Effect of siRNA-mediated LGR5 Inhibition on the Expression of Stemness-related Genes in Triple-negative Breast Cancer Spheroid Model

Abstract

Background:Approximately 10-15 of all breast cancer diagnoses are identified as triple-negative breast cancer (TNBC), a subtype known for its poor prognosis due to chemoresistance. TNBC lacks any receptors or proteins that are overexpressed, thus lacking targeted therapy. The protein LGR5, a marker for cancer stem cells and a promoter of Wnt signaling pathway activity, is notably upregulated in TNBC cases. LGR5 plays an important role in chemoresistance and tumorigenesis. Considering this, our research aim was to investigate the effect of LGR5 suppression by using specific siRNA in multicellular spheroid of the TNBC cell line and evaluation of stemness-related gene expression changes.Materials and Methods:Multicellular spheroids of the MDA-MB-231 TNBC cell line were prepared. The exosome was extracted from the human adipose mesenchymal stem cells (ADMSCs) and confirmed. Multicellular spheroids were separately transfected with siRNA, EXO-siRNA and, treated with cisplatin alone. Gene expression was studied using qRT-PCR.Results:Our findings revealed that inhibiting LGR5 with siRNA significantly reduces LGR5 expression. Furthermore, the results indicated a notable increase in the expression of stemness markers (SOX9, OCT4, and NANOG) in samples treated with cisplatin. However, the reduction of LGR5 expression via siRNA led to a marked decrease in SOX9 and OCT4 levels, while NANOG expression remained largely unchanged.Conclusions:In summary, the results indicated that LGR5 suppression is effective in reducing stemness-related genes and may be considered a good candidate for combination therapy along with chemotherapy.