Identification of a Novel Structural Class of H_V1 Inhibitors by Structure-Based Virtual Screening

The human voltage-gated proton channel, hH_V1, is highly expressed in various cell types including macrophages, B lymphocytes, microglia, sperm cells and also in various cancer cells. Overexpression of H_V1 has been shown to promote tumor formation by highly metastatic cancer cells, and has been associated with neuroinflammatory diseases, immune response disorders and infertility, suggesting a potential use of hH_V1 inhibitors in numerous therapeutic areas. To identify compounds targeting this channel, we performed a structure-based virtual screening on an open structure of the human H_V1 channel. Twenty selected virtual screening hits were tested on Chinese hamster ovary (CHO) cells transiently expressing hH_V1, with compound 13 showing strong block of the proton current with an IC₅₀ value of 8.5 μM. Biological evaluation of twenty-three additional analogs of 13 led to the discovery of six other compounds that blocked the proton current by more than 50% at 50 μM concentration. This allowed for an investigation of structure-activity relationships. The antiproliferative activity of the selected promising hH_V1 inhibitors was investigated in the cell lines MDA-MB-231 and THP-1, where compound 13 inhibited growth with an IC₅₀ value of 9.0 and 8.1 μM, respectively. The identification of a new structural class of H_V1 inhibitors contributes to our understanding of the structural requirements for inhibition of this ion channel and opens up the possibility of investigating the role of H_V1 inhibitors in various pathological conditions and in cancer therapy.