Abstract LB201: A novel combination of Listeria-based immunotherapy and repurposed drug combats colorectal cancer
Authors: Gaikwad Shreyas Ramchandra , Nguyen Hong My , Srivastava Sanjay K. , Wood Laurence 
Published: April 2023

Abstract
Background: Colorectal cancer (CRC) has been a difficult-to-treat cancer with a significant proportion of microsatellite stable (MSS-CRC) patients who do not respond to immunotherapy. Currently, various strategies are being investigated to sensitize MSS-CRC tumors by converting them to immunologically “hot” tumors and improve responsiveness to immunotherapy. In this study, we demonstrate a novel strategy of combining a repurposed FDA-approved compound, Pimavanserin tartrate (PVT), with a Listeria monocytogenes (Lm)-based vaccine targeting Interferon-Stimulated Gene 15(ISG15), Lm-LLO-ISG15, against MSS-CRC tumors. Our findings demonstrate that both agents synergize to produce CD8+ T cell-dependent therapeutic efficacy in immunotherapy-resistant MSS-CRC tumors.
Methods: The direct killing effect of PVT in vitro was confirmed by cytotoxic SRB assay. BALB/c mice bearing CT26, a murine model of MSS-CRC, were randomly assigned to receive either PBS, PVT, Lm-LLO-ISG15, or PVT+Lm-LLO-ISG15. Mice were monitored for tumor growth kinetics to study the anti-cancer effect of each monotherapy and the combination therapy. Changes in immune-related gene and protein expression were analyzed by qPCR and Western blot, respectively. Various immune cell subsets in the tumors and spleens were evaluated by multi-color flow cytometry and immunohistochemistry. In addition, organs such as the liver, kidney, heart, lungs, spleen, brain, and pancreas were collected for safety analysis.
Results: PVT efficiently induced apoptosis and directly killed both human and mouse MSS-CRC cell lines in vitro. Further, PVT treatment enhanced T-cell activation and induced ISG15 expression, thus, sensitizing the tumor cells to targeting by the CTL-mediated immunotherapy, Lm-LLO-ISG15. While each monotherapy failed to demonstrate anti-tumor efficacy, treatment with the combination therapy (PVT + Lm-LLO-ISG15) significantly controlled tumor burden and extended the median survival. The therapeutic efficacy of the combination approach was associated with a higher infiltration of CD8+ T cells in the TME and an increased population of effector memory T cells (CD4+CD44hiCD62Llo) in the spleen. The anti-tumor efficacy of combination therapy on subcutaneous CT26 tumor growth was completely abrogated after the depletion of CD8+ T cells. Moreover, the safety profile of dual therapy was comparable to that of each monotherapy and PBS.
Conclusions: MSS-CRCs are considered “immunologically cold” tumors that represent great challenges for any standalone therapies. In this study, we evaluated, for the first time, the therapeutic efficacy of a novel combination of PVT and Lm-LLO-ISG15 in an aggressive MSS-CRC mouse model. Combination therapy demonstrated a synergistic effect that led to a significant reduction of tumor burden and an extension of median survival. While the underlying mechanism for synergism requires further examination, our findings strongly suggest that combining PVT and Lm-LLO-ISG15 could be a promising approach for MSS-CRC patients.
Citation Format: Shreyas Ramchandra Gaikwad, Hong My Nguyen, Sanjay K. Srivastava, Laurence Wood. A novel combination of Listeria-based immunotherapy and repurposed drug combats colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr LB201.