Gene expression profiling of MC38 Zfx KO clones in vivo
A large fraction of patients with cancer exhibit an intrinsic or adaptive resistance to immunotherapies that aim at boosting T cell responses against tumor cells. To better understand resistance mechanisms in tumor cells, we used CRISPR-Cas9 whole-genome editing to identify mutant tumor cells that are resistant to T cell killing. Among the top hits, we identified the transcription factor zinc-finger protein X-linked (Zfx). Zfx knockout tumor cells are resistant to T cell killing both in vitro and in vivo. Mechanistically we showed that Zfx regulates expression of genes involved in apoptosis pathways and Caspase-3 is one of the central components regulated by Zfx. In addition, ZFX expression is decreased in several human cancer tissues compared to healthy tissues. Female kidney renal clear cell carcinoma (KIRC) patients with high ZFX expression showed significantly better survival compared to patients with low ZFX expression. Our results demonstrate a novel resistant mechanism in tumor cells and suggest that targeting tumor cell intrinsic resistance genes in combination with immune therapies could benefit cancer patients. - The "SAMPLE_ID" sample characteristic is a sample identifier internal to Genentech. Overall design: To delete Zfx from MC38 tumor cells, CRISPR ribonucleoprotein (RNP) complexes were transfected into cells using CRISPRMAX lipofectamine. The KO was confirmed by PCR. RNA was isolated from tumors with RNeasy (QIAGEN) following manufacturer's instructions, including DNase treatment steps. Quality control was performed with NanoDrop (Thermo Scientific) and 1 µg RNA was used for RNA sequencing.