Structure-Based Design\nof “Head-to-Tail”\nMacrocyclic PROTACs

Macrocyclization is a compelling strategy for conventional\ndrug\ndesign for improving biological activity, target specificity, and\nmetabolic stability, but it was rarely applied to the design of PROTACs\npossibly due to the mechanism and structural complexity. Herein, we\nreport the rational design of the first series of “Head-to-Tail”\nmacrocyclic PROTACs. The resulting molecule <b>SHD913</b> exhibited\npronounced Brd4 protein degradation with low nM DC<sub>50</sub> values\nwhile almost totally dismissing the “hook effect”, which\nis a general character and common concern of a PROTAC, in multiple\ncancer cell lines. Further biological evaluation revealed that the\ncompound exhibited positive cooperativity and induced <i>de novo</i> protein–protein interactions (PPIs) in both biophysical and\ncellular NanoBRET assays and outperformed macroPROTAC-1 that is the\nfirst reported macrocyclic Brd4 PROTAC, in cellular assays. <i>In vitro</i> liver microsomal stability evaluation suggested\nthat <b>SHD913</b> demonstrated improved metabolic stability\nin different species compared with the linear counterpart. The co-crystal\nstructure of Brd4<sup>BD2</sup>: <b>SHD913</b>: VCB (VHL, Elongin\nC and Elongin B) complex determination and molecular dynamics (MD)\nsimulation also elucidated details of the chemical-induced PPIs and\nhighlighted the crucial contribution of restricted conformation of <b>SHD913</b> to the ternary complex formation. These results collectively\nsupport that macrocyclization could be an attractive and feasible\nstrategy for a new PROTAC design.