New agents that block immune checkpoints have the potential to revolutionize the treatment of cancers. Anti-CTLA4 and anti-PD-1 antibodies show significant promise in clinical trials. However, the molecular determinants of response are unknown. The mechanism of checkpoint blockade remains one of the most important unanswered questions in oncology. Our preliminary data demonstrate that therapeutic response is determined strongly by tumor genome and homology with foreign antigens present in the microbiome. Here, we seek to define the molecular determinants that define T cell receptor sensitivity for neoantigen and the role of foreign pathogen antigens in stimulating those responses.
Checkpoint blockade immunotherapies are currently some of the most promising strategies for fighting cancer. PD-1/PD-L1 blockade induces measurable responses in 13-26% of patients with many types of cancer by increasing anti-tumor T cell responses 1. In some cases, PD-1/PD-L1 blockade results in rapid and durable anti-tumor responses. Despite the effectiveness of PD-1/PD-L1 blockade, many patients are unresponsive to therapy 2-4. There are two major factors that contribute to poor prognosis: 1) Tumors poorly elicit CD8 T cell responses and 2) if T cell responses are formed, they often fail to infiltrate the tumor or are functionally exhausted 5-7. T cell exhaustion is associated with increased T cell expression of inhibitory receptors such as PD-15.
Certain HIV-infected individuals, known as “elite suppressors,” are able to maintain undetectable viral loads without antiretroviral therapy. In effect, their immune responses to the virus induce a functional cure. Current efforts in HIV research seek to develop vaccines that can achieve this same kind of functional cure in all patients. One promising strategy recently reported in macaques has been to induce an effector memory CD8+ T cell response against SIV by using a Rhesus CMV (RhCMV) vector to deliver SIV proteins. To attempt to bypass the need to infect patients with CMV in order to achieve the effects of the RhCMV-SIV vaccine, we plan to expand non-canonically-restricted CD8+ T cells from the blood of healthy donors using artificial APCs (aAPCs). We will then test whether the HLA-E and Class II-restricted CD8+ Tcells are protective against HIV and if they can kill virally-infected targets.
In this proposal, we wish to demonstrate the feasibility of identifying and expanding neoantigen-specific T cells for adoptive transfer following PD-1 blockade, in order to show that the majority of patients can have melanoma neoantigen-specific T cells expanded from their peripheral blood. We also aim to optimize ex vivo conditions for expansion of functionally and phenotypically optimal T cells for transfer using analysis of T cell function, TCR diversity and affinity to identify optimal neoantigen-specific T cell populations that will allow us to design truly personalized immunotherapeutics in future studies. This is highly clinically significant and represents an outstanding translational project. The work described is quite novel in that for the first time we propose to use peripheral blood cells from patients treated with a PD-1 antibody that we have found are enriched for antigen specific T cells. Our preliminary data suggest that in the tumor and in the periphery, clonality of increased and cells within the tumor can be found in the peripheral blood at low levels. We will expand these infrequent clones with the use of artificial antigen presenting cells, a truly innovative approach to expanding rare T cells within the periphery.
During autoimmune disease, the body incorrectly identifies “self” molecules as foreign and mounts a chronic immune attack. Conventional therapies employ broad immunosuppression, which has provided significant benefits to patients, but can leave these individuals immunocompromised. This limitation, along with the lack of cures for most autoimmune diseases, has sparked intense interest in strategies that could control autoimmunity with vaccine-like specificity, leaving the rest of the immune system intact. Several pre-clinical reports and clinical trials have investigated this theory to combat multiple sclerosis (MS), a neurodegenerative disease in which myelin in the central nervous system (CNS) is attacked by the immune system. An important finding from these studies is that co-administration of myelin peptide and tolerizing immune signals can promote the development of regulatory T cells (TREGS) that ameliorate disease. New knowledge of how signal integration in lymph nodes (LNs) drives tolerance could help address limitations associated with current therapies, such as incomplete control of disease and non-specific suppression.
Periodontal disease affects over 78 million Americans and is considered the most pressing oral health concern today. Also known as periodontitis, this condition is characterized by destructive inflammation of the periodontium, including the gum tissue, supporting bone, and ligament. Importantly, this disease affects not only tooth loss, but also the incidence of cardiovascular disease, kidney disease, respiratory diseases, diabetes, and even premature childbirth. The current standard of care involves debridement of calculus and can be accompanied by local delivery of an antibiotic such as minocycline. These treatments temporarily kill pathogens, but neither protect against inevitable future infections nor address the sensitivity observed in patients disposed to immune dysfunction. Although invasive bacterial species are protagonists of the disease, tissue destruction is mediated by an adverse host inflammatory immune response. As the disease progresses, several populations of lymphocytes are recruited to the periodontium. A newlydiscovered subset of lymphocytes called regulatory T cells (Tregs) has been shown to play a critical role in the regulation of harmful aberrant inflammatory immune responses.
In multiple myeloma (and many other heme- and non heme malignancies), the bone marrow is enriched for cancer specific T cells. Moreover, the higher efficiency of antigen presentation and greater concentration of central memory T cells in the marrow, make MILs a unique type of cells for adoptive T cells therapy. MILs technology expands and activates these cells ex vivo and then re-infuses them in the patient. In TRD 4 we seek to metabolically reprogram cells to enhance their efficacy and persistence. Collaborating with Dr. Borrello’s group affords us an extraordinary opportunity to test this hypothesis in an ongoing clinical program of Adoptive Cellular Therapy for Cancer. In this regard, our findings are readily translatable. With this collaborative project we hope to not only further Adoptive Cellular Therapy but also develop a tool box for metabolic reprogramming that can be exploited for not only enhancing immunotherapy for cancer but for a vast array of immunologically engineered systems.
This collaboration will enable us to translate our findings regarding immunometabolism to this novel treatment modality for cancer. Aim 1 seeks to exploit the robust molecular biology of CAR T cell generation and T cell genome editing to make these cells more potent effector cells. Aim 2 will define novel media to promote T cell persistence and memory. Aim 3 will harness our small molecule approach to regulating metabolism in order to enhance effector function and promote memory. By developing these three Aims for CAR T cells, we hope to not only further advance adoptive T cell therapy but also develop a tool box for metabolic reprogramming that can be exploited to enhance cancer immunotherapy as well as a vast array of immunologically engineered systems.
The goal of SP #1 is to synthesize and validate artificial antigen presenting cells (aAPC) in preclinical AML models. For SP #1, we will push our engineered optimized aAPC, from SA1, for expansion of human CD8+ T cells which will be tested by NexImmune using their proprietary AML-specific antigens as well as shared AML antigens. In addition PLGA and other biocompatible aAPC developed in TR&D2 will be provided to NexImmune for their in vivo animal testing studies.
Specific Aim 1. Production and in vitro validation of aAPC particles based on iron dextran, and PLGA bead matrices (standard aAPC particles to be supplied by JHU). aAPC particles using iron dextran and PLGA matrices will be provided. Each matrix will be used to produce aAPC particles directed against the following AML targets, WT-1, PRAME, RHAMM and proprietary Neximmune identified antigen. The particles will be tested for functionality in T cell stimulation assays and populations will be phenotypically and functionally characterized. Long terms storage stability will be assessed by T cell proliferation assays.
Specific Aim 2. In Vivo efficacy studies oon PLGA and PLGA/PBAE based aAPC. These studies will be done using the murine B16 F10 melanoma model. As part of this Service Project, the Schneck lab along with Green lab will provide particles and expertise for protein coupling to nanoparticles
The goal of SP #2 is to establish new approaches for ex vivo expansion of circulating endogenous antigen-specific T cells for oncolytic immunotherapy. We will push TR&D1 optimized aAPC for expansion of tumor-specific T cells that Dr. Wan will study in their models of oncolytic virus therapy.
Specific Aim 1. Delivering a protocol that allows local manufacturing of T cells against broadly expressed tumor antigens. Use of leukapheresis products to optimize our culture conditions and evaluate T cell functions in vitro and in vivo (adoptive transfer to NRG/A2 mice followed by oncolytic vaccination).
Specific Aim 2. Further streamlining the enrichment/expansion (E+E) process with nano-artificial APC and simultaneously producing multi-antigen specific T cells. We have requested aAPC from Jonathan Schneck to develop an E+E strategy using nanoscale aAPC and forms the basis of this SP. This approach will be integrated with our culture conditions to optimize cell number and function
Pluripotent reprogramming and direct lineage reprogramming hold tremendous potential as powerful strategies for providing alternative autologous cell sources for cell therapeutics and disease modelling. Target lineage cells can be derived from human induced pluripotent stem cells (hiPSCs) or directly converted from human fibroblasts for cell therapy and tissue engineering. Using cellular reprogramming technology, disease models of many human disorders can also be reconstructed to elucidate previously unknown pathogenic mechanisms of disease development and to test new therapeutic targets. Our project will focus on the establishment of novel methodologies enabling clinically-relevant neuronal differentiation of hiPSCs and neuronal direct reprogramming.
There are very limited options currently available for the treatment of hepatocellular carcinoma (HCC), the most common type of liver cancer. HCC, like most forms of cancer, is dependent on angiogenesis, the growth of blood vessels. These and other tumors use VEGF and Ang2 to induce angiogenesis. The tumor also uses these factors to suppress the immune system. Anti-VEGF agents inhibit angiogenesis and also allow some activation of the immune system but they induce hypoxia which is itself immune suppressive. We have identified a peptide that simultaneously inhibits VEGF and activates Tie2, the receptor for Ang2. This peptide, AXT201, thus inhibits angiogenesis but promotes normalization of the remaining vasculature thus avoiding hypoxia and immune suppression. In addition, by inhibiting VEGF and activating Tie2, AXT201 could make the immune system more anti-tumorigenic by allowing more dendritic cell maturation, more T-cell proliferation and infiltration into the tumor, and reduced number of regulatory T cells in the tumor. We hypothesize that AXT201 will enhance the efficacy of an anti-PD-1 antibody by making the immune system more anti-tumorigenic.
The goal of SP#5 is engineering designer organoids with tunable ligand specificities, understanding antigen specific immune response, and establishing a link between integrin ligand specificity and cell cycle epigenetics of germinal center reaction. NIM technologies from TR&D2 and metabolic reprogramming approaches from TR&D3 will be transferred (push) to Dr. Singh so that he can utilize these technologies to better achieve his immunoengineering goals in ways not possible w/commercial technologies.
The overall goal of SP #6 is to develop a FACS-based system to interrogate metabolic status of cells. This SP will not develop any novel technology but rather push novel approaches to examining metabolic programming of cells using already technology. This FACS-based “individual” cell analysis will be developed (push) to interrogate cardiac HPSCS.
Specific Aim 1. Create standardized antibody panels and procedures for interrogating metabolic programing in cells using FACS.