Proteasome inhibitors restore the STAT1 pathway and enhance the expression of MHC class I on human colon cancer cells.

BACKGROUND: A new strategy, particularly a novel combination, for immunotherapy in microsatellite stable metastatic colorectal cancer (mCRC) treatment needs to be formulated. Studies on the interferon-γ (IFN-γ)/ Janus kinase (JAK)/ signal transducer and activator of transcription (STAT)1 pathway provide new directions in this regard.

METHODS: Our study applies three colon cancer cell lines, including microsatellite stable (MSS) cell lines, which are SW480 and SW620, and microsatellite instability-high (MSI-H) cell line, which is DLD-1. We compared the expressions of immune surface markers on colon cancer cells in response to IFN-γ. We elucidated these mechanisms, which involved the upregulation of immune surface markers. Furthermore, we examined real-world clinical samples using the PerkinElmer Opal multiplex system and NanoString analysis.

RESULTS: We established that the baseline expression of major histocompatibility complex (MHC) class I alleles and programmed death-ligand 1 (PD-L1) were generally low in cell line models. The immune surface markers were significantly increased after IFN-γ stimulation on SW480 but were notably unresponsive on the SW620 cell line. We discovered that STAT1 and phosphorylated STAT1 (pSTAT1) were downregulated in the SW620 cell line. We verified that the STAT1/pSTAT1 could be restored through the application of proteasome inhibitors, especially bortezomib. The expression of MHC class I as downstream signals of STAT1 was also up-regulated by proteasome inhibitors. The similar results were reproduced in DLD-1 cell line, which was also initially unresponsive to IFN-γ. In real-world samples of patients with mCRC, we found that higher STAT1 expression in tumor cells was strongly indicative of a highly immunogenic microenvironment, with significantly higher expression levels of MHC class I and PD-L1, not only on tumor cells but also on non-tumor cells. Furthermore, tumor infiltrating lymphocytes (TILs) were increased in the positive-STAT1 group. Through NanoString analysis, we confirmed that the mRNA expressions of IFN-γ, human leukocyte antigen (HLA)-A, HLA-E, and HLA-G were also significantly higher in the positive-STAT1 group than those in the negative-STAT1 group.

CONCLUSION: Our study provides a novel rationale for the addition of bortezomib, a proteasome inhibitor, into new immunotherapy combinations.