Src homology 2 (SH2) domain-containing phosphatase 2 (SHP2), encoded by PTPN11 , is a regulator of multiple signaling pathways and is implicated in various cancers. Gain-of-function mutations in PTPN11 account for ∼35% of juvenile myelomonocytic leukemia cases and occur in other malignancies. SHP2 contains a catalytic PTP domain and two SH2 domains (N-SH2 and C-SH2), which mediate interactions with phosphotyrosine-containing binding partners and regulate its enzymatic activity. Often SHP2 variants destabilize its autoinhibited conformation, leading to increased basal phosphatase activity and enhanced binding affinity: a key determinant of pathogenicity. Current therapeutic strategies, including allosteric inhibitors, often fail against these mutants, highlighting the need for alternative approaches. Experimental evidence suggests that inhibiting the interaction between SHP2 and its binding partners acting through SH2 domains is the most effective way to counteract pathogenic mutations. Clinical and research applications of SH2 ligands have been limited due to low affinity and selectivity. Here, we develop a systematic framework to generate peptide inhibitors of SHP2 protein-protein interactions (PPIs) by targeting its SH2 domains. By integrating data from natural ligands and peptide libraries with molecular dynamics simulations, we identified the structural determinants required for high affinity and selectivity. Several peptide sequences were then evaluated through diverse enhanced sampling methods to predict binding affinity, synthesized based on these predictions, and experimentally tested using fluorescence anisotropy to determine dissociation constant values. Iterative optimizations were performed until single-digit nanomolar affinities were achieved. Further optimization with unnatural amino acids enhanced peptide stability for intracellular and in vivo applications. Conjugation with cell-penetrating peptides enabled cytosolic delivery. Using this approach, we developed and patented a peptide targeting N-SH2, and we are extending the strategy to the C-SH2 domain. Overall, this integrated approach provides a method for developing PPI inhibitors targeting SH2 domains.
Innamorati et al. (Sun,) studied this question.