Highlights d Lung cancer development is associated with local dysbiosis and inflammation d Depletion of commensal microbiota suppresses lung adenocarcinoma development d Microbiota drive proliferation and activation of Vg6 + Vd1 + T cells in lung cancer d gd T cells promote neutrophil infiltration and tumor cell proliferation

Effective immunosurveillance of cancer requires the presentation of peptide antigens on major histocompatibility complex Class I (MHC-I). Recent developments in proteomics have improved the identification of peptides that are naturally presented by MHC-I, collectively known as the "immunopeptidome". Current approaches to profile tumor immunopeptidomes have been limited to in vitro investigation, which fails to capture the in vivo repertoire of MHC-I peptides, or bulk tumor lysates, which are obscured by the lack of tumor-specific MHC-I isolation. To overcome these limitations, we report here the engineering of a Cre recombinase-inducible affinity tag into the endogenous mouse MHC-I gene and targeting of this allele to the Kras LSL-G12D/+ ; p53 fl/fl (KP) mouse model (KP; K b Strep). This novel approach has allowed us to isolate tumor-specific MHC-I peptides from autochthonous pancreatic ductal adenocarcinoma (PDAC) and lung adenocarcinoma (LUAD) in vivo. With this powerful analytical tool, we were able to profile the evolution of the LUAD immunopeptidome through tumor progression and show that in vivo MHC-I presentation is shaped by post-translational mechanisms. We also uncovered novel, putative LUAD tumor associated antigens (TAAs). Many peptides that were recurrently presented in vivo exhibited very low expression of the cognate mRNA, provoking reconsideration of antigen prediction pipelines that triage peptides according to transcript abundance. Beyond cancer, the K b Strep allele is compatible with a broad range of Cre-driver lines to explore antigen presentation in vivo in the pursuit of understanding basic immunology, infectious disease, and autoimmunity.

Purpose: Despite the accumulation of extensive genomic alterations, many cancers fail to be recognized as "foreign" and escape destruction by the host immune system. Immunotherapies designed to address this problem by directly stimulating immune effector cells have led to some remarkable clinical outcomes, but unfortunately, most cancers fail to respond, prompting the need to identify additional immunomodulatory treatment options.Experimental Design: We elucidated the effect of a novel treatment paradigm using sustained, low-dose HSP90 inhibition in vitro and in syngeneic mouse models using genetic and pharmacologic tools. Profiling of treatment-associated tumor cell antigens was performed using immunoprecipitation followed by peptide mass spectrometry.Results: We show that sustained, low-level inhibition of HSP90 both amplifies and diversifies the antigenic repertoire presented by tumor cells on MHC-I molecules through an IFNgindependent mechanism. In stark contrast, we find that acute, high-dose exposure to HSP90 inhibitors, the only approach studied in the clinic to date, is broadly immunosuppressive in cell culture and in patients with cancer. In mice, chronic nonheat shock-inducing HSP90 inhibition slowed progression of colon cancer implants, but only in syngeneic animals with intact immune function. Addition of a single dose of nonspecific immune adjuvant to the regimen dramatically increased efficacy, curing a subset of mice receiving combination therapy.Conclusions: These highly translatable observations support reconsideration of the most effective strategy for targeting HSP90 to treat cancers and suggest a practical approach to repurposing current orally bioavailable HSP90 inhibitors as a new immunotherapeutic strategy.See related commentary

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Effective immunosurveillance of cancer requires the presentation of peptide antigens on major histocompatibility complex Class I (MHC-I). Recent developments in proteomics have improved the identification of peptides that are naturally presented by MHC-I, collectively known as the “immunopeptidome”. Current approaches to profile tumor immunopeptidomes have been limited to in vitro investigation, which fails to capture the in vivo repertoire of MHC-I peptides, or bulk tumor lysates, which are obscured by the lack of tumor-specific MHC-I isolation. To overcome these limitations, we report here the engineering of a Cre recombinase-inducible affinity tag into the endogenous mouse MHC-I gene and targeting of this allele to the KrasLSL-G12D/+; p53fl/fl (KP) mouse model (KP; KbStrep). This novel approach has allowed us to isolate tumor-specific MHC-I peptides from autochthonous pancreatic ductal adenocarcinoma (PDAC) and lung adenocarcinoma (LUAD) in vivo. With this powerful analytical tool, we were able to profile the evolution of the LUAD immunopeptidome through tumor progression and show that in vivo MHC-I presentation is shaped by post-translational mechanisms. We also uncovered novel, putative LUAD tumor associated antigens (TAAs). Many peptides that were recurrently presented in vivo exhibited very low expression of the cognate mRNA, provoking reconsideration of antigen prediction pipelines that triage peptides according to transcript abundance. Beyond cancer, the KbStrep allele is compatible with a broad range of Cre-driver lines to explore antigen presentation in vivo in the pursuit of understanding basic immunology, infectious disease, and autoimmunity.