Tyler J. Curiel - US grants

DESCRIPTION (adapted from the applicant's abstract): Infection with Toxoplasma gondii (Tg) is an important cause of morbidity and mortality in persons infected with HIV or who have other defects in cell-mediated immunity. Treatment of toxoplasmosis in HIV-infected patients is complicated by the need for life-long therapy, the increased tendency to allergic reactions to drugs in this population, and by the overlapping toxicities of some antiretroviral and anti-Tg drugs. Clearly, better therapeutic agents are needed. A better understanding of the immune response to this important intracellular pathogen might someday lead to an immunotherapeutic to serve as an adjunct to anti-Tg chemotherapy. The principal investigator's laboratory has investigated the human cellular immune response to Tg over the past four years, as a considerable body of evidence suggests that protection against toxoplasmosis is mediated primarily by cellular defenses. Unlike in mouse models, where Tg- specific, class I-restricted CD8+ cytotoxic T lymphocytes (CTL) have been shown to be pivotal effectors, human Tg- specific CTL generated in the applicant's laboratory are primarily class II-restricted and CD4+. Although most CTL described to date are CD8+ and HLA class-I restricted, recent evidence has demonstrated that in many infections, CD4+, class-II restricted CTL are important mediators of cellular immunity. These studies are relevant not only from the standpoint of understanding immunity to an important HIV- associated opportunistic pathogen, but also from the perspective of furthering the understanding of the role of CD4+ CTL in immunity. Intracellular tachyzoites form a structure known as the parasitophorous vacuole (PV), which does not fuse with phagosomes, does not acidify normally, and which acts as a molecular sieve, controlling the entry and egress of various molecules. Recent work in the applicant's laboratory suggests that Tg may use its PV to evade host immune responses by decreasing Tg antigen availability for class I processing. Accordingly, it is hypothesized that human anti-Tg CD4+ CTL arise because the Tg PV plays a role in exclusion of Tg antigens from class I processing. It is also likely that endogenous Tg is processed by, and presented in the context of, the class II pathway. The specific aims of the application are: 1) to evaluate the role of the Tg PV in the processing and presentation of Tg antigens; 2) to look for cytoplasmic processing of Tg antigen in the class II pathway; and 3) to examine the biology of CD4+ compared to CD8+ Tg-specific CTL.

DESCRIPTION: (Adapted from Applicant's Abstract) Immunity to HIV infection includes both cellular and humoral mechanisms. Dendritic cells (DCs) are good candidates to elicit HIV-specific immunity but optimal means to use them are not established. Three significant variables are i) the specific DC subset involved, 2) the antigens (Ags) used, and iii) the route of Ag acquisition by DCs. There are 3 well-characterized DC subsets identified in humans: Langerhans, interstitial, and lymphoid DCs. Several studies show that DCs elicit HIV-specific cellular immunity, but only studied monocyte-derived DCs. Ag formulation (apoptotic vs. necrotic) and acquisition pathway also influence elicited immunity, although little systematic study has been reported. A novel DC type, the macrophage (M)-derived DC, may have advantages over other DC subsets in eliciting HIV-specific immunity. The goal is to determine which DC subsets, acquiring Ag in specific contexts, elicit immune effector cells thought to be important in preventing HIV infection, and in controlling HIV replication. The focus on induction of cellular immunity because of the investigators expertise in that area. The hypothesize is that if optimal DC subsets and Ag acquisition pathways are identified in vitro, then ultimately specific targeting strategies may be used to make a convenient vaccine preparation that does not require the ex vivo growth or manipulation of autologous DCs. There are 3 specific aims: #1: to determine the differential ability of 3 DC subsets, interstitial or epidermal DCs derived from CD34+ HPCs and DCs produced from M-CSF derived Macrophages to activate HIV gag, pol, env or nef-specific T cells when Ags are acquired in two distinct contexts (endogenous production or exogenous capture). #2: to determine whether the specific effector cells elicited are those thought to prevent HIV infection, examining cytotoxicity, and cytokine/chemokine production (THl/TH2 and TCl/TC2 immunity); and #3: to determine the non-cytolytic capacity of these elicited immune cells to inhibit HIV

DESCRIPTION (provided by applicant): Hepatitis C is one of the worlds most pandemic and insidious diseases. Less than 41% of patients respond to the current treatment, and a large fraction is ineligible for therapy. Thus, there is an urgent need for new therapeutic strategies. Clearance of hepatitis C virus (HCV) is correlated with the level of HCV-specific CD4+ T cells, and viral escape mutations have been identified in immunodominant CD4+ T-cell epitopes. These results suggest that an immunotherapy designed to increase and broaden HCV-specific CD4+ T cells could provide a new therapeutic approach. Dendritic cells (DCs), the most important class of professional antigen presenting cells, possess the ability to elicit both humoral and cellular immune responses. These cells are poised to capture pathogens, migrate to draining lymph nodes, and select antigen-specific CD4+ T cells to regulate T, B, and NK cells, all of which may contribute to protective immunity. The objective of this proposal is to develop a novel vaccine strategy targeting the HCV nonstructural protein 3 (NS3) directly to DC subsets, e.g., Langerhans Cells (LCs). Recently we showed that LCs can be generated by culturing monocytes with GM-CSF+IL15. Such LCs induce significant T-cell activation in vivo. Furthermore, we have generated peptides that bind specifically to LCs or interstitial DCs from a phage display peptide library. We hypothesize that targeting NS3 directly to DCs will increase the level and duration of specific immune responses. Thus, we will target NS3 to DCs by coupling or fusing it to DC-specific peptides. We further hypothesize that NS3 can be structurally modified in order to eliminate the immunodominant epitopes and therefore recruit new T cells against HCV. Specific aims are: 1) To determine whether targeting NS3 specifically to DC subsets enhances specific immune responses against HCV by analyzing T-cell proliferation/activation in humanized SCID mice; and 2) To augment the development of IFN3 gamma-producing NS3-specific CD4+ T cells by engineering the three-dimensional structure of HCV NS3. Alternative modes of loading DC subsets will be explored, including via recombinant Lactobacillus sp. that express and secrete DC-targeted NS3. A needle-less and non-toxic immunotherapy would provide a treatment for hepatitis C patients who currently have none.

recombinant proteins; human therapy evaluation; neoplasm /cancer immunotherapy; immunopharmacology; intravenous administration; drug screening /evaluation; interleukin 2; clinical trial phase II; clinical trial phase I; helper T lymphocyte; liver function; kidney function; neoplasm /cancer immunology; longitudinal human study; patient oriented research; clinical research; biotechnology; human subject;

DESCRIPTION (provided by applicant): To study SDF-1/CXCR4 in human tumors, we isolated immune cells in ascites and lymph nodes of patients with ovarian carcinomas. We will present extensive evidence suggesting significant and previously unknown roles for SDF-1/CXCR4 signals in tumor immunopathogenesis through distinct immune effects on plasmacytoid dendritic cells (PDC) and regulatory T cells (Tregs).Thus, SDF-1/CXCR4 signals may be a final common pathway mediating significant and diverse tumor immunopathologic mechanisms. We hypothesize that SDF-1/CXCR4 interactions play a significant role in ovarian cancer pathogenesis by enhancing immune evasion, metastasis, tumor growth and neoangiogenesis. This hypothesis predicts that interrupting SDF-1/CXCR4 signals will be therapeutic through several distinct mechanisms, not just by blocking metastasis. Prior studies of chemokines have been hampered by promiscuity: one chemokine may bind several receptors, and one receptor may bind several distinct chemokines. However, SDF-1/CXCR4 is a unique ligand/receptor pair. Thus, study of this interaction is tractable, and offers significant opportunities to test the effects of blocking such interactions as an anti-tumor strategy in a model where definitive conclusions regarding ligand/receptor interactions can be drawn. This proposal focuses on immunological aspects of this interaction. Our over-arching hypothesis is that the net effect of blocking SDF-1/CXCR4 is to reduce tumor burden through augmenting tumor-specific immunity. Tumors also express cytokines (IL-l0 and TGF-beta) and ligands (B7-H1) that may induce Treg differentiation. Thus, we hypothesize that the tumor itself is an alternative or complementary source of Treg differentiation. Tumor SDF-1 may play a significant role in this regard. Our specific aims: 1. Test the hypothesis that SDF-1/CXCR4 signals mediate immunopathology through control of PDC migration, survival or function in vivo. 2. Test the hypothesis that SDF-1/CXCR4 signals mediate immunopathology through control of Treg migration, survival or function in ovarian cancer. 3. Test the hypothesis that the net effect of blocking SDF-1/CXCR4 is to reduce tumor burden through augmenting tumor-specific immunity using our well-defined TAA-specific systems. 4. Test the hypothesis that SDF-1/CXCR4 signals mediate Treg differentiation in tumors. We will use SDF-1( MMC and LM-1 tumors and their SDF-1- derivatives produced using siRNA technology.

The San Antonio Cancer Institute (SACI) represents the combined cancer research programs of a private, not-forprofit institution, the Cancer Therapy and Research Center (CTRC), and a state-supported academic medical center, The University of Texas Health Science Center at San Antonio (UTHSCSA). Since the inception of the CTRC in 1972 and its dedication in 1974, the two institutions have had a long and successful history of collaboration in cancer treatment and cancer research. The UTHSCSA brings significant attributes to the partnership, including its reputation as an outstanding academic institution, strong and diverse programs in the basic sciences, and equally strong clinical research capabilities. The UTHSCSA provides the facilities and faculty to train new cancer research investigators, particularly those interested in translational research. The CTRC contributes comprehensive outpatient cancer treatment, outreach, and education capabilities to the partnership. This nationally known treatment center registers more than 100,000 patient visits annually. The CTRC also includes the Institute for Drug Development as one of its subsidiaries. This world-class anticancer drug development organization is the largest such program capable of taking new anticancer agents through "first-in-man" (phase I) clinical testing and into early safety and efficacy trials (phase II). The seven research programs presented for review in this application include: Cancer Metastasis, Cancer Prevention and Health Promotion, DNA Repair and Tumor Suppressor Genes, Experimental Therapeutics, Prostate Cancer, Geriatric Oncology, Signal Transduction and Macromolecular Interaction. SACI members have access to fourteen proposed Shared Resources that provide technology and expertise to enhance research productivity and scientific collaborations within the Institute. The SACI Shared Resources presented for approval in this application include the following: Antigen and Antibody Production, Biostatistics and Medical Informatics, Clinical Protocol and Data Management, Cytogenetics and Genetics Resource, Flow Cytometry, Genetic Mouse Models, Laboratory Animal Resources, Macromolecular Structure, Mass Spectrometry, Microarray, Optical Imaging, Pathology, Pharmacology, and Protein-Protein Interactions. Three programs are targeted for development over the next five years: our emerging programs in pediatric oncology and gastrointestinal malignancies, and a new Molecular Therapeutics Program that will expand SACI?s medicinal chemistry and high throughput screening capabilities, and facilitate the in-house discovery of new anticancer agents that will be fed into SACI?s drug pipeline-from identification of the molecular target to screening, chemistry, formulation, testing in animal models, toxicology, and, ultimately, to testing in humans.

DESCRIPTION (provided by applicant): Two human dendritic cell (DC) subpopulations are recognized: myeloid (MDC) and plasmacytoid (PDC), which mediate distinct immunologic functions. Malignant ascites contained a striking accumulation of PDC, whereas MDC were entirely absent. Tumor draining lymph nodes (LN) contained both PDC and MDC although with marked phenotypic and functional differences compared to blood DC. Little regarding PDC mediated immunity is known. CD4+CD25+ regulatory T cells (Tregs) are elevated in human cancers where they inhibit TAA-specific immunity and predict poor survival. We recently demonstrated a role for PDC in immunopathogenesis of ovarian cancer, including induction of Tregs. We hypothesize that tumor PDC-induced Tregs inhibit tumor-associated antigen (TAA)-specific immunity and facilitate tumor growth, thus contributing to cancer immunopathogenesis. Tumor PDC express high-level B7-H1 that induces T cell IL-10. We will test the hypothesis that tumor PDC B7-H1 is a molecular mechanism for Treg differentiation. Factors inducing Treg differentiation are poorly understood. Understanding these processes is central to unraveling the immunopathologic basis of cancer and developing novel immune-based therapies. Our specific aims are: AIM 1: Test the hypothesis that tumor PDC-activated T cells block TAA-specific immunity. We will use well-characterized in vitro models for human immunity to test effects of naive T cell priming and on defined TAA-specific effector cell functions including cytokine secretion, proliferation, and cytolytic activity. We have preliminarily identified a PDC-activated CD8+ Treg which will be further studied. AIM 2: Test the hypothesis that PDC-activated T cells block immune-mediated tumor rejection in vivo. We developed novel chimeric SCID/NOD mouse models for these studies in which autologous human tumor, effector cells and Treg can be studied in vivo. This model allows testing of Treg function, TAA-specific immune suppression and tumor growth in vivo in an autologous setting. Mechanisms will be tested by blocking IL-10, CTLA-4 or other Treg effector arms in vivo.

[unreadable] DESCRIPTION (provided by applicant): We recently cloned the first T. gondii MAP kinase, MAPK1-Tg, which acts as a stress MAPK and has 40% homology to human p38 MAPK. MAPK1-Tg is expressed as the full-length 58 kDa protein in tachyzoites, and is the first member of a novel MAPK family. T. gondii tachyzoites expressing a dominant-negative MAPK1-Tg replicated significantly more slowly than parental parasites in vitro, and expressed significantly more bradyzoite antigens. Most strikingly, these tachyzoites were remarkably attenuated in mouse virulence. These data implicate MAPK1-Tg in control of tachyzoite proliferation, stage differentiation and virulence. We previously showed that pyridinylimidazole drugs designed to block human p38 MAPK activation also blocked T. gondii replication in vitro, cured T. gondii-infected mice, and induced stage differentiation in vitro. Human p38 MAPK inhibitors block MAPK1-Tg activity in vitro. We hypothesize that p38 MAPK inhibitors treat T. gondii infection through inhibition of MAPK1-Tg. This proposal will study MAPK1-Tg with two fundamental objectives: i) to exploit MAPK1-Tg as a novel drug discovery target, and ii) to use MAPK1-Tg as a tool to increase our understanding of T. gondii biology. 67657 is the prototypical p38 MAPK inhibitor used. It was safe in human Phase I trials in arthritis, and could be translated into a human clinical trial as an anti-parasitic agent rapidly. MAPK homologues with structural features of MAPK1-Tg were identified in genomic databases for Plasmodium, Leishmania and Trypanosoma. We hypothesize that parasite MAPKs represent novel, broad spectrum drug development target. Finally, T. gondii is a category B bioterror agent. Thus, these discoveries could lead to novel treatment approaches to combating bioterror. Our Specific aims are: 1 Test hypotheses regarding how MAPK1-Tg regulates replication and stage differentiation. The development of reagents to detect endogenously produced total and active MAPK1-Tg in wild-type parasites, and the development of recombinant parasites with dominant-negative MAPK1-Tg now permit definitive testing of hypotheses regarding how MAPK1-Tg activation regulates tachyzoite replication and stage differentiation, and effects on the cell cycle regulation. Test the hypothesis that 67657 blocks MAPK1-Tg activation. These studies help elucidate factors controlling parasite virulence. 2 Test the hypothesis that blocking MAPK1-Tg activation is a mechanism of therapeutic action of p38 MAPK inhibitors. 3 Test the hypothesis that blocking MAPK1-Tg augments efficacy of anti-Toxoplasma therapies in vivo. [unreadable] [unreadable] [unreadable] [unreadable] [unreadable]

The Cancer Development and Progression (CDP) research program, one of the three current and interactive programs of the Cancer Therapy &Research Center (CTRC) at the University of Texas Health Science Center at San Antonio (UTHSCSA), has evolved from the merging of essential elements of several cancer center programs since the last competitive renewal. The major thematic areas under CDP are: (1) Genomic Integrity, (2) Aging and Cancer, (3) Chronic Inflammation and Cancer, and (4) Women's Cancer. The reorganization reflects some significant changes in the previous program areas, including the departure of several key members and new cancer focuse areas brought to the Center by newly recruited cancer research scientists. The new programmatic structure provides a consolidated platform that is more conducive to collaborative and integrative cancer research. All of the above themes have the potential to develop or re-develop into strong, stand-alone programs in the future, and are expected to do so. The overarching scientific goals of the CDP Program are: (1) to integrate basic research in genomic integrity, age-related cancer susceptibility, tumor microenvironment, and hormone actions and therapeutic resistance in women's cancer to gain a deeper understanding of cancer development and progression;(2) to foster cross-disciplinary collaboration between CDP and the research programs in population studies and experimental and developmental therapeutics;and (3) to translate findings of genetic instability, tumor immunity, obesity/nutrition, and hormone resistance into better cancer prevention and treatment. Currently, the CDP research program has 23 key cancer-focused and funded members and another 25 funded members who conduct cancer-related research that contributes to the understanding of cancer development and progression. Cancer research in the CDP Program receives a total of $7.5 million in peer-reviewed funding (direct), of which $2.4 million is from the NCI (direct). In the current funding period, researchers in the CDP program have made a number of major accomplishments in many areas of cancer biology. These include discoveries of novel factors in double strand DNA break repair, new molecular links between genetic instability and aging, important insight into host-tumor interactions, and molecular interplay of hormone synthesis and actions in breast cancer development. With new cancer research focus, reconfigured interactive programmatic structure, and strong leadership, the CDP Program is well positioned to synergize and integrate further the multi-disciplinary cancer research ongoing in our Cancer Center.

The Cancer Prevention and Population Science (CPPS) research program, one of the three established research programs of the Cancer Therapy &Research Center (CTRC) at The University of Texas Health Science Center at San Antonio (UTHSCSA), has 17 members (13 primary, four secondary) who are focused on, and have made great strides in cancer prevention affecting the residents of San Antonio, South Texas, and the nation. The CPPS Program's key research themes are: eliminating cancer-related health disparities, especially among the region's predominantly Latino population;using cancer control to improve outcomes, especially focusing on children, adolescents, and young adults;and identifying the determinants of cancer and cancer risk and prognosis, especially focusing on prostate cancer. CPPS members have several vital accomplishments in these areas: developing a national research, training, and awareness network on Latino cancer;promoting tobacco prevention and cessation among children and Latinos; developing several patient navigation programs to increase Latino access to cancer care;testing video game technology to improve adherence to prescribed cancer treatment regimens for adolescents and young adults;assessing the relationship between obesity and childhood acute lymphoblastic leukemia in Latinos; creating a prostate cancer risk calculator tool;and identification of risk biomarkers for prostate and other cancers. Program members come from broad scientific backgrounds, such as behavioral science, epidemiology, basic science, chemoprevention, and others across many UTHSCSA departments and schools. CPPS researchers work collaboratively within the program and across the entire CTRC to translate their cancer prevention findings from the laboratory/clinic and population-based studies to bring improved methods of cancer reduction, early detection, diagnosis, treatment, and survival to South Texas and the nation. The program features severarmulti-Investigator projects, including a U10 and several U01 grants, as well as 42 overall projects, 19 of which are NCl-funded at more than $5.5 million in annual directs. Education and training is another important program component, both among membership (through regular CPPS member meetings and cancer prevention/control guest lecturers) and in projects, including several projects geared to training minorities in research and aiding their development in cancer-related fields at the doctoral level. In summary, the CPPS Program is a highly integrated multidisciplinary collaborative effort between diverse disciplines of scientists focused on cancer prevention and control.

The primary purpose of this shared resource is to generate novel polyclonal and monoclonal antibody reagents, to assist investigators in characterization and optimization of these reagents, and to provide other technical assistance for antibody-related research. Availability of high quality antibody reagents is critical for understanding protein function in normal and neoplastic cells and tissues. We aim to provide antibodies to cancer center members in a timely manner at a reduced cost, relative to commercial custom antibody services. We can also provide an expanded range of services that are not generally available from commercial sources, so that more investigators without expertise or equipment for molecular cell biology or biochemistry can successfully generate and characterize custom antibody reagents. Our most commonly used services include the generation of rabbit polyclonal antibodies, purification and/or testing of polyclonal antibody, generation of mouse polyclonal antibodies, and the generation, purification, characterization and testing of mouse monoclonal antibodies. Other services that we have provided include production and purification of bacterially expressed fusion protein antigens, antibody isotyping, cryopreservation and storage of hybridoma lines, preparation of Fab fragments from existing antibodies, adaptation of hybridoma lines to serum-free culture and recovery of poorly frozen hybridoma cell lines from other sources.

The Cancer Center requests $461,731 in annual Developmental Funds to allow the following: 1) Recruitment of new clinical oncologists especially related to Surgical Oncology and Medical Oncology, and basic and translation scientists for our three scientific programs. 2) Support for our pilot project program that is vital to our ability to grow the programs as needed. 3) Support for Shared Resources in Development that are crucial to our ability to provide Cancer Center investigators with the research infrastructure required to meet future programmatic goals. 4) Retention of critical members of the Cancer Center. 5) Bridging of funding for Cancer Center members to redirect their laboratories to a high priority cancer research area to increase their funding potential. The past developmental funds were used in response to the advice of our External Advisory Board (EAB) and our internal advisory committees. To continue to respond to the strategic planning process identified by both our EAB and internal advisory committees, the Cancer Center must invest in further recruitment of high priority senior basic scientists and physician-scientists with a strong cancer focus in organ-specific cancers. The Cancer Center considers continued support for the pilot project program, which was very successful during the past grant period, as key to programmatic growth and innovation.

Cancer Center Administration provides the overall administrative services that further the research and clinical missions of the P30 support grant. Administration has been extensively reorganized to address prior critiques, and to optimize its new position in the matrix Cancer Center formed following the December 2007 merger of the formerly independent CTRC with UTHSCSA. The new Associate Director of Administration has specific responsibilities commensurate with a senior leadership position within the Cancer Center. Administration provides a variety of critical functions in support of P30 operations, including: 1) providing oversight and coordination of P30-supported programs and shared resources, 2) serving as the direct liaison for, and supporting and coordinating the functions and meetings of key scientific and oversight committees, such as the Executive Committee, the Internal Advisory Committee, the External Advisory Board, program member meetings, and scientific retreats, 3) determining policy needs, and writing and implementing those policies for programs and shared resources, 4) providing financial accounting for all P30-related activities ncluding programs, shared resources, pilot programs, seminars, retreats and other activities, 5) coordinating and documenting use of P30-related laboratory, clinical and administrative space, 6) coordinating and overseeing P30-funded personnel, 7) coordinating communications among Cancer Center investigators and leadership, and UTHSCSA personnel, 8) serving as liaison with, and coordinator for funding sources including UTHSCSA and local and national agencies, 9) working closely with the Office of Research Administration to help ensure that clinical protocols and Material Transfer Agreements are executed, documented and maintained to appropriate standards and aligned well with UTHSCSA standards, and 10) coordinating informational communications between P30-supported scientific and clinical research arms and UTHSCSA, the region and the nation. Cancer Center Administration activities do not overiap with administrative functions supported elsewhere in the CTRC or in UTHSCSA, maximizing economies of scale.

The Cancer Center at the University of Texas Health Science Center at San Antonio requests $35,200 in support of the Planning and Evaluation process. The Cancer Center has both an External Advisory Board (EAB) and three Internal Advisory Committees, the Executive Committee (EC) for the day to day management, the Scientific Leadership Committee (SLC) for scientific programs and shared resources development and review and the Internal Advisory Committee for more global issues related to the various Schools and the UTHSCSA in general. The Cancer Center Director and leadership interact very closely with it's EAB through many telephone conferences, sending documents for review on a regular basis by on-site visits by the Chair, Dr. John C. Ruckdeschel, members of the EAB and by an annual visit of the EAB. the EAB reviews and advices Director on all activities of the Cancer Center. Their reviews, evaluations and criticisms have led us to develop much stronger Cancer Center Funds to support the EAB are requested ($25,000). The EC and the SLC work very closely together and with the EAB for Strategic planning and evaluations of the programs and shared resources. The Cancer Center uses center-wide retreats and program retreats to further develop strong and effective programs and shared resources and for future recruitment strategies. We request $10,000 to help with the Cancer Center and program retreats.

Senior Leadership are individuals with the requisite skill sets that in their totality, provide a balanced and complete perspective on our Cancer Center to ensure its optimal growth, function, resource utilization, and intra- and inter-programmatic interactions. These individuals also represent relevant aspects of Cancer Center affairs to the various clinical and research entities that comprise our matrix environment to ensure that the Cancer Center interacts to best advantage and can best capitalize on available external resources. Leadership includes the Cancer Center Executive Director (Dr. Curiel) and Deputy Director (Dr. Giles), our three scientific Associate Directors, Dr. Slaga (Basic and Translational Research), Dr. Naylor (Shared Resources and Internal Education) and Dr. Pollock (Cancer Prevention and Control). Dr. Ramirez serves as Senior Leadership owing to her directorship of the Institute for Health Promotion Research, which serves a critical cancer prevention and Hispanic outreach function for our Cancer Center. We also have an interim Associate Director of Administration with greatly augmented authority and a clear Senior Leadership role as requested in the prior review. A national search for a permanent Associate Director of Administration is undenway, with the expectation that the permanent position will be filled by the time of the site visit. The Senior Leadership ensures that our Cancer Center discharges its public trust through appropriate stewardship of NCI, NIH and related, institutional and philanthropic dollars to translate discoveries into practical applications that provide for the clinical and research cancer care needs of San Antonio, South Texas and the nation. All Senior Leadership members sit on the Executive Committee and other important committees that set current Cancer Center agendas, plan for future growth, challenges and opportunities and help set budgets, recruiting pnorities and scientific agendas that meet these various needs. Support is requested for Dr. Curiel (50%), Dr. Giles (5%), Dr. Naylor (10%), Dr. Ramirez (10%), Dr. Slaga (20%), and Ms. McCarroll (50%). Some Senior Leadership also serve as Program Leaders, but generally those duties are co-shared with Senior Leadership in terms of financial support. Thus, most support for the totality'of the efforts of these individuals comes from Senior Leadership, freeing our scare resources in the budget for additional, constructive uses in our Cancer Center.

DESCRIPTION (provided by applicant): Project Summary. We have shown that regulatory T cells (Tregs) defeat anti-cancer immunity and that their depletion can be therapeutic, but their rapid regeneration is problematic. This project uses our discoveries that B7-H1 immune co-signaling facilitates Treg generation and that estrogen receptor (ER)? signals inhibit Treg regeneration in relevant pre-clinical mouse models with proven substantial translational relevance. Initial studies focus on ovarian cancer (OC) and use melanoma models to demonstrate concepts in additional tumors, as our discoveries should be applicable to a wide variety of cancers. Effects of B7-H1 on ER? signaling and relations to immune pathology and clinical outcomes are studied. Our overarching objective is to identify novel and effective immune therapy for cancers, with a focus on OC. Our overarching hypothesis is that B7-H1 blockade will augment Treg depletion as cancer immunotherapy and that ER? signals will boost B7-H1 blockade effects. This hypothesis predicts novel approaches to immunotherapy for OC, greatly extends our understanding of its immunopathogenesis and allows development of a similar strategy for other cancers and in men. ER? agonists can be used in males as we have shown and avoid estrogen side effects. The specific aims are Aim 1 Test the hypothesis that ER? signals augment B7-H1 blockade effects in cancer and AIM 2 Test the hypothesis that dysfunctional B7-H1 signaling in cancer is dendritic cell-dependent. These aims are achieved using mice genetically null for signaling components, pharmacologic agents affecting key signaling pathways, and bone marrow chimeras to study hematopoietic versus non-hematopoietic B7-H1 signals. Relevance. We seek to improve treatment options for OC, which kills over 15,000 American women annually, and for which there is no curative option after first-line therapy fails, as it doe in most cases. Principles can be applied to a wide variety of cancers, including melanoma, studied here as a confirmatory second cancer. Our insights into tumor-associated immune dysfunction promise to help improve the efficacy of cancer immunotherapy, whose record of success to date has been only modest.

DESCRIPTION (provided by applicant): This application will define mechanisms of rapamycin-mediated tumor onset delay or prevention and will dissect mTOR effects directly on the tumor versus immune cell effects. It addresses Provocative Question 5: defining mechanisms of action of drugs used for other purposes. We challenge the paradigm that mTOR inhibition reduces or prevents cancer by direct effects on tumors through mTOR growth and metabolic effects, and explore the potential for mTOR inhibitors to prevent cancer through mTOR-mediated immune effects. We hypothesize that mTOR inhibition with oral rapamycin delays or prevents cancer onset in part through immune mechanisms, and will test concepts in a well-defined carcinogen-induced skin cancer model in which T cells and IFN-¿ are important protective agents and in which mTOR inhibition prevents cancer. Mice will have tumor induced with dimethylbenz(a)-anthracene (DMBA) plus the promoter 12-O- tetradecanoylphorbol-13-acetate (TPA) and will be treated with oral rapamycin or control. Time to tumor onset, malignant change and tumor size and effects on tumor immune surveillance will be studied as will mTOR signaling in tumor versus other cells. Aim 1 Test the hypothesis that T cells contribute to oral rapamycin- mediated cancer prevention. Aim 2 Test the hypothesis that IFN-¿ contributes to oral rapamycin- mediated cancer prevention. Aim 3 Test the hypothesis that rapamycin prevents cancer by direct effects on tumor cells. mTOR inhibition directly in tumors cell is not mutually exclusive with immune mechanisms. Relevance: Cancer is the number one killer in the US. Cure rates for advanced cancers have changed little in the past 50 years. Prevention is more cost effective and broadly applicable than treatments. We thus propose a novel, safe, broad spectrum approach to cancer prevention using rapamycin as a potential first-in-class agent.

ABSTRACT ? RESOURCE CORE (RC) 2 Pharmacological aging-modulating interventions that have been validated in animal models are ready to be translated into human clinical medicine. Among those interventions are the FDA-approved agents rapamycin, metformin and acarbose, thus, the time has arrived to test potential aging-modulating drugs in humans. The goal of the San Antonio Older Americans Independence Center (SA OAIC) is to advance discoveries made in animal models (invertebrates and rodents) towards relevant human trials. To support this goal, the mission of the Clinical Research Core [(henceforth dubbed Research Core 2 (RC2)] is to provide knowledge, skills, techniques, logistical, and clinical research support to foster an understanding of the functional and physiological effects of pharmacological and nutraceutical interventions on aging in older people. RC-2 capitalizes and builds upon the rich tradition and institutional support of aging research in San Antonio, as well as the dedicated commitment of our investigators to translational research, thereby ensuring the success of the Core. RC2 will provide clinical research guidance to support human trials that test interventions capable of modulating the aging process to improve and extend the healthspan and lifespan of older Americans. The Specific Aims of the Clinical Research Core are to: 1) Assist basic and clinical investigators in developing rigorous and appropriately powered clinical studies and trial concepts that will lead to innovative approaches to improve healthspan and lifespan; 2) Facilitate implementation and execution of translational human studies and clinical trials by investigators; 3) Provide expertise and coordinated access to resources and technology in both our OAIC and facilities throughout our institution to maximize the depth of phenotypic characterization relevant to aging in trial outcomes; 4) Provide training in clinical research for early-stage faculty and those new to clinical research. To achieve these aims, the RC2 Core will provide research project consultation and planning, assist with safety and regulatory compliance processes, facilitate subject recruitment and retention, and coordinate relationships with relevant OAIC Cores and other Core facilities of our institution. Importantly, RC2 will be critical to the OAIC?s goal of developing a clinical focus to conduct clinical research in geriatrics with emphasis on transformative translational research to advance discoveries from basic aging research to clinical trials and ultimately to clinical geriatrics. We will train early-career investigators, and partner with other OAICs, institutions, and the public to advance translational aging research. This Core will integrate other important OAIC core functions to ensure efficient trial design and execution, including integrating studies with the OAIC Biostatistics and Data Management Core (RC3), Research Career Development Core (RCDC), and the Pilot and Exploratory Studies Core (PESC). RC2 will work in synchrony with the Pre-Clinical Core (RC1) so that studies conducted by both cores are compatible and thereby facilitate translation into subsequent human trials.

Estrogen receptor (ER)? exhibits an antitumor activity in multiple cancer types in both tumor-intrinsic and -extrinsic manners. However, little is known as to how such activity can be harnessed with high efficacy and precision, nor is it clear which host cell type(s) mediates the tumor-extrinsic function of ER?. These major knowledge gaps hamper efforts to unleash ER? antitumor activity for cancer therapies. We recently discovered a phosphotyrosine-dependent signaling axis that controls ER? antitumor activity. Furthermore, using a novel knockin mouse model, we found that this phosphotyrosine switch plays a significant role in host cells to promote antitumor immunity. Our central hypothesis is that this ER?-centered signaling axis provides a previously unrecognized molecular handle for mobilizing tumor-extrinsic antitumor activity of ER? in immune cells. Armed with genetic and pharmacological tools that both specifically target this signaling circuit, our multi-PI team will validate this novel hypothesis through three Specific Aims. First, we will identify the exact immune cell type(s) that mediates tumor-extrinsic ER? signaling in antitumor immunity (Aim 1). We will then delineate the upstream regulators and downstream target genes of ER? signaling in immune cells (Aim 2). Lastly, we will assess the anticancer therapeutic potential of targeting this ER? signaling axis to boost current cancer immunotherapies. Findings from these experiments will shed light on a previously under-appreciated signaling pathway governing tumor-immune cell interactions in the tumor microenvironment. As immunotherapies are becoming an important pillar of cancer therapy, the proposed study will help improve clinical outcomes and efficacy of immunotherapy for larger numbers of cancer patients.

We respond to PQ3 with our data showing that tumor PD-L1 (CD274, B7-H1) is a major regulator of tumor inflammatory infiltrates. Our preliminary data show that melanoma PD-L1 regulates TIL through several previously unknown tumor-intrinsic and extrinsic mechanisms. We define novel effects of tumor intrinsic PD-L1 signaling on tumor proliferation, sensitivity to immune killing, in vivo growth independent of anti-tumor immunity, and regulation of mTOR signals. We identified intracellular PD-L1, including those whose surface expression is low or negative, and identified interactions with tumor PD-1. We hypothesize that melanoma intrinsic PD-L1-driven signals, particularly mTOR signals, alter tumor progression and treatment responses. The research team is comprised of tumor immunotherapy, tumor immunology and PD-L1 experts at UTHSCSA and Dartmouth. We focus on melanoma for scientific reasons and based on our expertise. Aim 1 Define how tumor PD-L1 alters tumor immune infiltrates and immunotherapy responses. We use control versus PD-L1lo (shRNA) B16 in a novel model to study differential treatment outcomes by tumor PD-L1 status. We generated PD-L1KO B16 by CRISPR for highly detailed follow up mechanistic studies, and to assess if PD-L1 null status differentially affects treatment versus PD-L1lo. Effects will also be tested in transplanted BrafV600E mutated D4M melanoma (PD-L1+) engineered to be PD-L1lo and PD-L1KO, in mice with induced BrafV600E melanomas, and in syngeneic skin grafts of skin from Braf/Pten versus PD-L1KO Braf/Pten mice. Aim 2 Test tumor PD-L1-driven mTOR signal effects on TIL and immunotherapy responses. We will test PD-L1 KO, PD-1 KO and double KO melanoma cells for mTOR signals, TIL and treatment effects. Cells will be engineered for defects in mTORC1/2 for mechanistic studies, complemented with mTOR inhibitor treatments. We will use engineered tumors that express cytoplasm-only versus cell surface-only PD-L1, to define novel, intracellular PD-L1 signals. Constructs with mutations in known PD-1 signal sites will be engineered into these tumors for a complete understanding of PD-L1/PD-1 interactions. Aim 3 Define cell-intrinsic PD-L1 effects in human melanoma. We use well-defined human melanoma lines that are basal PD-L1+ and/or PD-1+ and/or BrafV600E mutated. We will use human vectors to knock down or knock out PD-L1, PD-1 and mTORC1/2 genes. In vitro assessments of effects on proliferation, responses to mTOR inhibitors, ?PD-L1 and ?PD-1 will be assessed. In vivo effects in NSG mice will be assessed. Primary human melanoma lines will be studied to complement data from long-term lines.

This proposal combines a team with expertise in aging, tumor immunology, tumor immunotherapy, specific genetically modified animal models and early phase clinical trials with a computational team having great expertise in analyzing and modeling aging of the immune system. We will study age effects on PD-L1/PD-1 signaling in the host and the tumor focusing on melanoma with some bladder cancer work, two tumors that are highly responsive to ?PD-1 and/or ?PD-L1 as proofs-of-concept, and residing in distinct anatomic compartments. In Aim 1 we study tumor PD-L1 intrinsic effects on ?PD-L1 and ?PD-1 treatment in melanoma and bladder cancer using transplantable B16 and inducible Nras/Cdk2n melanoma models, and transplantable MB49 and BBN-induced tumors for bladder cancer studies. We also use novel melanoma and BC models with tumor cell- specific PD-L1KO. We study 3 cohorts of elderly versus younger humans getting ?PD-L1 or ?PD-1 for melanoma or bladder cancer for human validation. We measure high-dimensional cell phenotypes and signaling responses, proteins and genes to maximize the information collected from human samples and mice using 23-color FACS, CyTOF, Luminex, Nanostring and other approaches. In Aim 2 we use all the above models and analytic strategies in young and aged PD-L1KO mice and WT or bone marrow chimeras to test hematopoietic and non- hematopoietic (host) PD-L1 signals in treatment outcomes in melanoma and bladder cancer. In Aim 3 the Systems Immunology team will use their innovative and successful computational modeling to identify age- related co-predictors of immunotherapy response and to identify candidate mechanisms for responders and non- responders. We will define a trajectory of immune system aging in mice at ultra-high resolution by performing a systems level integrative analysis of aging in Collaborative Cross and BL6 mice tracked in a combined longitudinal and cross-sectional study. This trajectory will be used to understand how tumor response and treatment outcomes vary as a function of age, and to build a simple, low parameter (i.e., easily testable and clinically translated), predictive models of treatment response. We will test insights by analyzing immune data from aged versus young patients undergoing ?PD-L1 and ?PD-1 cancer immunotherapy in novel machine learning approaches that we pioneered to identify insights from mouse data that are relevant to humans. Coupling this disease information with the healthy human aging trajectory that we recently defined will allow us to adapt our mouse data to predict optimal treatments in humans based on chronological and immune aging. This combined trans-disciplinary approach will identify common age-related disabilities that reduce PD-L1/PD-1 based immunotherapy responses and suggest tailored treatments for optimal efficacy that could later be tested in validation sets. These data can also be applied to other types of immunotherapy as we will also test.