The H4 histamine receptor agonist, clobenpropit, suppresses human cholangiocarcinoma progression by disruption of epithelial mesenchymal transition and tumor metastasis

Abstract

Cholangiocarcinoma (CCA) is a biliary cancer arising from damaged bile ducts. Epithelialmesenchymal transition (EMT) occurs as epithelial cells begin to resemble mesenchymal cells leading to increased invasion potential as the extracellular matrix (ECM) degrades. Histamine exerts its effects by way of four receptors (H1-H4 HRs). Clobenpropit, a potent H4HR agonist, inhibits mammary adenocarcinoma growth. We have shown that (1) cholangiocytes and CCA cells express H1-H4 HRs and (2) the H3HR decreases CCA proliferation. We evaluated the effects of clobenpropit on CCA proliferation, invasion, EMT phenotypes, and ECM degradation. In vitro, we used CCA cell lines to study proliferation, signaling pathways, and the morphological invasive potential. Gene and protein expression of the hepatobiliary epithelial markers CK-7, CK-8, and CK-19, the focal contact protein paxillin, and the mesenchymal markers fibronectin, s100A4, and vimentin were evaluated. Cell invasion across an ECM layer was quantitated and matrix metalloproteinase-1, -2, -3, -9, and -11 gene and protein expression was examined. Evaluation of the specific role of H4HR was performed by genetic knockdown of the H3HR and overexpression of H4HR. Proliferation was evaluated by proliferating cellular nuclear antigen immunoblotting. In vivo, xenograft tumors were treated with either vehicle or clobenpropit for 39 days. Tumor volume was recorded every other day. Clobenpropit significantly decreased CCA proliferation by way of a Ca 21 -dependent pathway and altered morphological development and invasion. Loss of H3HR expression or overexpression of H4HR significantly decreased CCA proliferation. In vivo, clobenpropit inhibited xenograft tumor growth compared with controls. Conclusion: Modulation of H4HR by clobenpropit disrupts EMT processes, ECM breakdown, and invasion potential and decreases tumor growth. Interruption of tumorigenesis and invasion by histamine may add to therapeutic advances for CCAs.

Epithelial-mesenchymal transition in cholangiocarcinoma: From clinical evidence to regulatory networks

“…CCA cells treated with the specific H4HR agonist clobenpropit induced a down-regulation of fibronectin, vimentin and S100A4, while expression of epithelial markers CK7, CK8 and CK19 was maintained. Consequently, clobenpropit treatment reduced invasive and metastatic potential of CCA cells [94].…”

“…While H1HR and H2HR stimulate biliary hyperplasia and CCA growth, H3HR and H4HR decrease CCA progression [93]. H4HR is up-regulated in human CCA cells compared to non-malignant tissue [94]. CCA cells treated with the specific H4HR agonist clobenpropit induced a down-regulation of fibronectin, vimentin and S100A4, while expression of epithelial markers CK7, CK8 and CK19 was maintained.…”

Cholangiocarcinoma: current knowledge and future perspectives consensus statement from the European Network for the Study of Cholangiocarcinoma (ENS-CCA)

“…Among neuroendocrine factors that either inhibit proliferation or induce apoptosis secretin, gastrin, γ-aminobutyric acid, endothelin-1 and the endocannabinoid anandamide have been described 103,104,[106][107][108][109][110][111] . Although the activation of histamine H 3 and H 4 receptors (HRH 3 and HRH 4 ) inhibits CCA growth, histamine itself is considered proliferative as it sustains CCA growth by forming an autocrine loop 112,113 . The proliferative effects of histamine are in part mediated through HRH 1 , as shown by in vitro blocking experiments using a HRH 1 antagonist [112][113][114] .…”

“…Although the activation of histamine H 3 and H 4 receptors (HRH 3 and HRH 4 ) inhibits CCA growth, histamine itself is considered proliferative as it sustains CCA growth by forming an autocrine loop 112,113 . The proliferative effects of histamine are in part mediated through HRH 1 , as shown by in vitro blocking experiments using a HRH 1 antagonist [112][113][114] .…”

Inhibition of Mast Cell-Derived Histamine Decreases Human Cholangiocarcinoma Growth and Differentiation via c-Kit/Stem Cell Factor–Dependent Signaling

“…3 CAA is a metastatic cancer, and studies have found its potential to migrate outside of the biliary tract. 4 We have found that histamine via the H4 histamine receptor (HR) promotes epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) breakdown and that globally blocking histamine synthesis decreases tumor growth. 4,5 Because of its clinically silent nature, CCA is typically diagnosed in advanced stages, when chemotherapy is often ineffective and radical surgery may be contraindicated.…”