RESUMO
Voltage-gated Na+ channels (VGSCs) are expressed widely in human carcinomas and play a significant role in promoting cellular invasiveness and metastasis. However, human tissue-based studies and clinical characterization are lacking. In several carcinomas, including colorectal cancer (CRCa), the predominant VGSC is the neonatal splice variant of Nav1.5 (nNav1.5). The present study was designed to determine the expression patterns and clinical relevance of nNav1.5 protein in human CRCa tissues from patients with available clinicopathological history. The immunohistochemistry was made possible by the use of a polyclonal antibody (NESOpAb) specific for nNav1.5. The analysis showed that, compared with normal mucosa, nNav1.5 expression occurred in CRCa samples (i) at levels that were significantly higher and (ii) with a pattern that was more delineated (i.e., apical/basal or mixed). A surprisingly high level of nNav1.5 protein expression also occurred in adenomas, but this was mainly intracellular and diffuse. nNav1.5 showed a statistically significant association with TNM stage, highest expression being associated with TNM IV and metastatic status. Interestingly, nNav1.5 expression co-occurred with other biomarkers associated with metastasis, including hERG1, KCa3.1, VEGF-A, Glut1, and EGFR. Finally, univariate analysis showed that nNav1.5 expression had an impact on progression-free survival. We conclude (i) that nNav1.5 could represent a novel clinical biomarker ('companion diagnostic') useful to better stratify CRCa patients and (ii) that since nNav1.5 expression is functional, it could form the basis of anti-metastatic therapies including in combination with standard treatments.
RESUMO
Functional expression of voltage-gated Na+ channels (VGSCs) occurs in human carcinomas and promotes invasiveness in vitro and metastasis in vivo. Both neonatal and adult forms of Nav1.5 (nNav1.5 and aNav1.5, respectively) have been reported to be expressed at messenger RNA (mRNA) level in colorectal cancer (CRCa) cells. Here, three CRCa cell lines (HT29, HCT116 and SW620) were studied and found to express nNav1.5 mRNA and protein. In SW620 cells, adopted as a model, effects of gene silencing (by several small interfering RNAs [siRNAs]) selectively targeting nNav1.5 or aNav1.5 were determined on (a) channel activity and (b) invasiveness in vitro. Silencing nNav1.5 made the currents more "adult-like" and suppressed invasion by up to 73%. Importantly, subsequent application of the highly specific, general VGSC blocker, tetrodotoxin (TTX), had no further effect. Conversely, silencing aNav1.5 made the currents more "neonatal-like" but suppressed invasion by only 17% and TTX still induced a significant effect. Hypoxia increased invasiveness and this was also blocked completely by siRNA targeting nNav1.5. The effect of hypoxia was suppressed dose dependently by ranolazine, but its effect was lost in cells pretreated with nNav1.5-siRNA. We conclude that (a) functional nNav1.5 expression is common to human CRCa cells, (b) hypoxia increases the invasiveness of SW620 cells, (c) the VGSC-dependent invasiveness is driven predominantly by nNav1.5 under both normoxic and hypoxic conditions and (d) the hypoxia-induced increase in invasiveness is likely to be mediated by the persistent current component of nNav1.5.
Assuntos
Neoplasias Colorretais/genética , Regulação Neoplásica da Expressão Gênica/genética , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Invasividade Neoplásica/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Neoplasias do Colo/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Hipóxia/tratamento farmacológico , RNA Interferente Pequeno/genética , Tetrodotoxina/farmacologiaRESUMO
Breast cancer (BCa) was induced in vivo in female rats with 7,12-dimethylbenz(a)anthracene (DMBA). Two main questions were addressed. Firstly, would the carcinogenesis be accompanied by oxidative stress as signalled by superoxide dismutase, glutathione peroxidase, malondialdehyde and total nitrate? Secondly, would treating the rats additionally with a blocker of voltage-gated sodium channel (VGSC) activity, shown previously to promote BCa progression, affect the oxidative responses? The DMBA-induced increases in the antioxidant systems were completely blocked by the VGSC inhibitor RS100642, which also significantly prolonged the lifespan. We conclude that VGSC inhibition in vivo can significantly protect against oxidative stress and improve survival from tumour burden.