Abstract 585: AB-1015, a novel integrated circuit T cells containing an ALPG/MSLN logic gate and FAS/PTPN2 shRNA-miR, demonstrates specific and potent activity against ALPG/MSLN tumors
Authors: Santoro Stephen , Cooper Aaron , Bezman Natalie , Feng Jun , Chawla Kanika , McDevitt Jennifer , Mikkelsen Tarjei , Jun Susie , Haining W. Nicholas 
Published: June 2022

Abstract
In solid tumors, CAR T cell efficacy is limited by off-tumor toxicity, poor persistence, and suppression by the tumor microenvironment (TME). To address these challenges we have engineered AB-1015, an integrated circuit T cell (ICT cell) intended for the treatment of ovarian cancer. The AB-1015 transgene cassette encodes an “AND” logic gate designed to limit off-tumor toxicity through dual tumor antigen recognition and a dual shRNA-miR to resist TME suppression and improve ICT cell function and persistence. The AB-1015 DNA cassette is inserted into the T cell genome at a defined novel genomic site via CRISPR-based gene editing. The AB-1015 logic gate consists of a priming receptor that induces expression of an anti-mesothelin (MSLN) CAR upon engagement of a ALPG/P (alkaline phosphatase germ-line/placental). The dual-antigen specificity of the logic gate was assessed in mice engrafted with MSLN+ and ALPG/P+MSLN+ K562 tumors established on contralateral flanks. AB-1015 ICT cells eliminated ALPG/P+MSLN+ tumors, while sparing tumors that lacked ALPG/P. To assess the ability of AB-1015 to mediate killing of MSLN+ tumor cells in the context of heterogeneous cultures, we utilized an admixed co-culture system where ALPG/P+ target cells were spiked into cultures that were otherwise MSLN+. AB-1015 was able to eliminate admixed co-cultures where as few as 5-15% of the target cells expressed ALPG/P.The AB-1015 also contains a dual shRNA-miR that targets FAS and PTPN2, two critical mediators of T cells survival and function. FASL, the cognate ligand for FAS receptor, is expressed on the surface of activated T cells and is significantly overexpressed in the ovarian cancer TME. In vitro, AB-1015 demonstrated resistance to FAS-mediated apoptosis. Knockdown of PTPN2, a phosphatase involved in T cell proliferation and functional persistence, resulted in enhanced AB-1015 ICT cell expansion during repetitive stimulation over a period of 14 days, as well as a 30-fold reduction in tumor outgrowth compared with logic gated T cells alone. In summary, AB-1015 ICT cells are specific for ALPG/P+MSLN+, demonstrate superior potency, expansion, and persistence compared with logic gated T cells alone, and are resistant to ovarian TME suppression. These results support further evaluation of AB-1015 as a novel therapy for indications including ovarian, fallopian tube, or primary peritoneal cancer.
Citation Format: Stephen Santoro, Aaron Cooper, Natalie Bezman, Jun Feng, Kanika Chawla, Jennifer McDevitt, Tarjei Mikkelsen, Susie Jun, W. Nicholas Haining. AB-1015, a novel integrated circuit T cells containing an ALPG/MSLN logic gate and FAS/PTPN2 shRNA-miR, demonstrates specific and potent activity against ALPG/MSLN tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 585.