Association of Augmented Immune-Staining of G-Quadruplex Tertiary DNA Structure in Chemo-Tolerant TNBC with Downregulation of WNT/Epidermal Growth Factor Receptor Pathway receptor Genes: A Pilot Clinicopathological Study

Abstract

Purpose: The aim of the study is to understand the involvement of G-Quadruplex (G-Q) structures in altering the expression profile of WNT/epidermal growth factor receptor (EGFR) pathway receptor genes in chemo-tolerant Triple Negative Breast Cancer (TNBC) samples. Materials and Methods: At first, Gene Expression Omnibus datasets were mined where the expression profile of WNT/EGFR pathway genes in TNBC samples and MDA-MB-231, a TNBC cell line, were checked in response to doxorubicin, a chemotherapeutic drug. Next, to unveil the probable mechanism of regulation, the presence of G-Q structure was checked in in silico study and later validated by immunohistochemical analyses in our pool of sample. These observed results were correlated with patient's demography and survival status. Results: Expression of the receptors (FZD7, LRP6, EGFR) of the WNT/EGFR pathway were found to be differentially expressed in TNBC samples; further emphasized in our samples (n = 61). Notably, these G-Q structures were found in the promoter region of the WNT pathway receptor genes (FZD7, LRP6, and EGFR). Validating in our patient sample pool, a significant increase in G-Q immunostaining was observed in samples, after neoadjuvant chemotherapy (NACT) samples (n = 17) than the pretherapeutic samples (n = 44). Similar pattern of G-Q immunostaining was noticed in doxorubicin-treated MDA-MB-231 cell line. Intriguingly, low staining of G-Q among the pretherapeutic samples, but NACT TNBC samples, was found to be significantly correlated with lymph node metastasis. Conclusions: This study showed that the augmented immunostaining of G-Q structure might have an important involvement in regulating the expression pattern of the WNT/EGFR pathway genes in response to doxorubicin treatment of TNBC.