Two Is Better Than One: Adoptive T Cell Therapy Targeting Tumor Antigens And The Tumor Endothelium Potentiates T Cell Activation And Cytolytic Activity

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Background: T cells can be genetically modified to express chimeric antigen receptors (CAR) molecules on the surface. These CAR-bearing T cells combine the specificity of an antibody with the killing properties of a T cell for a targeted immunotherapy approach. Previously our lab has targeted the tumor antigen HER2, using HER2 specific CAR T cells. While these CAR T cells induced tumor regression in an orthotopic xenogeneic model, a subset of mice went on to recur. Tumor endothelium marker 8 (TEM8), is a tumor endothelium associated antigen that is conserved in mice and humans. Purpose: The purpose of this study is to determine whether CAR T cells specific for both TEM8 and HER2 would display increased cytolytic activity and/or activation. Procedure: We designed in silico, a single CAR molecule composed of TEM8 and HER2 specific exodomains in tandem. These domains were linked together via a glycine serine linker and shared a CD28 and CD3-zeta chain signaling domain, to form a TEM8-HER2 TanCAR. The construct was then synthesized, cloned into a retroviral vector and used to transduced T cells. New results: TanCARs were expressed on >90% of primary T cells; up 20% from a previous report of 70%. When tested against TEM8 and HER2 double negative, single positive and double positive cell lines TanCAR T cells were able to distinctly recognize and TEM8 and HER2 positive targets as indicated by standard 4 hour 51Cr release assays. Particularly, when TanCAR cells were cultured with TEM8 and HER2 double positive cell lines versus single positive (TEM8+ or HER2+) there was a 10-14% increase in cytolytic activity. Increased T cell activation was also indicated by the release of the immune-stimulatory cytokines interferon-gamma and interleukin-2 in in vitro coculture assays. Forced expression of TEM8 on the same HER2 positive cell line resulted in a >3 fold increase in release of interferon-gamma. Further, IL-2 release was only detected when TanCAR T cells were exposed to cell lines positive for both antigens. Conclusion: The distinct recognition and increased cytolytic activity and T cell activation demonstrated by TEM8-HER2 specific TanCAR T cells upon exposure to two target antigens warrants further investigation. This single CAR molecule could potentially have therapeutic benefit in targeting not only the tumor, but associated vasculature in HER2 positive solid tumors. Citation Format: Tiara Byrd, Kristen Fousek, Zakaria Grada, Kevin Aviles-Padilla, Kevin Bielamowicz, Stephen Gottschalk, Bradley St Croix, Bradley Fletcher, Meenakshi Hegde, Nabil Ahmed. Two is better than one: Adoptive T cell therapy targeting tumor antigens and the tumor endothelium potentiates T cell activation and cytolytic activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-264. doi:10.1158/1538-7445.AM2014-LB-264