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Comprehensive engineering of the tarantula venom peptide huwentoxin-IV to inhibit the human voltage-gated sodium channel hNav1.7.
J Biol Chem. 2020 01 31; 295(5):1315-1327.JB

Abstract

Pain is a significant public health burden in the United States, and current treatment approaches rely heavily on opioids, which often have limited efficacy and can lead to addiction. In humans, functional loss of the voltage-gated sodium channel Nav1.7 leads to pain insensitivity without deficits in the central nervous system. Accordingly, discovery of a selective Nav1.7 antagonist should provide an analgesic without abuse liability and an improved side-effect profile. Huwentoxin-IV, a component of tarantula venom, potently blocks sodium channels and is an attractive scaffold for engineering a Nav1.7-selective molecule. To define the functional impact of alterations in huwentoxin-IV sequence, we produced a library of 373 point mutants and tested them for Nav1.7 and Nav1.2 activity. We then combined favorable individual changes to produce combinatorial mutants that showed further improvements in Nav1.7 potency (E1N, E4D, Y33W, Q34S-Nav1.7 pIC50 = 8.1 ± 0.08) and increased selectivity over other Nav isoforms (E1N, R26K, Q34S, G36I, Nav1.7 pIC50 = 7.2 ± 0.1, Nav1.2 pIC50 = 6.1 ± 0.18, Nav1.3 pIC50 = 6.4 ± 1.0), Nav1.4 is inactive at 3 μm, and Nav1.5 is inactive at 10 μm We also substituted noncoded amino acids at select positions in huwentoxin-IV. Based on these results, we identify key determinants of huwentoxin's Nav1.7 inhibition and propose a model for huwentoxin-IV's interaction with Nav1.7. These findings uncover fundamental features of huwentoxin involved in Nav1.7 blockade, provide a foundation for additional optimization of this molecule, and offer a basis for the development of a safe and effective analgesic.

Authors+Show Affiliations

Neuroscience Discovery, Janssen Research and Development, LLC, San Diego, California 92121 rneff@its.jnj.com.Biologics Research, Janssen Research and Development, LLC, San Diego, California 92121.Structural Biology, Janssen Research and Development, LLC, Spring House, Pennsylvania 19477.Discovery Chemistry-Computational Chemistry, Janssen Research and Development, LLC, San Diego, California 92121.Neuroscience Discovery, Janssen Research and Development, LLC, San Diego, California 92121.Neuroscience Discovery, Janssen Research and Development, LLC, San Diego, California 92121.Biologics Research, Janssen Research and Development, LLC, San Diego, California 92121.Translational Medicine and Early Development Statistics, Janssen Research and Development, LLC, San Diego, California 92121.Translational Medicine and Early Development Statistics, Janssen Research and Development, LLC, San Diego, California 92121.Peptides International, Inc., Louisville, Kentucky 40299.Biologics Research, Janssen Research and Development, LLC, San Diego, California 92121.Molecular and Cellular Pharmacology, Janssen Research and Development, LLC, San Diego, California 92121.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

31871053

Citation

Neff, Robert A., et al. "Comprehensive Engineering of the Tarantula Venom Peptide huwentoxin-IV to Inhibit the Human Voltage-gated Sodium Channel HNav1.7." The Journal of Biological Chemistry, vol. 295, no. 5, 2020, pp. 1315-1327.
Neff RA, Flinspach M, Gibbs A, et al. Comprehensive engineering of the tarantula venom peptide huwentoxin-IV to inhibit the human voltage-gated sodium channel hNav1.7. J Biol Chem. 2020;295(5):1315-1327.
Neff, R. A., Flinspach, M., Gibbs, A., Shih, A. Y., Minassian, N. A., Liu, Y., Fellows, R., Libiger, O., Young, S., Pennington, M. W., Hunter, M. J., & Wickenden, A. D. (2020). Comprehensive engineering of the tarantula venom peptide huwentoxin-IV to inhibit the human voltage-gated sodium channel hNav1.7. The Journal of Biological Chemistry, 295(5), 1315-1327. https://doi.org/10.1074/jbc.RA119.011318
Neff RA, et al. Comprehensive Engineering of the Tarantula Venom Peptide huwentoxin-IV to Inhibit the Human Voltage-gated Sodium Channel HNav1.7. J Biol Chem. 2020 01 31;295(5):1315-1327. PubMed PMID: 31871053.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Comprehensive engineering of the tarantula venom peptide huwentoxin-IV to inhibit the human voltage-gated sodium channel hNav1.7. AU - Neff,Robert A, AU - Flinspach,Mack, AU - Gibbs,Alan, AU - Shih,Amy Y, AU - Minassian,Natali A, AU - Liu,Yi, AU - Fellows,Ross, AU - Libiger,Ondrej, AU - Young,Stephanie, AU - Pennington,Michael W, AU - Hunter,Michael J, AU - Wickenden,Alan D, Y1 - 2019/12/23/ PY - 2019/10/01/received PY - 2019/12/20/revised PY - 2019/12/25/pubmed PY - 2020/9/15/medline PY - 2019/12/25/entrez KW - Nav1.2 KW - Nav1.7 KW - analgesic KW - antagonist KW - molecular modeling KW - neurotoxin KW - pain KW - peptide biosynthesis KW - sodium channel KW - spider toxin SP - 1315 EP - 1327 JF - The Journal of biological chemistry JO - J Biol Chem VL - 295 IS - 5 N2 - Pain is a significant public health burden in the United States, and current treatment approaches rely heavily on opioids, which often have limited efficacy and can lead to addiction. In humans, functional loss of the voltage-gated sodium channel Nav1.7 leads to pain insensitivity without deficits in the central nervous system. Accordingly, discovery of a selective Nav1.7 antagonist should provide an analgesic without abuse liability and an improved side-effect profile. Huwentoxin-IV, a component of tarantula venom, potently blocks sodium channels and is an attractive scaffold for engineering a Nav1.7-selective molecule. To define the functional impact of alterations in huwentoxin-IV sequence, we produced a library of 373 point mutants and tested them for Nav1.7 and Nav1.2 activity. We then combined favorable individual changes to produce combinatorial mutants that showed further improvements in Nav1.7 potency (E1N, E4D, Y33W, Q34S-Nav1.7 pIC50 = 8.1 ± 0.08) and increased selectivity over other Nav isoforms (E1N, R26K, Q34S, G36I, Nav1.7 pIC50 = 7.2 ± 0.1, Nav1.2 pIC50 = 6.1 ± 0.18, Nav1.3 pIC50 = 6.4 ± 1.0), Nav1.4 is inactive at 3 μm, and Nav1.5 is inactive at 10 μm We also substituted noncoded amino acids at select positions in huwentoxin-IV. Based on these results, we identify key determinants of huwentoxin's Nav1.7 inhibition and propose a model for huwentoxin-IV's interaction with Nav1.7. These findings uncover fundamental features of huwentoxin involved in Nav1.7 blockade, provide a foundation for additional optimization of this molecule, and offer a basis for the development of a safe and effective analgesic. SN - 1083-351X UR - https://www.unboundmedicine.com/medline/citation/31871053/Comprehensive_engineering_of_the_tarantula_venom_peptide_huwentoxin_IV_to_inhibit_the_human_voltage_gated_sodium_channel_hNav1_7_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0021-9258(17)49888-7 DB - PRIME DP - Unbound Medicine ER -