Novel analgesics based on antagonism of TRPV1-AKAP79 binding

• Applicant: Dr. Christina Hanack
• Subject Area: Biochemistry ; Molecular Biology and Physiology of Neurons and Glial Cells
• Term: from 2015 to 2017
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 285735074

Chronic pain is triggered by injury or inflammation of peripheral sensory neurons of the pain pathway. The heat-sensitive ion channel TRPV1 is a major contributor to chronic pain and has been a target for the development of novel analgesics in many pharmaceutical companies. However, direct block of the channel causes hyperthermia and insensitivity to heat stimuli, both major side effects which have largely halted work on TRPV1 as an analgesic target. Work in the chosen research lab has found that inflammation activates kinases which sensitize TRPV1, i.e. lower its heat threshold, and that these kinases are positioned adjacent to TRPV1 by a scaffolding protein, AKAP79, which binds directly to TRPV1. The binding sites on both proteins have been identified, and short competitor peptides with a sequence based on the binding sites block the binding, preventing phosphorylation and in this way completely prevent sensitization of TRPV1. When coupled to a TAT domain peptide, which confers cell permeability, these peptides are comparable in their analgesic efficacy to direct blockers of TRPV1 in animal pain models of inflammatory and neuropathic pain in vivo, but without the major side effects of hyperthermia and thermal insensitivity. Excellent analgesic efficacy is observed in animal models of both inflammatory pain and of an intractable human pain condition, diabetic neuropathy. The aim of this project is to investigate the interaction between TRPV1 and AKAP79 in more detail and to elucidate whether antagonising TRPV1-AKAP79 binding may prove to be a novel strategy for inhibiting chronic pain in vivo. We have expressed and purified AKAP79 and the C-terminal fragment of TRPV1 and will use these purified proteins for NMR structural analysis in order to gain a 3-D-structure of the interaction site. Biophysical and biochemical assays will enable us to determine the affinity and kinetics of the interaction between AKAP79 and TRPV1. The knowledge gained by our NMR work will be used to optimize and construct modified peptide inhibitors effective with in vitro binding assays and in vivo animal models of inflammatory and neuropathic pain. Furthermore, we will extend the animal pain models to widen our knowledge of the efficacy of peptide blockers of the AKAP79-TRPV1 interaction in vivo. The proposed research project will extend our knowledge of an important protein-protein interaction site, and we anticipate that it will also provide information leading to the development of a new analgesic peptide which could in future be used to treat intractable human pain conditions such as diabetic neuropathy.

• DFG Programme: Research Fellowships
• International Connection: United Kingdom
• Host: Professor Peter McNaughton, Ph.D.