Abstract
Despite decades of progress, breast cancer remains the most commonly diagnosed malignancy in women worldwide. Of all its subtypes, triple-negative breast cancer (TNBC) is the most lethal, defined by aggressive proliferation, high metastatic potential, resistance to standard therapies, and an enduring absence of effective targeted treatments. Against this backdrop, ion channels have emerged as potential therapeutic targets for TNBC. The Epithelial Sodium Channel (ENaC), classically associated with sodium homeostasis, blood pressure and extracellular fluid volume regulation, has been linked to tumour suppression, via the downregulation of proliferation and migration markers in malignant breast cancer cells. RNA sequencing data revealed that overexpression of the α subunit of ENaC in breast cancer cells significantly alters gene expression of several pro-oncogenic genes, with the nerve growth factor receptor (NGFR) emerging as one of the most downregulated transcripts. Given NGFR’s documented roles in sustaining stem-like tumour states, driving proliferation and metastatic spread, and conferring therapy resistance in TNBC, these findings identify α-ENaC as a potential upstream brake on a key oncogenic axis that drives TNBC progression.
This project aimed to elucidate the regulatory role of α-ENaC in modulating NGFR expression in MDA-MB-231 TNBC cells. We hypothesized that increased α-ENaC would downregulate NGFR at both the transcript and protein levels, thereby suppressing the migratory capacity of TNBC cells. Consistent with this hypothesis, RT-qPCR revealed that α-ENaC upregulation significantly reduced NGFR mRNA expression (p<0.0001). In agreement with this transcriptional change, western blotting confirmed a corresponding decrease in NGFR protein levels (p<0.0121). Acute pharmacological inhibition of α-ENaC with amiloride produced no significant change in NGFR mRNA and protein levels, suggesting that the mechanism underlying NGFR downregulation is unlikely to be dependent on sodium ion conduction. Functionally, scratch wound assay revealed that α-ENaC overexpressing cells exhibited the lowest migratory capacity, an effect reversed by NGF treatment, which restored migration levels comparable with parental MDA-MB-231 cells. This study offers novel evidence of how α-ENaC influences the molecular landscape underlying metastatic behaviour in TNBC, highlighting its role in modulating genes and proteins associated with disease progression. By establishing α-ENaC as a regulator of NGFR, this study identifies a previously unrecognised pathway that may be exploited for therapeutic targeting in breast cancer. This mechanistic link expands our understanding of TNBC biology and suggests that modulating ENaC activity could provide a novel therapeutic strategy, particularly for treatment-resistant cases.