A Gender-Dependent Molecular Switch of Inflammation via MyD88/Estrogen Receptor-Alpha Interaction.

INTRODUCTION: Most Toll-like receptors and IL-1/IL-18 receptors activate a signaling cascade via the adaptor molecule MyD88, resulting in NF-κB activation and inflammatory cytokine and chemokine production. Females are less susceptible than males to inflammatory conditions, presumably due to protection by estrogen. The exact mechanism underlying this protection is unknown. METHODS: MCF7 cells expressing wild-type or mutated LXXLL motif were used to determine MyD88/estrogen receptor (ER)-a interaction by immunoprecipitation and cell activation by ELISA and luciferase reporter assay. IL-1b and/or E2 were used to activate MCF7 cells expressing normal or knocked down levels of PRMT1. Finally, in situ proximity ligation assay with anti-MyD88 and anti-methylated ER-a (methER-a) antibodies was used to evaluate MyD88/methylated ER-a interaction in THP1 cells and histological sections. RESULTS: We show that MyD88 interacts with a methylated, cytoplasmic form of estrogen receptor-alpha (methER-α). This interaction is required for NF-κB transcriptional activity and pro-inflammatory cytokine production, and is dissociated by estrogen. Importantly, we show a strong gender segregation in gametogenic reproductive organs, with MyD88/methER-α interactions found in testicular tissues and in ovarian tissues from menopausal women, but not in ovaries from women age 49 and less - suggesting a role for estrogen in disrupting this complex in situ. DISCUSSION: Collectively, our results indicate that the formation of MyD88/methER-α complexes during inflammatory signaling and their disruption by estrogen may represent a mechanism that contributes to gender bias in inflammatory responses.

MyD88 in DNA repair and cancer cell resistance to genotoxic drugs.

BACKGROUND: MyD88 is an adaptor molecule in Toll-like receptor and interleukin 1 receptor signaling implicated in tumorigenesis through proinflammatory mechanisms. We have recently reported that MyD88 also directly promotes optimal activation of the Ras/Erk pathway. Here we investigate MyD88 implication in the maintenance of the transformation of Ras-dependent tumors. METHODS: RNA interference was used to inhibit MyD88 expression in the colon cancer cell lines HCT116 and LS513. Apoptosis, DNA damage, p53 function, ERCC1 levels, and Ras and inflammatory signaling pathways were analyzed. Using in vitro assays and xenotransplantation in nude mice (five per group), HCT116 tumor growth was assessed following MyD88 knockdown in presence or absence of chemotherapy. RESULTS: MyD88 exerts antiapoptotic functions in colon cancer cells via the Ras/Erk, but not the NF-κB, pathway. MyD88 inhibition leads to defective ERCC1-dependent DNA repair and to accumulation of DNA damage, resulting in cancer cell death via p53. Furthermore, we show that knocking down MyD88 sensitizes cancer cells to genotoxic agents such as platinum salts in vitro and in vivo. Indeed, HCT116 tumor growth following treatment with a combination of suboptimal MyD88 inhibition and suboptimal doses of cisplatin (fold tumor increase = 5.4 ± 1.6) was statistically significantly reduced in comparison to treatment with doxycycline alone (12.4 ± 3.1) or with cisplatin alone (12.5 ± 2.6) (P = .005 for both, one-sided Student t test). CONCLUSIONS: Collectively, these results indicate a novel and original link between inflammation, DNA repair, and cancer, and provide further rationale for MyD88 as a potential therapeutic target in Ras-dependent cancers, in the context of concomitant genotoxic chemotherapy.

Small molecule inhibitors of ERCC1-XPF protein-protein interaction synergize alkylating agents in cancer cells.

The benefit of cancer chemotherapy based on alkylating agents is limited because of the action of DNA repair enzymes, which mitigate the damage induced by these agents. The interaction between the proteins ERCC1 and XPF involves two major components of the nucleotide excision repair pathway. Here, novel inhibitors of this interaction were identified by virtual screening based on available structures with use of the National Cancer Institute diversity set and a panel of DrugBank small molecules. Subsequently, experimental validation of the in silico screening was undertaken. Top hits were evaluated on A549 and HCT116 cancer cells. In particular, the compound labeled NSC 130813 [4-[(6-chloro-2-methoxy-9-acridinyl)amino]-2-[(4-methyl-1-piperazinyl)methyl]] was shown to act synergistically with cisplatin and mitomycin C; to increase UVC-mediated cytotoxicity; to modify DNA repair as indicated by the staining of phosphorylated H2AX; and to disrupt interaction between ERCC1 and XPF in cells. In addition, using the Biacore technique, we showed that this compound interacts with the domain of XPF responsible for interaction with ERCC1. This study shows that small molecules targeting the protein-protein interaction of ERCC1 and XPF can be developed to enhance the effects of alkylating agents on cancer cells.