Nav1.5-Mediated Sodium Influx Drives Breast Cancer Metastasis Through Glycolytic Acidification

Breast cancer remains the leading cause of cancer-related mortality among women globally, with most fatalities attributed to metastatic disease that arises from limited treatment options and resistance to therapy. Despite advancements in early detection and treatment, around 20-30% of patients diagnosed with primary breast cancer will eventually develop distant metastases. Once metastasis occurs, it significantly diminishes the prospects for a cure. This grim reality underscores the urgent need for improved therapeutic strategies that can effectively prevent or mitigate the spread of breast cancer. One emerging area of interest in cancer research is the role of ion channels, particularly voltage-gated sodium channels (VGSCs), in cancer progression. VGSCs are a family of ion channels traditionally associated with the propagation of action potentials in neurons and muscle cells. However, growing evidence indicates that these channels are aberrantly expressed in various cancer types, including breast, prostate, lung, and colon cancers, where they contribute to the invasive and metastatic behavior of cancer cells. Among the VGSC subtypes, Nav1.5 has garnered significant attention due to its pronounced upregulation in breast cancer cells compared to normal breast tissue. Previous studies have identified a correlation between Nav1.5 expression and the aggressiveness of breast cancer, suggesting that Nav1.5 may play a pivotal role in cancer cell invasion and metastasis. However, these studies were often limited by small sample sizes and lacked comprehensive analysis of the mechanisms by which Nav1.5 facilitates cancer progression. The main challenges in understanding the role of Nav1.5 in breast cancer are multifaceted. Firstly, the clinical relevance of Nav1.5 expression in breast cancer patients has not been fully elucidated, particularly regarding its impact on patient outcomes such as metastasis-free survival and overall survival. Secondly, the functional activity of Nav1.5 in cancer tissues, as opposed to cell lines, remains poorly characterized. Electrophysiological studies that could confirm the presence of functional Nav1.5 channels in breast cancer tissue are sparse, leaving a gap in our understanding of how these channels operate in the tumor microenvironment. Furthermore, the mechanisms by which Nav1.5 promotes cancer cell invasion and metastasis are incompletely understood. It has been proposed that Nav1.5-mediated sodium influx may alter intracellular and extracellular ionic homeostasis, driving processes such as extracellular acidification, which in turn facilitates the activity of proteases that degrade the extracellular matrix (ECM). However, the specific pathways and feedback loops involved in this process need to be clarified to develop targeted interventions.

New study published in Oncogene and led by Professor William Brackenbury from the University of York addressed critical gaps in our understanding of Nav1.5’s role in breast cancer. The researchers analyzed a large cohort of breast cancer patients, the study aims to determine the prognostic significance of Nav1.5 expression, correlating it with clinical outcomes such as metastasis-free survival and overall survival. The study also employed electrophysiological techniques to record sodium currents in fresh breast cancer tissue samples, providing direct evidence of Nav1.5 functional activity in a clinical context. By investigating the relationship between Nav1.5 activity, intracellular sodium dynamics, and extracellular acidification, the study seeks to uncover the mechanisms by which Nav1.5 facilitates cancer cell invasion and metastasis. This includes examining how Nav1.5-induced changes in cellular metabolism and ion homeostasis contribute to the invasive behavior of breast cancer cells.

The researchers began by analyzing a breast cancer tissue microarray (TMA) containing 1740 cases to determine the expression of Nav1.5 and its correlation with clinical outcomes. They found that high Nav1.5 protein expression was significantly associated with larger tumor size, lymph node positivity, higher Nottingham prognostic index, and higher tumor grade. More importantly, patients with high Nav1.5 expression had a marked reduction in metastasis-free survival and overall survival. This large-scale analysis underscored the clinical relevance of Nav1.5 as a negative prognostic indicator in breast cancer. To directly assess the functional activity of Nav1.5 in breast cancer, the team performed electrophysiological recordings on fresh tissue samples from three breast tumors. Voltage-sensitive inward and outward currents were detected in cells within the tumor slices and in dissociated primary cells. In particular, small inward currents, indicative of Nav1.5 activity, were observed in 2 out of 3 patient specimens and in a subset of breast cancer cell lines, including triple-negative breast cancer (TNBC) cells and cancer-associated fibroblasts (CAFs). This confirmed that Nav1.5 channels are functionally active in breast cancer tissues, contributing to the invasive properties of the cancer cells.  The researchers used RNA sequencing to compare gene expression in xenograft tumors with and without SCN5A (Nav1.5) knockdown. The knockdown of Nav1.5 resulted in significant changes in the expression of genes related to cellular migration and invasion. Gene set enrichment analysis revealed a substantial reduction in the transcriptional response associated with invasion in the Nav1.5 knockdown tumors, supporting the role of Nav1.5 in promoting breast cancer cell invasion.

To understand the metabolic implications of Nav1.5 activity, the team measured cellular ATP content and the extracellular acidification rate (ECAR) using the Seahorse XFe96 analyzer. They found that Nav1.5 activity increased ATP demand, likely due to heightened activity of the Na+/K+ ATPase (NKA). This increased demand for ATP was primarily met through glycolysis, resulting in elevated production of H+ ions and extracellular acidification. The addition of the VGSC inhibitor tetrodotoxin (TTX) significantly reduced the ECAR in Nav1.5-expressing cells, confirming that Nav1.5 activity drives glycolytic H+ production and extracellular acidification. The researchers also measured the extracellular pH (pHe) of murine xenograft tumors using pH-sensitive microelectrodes. They discovered that the pHe was lower at the periphery of the tumors, where cellularity and proliferation were higher, compared to the core, where apoptosis was more prevalent. This spatial variation in pHe was linked to the metabolic activity driven by Nav1.5. The lower pHe in the tumor periphery facilitated persistent Na+ influx through Nav1.5, creating a positive feedback loop that promotes tumor invasion. Further investigations into the effect of pHe on Nav1.5 activity revealed that lowering the extracellular pH from 7.2 to 6.2 increased the persistent Na+ current through Nav1.5. This increase in Na+ influx at acidic pHe suggested that the invasive edges of tumors, which tend to be more acidic, experience heightened Nav1.5 activity. This, in turn, would promote further extracellular acidification and invasion, reinforcing the positive feedback loop between Nav1.5 activity and the tumor microenvironment.

To validate their findings in a living organism, the researchers conducted in vivo experiments using a mouse model of breast cancer. They knocked down Nav1.5 expression in xenograft tumors and observed a significant reduction in the expression of invasion-regulating genes. This genetic intervention also led to a decrease in tumor growth and metastasis, further supporting the role of Nav1.5 in breast cancer progression.

The study led by Professor William Brackenbury significantly advances our understanding of the role of voltage-gated sodium channel Nav1.5 in breast cancer progression and metastasis. By elucidating the mechanisms through which Nav1.5 promotes invasion and metastasis, this research provides critical insights into the pathophysiology of breast cancer and identifies a promising therapeutic target. The correlation between high Nav1.5 expression and poor clinical outcomes such as reduced metastasis-free survival and overall survival underscores the potential of Nav1.5 as a prognostic marker. This marker can help identify patients at higher risk of metastasis who may benefit from more aggressive or targeted therapies. The study reveals that Nav1.5-mediated Na+ influx increases cellular ATP demand and glycolysis, leading to extracellular acidification. This acidification, in turn, enhances Nav1.5 activity, creating a positive feedback loop that promotes cancer cell invasion. These mechanistic insights provide a deeper understanding of how ion channel dysregulation contributes to cancer progression. Moreover, the identification of Nav1.5 as a critical player in breast cancer metastasis highlights it as a potential therapeutic target. VGSC inhibitors, such as phenytoin, ranolazine, and lidocaine, which have shown efficacy in preclinical models, could be repurposed for breast cancer treatment. Targeting Nav1.5 could disrupt the feedback loop of Na+ influx and extracellular acidification, thereby inhibiting tumor invasion and metastasis. Furthermore, the study’s findings support the development of personalized treatment strategies. Patients with high Nav1.5 expression could be selected for therapies involving VGSC inhibitors, potentially improving outcomes and reducing the risk of metastasis. This personalized approach aligns with the goals of precision medicine, where treatments are tailored to the molecular profile of individual tumors.

Implementation of Nav1.5 expression testing in clinical settings could help stratify breast cancer patients based on their risk of metastasis. This could inform decisions regarding the intensity of monitoring and the need for adjuvant therapies. In addition, the use of existing VGSC inhibitors, which are already approved for other indications, can expedite the process of clinical translation. Conducting clinical trials to evaluate the efficacy of these drugs in breast cancer patients with high Nav1.5 expression could lead to new therapeutic options in a shorter timeframe. The development of new, more specific Nav1.5 inhibitors could provide additional treatment options for breast cancer patients. These targeted therapies would aim to minimize the side effects associated with non-specific ion channel blockers while maximizing anti-cancer efficacy. It is worth mentioning that understanding the role of extracellular pH in promoting cancer cell invasion opens avenues for novel therapeutic strategies that modulate the tumor microenvironment. Therapies aimed at normalizing tumor pH could potentially disrupt the positive feedback loop between Nav1.5 activity and extracellular acidification, thereby inhibiting metastasis.