P2Y2 receptors mediate nucleotide-induced EGFR phosphorylation and stimulate proliferation and tumorigenesis of head and neck squamous cell carcinoma cell lines.

Abstract

OBJECTIVES:

To assess functional expression of the P2Y2 nucleotide receptor (P2Y2R) in head and neck squamous cell carcinoma (HNSCC) cell lines and define its role in nucleotide-induced epidermal growth factor receptor (EGFR) transactivation. The use of anti-EGFR therapeutics to treat HNSCC is hindered by intrinsic and acquired drug resistance. Defining novel pathways that modulate EGFR signaling could identify additional targets to treat HNSCC.

MATERIALS AND METHODS:

In human HNSCC cell lines CAL27 and FaDu and the mouse oral cancer cell line MOC2, P2Y2R contributions to extracellular nucleotide-induced changes in intracellular free Ca2+ concentration and EGFR and extracellular signal-regulated kinase (ERK1/2) phosphorylation were determined using the ratiometric Ca2+ indicator fura-2 and immunoblot analysis, respectively. Genetic knockout of P2Y2Rs using CRISPR technology or pharmacological inhibition with P2Y2R-selective antagonist AR-C118925 defined P2Y2R contributions to in vivo tumor growth.

RESULTS:

P2Y2R agonists UTP and ATP increased intracellular Ca2+ levels and ERK1/2 and EGFR phosphorylation in CAL27 and FaDu cells, responses that were inhibited by AR-C118925 or P2Y2R knockout. P2Y2R-mediated EGFR phosphorylation was also attenuated by inhibition of the adamalysin family of metalloproteases or Src family kinases. P2Y2R knockout reduced UTP-induced CAL27 cell proliferation in vitro and significantly reduced CAL27 and FaDu tumor xenograft volume in vivo. In a syngeneic mouse model of oral cancer, AR-C118925 administration reduced MOC2 tumor volume.

CONCLUSION:

P2Y2Rs mediate HNSCC cell responses to extracellular nucleotides and genetic or pharmacological blockade of P2Y2R signaling attenuates tumor cell proliferation and tumorigenesis, suggesting that the P2Y2R represents a novel therapeutic target in HNSCC.

• Authors: Lucas T Woods, Kimberly J Jasmer, Kevin Muñoz Forti, Vinit C Shanbhag, Jean M Camden, Laurie Erb, Michael J Petris, Gary A Weisman
• Journal: Oral oncology (Oral Oncol) Vol. 109 Pg. 104808 (Jun 12 2020) ISSN: 1879-0593 [Electronic] England
• PMID: 32540611 (Publication Type: Journal Article)
• Copyright: Copyright © 2020 Elsevier Ltd. All rights reserved.