Abstract 621: Tumor suppressor TUSC2 immunogene therapy is synergistic with anti-PD1 in lung cancer syngeneic mouse models
Authors: Meraz Ismail M. , Majidi Mourad , Shao RuPing , Feng Meng , Cao Xiaobo , Rice David , Sepesi Boris , Ji Lin , Roth Jack 
Published: July 2017

Abstract
TUSC2, a pro-apoptotic tumor suppressor gene whose expression is lost or decreased in most lung cancers, activates the innate immune system through initiation of broad spectrum cytokine secretion and natural killer (NK) cell activation. TUSC2 delivered systemically by nanovesicles has mediated tumor regression in metastatic non-small cell lung cancer clinical trials. We studied the effect of TUSC2 on immune cell populations and the anti-tumor activity of TUSC2 in combination with anti-PD1 checkpoint blockade in two syngeneic mouse models: C57BL/6 mice subcutaneously injected with murine lung adenocarcinoma cell line CMT/167-luc cells (KrasG12V mutation) and 344SQ (KrasG12D allele and a knock-in Trp53R172HΔG allele) adenocarcinomas which metastasize to the lung in 129S2 mice. Tumor growth was monitored by scoring ex-vivo luminescence using the IVIS Imaging System 200. Multi-color flow cytometry was used for immune profiling of circulating immune cells after nanovesicle mediated TUSC2 intravenous injection. Cytokine gene expression in response to TUSC2 in sorted immune subpopulations was determined by real-time PCR. Tumor growth was significantly reduced with TUSC2 treatment compared with no treatment in both subcutaneous and metastatic mouse models. Synergistic anti-tumor activity was observed when TUSC2 was combined with anti-PD1 verified in five independent experiments. In the lung metastasis model, mice treated with TUSC2 + anti-PD1 lived significantly longer than with single agent treatment. Circulating NK cells increased three fold following TUSC2 nanovesicle intravenous injection both in tumor free and tumor bearing mice which correlated with tumor regression and survival. Cytotoxic T lymphocyte responses were increased whereas Tregs and MDSCs decreased with TUSC2 alone and TUSC2+anti-PD1 treatment. The levels of T cell checkpoint markers PD1, CTLA-4, LAG-3, and TIM-3 evaluated by flow cytometry were decreased after TUSC2 treatment. TUSC2 anti-tumor response was abolished when NK cells were depleted indicating NK cells are important mediators of the TUSC2 treatment effect. Single cell suspension analysis by flow cytometry showed high numbers of NK cells infiltrating lung tumor metastases after TUSC2 treatment. The number of tumor nodules in the lung was significantly less following treatment with TUSC2 nanovesicles compared with control. IL-15 gene expression which mediates NK cell proliferation, was increased by TUSC2. In conclusion, systemic TUSC2 nanovesicle immunogene therapy combined with checkpoint blockade showed synergistic anti-tumor efficacy and activated the immune system through upregulation of NK cells and CTL and downregulation of regulatory cells.
Citation Format: Ismail M. Meraz, Mourad Majidi, RuPing Shao, Meng Feng, Xiaobo Cao, David Rice, Boris Sepesi, Lin Ji, Jack Roth. Tumor suppressor TUSC2 immunogene therapy is synergistic with anti-PD1 in lung cancer syngeneic mouse models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 621. doi:10.1158/1538-7445.AM2017-621