Identification of potent pan-ephrin receptor kinase inhibitors using DNA-encoded chemistry technology

• DOI: 10.1073/pnas.2322934121
• Publication Date: 2024-05-03T17:38:32Z
• Authors: Chandrashekhar Ma...; Zian Liao; Sydney E. Parks; Kiran L. Sharma; Kurt M. Bohren; Qiuji Ye; Feng Li; Murugesan Palania...; Zhi Tan; Fei Yuan; Chad J. Creighton; Suni Tang; Ramya P. Masand; Xiaoming Guan; Damian W. Young; Diana Monsivais; Martin M. Matzuk
• Keywords: 0301 basic medicine; 570; Medical Sciences; Biochemical Phenomena; Biophysics; Endometriosis; 610; EphA2; Biochemistry; Cell Line; Small Molecule Libraries; 03 medical and health sciences; DNA-encoded chemistry; Cell Movement; Receptors; Tumor Microenvironment; Medical Specialties; Medicine and Health Sciences; Humans; Biology; Protein Kinase Inhibitors; Kynurenine; Cell Proliferation; and Nutrition; Receptors, Eph Family; Neoplastic; Tumor; Receptor, EphA2; Eph Family; Carcinoma; Life Sciences; Obstetrics and Gynecology; DNA; Biological Sciences; Medical Cell Biology; Pancreatic Neoplasms; Metabolism; Gene Expression Regulation; Pancreatic Ductal; and Structural Biology; RNA; Women's Health; Long Noncoding; Female; Public Health; ephrin receptor kinase inhibitors; Medical Genetics; Receptor

EPH receptors (EPHs), the largest family of tyrosine kinases, phosphorylate downstream substrates upon binding of ephrin cell surface–associated ligands. In a large cohort of endometriotic lesions from individuals with endometriosis, we found thatEPHA2andEPHA4expressions are increased in endometriotic lesions relative to normal eutopic endometrium. Because signaling through EPHs is associated with increased cell migration and invasion, we hypothesized that chemical inhibition of EPHA2/4 could have therapeutic value. We screened DNA-encoded chemical libraries (DECL) to rapidly identify EPHA2/4 kinase inhibitors. Hit compound, CDD-2693, exhibited picomolar/nanomolar kinase activity against EPHA2 (Ki: 4.0 nM) and EPHA4 (Ki: 0.81 nM). Kinome profiling revealed that CDD-2693 bound to most EPH family and SRC family kinases. Using NanoBRET target engagement assays, CDD-2693 had nanomolar activity versus EPHA2 (IC50: 461 nM) and EPHA4 (IC50: 40 nM) but was a micromolar inhibitor of SRC, YES, and FGR. Chemical optimization produced CDD-3167, having picomolar biochemical activity toward EPHA2 (Ki: 0.13 nM) and EPHA4 (Ki: 0.38 nM) with excellent cell-based potency EPHA2 (IC50: 8.0 nM) and EPHA4 (IC50: 2.3 nM). Moreover, CDD-3167 maintained superior off-target cellular selectivity. In 12Z endometriotic epithelial cells, CDD-2693 and CDD-3167 significantly decreased EFNA5 (ligand) induced phosphorylation of EPHA2/4, decreased 12Z cell viability, and decreased IL-1β-mediated expression of prostaglandin synthase 2 (PTGS2). CDD-2693 and CDD-3167 decreased expansion of primary endometrial epithelial organoids from patients with endometriosis and decreased Ewing’s sarcoma viability. Thus, using DECL, we identified potent pan-EPH inhibitors that show specificity and activity in cellular models of endometriosis and cancer.