Base editing screens map mutations affecting interferon-y signaling in cancer

Interferon-y (IFN-y) signaling mediates host responses to infection, inflammation and anti-tumor immunity. Mutations in the IFN-y signaling pathway cause immunological disorders, hematological malignancies, and resistance to immune checkpoint blockade (ICB) in cancer; however, the function of most clinically observed variants remains unknown. Here, we systematically investigate the genetic determinants of IFN-y response in colorectal cancer cells using CRISPR-Cas9 screens and base editing mutagenesis. Deep mutagenesis of JAK1 with cytidine and adenine base editors, combined with pathway-wide screens, reveal loss-of-function and gain-of-function mutations, including causal variants in hematological malig-nancies and mutations detected in patients refractory to ICB. We functionally validate variants of uncertain significance in primary tumor organoids, where engineering missense mutations in JAK1 enhanced or reduced sensitivity to autologous tumor-reactive T cells. We identify more than 300 predicted missense mu-tations altering IFN-y pathway activity, generating a valuable resource for interpreting gene variant function.

Automated summary

In this study, the genetic determinants of IFN-γ response in colorectal cancer cells were systematically investigated using CRISPR-Cas9 screens and base editing mutagenesis. The researchers found loss-of-function and gain-of-function mutations in the JAK1 gene, which were associated with hematological malignancies and resistance to immune checkpoint blockade in cancer patients. Additionally, functional validation of uncertain variants in primary tumor organoids showed that missense mutations in JAK1 could enhance or reduce sensitivity to autologous tumor-reactive T cells. This study provides a valuable resource for interpreting gene variant function, identifying over 300 predicted missense mutations altering IFN-γ pathway activity.