Neuropilin-1 inhibition suppresses nerve growth factor signaling and nociception in pain models
Chloe J. Peach, … , Rajesh Khanna, Nigel W. Bunnett
Chloe J. Peach, … , Rajesh Khanna, Nigel W. Bunnett
Published November 26, 2024
Citation Information: J Clin Invest. 2025;135(4):e183873. https://doi.org/10.1172/JCI183873.
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Research Article Cell biology Neuroscience

Neuropilin-1 inhibition suppresses nerve growth factor signaling and nociception in pain models

Abstract

Nerve growth factor (NGF) monoclonal antibodies inhibit chronic pain, yet failed to gain approval due to worsened joint damage in osteoarthritis patients. We report that neuropilin-1 (NRP1) is a coreceptor for NGF and tropomyosin-related kinase A (TrkA) pain signaling. NRP1 was coexpressed with TrkA in human and mouse nociceptors. NRP1 inhibitors suppressed NGF-stimulated excitation of human and mouse nociceptors and NGF-evoked nociception in mice. NRP1 knockdown inhibited NGF/TrkA signaling, whereas NRP1 overexpression enhanced signaling. NGF bound NRP1 with high affinity and interacted with and chaperoned TrkA from the biosynthetic pathway to the plasma membrane and endosomes, enhancing TrkA signaling. Molecular modeling suggested that the C-terminal R/KXXR/K NGF motif interacts with the extracellular “b” NRP1 domain within a plasma membrane NGF/TrkA/NRP1 of 2:2:2 stoichiometry. G α interacting protein C-terminus 1 (GIPC1), which scaffolds NRP1 and TrkA to myosin VI, colocalized in nociceptors with NRP1/TrkA. GIPC1 knockdown abrogated NGF-evoked excitation of nociceptors and pain-like behavior. Thus, NRP1 is a nociceptor-enriched coreceptor that facilitates NGF/TrkA pain signaling. NRP binds NGF and chaperones TrkA to the plasma membrane and signaling endosomes via the GIPC1 adaptor. NRP1 and GIPC1 antagonism in nociceptors offers a long-awaited nonopioid alternative to systemic antibody NGF sequestration for the treatment of chronic pain.

Authors

Chloe J. Peach, Raquel Tonello, Elisa Damo, Kimberly Gomez, Aida Calderon-Rivera, Renato Bruni, Harsh Bansia, Laura Maile, Ana-Maria Manu, Hyunggu Hahn, Alex R.B. Thomsen, Brian L. Schmidt, Steve Davidson, Amedee des Georges, Rajesh Khanna, Nigel W. Bunnett

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Figure 1

NGF/NRP1/TrkA interactions.

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NGF/NRP1/TrkA interactions. (A) CendR motifs (highlighted) of NGF C-term...
(A) CendR motifs (highlighted) of NGF C-terminus numbered according to mature βNGF (1–120 equivalent to proNGF 122–241). h, Homo sapiens; r, Rattus norvegicus; m, Mus musculus. (B) MST interaction assay between fluorescent human βNGF and NRP1 (residues 22–644) or staphylokinase (SK, negative control). n = 2 or 4 independent experiments. Data are represented as mean ± SEM. (C and D) BRET assay of VEGF or NGF proximity to NRP1 in HEK293T cells. Supernatant from cells secreting HiBiT-tagged VEGF165a or NGF was reconstituted with recombinant LgBiT and furimazine, forming the bioluminescent donor. HEK293T cells expressing WT SnapTag-NRP1 or VEGF165a binding-dead mutant (Y297A) were labeled with SNAPTag-Alexa Fluor 488 (AF488) and incubated with supernatant. BRET was measured between HiBiT-VEGF165a and SnapTag-NRP1 (WT or Y297A) or HiBiT-tagged NGF and SnapTag-NRP1 (WT). Cells were preincubated with vehicle or unlabeled VEGF165a followed by luminescent growth factor. BRET was compared with negative control (HiBiT/LgBiT only lacking AF488). n = 4 independent experiments. Data are represented as mean ± SEM. *P < 0.05; ****P < 0.0001, 1-way ANOVA, Šídák’s multiple comparisons. (E and F) Ternary complex of human NGF/TrkA/NRP1 generated using constraint-driven computational docking. Cartoon and surface representation of TrkA (gray), NGF (blue), and NRP1 (pink) are shown. (E) NGF/TrkA/NRP1 model and conserved interactions at the NGF/NRP1 interface suggest a 2:2:2 stoichiometry with 1 NRP1 molecule interacting with 1 TrkA molecule and the NGF dimer. Views of the NRP1/TrkA complex in surface representation represent complementarity between NRP1 and TrkA. The inset shows binding of NGF C-terminal R118 (blue) to conserved residues (pink, Y297, D320, S346, Y353) in C-terminal arginine-binding pocket of the NRP1 b1 domain (predicted hydrogen bonds in green). (F) Proposed cell surface NGF/TrkA/NRP1 complex. Membrane proximal MAM NRP1 domains are included to propose a sterically feasible membrane-tethered NGF/TrkA/NRP1 complex. Membrane linkers and transmembrane regions are not derived from structures and are not to scale.