tumor efficacy, pharmacokinetics and biodistribution of innovative anti-tumor agents. The Preclinical Animal Core assists in experimental design involving animals, performs technical aspects of pre-clinical testing in vivo, maintains a well-characterized repository of breast cancer cell lines and primary tumors optimized for passage in animals, develops and incorporates new tools and resources for preclinical testing, and fosters inter-SPORE collaborations involving in vivo tumor models. The availability of centralized cell and animal resources, and personnel with expertise in tumor cell propagation in vitro and in vivo, drug delivery and measurement endpoints, ensures appropriate experimental design and data reproducibility, compliance with local and federal regulatory guidelines for tumor-bearing animals, and maximum resource utilization through coordinated animal purchasing and housing. Human tumor xenografts implanted into the mammary fat pads of immune-deficient mice ( nu/nu ) are the primary pre-clinical model used by most SPORE investigators. Cell xenografts are selected based upon the genomic and phenotypic features desired for hypothesis testing and agent evaluation. The Core also develops and explores new assays and resources to enhance the ability of SPORE investigators to quantify induce-perturbations at multiple levels such as proliferation, invasion and metastases. The main services/resources provided by the Preclinical Animal Core are to: (1) Design and conduct preclinical studies using xenografts to support the development and translation of innovative anticancer therapies; (2) Enhance investigator capabilities to investigate the anti-proliferative effects of novel agents on early tumors and disseminated disease by optimizing assays to quantify tumor dissemination using species-specific DNA probes and in vivo tumor cell imaging with bioluminescent reporters; (3) Consult and assist SPORE investigators on potential uses of xenograft and other animal models of human breast cancer and promote inter-SPORE collaborations with investigators using pre-clinical animal tumor models.
Advocacy Core The Breast SPORE Advocacy Core (BSAC) is comprised of 12 members, 25 associates and >65 supporters who have experienced breast cancer. These individuals support Bay Area Breast Cancer SPORE projects and cores by infusing relevant patient experiences into the SPORE, and by addressing recurrent barriers to translational research that may not otherwise be addressed. The BSAC provides specific services through four advocate-led teams: An Education Team develops forums that promote communication, education, and awareness between researchers, breast cancer patient advocates, and the public to make SPORE projects applicable to people with cancer. A Clinical Trials Team provides clinical trial assistance on protocol and consent form development, accrual, and education for approved Breast Care Center-initiated clinical protocols, and on each SPORE project as it reaches preclinical development. A Tissue Team facilitates increased specimen collection by improving communication and helping resolve issues between the Breast SPORE and pathologists, surgeons and researchers at each collection site. A Policy Issues Team develops strategy sessions for the Breast SPORE program on emerging policy and ethical issues in translational research. The BSAC has also initiated inter-SPORE activities that include establishing advocate participation in other SPORE programs, providing tools to accelerate accrual to an inter-SPORE clinical trial, and facilitating inter-SPORE patient advocate discussions and activities. Bay Area Breast Cancer SPORE Investigators Joe W. Gray, PhD Amy Akbarian Christopher Benz, MD Elizabeth Blackburn, PhD Imok Cha, MD Karen Chew, ASCP Koei Chin, MD, PhD Colin Collins, PhD Shanaz Dairkee, PhD Peggy Devine, CLS Laura Esserman, MD, MBA Byron Hann, MD, PhD I. Craig Henderson, MD E. Shelley Hwang, MD Ajay Jain, PhD Karla Kerlikowske, MD Britt-Marie Ljung, MD Carolene Marks James D. Marks, MD, PhD Frank McCormick, PhD Catherine Park, MD John W. Park, MD Jane Po Hope S. Rugo, MD Susan Samson, MPH Bambi Schwartz Thea Tlsty, PhD Linda Vincent, MPH Frederic Waldman, MD, PhD |
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