Extracellular Proton Sensing GPR68 Mediates Acid Signaling in Development and Cancer

Abstract

Although an acidic microenvironment is a hallmark of cancer that promotes cancer progression, the underlying signaling pathways are not well understood. Here, in a chemical genetic screen for compounds that perturb zebrafish neural crest development, we identified ogremorphin, a novel small molecule inhibitor of the extracellular proton-sensing G protein-coupled receptor GPR68 that blocks all known modes of GPR68 activation, including acidity, flow and mechanical stretch. Using ogremorphin and genetic tools, we demonstrate that activation of GPR68 promotes cell migration and invasion in vitro and in vivo, by modulating formation of filipodia and focal adhesions. Next, we show that, even in globally neutral pH conditions, like the zebrafish embryo, there exists dynamic extracellular zones of local acidification, which we term proton flares. At the individual cell level, the transient proton flares are associated with new focal adhesions, and in tumor spheroids, a larger and longer lasting acidic domains are observed, recapitulating acidic tumor milieu in vitro. Using genetic and chemical biological approaches, we demonstrate that GPR68 signaling promotes glioblastoma cell survival and activates two important pathogenic pathways: O6-methylguanine-DNA methyltransferase, which confers resistance to temozolomide, and prosurvival and immune evasion chemokine interleukin 8 (IL8). Thus, GPR68 mediates the pathogenic signaling of the acidic tumor microenvironment, indicating ogremorphin and its analogs represent an attractive, novel therapeutic class.

Keywords: Filopodia, migration, cancer, GBM, GPR68, OGR1, glioblastoma, Zebrafish, chemical biology, drug discovery, acid sensing, proton sensing