Exploring Macrocyclic Peptides for TrkA-NGF Inhibition: A Novel Approach to Analgesics, with Intriguing Screening Tricks of the Trade
In recent years, macrocyclic peptides have gained traction as promising drug candidates due to their unique structural properties. Their large surface area allows them to cover extensive binding interfaces, making them well-suited for inhibiting challenging targets such as protein–protein interactions (PPIs). Additionally, these peptides can be identified using mRNA-display screening techniques. However, their broad interaction surfaces can also pose a challenge, as directing them to specific target regions within a PPI interface can be difficult.
A recent article by Mikamiyama and Yamada (https://pubs.acs.org/doi/10.1021/acs.jmedchem.4c00715) focuses on this problem, investigating the inhibition of the tropomyosin receptor kinase A (TrkA) and nerve growth factor (NGF) interaction. Blocking this interaction is postulated to provide a novel analgesic pathway, potentially avoiding some adverse effects associated with opioid-based treatments.
The authors began by screening a library of 19-mer macrocyclic peptides for their binding affinity to TrkA. In their initial screening phase, they identified a series of peptides with strong binding affinity for TrkA. However, when they synthesized and tested these lead peptides, they observed that although these peptides bound strongly to TrkA, they did not inhibit the TrkA–NGF interaction. Structural analysis through co-crystallization of TrkA with one of these peptides revealed that the peptide was binding to an allosteric site on TrkA, not the intended PPI interface with NGF. This off-target binding suggested that the identified site had an intrinsically favorable surface for peptide binding, redirecting the library selection away from the target site.
To address this, the team implemented an innovative strategy: they pre-incubated TrkA with a high-affinity peptide binder to block the allosteric site, thereby “shielding” it from subsequent rounds of screening. With this pre-incubation step, they could steer the screening library toward binding regions closer to the intended TrkA–NGF interaction surface. Through rounds of optimization, including libraries enriched with unnatural amino acids for improved binding, they successfully identified a peptide that bound to the TrkA–NGF interaction site, exhibiting both high affinity for TrkA and inhibitory activity against NGF binding.
A second, equally fascinating part of the study focused on improving the serum half-life of the selected peptides, as their initial lead compound displayed rapid degradation in serum. To address this, the authors employed an intriguing structure–activity relationship (SAR) approach: they incubated the library with a cocktail of proteases to degrade any unstable peptides before proceeding with the binding step. This screening method enabled the selection of peptides with increased stability in serum while retaining strong binding affinity and inhibitory activity.
As a synthetic chemist, I find this study an exciting example of how tailored screening approaches can guide the identification of high-affinity binders for challenging targets and address pharmacokinetic limitations through clever, functional SAR. Mikamiyama and Yamada’s work not only advances our understanding of TrkA–NGF interaction inhibitors but also highlights strategies that could be broadly applicable to the design of other macrocyclic peptide-based therapeutics.
Discovery and Hit to Lead Optimization of Macrocyclic Peptides as Novel Tropomyosin Receptor Kinase A Antagonists, Yamada T, Mihara K., Ueda T., Yamauchi D., Shimizu M., Ando A., Mayumi K., Nakata Z., and Mikamiyama H., Journal of Medicinal Chemistry 2024 67 (13), 11197-11208.
DOI: https://pubs.acs.org/doi/10.1021/acs.jmedchem.4c00715
Matteo Villain
Vice President of Peptides and Oligonucleotides, Veranova
Member, BPF Scientific Advisory Board
https://www.linkedin.com/in/matteo-villain/
Read previous editions of the BPF Journal Club series: https://www.boulderpeptide.org/journal-club