OBX-115 membrane-bound IL15 (mbIL15)-expressing engineered tumor-infiltrating lymphocytes (TIL) overcome target cell immune evasion related to major histocompatibility complex (MHC) silencing through both direct killing and transactivation of NK cells in patient-derived autologous non-small cell lung cancer (NSCLC) models

Background: Non-engineered TIL cell therapy has demonstrated antitumor activity in NSCLC [Schoenfeld Cancer Discov 2024, Creelan Nat Med 2021]. MHC loss remains a key mechanism of resistance to TIL cell therapy and other immunotherapies in NSCLC [Wang Nat Cancer 2025]. Tumoricidal CD8+ TIL have been shown to demonstrate MHC-independent cytotoxicity through lymphotoxin in melanoma, and are enriched in patients responsive to TIL therapy [Xie J Immunother Cancer 2025]. OBX-115 engineered TIL express mbIL15 under pharmacologic regulation using the FDA-approved small-molecule drug acetazolamide (ACZ). We hypothesize that OBX-115 will retain cytotoxicity and reactivity following MHC loss in NSCLC due to mbIL15-mediated NK cell transactivation. Methods: OBX-115 TIL were generated from NSCLC tumor samples as previously described [Bott SITC 2024]. Autologous NK cells were expanded from donor-matched peripheral blood mononuclear cells. Autologous patient-derived cell lines (PDc) were genetically modified to knock-out (KO) the B2M locus; B2M KO was confirmed using flow cytometry and Sanger sequencing. After co-culture with OBX-115 TIL and ACZ, NK cell transactivation was assessed by detection of phosphorylated STAT5 (pSTAT5) via flow cytometry. OBX-115 TIL reactivity and cytotoxicity against dye-labeled PDc were functionally assessed by quantitation of IFNgamma secretion and live-cell imaging. Results: B2M KO efficiency was 98.1% ± 0.7% (% of MHC-I-negative cells in the modified PDc). OBX-115 TIL partially retained reactivity against autologous tumor cell lines with the genetic B2M KO (IFNgamma secretion 821.2 ± 562.6 pg/mL against autologous tumor cell lines and 762.4 ± 545.4 pg/mL against corresponding B2M KO cell lines, n=4, p=0.27). Cytotoxicity against autologous B2M KO models was also partially retained as demonstrated in an orthogonal 3D spheroid co-culture model. Upon co-culture with autologous NK cells, OBX-115 TIL were able to transactivate the NK cells (pSTAT5+%: NK alone, 0.6% ± 0.3%; NK + OBX-115 + ACZ, 21.8% ± 9.5%, n=3, p=0.02). In a tri-culture system using patient-specific TIL, NK cells, and B2M-KO tumor cells, the TIL/NK mixture elicited higher ACZ-dependent cytotoxicity and IFNgamma secretion than TIL alone (23% higher IFNgamma secretion and 65% higher cytotoxicity by caspase staining, n=1). Conclusions: Our data demonstrate that, similar to previously reported MHC-I-independent tumoricidal effect observed in melanoma CD8+ T cells, OBX-115 engineered mbIL15-expressing NSCLC TIL partially retain cytotoxicity and reactivity toward autologous tumor cells in MHC-loss models. Further, transactivation of autologous NK cells via mbIL15 uniquely enhances activity.