Antagonism of the vasoactive intestinal peptide and PI3K δ signaling pathways in T cell-based therapies for leukemia and lymphoma Open Access

Petersen, Christopher (Fall 2017)

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The critical role of T cells in tumor surveillance and eradication in a variety of cancers has been established by recent advances in the understanding of T cell-tumor interactions. These advances have led to the development of a wide array of immunotherapeutic strategies that have seen varying degrees of clinical success. The use of T cell-based immunotherapies has largely been explored in settings of solid tumors whose high mutational burden provide a large amount of neo-antigenic targets. Unlike solid tumors, hematological malignancies such as leukemia and lymphoma have a relatively low mutational burden that severely limits their potential to be recognized and eliminated by autologous T cells. In addition to problems with tumor cell recognition, T cell responses to both solid and hematological malignancies are also inhibited by active and passive immune evasion strategies. While T cell responses to solid tumors largely fail as a result of exhaustion and inhibitory signals in the tumor microenvironment, endogenous antigen-specific T cells recognizing leukemia and lymphoma cells may be limited by the failure of antigen-specific T cells to fully activate in vivo leading to their subsequent deletion. This failure of primary T cell activation is attributed to the disseminated nature of hematological malignancies that results in decreased antigen and danger signal densities. Thus, the failure of T cells to eliminate solid tumors is an issue of exhaustion while the failure to eliminate leukemia and lymphoma cells is an issue of immunological ignorance or anergy. Despite these fundamental differences, similar strategies to enhance the anti-tumor T cell response have been explored in both disease settings. Hematological malignancies served as the prototypical target of immunotherapies with the advent of stem cell transplants and monoclonal antibodies. While these therapies remain powerful tools for the treatments of these diseases, many emerging therapies focus on enhancing autologous T cell responses. Some of these treatments include checkpoint blockade, cytokine therapies, and therapeutic and prophylactic vaccinations. The results of immunotherapies designed to bolster autologous anti-tumor T cell responses have been less promising in hematological malignancies due in large part to the reduced immunogenicity of the malignant cells which renders T cells anergic or senescent. In spite of this large obstacle, T cell-based therapies hold tremendous promise for leukemia and lymphoma treatment, and as such, it is critical to explore additional avenues for strengthening the anti-tumor response in these settings.

We addressed the issues of T cell anergy and senescence in settings of leukemia and lymphoma by using antagonists to the vasoactive intestinal peptide (VIP) and PI3K δ pathways to improve T cell activation and reverse or prevent senescence in settings of murine model systems of leukemia and lymphoma and pre-clinical studies of T cell expansion and CAR T cell manufacturing using clinical samples obtained from lymphoma patients. Using a peptide VIP antagonist, VIPhyb, as a therapeutic in leukemic mice we found that small subcutaneous doses led to a significant reduction in tumor burden and a significant survival benefit. The effect of VIPhyb was T cell-mediated as VIPhyb-treated mice had increased frequencies of effector CD8 T cells with reduced expression of PD-1 at later time points during tumor development, and the anti-tumor promoting activity of VIPhyb required the presence of CD8+ T cells in treated mice. Furthermore, T cells from VIPhyb-treated mice conferred protection from acute myeloid leukemia challenge following adoptive transfer to Rag1 knockout recipients. Based on these results, we explored the use of VIPhyb in combination with the PI3K δ inhibitor Idelalisib in the setting of ex vivo anti-CD3/CD28-mediated T cell expansion for chimeric antigen receptor T cell manufacture. Expansion of T cells from heavily treated DLBCL patients in the presence of VIPhyb and Idelalisib resulted in significantly increased yields as well as a preservation of the naïve and central memory compartments. T cells expanded in the presence of Idelalisib and VIPhyb had the greatest in vivo persistence in murine xenografts and significantly enhanced antigen-specific cytotoxic activity in an OVA-expressing murine tumor model. The results of our murine and human studies highlight the role of the VIP and PI3K δ pathways in T cell dysfunction in settings of leukemia and lymphoma. The addition of these inhibitors to clinical expansion cultures of T cells for adoptive cell therapies may provide a much-needed tool that is currently missing in immunotherapeutic strategies for the treatment and management of hematological malignancies.

Table of Contents

Chapter 1: Introduction

1-1-0   Hematological Malignancies and the Immune System                                          

1-1-1   Overview of Hematological Malignancies                                           2                                                

1-1-2   History of Immunotherapies for Leukemia and Lymphoma           3                             

1-1-3   Emerging Immunotherapies for Leukemia and Lymphoma         6     

1-2-0   T Cell Responses and Tumor Immune Evasion Strategies           

1-2-1   Tumor Antigen Recognition                                                                 10                                                    

1-2-2   T Cell Activation and Checkpoint Molecules                                   12                       

1-2-3   Immune Evasion by Leukemia and Lymphoma Cells                  14            

1-3-0   Vasoactive Intestinal Peptide       

1-3-1   Receptors, Signaling, and Non-immune Functions                      18               

1-3-2   Effects of VIP on T Cell Function                                                        20                                                  

1-3-3   VIP and Cancer                                                                                      23

1-4-0   Phosphoinositide 3-Kinase δ (PI3K δ)

1-4-1   Overview of PI3K δ                                                                                 25

1-4-2   Role of PI3K δ in T Cell Development and Function                      27

1-4-3   Idelalisib: a PI3K δ-Specific Inhibitor                                                 30

Chapter 2: Administration of a Vasoactive Intestinal Peptide Antagonist Significantly Enhances the Autologous Anti-Leukemia Response in Murine Models of Acute Leukemia

           2-1-0 Abstract                                                                                                            34

           2-2-0 Introduction                                                                                                       34

           2-3-0 Materials and Methods                                                                                   37

           2-4-0 Results                                                                                                             43

           2-5-0 Discussion                                                                                                       50

           2-6-0 Figure Legends                                                                                               54

           2-7-0 Figures                                                                                                              60

Chapter 3: Improving ex vivo T Cell Expansion from DLBCL Patients for T Cell Therapies via Antagonism of PI3K δ and VIP

           3-1-0 Abstract                                                                                                             67

           3-2-0 Introduction                                                                                                       68

           3-3-0 Materials and Methods                                                                                  70

           3-4-0 Results                                                                                                             75

           3-5-0 Discussion                                                                                                      81

           3-6-0 Figure Legends                                                                                             85

           3-7-0 Figures                                                                                                             91

Chapter 4: Discussion

          4-1-0 Summary                                                                                                        103

           4-2-0 Conclusions and Perspectives                                                                  105

           4-3-0 Future Directions                                                                                        109

           4-4-0 Model                                                                                                               113

References                                                                                                            116

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