Abstract 1822: IFN-γ and TRAIL involvement in androgen signaling-mediated NK cell killing of prostate cancer cells and further enhancement by NKG2A checkpoint inhibition with monalizumab
Authors: Schwermann Maximilian , Carlsen Lindsey , Huntington Kelsey E. , Zhou Lanlan , George Andrew , Srinivasan Praveen , Tajiknia Vida , Cruz Arielle De La , De Souza Andre , Mega Anthony E. , Safran Howard P. , Carneiro Benedito A. , El-Deiry Wafik S. 
Published: April 2023

Abstract
Although blocking the androgen receptor (AR) pathway is a core treatment strategy in early prostate cancer, most patients progress to metastatic castration-resistant prostate cancer(mCRPC). Immunotherapy with PD-1/PD-L1 inhibition has limited activity in mCRPC. AR signaling modulates CD8+ T cells and antitumor immune responses in mCRPC. In a companion presentation (Schwermann et al., submitted AACR 2023), we demonstrate that AR blockade activates NK cell killing of prostate cancer (PC) cells. We hypothesize this activation requires IFN-γ and TRAIL pathways. We investigated the mechanisms of AR inhibitors (ARi)enzalutamide (Enza) and darolutamide (Daro) on the antitumor function of NK cells and potential enhancement with checkpoint inhibitors. Material and Methods: We performed ATP-based CellTiterGlo viability assays to investigate the effects of Enza and Daro on PC cells (LNCap, 22Rv1 [ARv7mutation], DU145, PC3 [AR-]) and NK-92 cells. We performed co-culture experiments with PC, NK, and T cells, at a 1:1 ratio, using IFN-γ blocking mAb (10 μg/ml), RIK-2 (TRAIL blocking mAb- 10 μg/ml) and analyzed immune cell-mediated tumor cell killing (ImageXpress Confocal HT). The NK cell immune checkpoint inhibitor monalizumab that targets NKG2A was combined with ARi.
Results: Co-cultures of PC plus NK-92 cells with ARi significantly increased immune-mediated PCkilling within 24hrs (control[C]: 20%±1.9; Enza: 40%±4.3; Daro: 36%±3.72, p=0.0001). This immune enhancement effect was abolished by IFN-γ mAb. The combination of TRAIL inhibitory mAb with Enza decreased NK immune-mediated killing by 16±2.4% ([C]: 40%±4.3; Enza+RIK-2:25.6%±2.1, p=0.002). Treatment of PC cells plus NK cells with Enza, IFN-γ mAb, and TRAIL-blocking RIK-2 significantly reduced NK cell-mediated killing of PCs (control[C]: 20%±1.9, [T]:3.2%±2.6, p=0.003). ARi immune enhancement effect was increased by NKG2A blockade with monalizumab ([C]: 31.2%±3.1, [T]: 43.92%±4.52, p=0.0045). Conclusions: ARi promotes NK cell killing of PC cells via IFN-γ gamma and TRAIL. NK cell activation is enhanced by the combination of ARi and the checkpoint inhibitor monalizumab. Validation of these results in patient-derived organoids paired with tumor samples from patients with PC is ongoing to further understand the innate immune response and direct novel therapeutic strategies.
Citation Format: Maximilian Schwermann, Lindsey Carlsen, Kelsey E. Huntington, Lanlan Zhou, Andrew George, Praveen Srinivasan, Vida Tajiknia, Arielle De La Cruz, Andre De Souza, Anthony E. Mega, Howard P. Safran, Benedito A. Carneiro, Wafik S. El-Deiry. IFN-γ and TRAIL involvement in androgen signaling-mediated NK cell killing of prostate cancer cells and further enhancement by NKG2A checkpoint inhibition with monalizumab [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1822.