Nanoscale Direct-to-Biology Optimization of Cdk2 Inhibitors

  • J Med Chem. 2026 Apr 23;69(8):9142-9162. doi: 10.1021/acs.jmedchem.5c03614.
James L Douthwaite  1 ,  Damian J Houde  2 ,  Eneida Pardo  2 ,  Mark Moran  2 ,  Jason Baird  2 ,  Sophia R Meyer  1 ,  Babak Mahjour  1 ,  Qiyuan Zhao  1 ,  Jay F Larrow  2 ,  Yu-Pu Juang  1 ,  Michael J Holliday  2 ,  Calvin Han  2 ,  Brian Kelley  2 ,  David Dunstan  2 ,  Katelyn Billings  2 ,  Mary M Mader  2 ,  Alexander M Taylor  2 ,  Jonathan Z Sexton  1  3 ,  Alessandro A Boezio  2 ,  Tim Cernak  1
Affiliations
  • 1. Department of Medicinal Chemistry, College of Pharmacy, University of Michigan, Ann Arbor, Michigan 48109, United States.
  • 2. Relay Therapeutics, 60 Hampshire Street, Cambridge, Massachusetts 02139, United States.
  • 3. Department of Internal Medicine, Gastroenterology and Hepatology, Michigan Medicine at the University of Michigan, Ann Arbor, Michigan 48109, United States.
PMID: 41954971 DOI: 10.1021/acs.jmedchem.5c03614
Abstract

Modern hit-to-lead optimization winnows down vast chemical spaces of virtual compounds into a selection of potent and selective compounds that can be further profiled with in vitro assays. Today, miniaturized Chemical Synthesis can be performed in high-throughput, shifting the bottleneck to compound purification. Direct-to-biology (D2B) approaches seek to overcome this hurdle by omitting the purification step and submitting reaction mixtures directly to bioassays. Here, we explore nanoscale hit-to-lead optimization through the multistep synthesis of a library of CDK2/CycE inhibitors, utilizing ultrahigh-throughput experimentation (ultraHTE) in 1,536-well plates. Library performance is assessed by D2B in functional biochemical, bioaffinity, and X-ray crystallographic assays. A selection of potent lead CDK2/CycE inhibitors identified by D2B was submitted to a phenotypic cell painting assay, which showed cell cycle arrest at G0, consistent with CDK2 inhibition. This miniaturized workflow allows the upper tiers of a typical optimization screening cascade to be performed in a single experiment.

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