26P - 3MST: A potential workhorse in H2S signaling trimmed by microRNA-548 in breast cancer

Background

Gasotransmitters have been established as key players in cancer pathophysiology including breast cancer (BC). Hydrogen sulfide (H₂S), being the newest member of gasotransmitters family, was found to be highly elevated in BC tissues. Cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (3MST) are the endogenous producers of H₂S in viable cells. High expression of CBS and CSE in BC tissues was noticed. However, 3MST received less attention. Thus, the aim of this work is to unravel the expression pattern and regulation of 3MST in BC patients and cell lines.

Methods

BC female patients (n=20) were recruited. Patients’ clinical features showed that 65% of patients had lymph node metastasis, 15% were in stage 3-4 of BC, 25% had tumor size ≥5 cm, and 45% were in premenopausal status. Total RNA was extracted using Biazol, reverse transcribed and quantified using qRT-PCR. Western blot analysis was used to quantify the protein. In-silico analysis was used to identify microRNAs (miRNAs) targeting 3MST. MDA-MB-231 cells were cultured and transfected using oligonucleotides. H₂S levels were measured by fluorescence probe (AzMc). Functional analysis experiments such as trans-well migration, MTT and colony forming assays were performed in MDA-MB-231 cells following ectopic miR-548 expression.

Results

3MST was found to be significantly up-regulated in BC tissues compared to the non-cancerous tissues. Significant high expression for 3MST was correlated to tumor size ≥5 cm, albeit not to lymph node metastasis nor to menopause. MiR-548 was listed among the top potential regulators of 3MST using 6 different bioinformatics softwares. Ectopic expression of miR-548 in MDA-MB-231 cells resulted in a marked repression of 3MST on both mRNA and protein levels. H₂S levels were significantly repressed by 3MST siRNAs and miR-548 mimics. On the functional level, miR-548 markedly reduced cellular viability, migration and colony forming ability of MDA-MB-231 cells.

Conclusions

This study sheds light onto the significant involvement of 3MST in H₂S signaling and its positive correlation with BC aggressiveness. Moreover, it highlights its potential target-ability using miR-548 as a novel tumor suppressor therapeutic approach in BC.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

Swiss National Science Foundation (SNSF), grant SNF IZSTZ0_198887.

Disclosure

All authors have declared no conflicts of interest.