Title

Modulation of T Cell Response Hierarchy by Coinhibition and Costimulation with Cancer Immunotherapy

Date of Award

Fall 10-1-2021

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Investigative Medicine

First Advisor

Chen, Lieping

Abstract

Cancer is a significant cause of morbidity and mortality worldwide. Cancer immunotherapy with coinhibition antagonist antibodies blocking the Program Death 1 (PD-1) and Program Death Ligand 1 (PD-L1) coinhibitory pathway (anti-PD therapy) have improved the survival for millions of patients with advanced cancers providing the possibility of achieving a prolonged durable response. However, few patients are considered to be clinically cured with anti-PD therapy. Many patients who initially respond to anti-PD therapy eventually develop tumor progression or recurrence in a clinical observation collectively called acquired immune resistance (AIR). Many potential mechanisms of AIR are being studied, such as the loss of tumor antigen expression, the loss of antigen processing and presentation machinery, loss of responsiveness to IFNg, the presence of other immune inhibitory pathways in the tumor, the abundance of inhibitory immune cells such as Tregs and MDSCs in the tumor, etc., but there is ample evidence that these mechanisms may only account for a small portion of patients who develop AIR.CD8+ cytotoxic T lymphocytes (CTL) play an important role in tumor immunity through their ability to recognize specific antigens and to directly kill the tumor cells expressing those antigens. Since there is tumor antigen heterogeneity within the tumor, the simultaneous generation of an unbiased polyclonal CTL response to a broad spectrum of tumor antigens is ideal as it may prevent tumor variants from escaping the immune response. However, CTL responses to the individual antigens from complex multiantigen organisms/cells such as viruses, bacteria, and tumors, are not all created equal but occur in a biased manner. CTL responses are only made to a few epitopes, and there is a biased response hierarchy. This poorly understood phenomenon is called immune dominance. In an immune response against cancer, CTL responses also only occur to a few epitopes, where the dominant antigen produces a strong T cell response and subdominant antigens produce far weaker T cell responses. This limitation on the breadth of the T cell response to tumor antigens may also be contributing to the development of AIR. Currently, there is a gap in knowledge in how anti-PD therapy affects the T cell response hierarchy. Our prior studies show that anti-PD therapy combined with a multi-antigen vaccine affected immune dominance by selectively increasing the T cell response to the dominant antigen and suppressing the T cell response to the subdominant antigen. This skewed response led to poor tumor control and promoted the growth of tumor variants that do not express the dominant antigen, suggesting that immune dominance may be an important mechanism underlying the development of the AIR with anti-PD therapy. However, there is no consensus on how anti-PD therapy affects T cell response hierarchy. Another study had shown anti-PD therapy can increase the subdominant T cells response. Both these prior studies evaluated the effect of anti-PD therapy on the systemic T cell response to antigens. Currently, it is unclear how anti-PD therapy affects the T cell response hierarchy within the tumor microenvironment (TME) and is thus, being evaluated in this thesis work. Because AIR may occur from the failure to develop a broad antigen response, strengthening the subdominant T cell responses is of interest and may help prevent tumors from escaping immunity. Since 4-1BB (CD137) costimulation agonist immunotherapy provides T cells costimulation and is known to expand the CTLs responsible for tumor killing and maintain CTL survival, 4-1BB agonist is being evaluated to determine if it can strengthen subdominant CTL responses in the TME. An MC38 mouse tumor model that expresses a dominant viral antigen p15E and a subdominant Dpagt neoantigen was used to study the effect of anti-PD therapy and that of a 4-1BB agonist therapy on the immune dominance and the diversity of the antigen-specific T-cell response within the TME of the syngeneic C57BL/6 mice. I found that anti-PD therapy selectively boosted the dominant T cell response in the TME. Interestingly, dominant T cells are found to have higher inhibitory PD-1 receptor expression and are likely receiving stronger inhibitory PD-1 signaling than subdominant T cells. Dominant T cells also have higher T Cell Receptor (TCR) avidity and receive stronger TCR signaling than the subdominant T cells. My findings suggest that anti-PD therapy removes the potent inhibition on the dominant T cells and allows them to receive stronger stimulatory signals, such as signals from the TCR, and potentially also from costimulation and cytokines, than the subdominant T cells, leading to preferential expansion of the dominant T cells. This further skewing of the T cell diversity with anti-PD therapy, while rapidly promotes the elimination of tumor cells expressing dominant antigens, may promote the development of AIR due to weak subdominant CTL response. However, this caveat of anti-PD therapy may be overcome by 4-1BB agonist treatment since my experiments showed that 4-1BB agonist antibody could unbiasedly promote CTL responses towards both dominant and subdominant T cells. By enhancing the CTL response to a broad number of tumor antigens, 4-1BB agonist treatment may potentially prevent tumor escape. Perhaps immunotherapy targeting T cell costimulation can be useful as a sequential therapy in treating patients who develop AIR to anti-PD therapy. Since anti-PD and 4-1BB agonist therapies function through different mechanisms, the combination of the two therapies was evaluated and was found to be more effective in treating MC38 tumors and further increased the number of dominant and subdominant T cells in the tumor than the single treatments. However, this improved response came with the cost of worsening the hepatotoxicity associated with 4-1BB agonist treatment and is not a currently feasible combination as hepatoxicity has already limited the development of 4-1BB agonists clinically. My findings point out the promise and limitation of using this combination therapy. Further studies to prevent the hepatotoxicity associated with 4-1BB agonist antibody will be critical before 4-1BB agonist can be developed as a single agent or as this promising combination immunotherapy. In summary, my thesis study reveals that cancer immunotherapies modulating T cell coinhibition and costimulation could differentially affect the T cell responses to tumor-specific antigens based upon their response hierarchies. These findings provide us the rationale and a scientific basis for selecting therapies to combine or be given sequentially to improve immunotherapy efficacy and duration of response.

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