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In vivo discovery of immunotherapy targets in the tumour microenvironment.

ABSTRACT: Recent clinical trials showed that targeting of inhibitory receptors on T cells induces durable responses in a subset of cancer patients, despite advanced disease. However, the regulatory switches controlling T-cell function in immunosuppressive tumours are not well understood. Here we show that such inhibitory mechanisms can be systematically discovered in the tumour microenvironment. We devised an in vivo pooled short hairpin RNA (shRNA) screen in which shRNAs targeting negative regulators became highly enriched in murine tumours by releasing a block on T-cell proliferation upon tumour antigen recognition. Such shRNAs were identified by deep sequencing of the shRNA cassette from T cells infiltrating tumour or control tissues. One of the target genes was Ppp2r2d, a regulatory subunit of the PP2A phosphatase family. In tumours, Ppp2r2d knockdown inhibited T-cell apoptosis and enhanced T-cell proliferation as well as cytokine production. Key regulators of immune function can therefore be discovered in relevant tissue microenvironments.

SUBMITTER: Zhou P

PROVIDER: S-EPMC4052214 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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In vivo discovery of immunotherapy targets in the tumour microenvironment.

Zhou Penghui P Shaffer Donald R DR Alvarez Arias Diana A DA Nakazaki Yukoh Y Pos Wouter W Torres Alexis J AJ Cremasco Viviana V Dougan Stephanie K SK Cowley Glenn S GS Elpek Kutlu K Brogdon Jennifer J Lamb John J Turley Shannon J SJ Ploegh Hidde L HL Root David E DE Love J Christopher JC Dranoff Glenn G Hacohen Nir N Cantor Harvey H Wucherpfennig Kai W KW

Nature 20140129 7486

Recent clinical trials showed that targeting of inhibitory receptors on T cells induces durable responses in a subset of cancer patients, despite advanced disease. However, the regulatory switches controlling T-cell function in immunosuppressive tumours are not well understood. Here we show that such inhibitory mechanisms can be systematically discovered in the tumour microenvironment. We devised an in vivo pooled short hairpin RNA (shRNA) screen in which shRNAs targeting negative regulators bec ...[more]

PMID: 24476824

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Project description:The immune system influences the fate of developing cancers by not only functioning as a tumour promoter that facilitates cellular transformation, promotes tumour growth and sculpts tumour cell immunogenicity, but also as an extrinsic tumour suppressor that either destroys developing tumours or restrains their expansion. Yet, clinically apparent cancers still arise in immunocompetent individuals in part as a consequence of cancer-induced immunosuppression. In many individuals, immunosuppression is mediated by cytotoxic T-lymphocyte associated antigen-4 (CTLA-4) and programmed death-1 (PD-1), two immunomodulatory receptors expressed on T cells. Monoclonal-antibody-based therapies targeting CTLA-4 and/or PD-1 (checkpoint blockade) have yielded significant clinical benefits-including durable responses--to patients with different malignancies. However, little is known about the identity of the tumour antigens that function as the targets of T cells activated by checkpoint blockade immunotherapy and whether these antigens can be used to generate vaccines that are highly tumour-specific. Here we use genomics and bioinformatics approaches to identify tumour-specific mutant proteins as a major class of T-cell rejection antigens following anti-PD-1 and/or anti-CTLA-4 therapy of mice bearing progressively growing sarcomas, and we show that therapeutic synthetic long-peptide vaccines incorporating these mutant epitopes induce tumour rejection comparably to checkpoint blockade immunotherapy. Although mutant tumour-antigen-specific T cells are present in progressively growing tumours, they are reactivated following treatment with anti-PD-1 and/or anti-CTLA-4 and display some overlapping but mostly treatment-specific transcriptional profiles, rendering them capable of mediating tumour rejection. These results reveal that tumour-specific mutant antigens are not only important targets of checkpoint blockade therapy, but they can also be used to develop personalized cancer-specific vaccines and to probe the mechanistic underpinnings of different checkpoint blockade treatments.

Project description:Cancer immunotherapy targets the interplay between immune and cancer cells. In particular, interactions between cytotoxic T lymphocytes (CTLs) and cancer cells, such as PD-1 (PDCD1) binding PD-L1 (CD274), are crucial for cancer cell clearance. However, immune checkpoint inhibitors targeting these interactions are effective only in a subset of patients, requiring the identification of novel immunotherapy targets. Genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) screening in either cancer or immune cells has been employed to discover regulators of immune cell function. However, CRISPR screens in a single cell type complicate the identification of essential intercellular interactions. Further, pooled screening is associated with high noise levels. Herein, we propose intercellular CRISPR screens, a computational approach for the analysis of genome-wide CRISPR screens in every interacting cell type for the discovery of intercellular interactions as immunotherapeutic targets. We used two publicly available genome-wide CRISPR screening datasets obtained while triple-negative breast cancer (TNBC) cells and CTLs were interacting. We analyzed 4825 interactions between 1391 ligands and receptors on TNBC cells and CTLs to evaluate their effects on CTL function. Intercellular CRISPR screens discovered targets of approved drugs, a few of which were not identifiable in single datasets. To evaluate the method's performance, we used data for cytokines and costimulatory molecules as they constitute the majority of immunotherapeutic targets. Combining both CRISPR datasets improved the recall of discovering these genes relative to using single CRISPR datasets over two-fold. Our results indicate that intercellular CRISPR screens can suggest novel immunotherapy targets that are not obtained through individual CRISPR screens. The pipeline can be extended to other cancer and immune cell types to discover important intercellular interactions as potential immunotherapeutic targets.

Dataset Information

In vivo discovery of immunotherapy targets in the tumour microenvironment.

Publications

In vivo discovery of immunotherapy targets in the tumour microenvironment.

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