Using a genome-wide shRNA library targeting 39,000 transcripts in a murine triple negative breast cancer cell line, Shuptrine et al. employed a novel strategy of differential target gene enrichment or depletion in tumors growing in immune-competent compared to immune-incompetent mice to identify genes involved in immune pathways affecting tumor growth. Both previously known critical regulators (TGFβ, CD47) and novel regulators (Sgpl1, Tex9, Pex 14) were identified and validated.

The clinical successes of immune checkpoint therapies for cancer make it important to identify mechanisms of resistance to anti-tumor immune responses. Numerous resistance mechanisms have been identified employing studies of single genes or pathways, thereby parsing the tumor microenvironment complexity into tractable pieces. However, this limits the potential for novel gene discovery to in vivo immune attack. To address this challenge, we developed an unbiased in vivo genome-wide RNAi screening platform that leverages host immune selection in strains of immune-competent and immunodeficient mice to select for tumor cell-based genes that regulate in vivo sensitivity to immune attack. Utilizing this approach in a syngeneic triple-negative breast cancer (TNBC) model, we identified 709 genes that selectively regulated adaptive anti-tumor immunity and focused on five genes (CD47, TGFbeta1, Sgpl1, Tex9 and Pex14) with the greatest impact. We validated the mechanisms that underlie the immune-related effects of expression of these genes in different TNBC lines, as well as tandem synergistic interactions. Furthermore, we demonstrate the impact of different genes with previously unknown immune functions (Tex9 and Pex14) on anti-tumor immunity. Thus, this innovative approach has utility in identifying unknown tumor-specific regulators of immune recognition in multiple settings to reveal novel targets for future immunotherapies.

Author Info: (1) Department of Oncology and Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, 3970 Reservoir Road NW, Washington DC, 20057, USA. Department of Surgery,

Author Info: (1) Department of Oncology and Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, 3970 Reservoir Road NW, Washington DC, 20057, USA. Department of Surgery, Duke University Medical Center, Durham, NC, USA. (2) Department of Oncology and Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, 3970 Reservoir Road NW, Washington DC, 20057, USA. (3) Department of Oncology, Division of Biostatistics, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University, Baltimore, MD, USA. (4) Department of Oncology and Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, 3970 Reservoir Road NW, Washington DC, 20057, USA. (5) Department of Surgery, Duke University Medical Center, Durham, NC, USA. (6) Department of Surgery, Duke University Medical Center, Durham, NC, USA. (7) Department of Oncology and Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, 3970 Reservoir Road NW, Washington DC, 20057, USA. weinerl@georgetown.edu.