TY - JOUR
T1 - Legumain induces oral cancer pain by biased agonism of protease-activated receptor-2
AU - Tu, Nguyen Huu
AU - Jensen, Dane D.
AU - Anderson, Bethany M.
AU - Chen, Elyssa
AU - Jimenez-Vargas, Nestor N.
AU - Scheff, Nicole N.
AU - Inoue, Kenji
AU - Tran, Hung D.
AU - Dolan, John C.
AU - Meek, Tamaryn A.
AU - Hollenberg, Morley D.
AU - Liu, Cheng Z.
AU - Vanner, Stephen J.
AU - Janal, Malvin N.
AU - Bunnett, Nigel W.
AU - Edgington-Mitchell, Laura E.
AU - Schmidt, Brian L.
N1 - Funding Information:
This work was supported by National Institutes of Health Grants NS102722, DE026806, DK118971, and DE029951 (to N.W.B., B.L.S.) and the Department of Defense Grant W81XWH1810431 (to N.W.B., B.L.S.). L.E.E.-M. was supported by the Priority-Driven Collaborative Cancer Research Grant GNT1157171 (co-funded by Cancer Australia and Cure Cancer), a Grimwade Fellowship from the Russell and Mab Grimwade Miegunyah Fund at The University of Melbourne, and the Australian Research Council Discovery Early Career Researcher Award Fellowship DE180100418.
Funding Information:
N.W.B. is a founding scientist of Endosome Therapeutics Inc, and research in N.W.B.’s laboratory is partly supported by Takeda Pharmaceuticals International. J.C.D. fabricates dolognawmeter assay devices through Gnatheon Scientific LLC. All other authors declare no competing financial interests.
Publisher Copyright:
Copyright © 2021 the authors.
PY - 2021/1/6
Y1 - 2021/1/6
N2 - 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 (PAR2) 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 NaV1.8-positive neurons (Par2Nav1.8), 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. Par2Nav1.8 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.
AB - 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 (PAR2) 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 NaV1.8-positive neurons (Par2Nav1.8), 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. Par2Nav1.8 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.
KW - Asparaginyl endopeptidase
KW - Cancer pain
KW - Legumain
KW - Oral cancer
KW - Protease
KW - Protease-activated receptor-2
UR - http://www.scopus.com/inward/record.url?scp=85100279772&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85100279772&partnerID=8YFLogxK
U2 - 10.1523/JNEUROSCI.1211-20.2020
DO - 10.1523/JNEUROSCI.1211-20.2020
M3 - Article
C2 - 33172978
AN - SCOPUS:85100279772
VL - 41
SP - 193
EP - 210
JO - Journal of Neuroscience
JF - Journal of Neuroscience
SN - 0270-6474
IS - 1
ER -