Inhibition of Pannexin 1 Reduces the Tumorigenic Properties of Human Melanoma Cells.

Taylor Freeman,Aaron Grant,Silvia Penuela,Zameena Lakhani,Kevin Barr,Danielle Johnston,Rafael Sanchez-Pupo,Kenneth Huang,Daniel Nouri-Nejad,Brooke O’Donnell,Luke Harland,Samar Sayedyahossein,Steven Latosinsky,Lina Dagnino

doi:10.3390/cancers11010102

Abstract

Pannexin 1 (PANX1) is a channel-forming glycoprotein expressed in many tissues including the skin. PANX1 channels allow the passage of ions and molecules up to 1 kDa, including ATP and other metabolites. In this study, we show that PANX1 is highly expressed in human melanoma tumors at all stages of disease progression, as well as in patient-derived cells and established melanoma cell lines. Reducing PANX1 protein levels using shRNA or inhibiting channel function with the channel blockers, carbenoxolone (CBX) and probenecid (PBN), significantly decreased cell growth and migration, and increased melanin production in A375-P and A375-MA2 cell lines. Further, treatment of A375-MA2 tumors in chicken embryo xenografts with CBX or PBN significantly reduced melanoma tumor weight and invasiveness. Blocking PANX1 channels with PBN reduced ATP release in A375-P cells, suggesting a potential role for PANX1 in purinergic signaling of melanoma cells. In addition, cell-surface biotinylation assays indicate that there is an intracellular pool of PANX1 in melanoma cells. PANX1 likely modulates signaling through the Wnt/β-catenin pathway, because β-catenin levels were significantly decreased upon PANX1 silencing. Collectively, our findings identify a role for PANX1 in controlling growth and tumorigenic properties of melanoma cells contributing to signaling pathways that modulate melanoma progression.

Highlights

Pannexins (Panx1, Panx2, Panx3) are a family of glycoproteins [1,2,3,4] that form large-pore channels at the cell membrane [4,5]
We previously reported that Panx1 expression was significantly up-regulated in B16-BL6 mouse melanoma cells compared to normal melanocytes
Our results suggest that high levels of Pannexin 1 (PANX1) found in human melanomas at different stages of disease progression may be associated with their malignant behavior, making PANX1 a potential new target for therapeutic intervention

Summary

Introduction

Pannexins (Panx, Panx, Panx3) are a family of glycoproteins [1,2,3,4] that form large-pore channels at the cell membrane [4,5]. Pannexin 1 channels (Panx in rodents, or PANX1 in humans) allow the passage of molecules up to 1 kDa in size important for paracrine and autocrine signaling, including adenosine triphosphate (ATP), which they release at the cell surface [1,4,7,8]. PANX1 is involved in multiple disease states, including cancer [11]. Panx expression positively correlates with the onset or progression of the disease [12]. Human leukemic lymphocytes show higher levels of PANX1 in comparison to normal T cells, and high expression of PANX1 has been reported across aggressive multiple myeloma cell lines [13,14]. A study demonstrated high PANX1 mRNA expression in human glioma cell lines. The authors found that rat C6 glioma cells lacked Panx, and upon overexpressing green fluorescent protein (EGFP)

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