1. Legumain Induces Oral Cancer Pain by Biased Agonism of Protease-Activated Receptor-2.
- Author
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Tu NH, Jensen DD, Anderson BM, Chen E, Jimenez-Vargas NN, Scheff NN, Inoue K, Tran HD, Dolan JC, Meek TA, Hollenberg MD, Liu CZ, Vanner SJ, Janal MN, Bunnett NW, Edgington-Mitchell LE, and Schmidt BL
- Subjects
- Aged, Aged, 80 and over, Animals, Arrestin metabolism, Cancer Pain psychology, Cyclic AMP-Dependent Protein Kinases drug effects, Endocytosis drug effects, Enzyme Activation drug effects, Female, Humans, Male, Mice, Mice, Inbred C57BL, Mice, Knockout, Middle Aged, Protein Kinase C drug effects, Protein Kinase Inhibitors pharmacology, Receptor, PAR-2 genetics, Tumor Microenvironment drug effects, Cancer Pain chemically induced, Carcinoma, Squamous Cell complications, Cysteine Endopeptidases administration & dosage, Mouth Neoplasms complications, Receptor, PAR-2 agonists
- Abstract
Oral squamous cell carcinoma (OSCC) is one of the most painful cancers, which interferes with orofacial function including talking and eating. We report that legumain (Lgmn) cleaves protease-activated receptor-2 (PAR
2 ) in the acidic OSCC microenvironment to cause pain. Lgmn is a cysteine protease of late endosomes and lysosomes that can be secreted; it exhibits maximal activity in acidic environments. The role of Lgmn in PAR2 -dependent cancer pain is unknown. We studied Lgmn activation in human oral cancers and oral cancer mouse models. Lgmn was activated in OSCC patient tumors, compared with matched normal oral tissue. After intraplantar, facial or lingual injection, Lgmn evoked nociception in wild-type (WT) female mice but not in female mice lacking PAR2 in NaV 1.8-positive neurons ( Par2 Nav 1.8 ), nor in female mice treated with a Lgmn inhibitor, LI-1. Inoculation of an OSCC cell line caused mechanical and thermal hyperalgesia that was reversed by LI-1. Par2 Nav 1.8 and Lgmn deletion attenuated mechanical allodynia in female mice with carcinogen-induced OSCC. Lgmn caused PAR2 -dependent hyperexcitability of trigeminal neurons from WT female mice. Par2 deletion, LI-1, and inhibitors of adenylyl cyclase or protein kinase A (PKA) prevented the effects of Lgmn. Under acidified conditions, Lgmn cleaved within the extracellular N terminus of PAR2 at Asn30 ↓Arg31 , proximal to the canonical trypsin activation site. Lgmn activated PAR2 by biased mechanisms in HEK293 cells to induce Ca2+ mobilization, cAMP formation, and PKA/protein kinase D (PKD) activation, but not β-arrestin recruitment or PAR2 endocytosis. Thus, in the acidified OSCC microenvironment, Lgmn activates PAR2 by biased mechanisms that evoke cancer pain. SIGNIFICANCE STATEMENT Oral squamous cell carcinoma (OSCC) is one of the most painful cancers. We report that legumain (Lgmn), which exhibits maximal activity in acidic environments, cleaves protease-activated receptor-2 (PAR2 ) on neurons to produce OSCC pain. Active Lgmn was elevated in OSCC patient tumors, compared with matched normal oral tissue. Lgmn evokes pain-like behavior through PAR2 Exposure of pain-sensing neurons to Lgmn decreased the current required to generate an action potential through PAR2 Inhibitors of adenylyl cyclase and protein kinase A (PKA) prevented the effects of Lgmn. Lgmn activated PAR2 to induce calcium mobilization, cAMP formation, and activation of protein kinase D (PKD) and PKA, but not β-arrestin recruitment or PAR2 endocytosis. Thus, Lgmn is a biased agonist of PAR2 that evokes cancer pain., (Copyright © 2021 the authors.)- Published
- 2021
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