Base editing screens map mutations affecting interferon-γ signaling in cancer.

Area of Science:

• Immunology
• Genetics
• Oncology

Background:

• Interferon-gamma (IFN-γ) signaling is crucial for immune responses against infections, inflammation, and tumors.
• Mutations in the IFN-γ pathway are linked to immune disorders, blood cancers, and resistance to immune checkpoint blockade (ICB) in cancer.
• The functional impact of many identified IFN-γ pathway variants remains largely unknown.

Purpose of the Study:

• To systematically identify genetic factors influencing IFN-γ response in colorectal cancer cells.
• To characterize the functional effects of JAK1 mutations on IFN-γ signaling and anti-tumor immunity.
• To create a resource for interpreting the clinical significance of gene variants in the IFN-γ pathway.

Main Methods:

• Utilized CRISPR-Cas9 screens and base editing mutagenesis for deep mutagenesis of JAK1.
• Performed pathway-wide screens to assess IFN-γ response across various genetic modifications.
• Functionally validated selected variants using primary tumor organoids and engineered missense mutations in JAK1.

Main Results:

• Identified both loss-of-function and gain-of-function mutations in JAK1, including variants relevant to hematological malignancies and ICB resistance.
• Demonstrated that engineered JAK1 missense mutations can alter sensitivity to tumor-reactive T cells in primary organoids.
• Predicted over 300 missense mutations that affect IFN-γ pathway activity.

Conclusions:

• JAK1 mutations significantly modulate IFN-γ pathway activity and cellular responses in colorectal cancer.
• This study provides critical insights into the genetic basis of IFN-γ signaling dysfunction and its implications for cancer immunity and therapy.
• The generated data serves as a valuable resource for understanding gene variant function in clinical contexts.