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Last Updated: 10/17/18

SPORE in Skin Cancer

Yale University

Principal Investigators:
Marcus Bosenberg, M.D., Ph.D. and Harriet Kluger, M.D.

PRINCIPAL INVESTIGATOR CONTACT INFORMATION

Marcus Bosenberg, M.D., Ph.D.
Laboratory for Medicine and Pediatrics (LMP)
15 York Street, Ste 5038A
New Haven, CT 06510
Phone: (203) 737-3484
Email: marcus.bosenberg@yale.edu

Harriet Kluger, M.D.
Yale Cancer Center
Yale New Haven Hospital
20 York Street, Ste North Pavilion 4
New Haven, CT 06510
Phone: (203) 200-6622
Email: harriet.kluger@yale.edu

OVERALL ABSTRACT

The goal of the Yale SPORE in Skin Cancer (YSPORE) is to decrease mortality from melanoma by developing methods to prevent the disease and treat patients once it metastasizes. YSPORE leans on the scientific and clinical strengths at Yale School of Medicine and Yale Cancer Center including immunobiology, genetics, epigenetics, quantitative sciences, immuno-oncology, nanoparticles, animal models, and digital pathology. YSPORE investigators have made major contributions in recent years to understanding the effects of ultraviolet damage on melanocytes, determining the molecular events that lead to melanoma progression and developing novel immune therapy approaches for melanoma patients with advanced disease. Recent advances in systemic therapies for advanced melanoma, particularly immune checkpoint inhibitors, that prolong survival, pose a new set of challenges for clinicians which will be addressed by the proposed research program. The YSPORE translational research team proposes to accomplish the objective of decreasing morbidity and mortality from skin cancer through five specific aims: Specific Aim 1: Develop novel bioadhesive sunscreens and triplet-state quenchers for prevention of melanoma and test them in pilot clinical trials; Specific Aim 2: Develop liquid biopsy approaches for melanomas that metastasize to sites that are difficult to biopsy. Liquid biopsies will then be studied in patients treated with PD-1 inhibitors to identify predictors of response and determine mechanisms of resistance; Specific Aim 3: Study novel drugs that target the epigenetic modifier KDM5, to enhance T cell infiltration in tumors that are unresponsive to immune checkpoint inhibitors; Specific Aim 4: Develop new approaches to overcoming resistance to PD-1 inhibitors by co-targeting the innate and adaptive immune systems; and Specific Aim 5: Develop new research directions to decrease mortality from melanoma and nurture the next generation of translational investigators focusing on skin cancer through a Developmental Research Program and a Career Enhancement Program. We propose three cores (Administrative Core, Biospecimen Core and Biostatistics and Bioinformatics Core) to support the projects, their clinical aims, mechanistic studies, and biomarker development for clinical application. The Projects and Cores are designed to be highly coordinated with the goal of maximizing resources and potential impact. New collaborations will be established during the funding period through the Developmental Research Program with the support of the Administrative Core. Collaborations with other institutions and skin cancer SPORE sites will be fostered by the Administrative Core. These coordinated efforts will enhance analysis of patient samples, use of cell cultures and animal models, and development of predictive biomarker assays. Our purpose is to translate the innovative approaches proposed here to clinical therapies for prevention and treatment of skin cancer.

PROJECT 1: BIOADHESIVE SUNSCREENS AND TRIPLET-STATE QUENCHERS FOR MELANOMA

Project Co-Leaders:
Douglas Brash, Ph.D. (basic)
Michael Girardi, M.D. (clinical)
Mark Saltzman, Ph.D. (translational)

Ultraviolet radiation (UVR) exposure is the major environmental risk factor for melanoma development; thus, limiting UVR exposure and its damaging effects is a logical strategy for melanoma prevention. Wehypothesize that early and fundamental processes of melanomagenesis can be inhibited via complementary approaches that reduce both direct and indirect UVR-induced melanocyte genotoxicity. While previous topical formulations have attempted to address direct damage pathways (using sunscreens) and indirect damage pathways (using antioxidants), recent breakthroughs from our research programs provide us with unique opportunities for further technological innovation in preventing malignant melanoma. We have developed a novel sunscreen delivery platform for the prevention of UVR-damage that uses bioadhesive nanoparticles (BNPs). UVR-absorbing compounds are stably encapsulated within particles that have a biodegradable polymer core and a surface coating that allows them to adhere covalently to the stratum corneum, increasing sunscreen agent efficiency and durability while mitigating safety concerns attributable to penetration of agents into the epidermis, e.g. contact allergy, increased generation of ROS, and hormone receptor binding. We have also identified a novel pathway of indirect damage that continues long after UVR-exposure, via triplet-state electrons in melanin fragments that also drive formation of cyclobutane pyrimidine dimers (CPDs). This process of chemiexcitation occurs over the course of hours, suggesting several opportunities to prevent or dissipate the high energy excited states involved, by using agents that have not yet been examined for this novel activity. Accordingly, we have demonstrated the potential of triplet-state quenchers (TSQs) and triplet-state preventers (TSPs) to avert formation of these delayed CPDs. The overall goal of this proposal is to systematically optimize these approaches for melanoma prevention. To accomplish this, we will develop both a novel broad-spectrum BNP-sunscreen and a novel TSQ/TSP topical formulation that, in combination, we hypothesize will provide optimized prevention of the direct and indirect UVR-induced melanocyte genotoxicity that fundamentally drives melanomagenesis.

PROJECT 2: LIQUID BIOPSIES FOR MELANOMA THAT METASTASIZE TO INACCESSIBLE SITES

Project Co-Leaders:
David Hafler, M.D. (basic)
Harriet Kluger (clinical)

With the rise in incidence of metastatic melanoma and prolonged survival of patients with this disease, an ever increasing number of patients are living with metastatic melanoma and requiring therapy. Not all patients respond to PD-1 inhibitors, and there is therefore an urgent need for means to predict response to this class of agents, to monitor patients for response, and to understand mechanisms of sensitivity and resistance to these drugs that might be harnessed for designing effective combination regimens. Although tumor cell properties, such as PD-L1 expression, are somewhat associated with response, mounting evidence generated suggests that properties and quantity of tumor infiltrating inflammatory cells might be more informative in terms of predicting and monitoring response and can enable us to determine approaches to overcome resistance. However, melanomas often metastasize to areas that are difficult to biopsy, such as the brain, spleen, small bowel and bone, and we therefore propose approaches to studying properties of tumor infiltrating lymphocytes (TIL) by isolating and profiling T cells that originated in the tumor but can be identified in the circulation. In previous studies we demonstrated that these circulating T cells, which express both PD-1 and TIM-3, are specific to the tumor microenvironment, and are not found in the circulation of healthy individuals or patients with autoimmune disease. We propose to conduct single cell analysis of these TIM3+PD-1+ circulating CD4 and CD8 cells that have been in tumor tissue using state-of-the-art technologies. We have assembled a team of interdisciplinary researchers with the goal of developing liquid biopsy methods to determine mechanisms of resistance to immune checkpoint inhibitors. We will start by conducting single cell CyTOF and functional cytokine analysis of CD4 and CD8 cells from the circulation and tumor tissue to determine similarities and differences at the molecular level. We will then barcode TIM3+PD-1+ cells using TcR chain sequences to identify circulating TIL by the presence of TcR sequences that are identical to TIL isolated from tumors representing sister clones. Using single cell RNA-seq, we will therefore be able to assess TIL function by studying function in this population of circulating TIL. This will be done on prospectively collected matched samples from melanoma patients undergoing resection of metastases, including brain metastases, biopsies from other sites and blood samples. Our liquid biopsy technique then be studied on blood samples from patients treated with PD-1 inhibitors before and on therapy, with complete clinical annotation including response to therapy. If successful, our liquid biopsy approach can be applied to other treatment settings and other tumor types, and thus has the potential to significantly impact patient care by enabling improved patient selection and avoiding the morbidity and expense associated with repeat tumor biopsies. Moreover, the single cell analysis will enable us to better understand mechanisms of resistance, which in turn can facilitate drug development for patients with tumors resistant to PD-1 inhibitors.

PROJECT 3: KDM5 HISTONE DEMETHYLASES IN MELANOMA GROWTH AND TUMOR IMMUNITY

Project Co-Leaders:
Qin Yan, Ph.D. (basic)
Marcus Bosenberg, M.D., Ph.D. (translational)
Mario Sznol, M.D. (clinical)

Despite remarkable recent progress, prognosis for patients with advanced melanoma remains poor. Specifically, primary and acquired drug resistance often develops for targeted therapy, and only a subset of patients respond to immunotherapy. These challenges highlight an urgent need to develop novel therapeutic methods, improve current treatments against melanoma, and develop biomarkers that predict response. Emerging evidence suggests that the epigenetic regulator KDM5B is an attractive target and biomarker for melanoma treatment. The long-term goal is to translate our findings of novel mechanisms involved in melanoma formation and progression to the clinic. The objective of this project is to evaluate the therapeutic potential of targeting the KDM5 histone demethylases in melanoma and to develop biomarkers to predict response to PD-1 pathway blockade and combined KDM5 inhibition/immunotherapy. Our central hypothesis is that KDM5 targeting results in direct anti-tumor effects and indirect effects by converting immunologically “cold” tumors into “hot” tumors, which are more likely to be infiltrated by lymphocytes and to respond to immune checkpoint blockade. Preliminary data suggests that this effect may be mediated by the STING pathway. The hypothesis is supported by previous studies as well as our own preliminary data from patient-derived melanomas and preclinical melanoma models. The rationale is that better understanding of KDM5 histone demethylase function in melanoma growth and anti-tumor immune responses will result in new and innovative approaches to treat melanoma. The hypotheses will be tested in two Specific Aims: 1) Evaluate the therapeutic potential of targeting KDM5 in melanoma; 2) Evaluate KDM5B as a biomarker in human melanoma. The proposed research is conceptually and translationally innovative, because it aims to determine whether KDM5 inhibition can convert melanomas from an immunologically “cold” to “hot” state, and to evaluate tumor KDM5B level as a new biomarker for melanoma. The results from these studies could impact the treatment of patients with melanoma and increase our understanding of the factors that regulate anti-tumor immune responses.

PROJECT 4: MODULATING INNATE IMMUNITY TO OVERCOME RESISTANCE TO PD-1/PD-L1 BLOCKADE

Project Co-Leaders:
Harriet Kluger, M.D. (clinical)
Susan Kaech, Ph.D. (basic)

Dramatic progress has been made in recent years in treating patients with advanced melanoma, particularly with inhibitors of PD-1 or PD-L1, resulting in prolonged survival and a sharp rise in the number of patients living with metastatic melanoma. However, over half the patients do not respond to PD-1/-L1 inhibitors, and tumor regrowth is seen in about half the patients by two years. Thus, there is great need to understand mechanisms of resistance and to develop new approaches to overcome resistance. A well-documented mechanism of resistance to PD-1/L1 inhibitors is paucity of tumor infiltrating T cells (TILs). We have found that targeting tumor associated macrophages by inhibition of the colony stimulating factor-1 receptor (CSF1R) results in tumor regression in a T cell independent fashion and induces increased TNFa, IL-1Β, IFN? expression and neutrophil recruitment. Similarly, antibodies that activate CD40 also result in macrophage modulation and tumor regression and synergize with CSF1R. When used together in mice, these treatments stimulate a T cell-dependent anti-tumor response. We hypothesize that triple therapy (inhibitors of CSF1R and PD1 with CD40 agonists) may be even more effective at eliciting anti-tumor responses to tumors that were at one time poorly infiltrated and/or recognized by T cells. We will test the key hypothesis that stimulating both innate and adaptive immunity leads to more robust anti-tumor immunity than those that preferentially target T cells or myeloid cells alone. We will utilize a series of novel immune competent murine models of resistance to PD-1/PD-L1 inhibitors generated at Yale with driver mutations that mimic human melanomas (e.g., BrafV600E, NrasQ61R, loss of Cdkn2a, Pten, p53, and gain of Β-catenin). We will study the effects of CSF1R inhibitors and CD40 agonists with and without PD-1 inhibitors to determine activity and toxicity of doublet and triplet regimens in murine models with various genetic backgrounds (Aim 1). In Aim 2 we will study the mechanisms of sensitivity or resistance to the various combinations to facilitate predictive biomarker studies for humans treated with these regimens. We will conduct a phase I/IB clinical trial (Aim 3) of a CSF1R mAb in combination with a CD40 agonist with concurrent dose escalation of a triplet regimen that will include the addition of nivolumab. Expansion of the optimal regimen will be conducted using a Simon two stage design. Patients eligible for the trial will have advanced melanoma that has progressed on prior PD-1/PD-L1 based regimens. Tumors from patients treated on this trial will be characterized for tumor associated macrophage subsets and TIL subsets using state-of-the-art technologies. The collaborative team, availability of clinically-relevant animal models and access to patients and their tumors will thus enable us to go back and forth between the clinic and the lab, facilitating personalized approaches for patients with tumors poorly responsive to PD-1/PD-L1 inhibitors. Results from these studies will be helpful for overcoming resistance to PD-1/PD-L1 inhibitors in other tumor types as well.

ADMINISTRATIVE CORE

Core Directors:
Marcus Bosenberg, M.D., Ph.D.
Harriet Kluger, M.D.

Core Co-Director:
Mario Sznol, M.D.

The Administrative Core (Core 1) is critical to the success of the Yale SPORE in Skin Cancer (YSPORE). The primary purpose of the Core is to support and facilitate transdisciplinary research efforts in skin cancer and identify new challenges and opportunities as they emerge. The Administrative Core serves as the central coordination point for YSPORE investigators, with responsibility for monitoring the progress of all projects and cores toward a translational/clinical endpoint. These services include provision of (a) fiscal management, (b) clerical and organizational support, (c) mechanisms for internal and external review, (d) support for enhancement of scientific interactions and collaborations among the Project/Core leaders, (e) mechanisms to monitor projects and cores for scientific progress, (f) coordination of outreach efforts, and (g) mechanisms to expand research in skin cancer beyond the activities of the SPORE, both within Yale and outside of Yale. Core 1 will also be responsible for ensuring full integration of activities of the Yale SPORE in Skin Cancer with those of Yale Cancer Center, with the goal of avoiding overlap and duplication of efforts while maximizing synergy and use of resources. Any conflicts that might arise between SPORE investigators will be addressed by the Core 1 Directors, with input from the Executive Committee and the External and Internal Advisory Boards and Charles Fuchs, MD if needed. Core 1 will work closely with the Bioinformatics and Biostatistics Core (Core 3) to ensure the integration of data generated by the SPORE program into that of Yale Cancer Center and vice versa, and to oversee sharing of data with other skin cancer/melanoma SPORE sites, and when appropriate, with the Yale SPORE in Lung Cancer. Joint scientific research in progress meetings will be held with the SPORE researchers, organized by Core 1, to ensure bi-directional flow of ideas and sharing of resources. Investigators involved in this program will also participate in the annual inter-SPORE workshops, organized by Core 1 in collaboration with Administrative Cores at other sites. YSPORE hosted the 2017 inter-SPORE workshop in New Haven, CT. The Core will be directed by Drs. Marcus Bosenberg and Harriet Kluger and the Co-Director is Dr. Mario Sznol. Program oversight by Core 1 will be done with input from the Executive Committee, and input for scientific and administrative functions will be sought from the Internal and External Advisory Boards. Annual meetings with the Internal and External Advisory Boards will be organized by Core 1. The Core will also organize research in progress seminars, talks by outside investigators, and patient and community outreach.

BIOSPECIMEN CORE

Core Directors:
Ruth Halaban, Ph.D.
Marcus Bosenberg, M.D., Ph.D.
Harriet Kluger, M.D.

The Biospecimen Core (Core 2) is the cornerstone of all Yale SPORE in Skin Cancer (YSPORE) activities. It addresses the broad melanoma patient specimen needs of all the projects that are not met by current shared facilities at Yale. In addition to collecting, storing and distributing a wide range of specimens and reagents, the core performs quality assurance testing and a wide range of molecular analyses of specimens. More specifically, the core: a) collects a large repertoire of specimens for translational and preclinical studies in melanoma, including melanocytic lesions, melanoma tumors and cells, normal skin, serum, and circulating lymphocytes; b) ensures high quality control, proper long-term storage, detailed annotation, and timely distribution of specimens to YSPORE investigators; c) establishes and maintains a central database of essential pathological, clinical, epidemiological, and follow-up information and basic research data generated by the YSPORE projects that is integrated with the Biostatistics and Bioinformatics Core; d) provides special services such as the analysis of specimens from clinical trials, mutations, chromatin modification, and collection of TIL (tumor infiltrating lymphocytes); e) maintains and distributes validated reagents (antibodies, oligonucleotides for PCR, DNA, RNA, plasmids, cell extracts) needed for molecular analyses of tumors by different YSPORE investigators; f) establishes links with shared facilities at Yale and similar resources in other institutions, including other SPOREs; g) distributes samples to collaborating investigators outside of Yale; and h) provides specimens for larger NCI objectives, such as melanoma specimens, peripheral blood lymphocytes, and annotated clinical data for The Cancer Genome Atlas (TCGA). The core interacts extensively with investigators in each project, the Bioinformatics and Biostatistics Core, the Clinical Trial Office, and Yale Cancer Center shared resource cores such as Yale Pathology Tissue Services and Yale Center for Genome Analysis. The services of the Biospecimen Resource Core enhance the efficient operation of the translational studies by YSPORE investigators in a cost-effective manner, and expedite the application of discoveries from the bench to clinical practice, and clinical results to basic research.

BIOSTATISTICS AND BIOINFORMATICS CORE

Core Directors:
Shuangge Ma, Ph.D.
Michael Krauthammer, Ph.D.

The goals of the Biostatistics and Bioinformatics Core are to address the study design and analysis needs of the Yale SPORE in Skin Cancer (YSPORE) projects, the Developmental Research Program (DRP), and the Career Development Program (CDP), and to address the analytical and data management needs of the Biospecimen Core. Drs. Krauthammer and Ma will continue to serve as Co-Directors of this Core, and will be assisted by statisticians and bioinformatitions with unique expertise and extensive experience. The Specific Aims of the Biostatistics and Bioinformatics Core are as follows. Aim 1: Provide biostatistical and bioinformatics support to all YSPORE projects and investigators. The Core will provide timely and comprehensive support to address analytic questions arising from the YSPORE projects. Our service will cover the whole spectrum of the studies, including study design, data management and analysis, result interpretation, and preparation of manuscripts, abstracts, posters, and applications. The Core will have regular research meetings with YSPORE investigators and maintain an open door policy for all analytic needs. Aim 2: Provide effective data management for YSPORE projects. The Core will offer data management services to all projects and investigators and ensure that all NIH data-sharing regulations are properly followed. This includes deposit of properly curated data to repositories such as GEO, SRA, dbGaP, and others. Aim 3: Develop innovative biostatistical and bioinformatics methods tailored to melanoma research. Data with more complex characteristics will be generated by YSPORE, TCGA, and other studies. The Core will develop new analysis methods which will facilitate more effective utilization of data and foster melanoma analytic research. Aim 4: Promote program-specific research and training opportunities involving biostatistics and bioinformatics trainees and faculty. The Core will promote opportunities within existing Yale programs and centers, including the Yale Graduate Program for Computational Biology and Bioinformatics (CBB), Yale Center for Medical Informatics (YCMI), Yale Center for Analytic Sciences (YCAS), and others. During the last two funding period, the Core has made significant contributions to research and training, which is partly reflected in our strong publication record and successes in training postdoc associates and graduate students. We will continue to do so in the proposed funding period. The Core will borrow strength while not overlap with that of other SPORE(s) and Yale Comprehensive Cancer Center (YCC). The Core will also be a key partner in the YSPORE with representation on the Senior Leadership Team and participation in all key meetings.