Speckle-type POZ protein functions as a tumor suppressor in non-small cell lung cancer due to DNA methylation

Background: Tumor suppressor epigenetic silencing plays an important role in non-small cell lung cancer (NSCLC) development and progression. Previously, the expression of speckle-type POZ protein (SPOP) has been found to be significantly inhibited in NSCLC. Our research aimed to investigate the molecular mechanisms, clinical significance and epigenetic alteration of SPOP in NSCLC.

Materials and methods: Bisulfite sequencing PCR and methylation-specific PCR were performed to test gene methylation. Chromatin immunoprecipitation (ChIP) was performed to detect transcription factor C/EBPα combinations and the promoter of the SPOP gene. Furthermore, we evaluated the effects of C/EBPα siRNA on SPOP expression, tumor cell migration and proliferation via MTT and Transwell assays in vitro and tumor growth in vivo. The relationship between the methylation status of the SPOP gene and clinicopathologic characteristics was investigated.

Results: Hypermethylation was found in the CpG island of the SPOP gene promoter in NSCLC tissues, and this methylation was found to be correlated with SPOP expression. SPOP promoter methylation was associated with the pathology grade. The transcriptional activities were significantly inhibited by the hypermethylation of specific CpG sites within the SPOP gene promoter, while 5-aza-2'-deoxycytidine significantly increased SPOP gene expression. C/EBPα also played a key role in SPOP regulation. Five C/EBPα binding sites in the CpG island of the SPOP gene promoter were identified by ChIP. Inhibition of C/EBPα significantly reduced SPOP expression. SPOP mediated the C/EBPα-regulated suppression of invasion, migration and proliferation in vitro and tumor growth in vivo.

Conclusions: SPOP function and expression in NSCLS were regulated by DNA methylation and C/EBPα transcriptional regulation combination effects, indicating that the SPOP promoter methylation status could be utilized as an epigenetic biomarker and that the C/EBPα-SPOP signaling pathway could be a potential therapeutic target in NSCLC.