Abstract Protein tyrosine kinases (PTKs) have been perceived mostly as proto-oncogenes. PTK mutations that have been studied in human cancer usually have increased kinase activities to drive malignant phenotypes. Thus, these PTK mutations are gain-of-function (GOF) mutations. By analyzing the targeted exome sequencing data of 3,274 human tumors from 48 different tissues in the Total Cancer Care (TCC) tumor bank, we have found JAK1 truncating mutations in 36 of 635 gynecologic tumors. The highest JAK1 truncating mutation rate (9.5%) was found in endometrial cancer. These truncating mutations result in the loss of the JAK1 PTK domain. Thus, they are loss-of-function (LOF) mutations. JAK1 truncating mutations in the Cancer Cell Line Encyclopedia (CCLE) databank also occur most often in endometrial cancer cell lines. Analysis of mutation sites identified three mutation hot spots. Re-sequencing of cancer cell lines, primary tumors, and matched normal tissues confirmed JAK1 mutations and revealed that these JAK1 mutations are somatic. JAK1 and JAK2 mediate interferon (IFN)-γ-regulated tumor immune surveillance. Recurrent JAK2 LOF mutations were also found in TCC tumors but at lower rates. Similar to JAK1, the highest rate of LOF JAK2 mutations was found in endometrial/uterine cancer. No JAK1 or JAK2 truncating mutation was identified in 130 samples from hematological malignancies. Functional assays show that JAK1-deficient cancer cells are defective in IFN-γ-induced expression of tumor antigen processing machinery proteins LMP2 and TAP1 and cell surface expression of HLA molecules. These data identify recurrent JAK1 deficiency in endometrial cancer that could impair tumor antigen presentation by the MHC class I complex to cytotoxic T lymphocytes. Our findings suggest that JAK1 is a PTK tumor suppressor and reveal a novel mechanism of tumor immune evasion via LOF mutations in a PTK. Citation Format: Yuan Ren, Yonghong Zhang, Richard Z. Liu, David A. Fenstermacher, Kenneth L. Wright, Jamie K. Teer, Jie Wu. Loss-of-function JAK1 mutations reveal a new role of protein tyrosine kinase mutations in human cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1560. doi:10.1158/1538-7445.AM2014-1560