Directed Molecular Engineering of Mig6 Peptide Selectivity between Proto-oncogene ErbB Family Receptor Tyrosine Kinases

다국어 초록 (Multilingual Abstract)

The ErbB signaling pathway plays important roles in normal physiology and cancer, which consists of four proto-oncogene receptor tyrosine kinases ErbB1/EGFR, ErbB2/Her2, ErbB3/Her3, and ErbB4/Her4. Selective targeting of different ErbB kinases would result in distinct therapeutic effects, but traditional small-molecule inhibitors generally exhibit a strong cross-reactivity over these kinases due to the very high conservation in kinase’s active site.
Instead of developing small-molecule drugs to selectively target the conserved active site of ErbB kinases, we herein attempt to design peptide agents for selectively disrupting the dimerization event of these kinases at their asymmetric dimer interfaces that have a relatively low homology.
Three hotspot peptides S1P1, S1P2, and S1P3 are split from the functional segment 1 (Seg1) of mitogen-inducible gene 6 (Mig6), a natural EGFR-inhibitory protein that has been observed to inactivate the kinase by disrupting its dimerization. We demonstrate that the Mig6 peptides not only inhibit EGFR, but also bind Her2, Her3, and Her4, although the peptide affinities to the four ErbB kinases are different considerably, exhibiting a typical selectivity. The S1P2 peptide locates in the core binding region of Mig6 Seg1 and contributes significantly to the segment interaction with kinases. An iteration algorithm is employed to guide the directed molecular engineering of S1P2 peptide selectivity towards each of the four kinases. Hundreds of parallel evolution running yield a series of peptide candidates with potential selectivity, which are then substantiated by fluorescence-based assays. The designed EGFR-selective peptide S1P2-p1EGFR is determined to have a moderate affinity to EGFR (Kd = 56 μM) and a high selectivity for EGFR over Her2, Her3, and H4 (FEGFR = 10.1-fold), which is improved considerably relative to wild-type S1P2 peptide (FEGFR = 2.7-fold). Structural examination observes different noncovalent interaction modes at the complex interfaces of S1P2-p1EGFR with EGFR and other three kinases, revealing a molecular origin of the peptide selectivity.

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