**3.5. hEAG1 channels as pharmacological targets in breast cancer**

Among the active compounds that act on TRP channels, polyunsaturated fatty acids (PUFA) have been evidenced to act on TRPC3, to reduce MCF-7 breast cancer cell proliferation and migration and to inhibit the TRPC-mediated calcium entry [80]. These compounds are particularly interesting as they are common in the diet, and previous studies have already shown their positive effects in cardiovascular diseases, including atherosclerosis, arrhythmias, etc. [121, 122]. The increased tumor vascularization is among the typical alterations and some PUFA have been demonstrated to exert effects against angiogenesis, inflammation, and cancer [123]. In this context, the action of PUFA on TRP channels is explaining their effects against breast tumors, either by diminishing angiogenesis in tumors or by reducing the proliferation/migration of tumor cells. The Rho-associated kinases, ROCK1 and ROCK2, are considered as important therapeutic targets in breast cancer, being already demonstrated their critical role in cancer cell migration and invasion [124]. Identifying efficient Rho-kinase inhibitors is of particular interest in several subtypes of cancer, including breast cancer. Some Rho-kinase inhibitors have been identified to inhibit TRP channels. To date, fasudil, a Rho-kinase inhibitor, upregulates the expression of TRPC1 and TRPV2 in breast cancer cell lines (e.g., ZR-75-1, MCF7, and MDA-MB-231) [125]. Additionally, another Rho-kinase inhibitor, Y-27632, was demonstrated to downregulate

TRPV1 seems to be one of the most promising TRP channels as a pharmacological target in breast cancer. To date, capsaicin inhibits cancer growth in SUM149PT breast cancer cells, by triggering apoptotic/necrotic mechanisms, while capsazepine diminishes these effects [88]. MRS1477, a positive allosteric modulator of TRPV1, when co-applied with capsaicin, diminished the fraction of MCF7 breast cancer apoptotic cells but was ineffective against tumor growth in MCF7 tumor-bearing immunodeficient mice [89]. Another study demonstrated that doxorubicin and melatonin exert synergistic effects against apoptosis and mitochondrial oxidative stress in MCF-7 breast cancer cells by activating TRPV1 [126]. The chemotherapy agent, 5-fluorouracil, also exerts its apoptotic effect in MCF7 breast cancer cells via TRPV1 channels [127]. The anticancer effects of cisplatin mediated by TRPV1 have to be potentiated by co-application of selenium or alpha-lipoic acid on MCF-7 breast cancer cells [128, 129]. Interestingly, selenium was shown to diminish the electromagnetic radiation (900 MHz) effects

Although, tamoxifen is commonly known as a selective estrogen receptor modulator and used in estrogen receptor-positive breast cancer cells, it was also shown to be effective in estrogen-receptors negative tumors. Recent data indicate that it exerts antiproliferative effects in MCF7 breast cancer cells by an estrogen receptor-independent pathway that involves

N-(3-aminopropyl)-2-([(3-methylphenyl)methyl]oxy)(20)-N-(2-thienylmethyl)benzamide (AMTB), an inhibitor of TRPM8, was also described to diminish the proliferation and migration of MDA-MB-231 and SK-BR-3 breast cancer cells via a TRPM8-independent mechanism

Blocking Ca2+ influx with EGTA, Ni2+, or SKF96365 in STIM1 siRNA-, or Orai1 siRNA-treated MDA-MB-231 human breast tumor cells decreased the number of invasive tumor cells [95].

TRPM6 and to upregulate TRPC7 in the same breast cancer cells [125].

in MDA-MB-231 breast cancer cell line mediated by TRPV1 activation [130].

TRPV6 channels [87].

178 Calcium and Signal Transduction

involving voltage-gated sodium channels [131].

**3.4. STIM and Orai as pharmacological targets in breast cancer**

Astemizole, an antihistaminic drug, was shown to block hEAG1 channels in MDA-MB-231 breast cancer cells [97]. The authors have demonstrated that blocking hEag1 with astemizole or silencing induces the breast cancer depolarization and consequently reduces the calcium influx and the cell migration without any influence on the cell proliferation [97]. These data are clinically valuable as hEAG1 are overexpressed in invasive ductal carcinoma breast cancer or metastatic lymph nodes [97, 132] and their co-expression with HIF-1α is correlated with tumor size, lymph node status, and tumor stage [132], and the possibility of pharmacologically blocking these channels might represent a promising therapy. Moreover, astemizole may be used to pharmacologically discriminate hEAG from the related hERG potassium channels in MCF-7 breast cancer cells [133].

Insulin-like growth factor-1 (IGF1) is known to ubiquitary stimulate the growth of various cells in the human body, and also to strongly inhibit the programmed cell death [134]. IGF1 was shown to activate hEAG1 channels in breast cancer cells via an Akt-dependent signaling pathway [135]. Corroborating the antiapoptotic activity of IGF1 with its activatory effect exerted on hEAG1, we can conclude that hEAG1 plays an important role in breast cancer proliferation and its blocking is of particular interest in finding an efficient anticancer therapy.
