Cancer is a disease with marked heterogeneity in both response to therapy and survival. Clinical and histopathological characteristics have long determined prognosis and therapy. The introduction of molecular diagnostics has heralded an explosion in new prognostic factors. Overall, histopathology, immunohistochemistry and molecular biology techniques have described important new prognostic subgroups in the different cancer categories. Ion channels and transporters (ICT) are a new class of membrane proteins which are aberrantly expressed in several types of human cancers. Besides regulating different aspect of cancer cell behavior, ICT can now represent novel cancer biomarkers. A summary of the data obtained so far and relative to breast, prostate, lung, colorectal, esophagus, pancreatic and gastric cancers are reported. Special emphasis is given to those studies aimed at relating specific ICT or a peculiar ICT profile with current diagnostic methods. Overall, we are close to exploit ICTs for diagnostic, prognostic or predictive purposes in cancer. This article is part of a Special Issue entitled: Membrane channels and transporters in cancers.

Background:hERG1 channels are aberrantly expressed in human cancers. The expression, functional role and clinical significance of hERG1 channels in pancreatic ductal adenocarcinoma (PDAC) is lacking.Methods:hERG1 expression was tested in PDAC primary samples assembled as tissue microarray by immunohistochemistry using an anti-hERG1 monoclonal antibody (α-hERG1-MoAb). The functional role of hERG1 was studied in PDAC cell lines and primary cultures. ERG1 expression during PDAC progression was studied in Pdx-1-Cre,LSL-KrasG12D/+,LSL-Trp53R175H/+ transgenic (KPC) mice. ERG1 expression in vivo was determined by optical imaging using Alexa-680-labelled α-hERG1-MoAb.Results:(i) hERG1 was expressed at high levels in 59% of primary PDAC; (ii) hERG1 blockade decreased PDAC cell growth and migration; (iii) hERG1 was physically and functionally linked to the Epidermal Growth Factor-Receptor pathway; (iv) in transgenic mice, ERG1 was expressed in PanIN lesions, reaching high expression levels in PDAC; (v) PDAC patients whose primary tumour showed high hERG1 expression had a worse prognosis; (vi) the α-hERG1-MoAb could detect PDAC in vivo.Conclusions:hERG1 regulates PDAC malignancy and its expression, once validated in a larger cohort also comprising of late-stage, non-surgically resected cases, may be exploited for diagnostic and prognostic purposes in PDAC either ex vivo or in vivo.

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We have studied how the macrolide antibiotic Clarithromycin (Cla) regulates autophagy, which sustains cell survival and resistance to chemotherapy in cancer. We found Cla to inhibit the growth of human colorectal cancer (CRC) cells, by modulating the autophagic flux and triggering apoptosis. The accumulation of cytosolic autophagosomes accompanied by the modulation of autophagic markers LC3-II and p62/SQSTM1, points to autophagy exhaustion. Because Cla is known to bind human Ether-à-go-go Related Gene 1 (hERG1) K + channels, we studied if its effects depended on hERG1 and its conformational states. By availing of hERG1 mutants with different gating properties, we found that fluorescently labelled Cla preferentially bound to the closed channels. Furthermore, by sequestering the channel in the closed conformation, Cla inhibited the formation of a macromolecular complex between hERG1 and the p85 subunit of PI3K. This strongly reduced Akt phosphorylation, and stimulated the p53-dependent cell apoptosis, as witnessed by late caspase activation. Finally, Cla enhanced the cytotoxic effect of 5-fluorouracil (5-FU), the main chemotherapeutic agent in CRC, in vitro and in a xenograft CRC model. We conclude that Cla affects the autophagic flux by impairing the signaling pathway linking hERG1 and PI3K. Combining Cla with 5-FU might be a novel therapeutic option in CRC.

Monoclonal antibodies (mAbs), either mono-or bispecific (bsAb), represent one of the most successful approaches to treat many types of malignancies. However, there are certain limitations to the use of full length mAbs for clinical applications, which can be overcome by engineered antibody fragments. The aim of the present study was to develop a small bsAb, in the format of a single-chain diabody (scDb), to efficiently target two proteins, the hERG1 potassium channel and the 1 subunit of integrin receptors, which specifically form a macromolecular complex in cancer cells.We provide evidence that the scDb we produced binds to the hERG1/1 complex in cancer cells and tissues, whereas does not bind to the hERG1 channel in non-pathological tissues, in particular the heart. The scDb-hERG1-1 (1) downregulates the formation of the hERG1/1 complex, (2) inhibits Akt phosphorylation and HIF-1 expression and (3) decreases cell survival, proliferation and migration in vitro. These effects only occur in cancer cells (either colon, pancreatic or breast), but not in normal cells. In vivo, the scDb-hERG1-1 shows a good pharmacokinetic profile, with a half-life of 13.5 hours and no general, cardiac or renal toxicity when injected intravenously up to the dose of 8 mg/Kg. The scDb-hERG1-1 accumulates into subcutaneous xenografted tumors, arising from either colon or pancreatic human cancer cells, and induces a reduction of tumor growth and vascularization.Overall, the scDb-hERG1-1 represents an innovative single-chain bispecific antibody for therapeutic applications in solid cancers which over express the hERG1/1 integrin signaling complex.

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Increasing evidence indicates that ion channels and transporters cooperate in regulating different aspects of tumor pathophysiology. In cancer cells, H + /HCO 3 transporters usually invert the transmembrane pH gradient typically observed in non-neoplastic cells, which is thought to contribute to cancer malignancy. To what extent the pH-regulating transporters are functionally linked to K + channels, which are central regulators of cell membrane potential (V m ), is unclear. We thus investigated in colorectal cancer cells the implication of the pH-regulating transporters and K V 11.1 (also known as hERG1) in the pH modifications stimulated by integrin-dependent cell adhesion. Colorectal cancer cell lines (HCT 116 and HT 29) were seeded onto b1 integrin-dependent substrates, collagen I and fibronectin. This led to a transient cytoplasmic alkalinization, which peaked at 90 min of incubation, lasted approximately 180 min, and was inhibited by antibodies blocking the b1 integrin. The effect was sensitive to amiloride (10 µM) and cariporide (5 µM), suggesting that it was mainly caused by the activity of the Na + /H + antiporter NHE1. Blocking K V 11.1 with E4031 shows that channel activity contributed to modulate the b1 integrindependent pH i increase. Interestingly, both NHE1 and K V 11.1 modulated the colorectal cancer cell motility triggered by b1 integrin-dependent adhesion. Finally, the b1 integrin subunit, K V 11.1 and NHE1 co-immunoprecipitated in colorectal cancer cells seeded onto Collagen I, suggesting the formation of a macromolecular complex following integrinmediated adhesion. We conclude that the interaction between K V 11.1, NHE1, and b1 integrin contributes to regulate colorectal cancer intracellular pH in relation to the tumor microenvironment, suggesting novel pharmacological targets to counteract pro-invasive and, hence, pro-metastatic behavior in colorectal cancer.

Ion channels are progressively emerging as a novel class of membrane proteins expressed in several types of human cancers and regulating the different aspects of cancer cell behavior. The metabolism of cancer cells, usually composed by a variable proportion of respiration, glycolysis, and glutaminolysis, leads to the excessive production of acidic metabolic products. The presence of these acidic metabolites inside the cells results in intracellular acidosis, and hinders survival and proliferation. For this reason, tumor cells activate mechanisms of pH control that produce a constitutive increase in intracellular pH (pH i) that is more acidic than the extracellular pH (pH e). This condition forms a perfect microenvironment for metastatic progression and may be permissive for some of the acquired characteristics of tumors. Recent analyses have revealed complex interconnections between oncogenic activation, ion channels, hypoxia signaling and metabolic pathways that are dysregulated in cancer. Here, we summarize the molecular mechanisms of the Warburg effect and hypoxia and their association. Moreover, we discuss the recent findings concerning the involvement of ion channels in various aspects of the Warburg effect and hypoxia, focusing on the role of Na + and K + channels in hypoxic and metabolic reprogramming in cancer.