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Immune checkpoint blockade therapy aims to activate the immune system to eliminate cancer cells. However, clinical benefits are only recorded in a subset of patients. Here, we leverage genome-wide CRISPR/Cas9 screens in a Tumor-Immune co-Culture System focusing on triple-negative breast cancer (TNBC). We reveal that NEDD8 loss in cancer cells causes a vulnerability to nivolumab (anti-PD-1). Genetic deletion of NEDD8 only delays cell division initially but cell proliferation is unaffected after recovery. Since the NEDD8 gene is commonly essential, we validate this observation with additional CRISPR screens and uncover enhanced immunogenicity in NEDD8 deficient cells using proteomics. In female immunocompetent mice, PD-1 blockade lacks efficacy against established EO771 breast cancer tumors. In contrast, we observe tumor regression mediated by CD8+ T cells against Nedd8 deficient EO771 tumors after PD-1 blockade. In essence, we provide evidence that NEDD8 is conditionally essential in TNBC and presents as a synergistic drug target for PD-1/L1 blockade therapy. NEDD8 is a ubiquitin-like protein that governs protein neddylation, previously demonstrated to be essential for cell survival. Here the authors show that NEDD8 loss in breast cancer cells is associated with enhanced immunogenicity and increased sensitivity to PD-1 blockade in preclinical cancer models.

The cancer-intrinsic network controlling the response to immune checkpoint blockade therapy remains elusive. Using a published single-cell RNA sequencing (scRNA-seq) dataset from patients with breast cancer, we found that pretherapy mRNA expression of NEDD8, which encodes the regulatory protein essential for neddylation, in cancer cells was negatively associated with T-cell expansion upon pembrolizumab treatment. Genetic ablation of protein neddylation in murine tumors induced a curative response to PD-1 or PD-L1 blockade in mice. Combined analysis of neddylation-deficient murine tumors using scRNA-seq and high-resolution spatial transcriptomics revealed distinct spatial immune co-localization and immune cell signatures. In a high-content phenotypic screen of known compounds, protein neddylation was required for the induction of phenotypic changes by lipid inhibitors and ferroptosis inducers in human breast cancer cells. In accordance, these compounds enhanced the efficacy of PD-1 blockade and protected mice from tumor rechallenge. Altogether, we delineate the unknown functions of protein neddylation on antitumor immunity and provide therapeutic options that can enhance immunotherapy.