Silencing of matrix metalloproteinase MMP9 nurtures tumor microenvironment in chronic inflammation driven colon cancer

Abstract

IntroductionChronic inflammation nurtures tumor microenvironment due to the release of various biomolecules such as cytokines, chemokines and growth factors which are the first response of the cell against the inflammation. These biomolecules contribute to the accumulation of reactive oxygen species (ROS) which leads to DNA damage. Matrix Metalloproteinases (MMPs) are zinc dependent endopeptidases which mediate inflammation, tissue remodeling, and tumorigenesis. Among them MMP9 is the unique one because it is undetectable in healthy tissues and highly upregulated during inflammation and cancer. We have previously shown that MMP9 plays a protective role in chronic inflammation associated colon cancer (CAC) which is opposite to its conventional role of mediating acute inflammation and sporadic colon cancer. Therefore, we hypothesize that in chronic inflammatory conditions of colon direct and/or indirect silencing of MMP9 should be avoided.AimAim of this study is to understand the mechanistic explanation for the failure of CAC therapeutics via MMP9 silencing (such as failure of clinical trial by Geliad with GS5745).MethodsWe used MMP9 small interfering RNA (siRNA) loaded nanoparticles and gavaged 10 weeks old WT (C57/B6) mice to inhibit MMP9 expression in colonic epithelium. CAC was induced by three cycles of dextran sodium sulfate. We also used C57/B6 transgenic mice with constitutive expression of MMP9 under villin promoter (TgM9) and their wild type (VWT) littermates.ResultsWT mice treated with MMP9 siRNA loaded nanoparticles (WTnpM9) showed worsened CAC compared to WT mice given scrambled siRNA loaded nanoparticles (control group) as reflected by more weight loss, higher clinical score and more polyps and dysplastic lesions. In vivo ROS estimation and immunofluorescence (IF) staining showed increased ROS levels and expressions of ROS marker‐8OHDG and DSBs (double stand breaks) marker SOD among WTnpM9 compared to control group in CAC. Western blot (WB) data showed higher levels of γH2AX but decreased levels of mismatch repair (MMR) protein MLH1. Our proof of principle model of TgM9 mice confirmed that in CAC, MMP9 keeps a check on ROS levels by maintaining the microbial population (QPCR analysis of 16SrRNA, Firmicutes, Bacteroidetes, Akkermansia muciniphila and Fusobacterium). This results in maintenance of genomic stability through activation of MMR proteins (WB analysis) in CAC. Enteroids isolated from TgM9 and VWT mice were analyzed by IF staining for proliferation marker (PCNA) and antioxidant marker (SEPP1). The data suggested that MMP9 regulates proliferation and ROS levels.ConclusionOur study showed that MMP9 expression correlates with the reduced levels of ROS, decreased DNA damage, and activation of MMR proteins. Our study recognizes that despite being a proteinase MMP9 has a novel beneficial role of a tumor suppressor in CAC. Therefore in the setting of chronic inflammation MMP9 silencing should be avoided for CAC patients.Support or Funding InformationNIDDK‐R01 Grant DK064711CCFA career development grant, award number 3057NIDDK‐R01 Grant DK097256This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.