ANO1 (TMEM16A) in pancreatic ductal adenocarcinoma (PDAC)

Abstract

Pancreatic ductal adenocarcinoma (PDAC) has one of the worst survival rates of all cancers. ANO1 (TMEM16A) is a recently identified Ca2+-activated Cl− channel (CaCC) that is upregulated in several tumors. Although ANO1 was subject to extensive studies in the recent years, its pathophysiological function has only been poorly understood. The aim of the present study is to establish the significance of ANO1 in PDAC behavior and demarcate its roles in PDAC from those of the volume-regulated anion channel (VRAC). We performed qPCR and Western blot measurements on different PDAC cell lines (Panc-1, Mia PaCa 2, Capan-1, AsPC-1, BxPC-3) and compared the results to those obtained in a human pancreatic ductal epithelium (HPDE) cell line. All cancer cell lines showed an upregulation of ANO1 on mRNA and protein levels. Whole-cell patch-clamp recordings identified large Ca2+ and voltage-dependent Cl− currents in PDAC cells. Using siRNA knockdown of ANO1 and three ANO1 inhibitors (T16Ainh-A01, CaCCinh-A01, and NS3728), we found that ANO1 is the main constituent of CaCC current in PDAC cells. We further characterized these three inhibitors and found that they had unspecific effects on the free intracellular calcium concentration. Functional studies on PDAC behavior showed that surprisingly inhibition of ANO1 did not influence cellular proliferation. On the other hand, we found ANO1 channel to be pivotal in PDAC cell migration as assessed in wound healing experiments.Electronic supplementary materialThe online version of this article (doi:10.1007/s00424-014-1598-8) contains supplementary material, which is available to authorized users.

CFTR Involvement in Cell Migration and Epithelial Restitution

“…Various pancreatic ductal adenocarcinoma cells have also been shown to have increased expression of ANO1 and enhanced CaCC activity. Knockdown of ANO1 or inhibition by CaCC blockers including CaCCinh-A01, and NS3728 delay migration in BxPC-2 cells, however, T16Ainh-A01 exhibited no effect [77]. The authors speculated that activation of ANO1 was important for cell volume changes necessary to control cell shape and that the channel may serve as a potential target for reducing the metastatic potential of pancreatic tumor cells.…”

Role of volume-regulated and calcium-activated anion channels in cell volume homeostasis, cancer and drug resistance

“…13) 120 Concomitantly, the time-dependent component of ANO1 activation disappears. 1,3,5,119 ANO1 -Pharmacological profile Verkmanns group identified 2 ANO1 inhibitors (T16 inh -A01; CaCC inh -A01), 50,51 which have no effect on VRAC (Fig.…”

“…3) although some side effects of CaCC inh -A01 on Ca 2C signaling was reported. 120,121 In addition the VRAC inhibitor NS3728 was found to be an inhibitor of ANO1 (IC 50 Figure 15. ANO6 is involved in Cisplatin induced cells death.…”

“…Figure adapted from. 120 currents. 52,53 Native CaCC in rabbit pulmonary artery myocytes exhibit 2 conductance states with 1.8 and 3.5 pS when examined in a single-channel patch-clamp experiment.…”

“…118,[135][136][137][138] ANO1 is upregulated in several cancer tissues including head and neck squamous cell carcinoma, prostate, breast, pancreatic cancer and gastrointestinal stromal tumor cells. 120,[139][140][141][142] Figure 14A and 14B show overexpression of ANO1 at the mRNA level and the protein level, respectively, in PDAC cells compared to normal human pancreatic ductal epithelium cells (HPDE). Finally, the functional expression of ANO1 was assessed using the whole-cell patch-clamp technique (steady-state I-V relationship) and cells transfected with scrambled siRNA or siRNA targeting ANO1.…”