Blockade of IL-6 inhibits tumor immune evasion and improves anti–PD-1 immunotherapy
Authors: Ma Wenhui , Li Wenyi , Wu Zhaokun , Zhang Chaoting , Cheng Mingzhen , Chen Yuehong , Zou Yini , Li Kejun , Lin Simin , Xiong Wenjun , Wang Ying , Lin Yixiong , Meng Weidong , Zhou Weijie 
Published: March 2022

Abstract
BackgroundLong-standing inflammatory bowel disease predisposes to the development of colorectal cancer (CRC). Interleukin (IL) -6, a pivotal link between chronic inflammation and tumor progression, has recently been recognized as a potential target. The effect of IL-6 on proliferation and metastasis of CRC by activating STAT3 pathway has been widely demonstrated in recent years, but few on mediating tumor immune evasion has been reported. MethodsIn this study, we analyzed IL-6 gene expression in human CRC tissues and carried out survival probability in CRC patients with TCGA Data portal and GEO Database. We studied CRC tumorigenesis in vivo by inoculating MC38 tumors and induced-CRC model via AOM/DSS （azoxymethan / dextran sulfate sodium）in (IL-6 deficient) IL-6-/- and (wild type) WT mice. We measured tumor growth and detected CD8+ T cells via immunofluorescence, we validated the function of IL-6 in tumorigenesis and tumor immune evasion in subcutaneous transplantation models. Using endoscopy monitoring and immunofluorescence, we validated the function of IL-6 in tumor immune evasion in AOM/DSS models. We conducted colitis model with DSS reveal the relationship of IL-6 and MHC-1 molecules by qRT-PCR and bioinformatics analysis. Organoid and CRC cells culture, western blot analysis, and qRT-PCR were used to investigate the expression and significance of MHC-1 molecules in vitro without IL-6 stimulation. MHC-1 molecules levels were also detected in tumors of IL-6-/- and WT mice from MC38-bearing tumor models and AOM/DSS models. Subcutaneous MC38 implantation models in C57BL/6 J mice were established to investigate anti-IL-6 therapy on CRC tumorigenesis. In vivo efficacy were conducted using antibodies blocking IL-6 and PD-1 in mice subcutaneous-bearing different types of tumors. Results Bio-information analysis and results of our center founded that IL-6 was remarkably overexpressed in CRC and its elevation was associated with poor prognosis. We observed that IL-6-/- mice were less susceptible to develop tumors, compared to (wild type) WT mice. Interestingly, infiltrated CD8+ T cells were found to exhibit high expression in tumor of IL-6-/- mice. High level of IL-6 was found in colitis model, with down-regulation of MHC-1molecules. Moreover, in vitro experiments, we found that IL-6 may act as a negative regulator in IFNγ-STAT1-MHC-1 signaling. In addition, vivo trials also confirmed that MHC-1 mRNA levels are negatively related to the existence of IL-6. Furthermore, blockade of IL-6 also activated CD8+ T-cell accumulation and led to the high PDL-1 expression in CRC, which can sensitize animals to anti-PD-1 therapy. Importantly, compared to treatment with anti-IL-6 alone and single treatment of anti-PD-1, combined IL-6 and PD-1 blockade greatly improved the therapeutic effect, not only in CRC but also melanoma and breast cancer.Conclusions Our study demonstrated that IL-6 act as a negative regulator in immunological surveillance to promote tumor immune evasion by inhibiting the response of cell to IFNγ. Blocking IL-6 enhanced tumor response to anti-PD-1 therapy via infiltration of CD8+ T cell and high expression of PDL-1. Based on the findings of our study, IL-6 is a novel target to improve the efficacy of immunotherapy.