The VRAC blocker DCPIB directly gates the BK channels and increases intracellular Ca2+ in melanoma and pancreatic duct adenocarcinoma cell lines

Abstract

Background and purpose: The volume regulated anion channel (VRAC) is known to be involved in different aspects of cancer cell behaviour and response to therapies. For this reason, we investigated the effect of DCPIB, a presumably specific blocker of VRAC, in two types of cancer: pancreatic duct adenocarcinoma (PDAC) and melanoma.

Experimental approach: We used patch-clamp electrophysiology, supported by Ca²⁺ imaging, gene expression analysis, docking simulation and mutagenesis. We employed two PDAC lines (Panc-1 and MiaPaCa-2), as well as a primary (IGR39) and a metastatic (IGR37) melanoma line.

Key results: DCPIB markedly increased whole-cell currents in Panc-1, MiaPaca2 and IGR39, but not in IGR37 cells. The currents were mostly mediated by K_Ca 1.1 channels, commonly known as BK channels. We confirmed DCPIB activation of BK channels also in HEK293 cells transfected with α subunits of this channel. Further experiments showed that in IGR39, and to a smaller degree also in Panc-1 cells, DCPIB induced a rapid Ca²⁺ influx. This, in turn, indirectly potentiated BK channels and, in IGR39 cells, additionally activated other Ca²⁺ -dependent channels. However, Ca²⁺ influx was not required for activation of BK channels by DCPIB, as such activation involved the extracellular part of the protein and we have identified a residue crucial for binding.

Conclusion and implications: DCPIB directly targeted BK channels and, also, acutely increased intracellular Ca²⁺ . Our findings extend the list of DCPIB effects that should be taken into consideration for future development of DCPIB-based modulators of ion channels and other membrane proteins.

Keywords: calcium-activated potassium channel; intracellular calcium; melanoma; pancreatic duct adenocarcinoma; potassium channel openers; volume regulated anion channel.