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Fstl1/DIP2A/MGMT signaling pathway plays important roles in temozolomide resistance in glioblastoma.

ABSTRACT: Temozolomide was recognized as the first-line therapy for glioblastoma to prolong the survival of patients noticeably, while recent clinical studies found that some patients were not sensitive to temozolomide treatment. The possible mechanisms seemed to be methylguanine-DNA-methyltransferase (MGMT), mismatch repair, PARP, etc. And the abnormal expression of MGMT might be the most direct factor. In this study, we provide evidence that Fstl1 plays a vital role in temozolomide resistance by sequentially regulating DIP2A protein distribution, H3K9 acetylation (H3K9Ac), and MGMT transcription. As a multifunctional protein widely distributed in cells, DIP2A cooperates with the HDAC2-DMAP1 complex to enhance H3K9Ac deacetylation, prevent MGMT transcription, and increase temozolomide sensitivity. Fstl1, a glycoprotein highly expressed in glioblastoma, competitively binds DIP2A to block DIP2A nuclear translocation, so as to hinder DIP2A from binding the HDAC2-DMAP1 complex. The overexpression of Fstl1 promoted the expression of MGMT in association with increased promoter H3K9Ac. Upregulation of Fstl1 enhanced temozolomide resistance, whereas Fstl1 silencing obviously sensitized GBM cells to temozolomide both in vivo and in vitro. Moreover, DIP2A depletion abolished the effects of Fstl1 on MGMT expression and temozolomide resistance. These findings highlight an important role of Fstl1 in the regulation of temozolomide resistance by modulation of DIP2A/MGMT signaling.

SUBMITTER: Nie E

PROVIDER: S-EPMC6484760 | biostudies-literature | 2019 Apr

REPOSITORIES: biostudies-literature

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Project description:BackgroundOur previous studies have indicated that miR-198 reduces cellular methylguanine DNA methyltransferase (MGMT) levels to enhance temozolomide sensitivity. Transforming growth factor beta 1 (TGF-β1) switches off miR-198 expression by repressing K-homology splicing regulatory protein (KSRP) expression in epidermal keratinocytes. However, the underlying role of TGF-β1 in temozolomide resistance has remained unknown.MethodsThe distribution of KSRP was detected by western blotting and immunofluorescence. Microarray analysis was used to compare the levels of long noncoding RNAs (lncRNAs) between TGF-β1-treated and untreated cells. RNA immunoprecipitation was performed to verify the relationship between RNAs and KSRP. Flow cytometry and orthotopic and subcutaneous xenograft tumor models were used to determine the function of TGF-β1 in temozolomide resistance.ResultsOverexpression of TGF-β1 contributed to temozolomide resistance in MGMT promoter hypomethylated glioblastoma cells in vitro and in vivo. TGF-β1 treatment reduced cellular MGMT levels through suppressing the expression of miR-198. However, TGF-β1 upregulation did not affect KSRP expression in glioma cells. We identified and characterized 2 lncRNAs (H19 and HOXD-AS2) that were upregulated by TGF-β1 through Smad signaling. H19 and HOXD-AS2 exhibited competitive binding to KSRP and prevented KSRP from binding to primary miR-198, thus decreasing miR-198 expression. HOXD-AS2 or H19 upregulation strongly promoted temozolomide resistance and MGMT expression. Moreover, KSRP depletion abrogated the effects of TGF-β1 and lncRNAs on miR-198 and MGMT. Finally, we found that patients with low levels of TGF-β1 or lncRNA expression benefited from temozolomide therapy.ConclusionsOur results reveal an underlying mechanism by which TGF-β1 confers temozolomide resistance. Furthermore, our findings suggest that a novel combination of temozolomide with a TGF-β inhibitor may serve as an effective therapy for glioblastomas.

Project description:BackgroundThe use of temozolomide (TMZ) has improved the prognosis for glioblastoma multiforme patients. However, TMZ resistance may be one of the main reasons why treatment fails. Although this resistance has frequently been linked to the expression of O6-methylguanine-DNA methyltransferase (MGMT) it seems that this enzyme is not the only molecular mechanism that may account for the appearance of drug resistance in glioblastoma multiforme patients as the mismatch repair (MMR) complex, P-glycoprotein, and/or the presence of cancer stem cells may also be implicated.MethodsFour nervous system tumor cell lines were used to analyze the modulation of MGMT expression and MGMT promoter methylation by TMZ treatment. Furthermore, 5-aza-2'-deoxycytidine was used to demethylate the MGMT promoter and O(6)-benzylguanine to block GMT activity. In addition, MMR complex and P-glycoprotein expression were studied before and after TMZ exposure and correlated with MGMT expression. Finally, the effect of TMZ exposure on CD133 expression was analyzed.ResultsOur results showed two clearly differentiated groups of tumor cells characterized by low (A172 and LN229) and high (SF268 and SK-N-SH) basal MGMT expression. Interestingly, cell lines with no MGMT expression and low TMZ IC50 showed a high MMR complex expression, whereas cell lines with high MGMT expression and high TMZ IC50 did not express the MMR complex. In addition, modulation of MGMT expression in A172 and LN229 cell lines was accompanied by a significant increase in the TMZ IC50, whereas no differences were observed in SF268 and SK-N-SH cell lines. In contrast, P-glycoprotein and CD133 was found to be unrelated to TMZ resistance in these cell lines.ConclusionsThese results may be relevant in understanding the phenomenon of TMZ resistance, especially in glioblastoma multiforme patients laking MGMT expression, and may also aid in the design of new therapeutic strategies to improve the efficacy of TMZ in glioblastoma multiforme patients.

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Fstl1/DIP2A/MGMT signaling pathway plays important roles in temozolomide resistance in glioblastoma.

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