Osteopontin alters DNA methylation through up-regulating DNMT1 and sensitizes CD133+/CD44+ cancer stem cells to 5 azacytidine in hepatocellular carcinoma

Abstract

BackgroundIn hepatocellular carcinoma (HCC), CD133+/CD44+ cells are one subgroup with high stemness and responsible for metastatic relapse and resistance to treatment. Our previous studies have demonstrated that osteopontin (OPN) plays critical roles in HCC metastasis. We further investigated the molecular mechanism underlying the role of OPN in regulating the stemness of HCC epigenetically and explored possible targeting strategy.MethodsCD133+/CD44+ subgroup sorting from HCC cell lines and HCC tissues was used to investigate the effects of OPN knockdown on stemness. iTRAQ and MedIP-sequencing were applied to detect the protein profile and epigenetic modification of CD133+/CD44+ subgroup with or without OPN knockdown. The antitumor effects of 5 Azacytidine were examined in cultured HCC cells and patient derived xenograft (PDX) models.ResultsOPN was accumulated in CD133+/CD44+ subgroup of HCC cells. Knocking down OPN significantly inhibited the sphere formation and stemness-related genes expression, and delayed tumor initiation of CD133+/CD44+ subgroup of HCC cells. Employing MedIP-sequencing, dot blot and iTRAQ analyses of CD133+/CD44+ SCR and CD133+/CD44+ shOPN cells, we found that OPN knockdown leaded to reduction in DNA methylation with particular enrichment in CGI. Meanwhile, DNA (cytosine-5)-methyltransferase 1 (DNMT1), the main methylation maintainer, was downregulated via proteomics analysis, which mediated OPN altering DNA methylation. Furthermore, DNMT1 upregulation could partially rescue the properties of CD133+/CD44+ shOPN cells. Both in vitro and in vivo assays showed that CD133+/CD44+ cells with high OPN levels were more sensitive to DNA methylation inhibitor, 5 Azacytidine (5 Aza). The above findings were validated in HCC primary cells, a more clinically relevant model.ConclusionsOPN induces methylome reprogramming to enhance the stemness of CD133+/CD44+ subgroup and provides the therapeutic benefits to DNMT1 targeting treatment in HCC.Electronic supplementary materialThe online version of this article (10.1186/s13046-018-0832-1) contains supplementary material, which is available to authorized users.

Identification of a Subtype of Hepatocellular Carcinoma with Poor Prognosis Based on Expression of Genes within the Glucose Metabolic Pathway

“…CA9 and PROM1 were also significantly overexpressed in Cluster2 even though these are not the most overexpressed genes (Table S7). Notably, SPP1 is one of the top upregulated genes in Cluster2 patients in both data sets ( Figure 7A, Tables S6 and S7), which was reported to enhance the stemness of HCC cells [19] and to be involved in PD-L1-mediated immune escape of HCC [20]. The characteristics of Cluster 2 identified by us is very similar to those of iCl1/C1 cluster, as reported by Woo et al [17].…”

The DNMT1/miR-34a/FOXM1 Axis Contributes to Stemness of Liver Cancer Cells

“…Recently, silencing tumor repressors or suppressive microRNAs in precancerous cells by epigenetic regulation has attracted increasing attention [31]. DNMT1 is abnormally activated in tumor tissues and cancer stem cells and functions by catalyzing DNA methylation [32,33]. In addition, DNMT1 is required for maintaining CSLC characteristics [33].…”

“…DNMT1 is abnormally activated in tumor tissues and cancer stem cells and functions by catalyzing DNA methylation [32,33]. In addition, DNMT1 is required for maintaining CSLC characteristics [33]. In breast cancer, epigenetic modifying agents, such as 5-azacytidine, could markedly reduce the CSC subpopulation [34].…”

TCA Cycle Rewiring as Emerging Metabolic Signature of Hepatocellular Carcinoma

“…Since its overexpression is due to epigenetic regulation, using epigenetic modifier could be tested. It is widely demonstrated that epigenetic alterations are associated with different mechanisms of proliferation and metastasis in several types of cancer, including HCC [139,[161][162][163]. Histone acetylation is one of most important epigenetic mechanisms, regulating cellular events such as differentiation, proliferation, and cell cycle.…”