Identification of tetracycline combinations as EphB1 tyrosine kinase inhibitors for treatment of neuropathic pain

Abstract

Previous studies have demonstrated that the synaptic EphB1 receptor tyrosine kinase is a major mediator of neuropathic pain, suggesting that targeting the activity of this receptor might be a viable therapeutic option. Therefore, we set out to determine if any FDA-approved drugs can act as inhibitors of the EphB1 intracellular catalytic domain. An in silico screen was first used to identify a number of tetracycline antibiotics which demonstrated potential docking to the ATP-binding catalytic domain of EphB1. Kinase assays showed that demeclocycline, chlortetracycline, and minocycline inhibit EphB1 kinase activity at low micromolar concentrations. In addition, we cocrystallized chlortetracycline and EphB1 receptor, which confirmed its binding to the ATP-binding domain. Finally, in vivo administration of the three-tetracycline combination inhibited the phosphorylation of EphB1 in the brain, spinal cord, and dorsal root ganglion (DRG) and effectively blocked neuropathic pain in mice. These results indicate that demeclocycline, chlortetracycline, and minocycline can be repurposed for treatment of neuropathic pain and potentially for other indications that would benefit from inhibition of EphB1 receptor kinase activity.

Hit Identification of a Novel Quinazoline Sulfonamide as a Promising EphB3 Inhibitor: Design, Virtual Combinatorial Library, Synthesis, Biological Evaluation, and Docking Simulation Studies

“…The crystal structures of EphB1 (PDB code: 6UMW, Resolution: 1.98 Å) [41], EphB2 (PDB code: 2HEN, Resolution: 2.60 Å) [42], EphB3 (PDB ID: 5L6P, Resolution: 2.26 Å) [15], EphB4 (PDB ID: 3ZEW, Resolution: 2.50 Å) [43], VEGFR2 (PDB code: 3VHE, Resolution: 1.55 Å) [44], Aurora A (PDB code: 2X6E, Resolution: 3.35 Å) [45], HER2 (PDB code: 3PP0, Resolution: 2.25 Å) [46] and CDK2 (PDB code3PY0, Resolution: 1.75 Å) [47] were downloaded from protein data bank (www.pdb.org, accessed date: 18 April 2021). To ensure the validity of the downloaded crystal structures, PROCHECK server was employed to generate a Ramachandran plot for all four downloaded crystal structures to ensure their validity [30].…”

Identification of Atovaquone and Mebendazole as Repurposed Drugs with Antiviral Activity against SARS-CoV-2 (Version 5)

“…The energy-minimized drugs underwent protonation states to add the missing hydrogens for proper ionization states. MOE docking module used to evaluate the favorable binding conformers based on London dG scoring method to estimate energy profile based on the binding affinity with respect to hydrophobic-hydrophobic interactions, hydrogen bonding, pi-pi interactions, and ionic interactions [57,58]. Each drug gave 10 possible docked poses.…”

Drug Repurposing to Target Neuroinflammation and Sensory Neuron-Dependent Pain

“…A common ceftriaxone dose was used for this study [ 48 ] Demeclocycline Chlortetracycline Antibiotics (tetracyclines) In silico screen, capsaicin-induced pain model, CFA model, formalin model in mice Combination of chlortetracycline 7 mg/kg/day PO and demeclocycline 20 mg/kg/day PO 3 days in advance Chlortetracycline binds and inhibits the EphB1 activation and reduces phosphorylation in the brain, spinal cord, and dorsal root ganglion (DRG) Combination of minocycline, chlortetracycline and demeclocycline reduces mechanical and thermal hypersensitivity. Likely off-target due to high dose Chlortetracycline is used ophthalmic or local (ointment) 1–3% (w/w) Demeclocycline is used at 100 mg orally/d [ 36 ] Fingolimod S1P-modulator Cancer-induced bone pain (CIBP) model in mice 1 mg/kg/day IP or SC (continuous infusion with osmotic minipump) for 7 consecutive days Fingolimod treatment reduces flinching and guarding behavior. Possibly off-target due to high dose 0.5 mg PO once daily [ 55 ] Mouse model of multiple sclerosis-induced neuropathic pain (EAE model) 0.1–1 mg/kg IP daily from days 15–34 and 45–52 post-EAE induction; 0.001–1 mg/kg IP daily from days 15–37 post-EAE induction Reduces mechanical hypersensitivity and cold allodynia, reduces pERK activation in the spinal cord via S1P1R and reduces the number of activated spinal microglia and astrocytes.…”

“…A recent study identified that demeclocycline, chlortetracycline, and minocycline bind and inhibit the tyrosine kinase ephrin type-B receptor 1 (EphB1) in the brain, spinal cord, and dorsal root ganglia (DRG). Treating mice with a combination of these three tetracyclines caused a reduction of mechanical and thermal hypersensitivity in capsaicin-, CFA- and formalin-induced pain [ 36 ]. Mechanistically, minocycline seems to reduce the production of reactive oxygen species in microglia and the release of proinflammatory and prologetic cytokines and chemokines, such as TNFα, IL-1β, and CXCL1 [ 37 ] (Fig.…”

“…Likely off-target due to high dose Chlortetracycline ophthalmic or local (ointment) 1–3% (w/w). Demeclocycline is used at 100 mg orally/d [ 36 ] Dimethyl fumarate Anti-multiple sclerosis drug Oxaliplatin-induced neuropathic pain in rats 100 or 200 mg/kg PO 5 times per week for 4 weeks Reduces axonal degradation mechanical but not cold hypersensitivity in rats caused by oxaliplatin. Likely off-target due to high dose Initial dose: 120 mg PO twice daily for 7 days, then 240 mg PO twice daily [ 104 ] Donepezil Anti-dementia drug Oxaliplatin-induced neuropathic pain in rats 0.3 or 1 mg/kg PO 5 times a week for 4 weeks Reduces neurite shortening and mechanical hypersensitivity in rats caused by oxaliplatin possibly by restoring the activity of superoxide dismutase (SOD) and neuroprotection.…”