Discovery and Synthesis of a Naturally Derived Protein Kinase Inhibitor that Selectively Inhibits Distinct Classes of Serine/Threonine Kinases
Lin Du, Brice A. P. Wilson, Ning Li, Rohan Shah, Masoumeh Dalilian, Dongdong Wang, Emily A. Smith, Antony Wamiru, Ekaterina I. Goncharova, Ping Zhang, Barry R. O’Keefe
Abstract
High Resolution Image Download MS PowerPoint Slide The DNAJB1–PRKACA oncogenic gene fusion results in an active kinase enzyme, J-PKAcα, that has been identified as an attractive antitumor target for fibrolamellar hepatocellular carcinoma (FLHCC). A high-throughput assay was used to identify inhibitors of J-PKAcα catalytic activity by screening the NCI Program for Natural Product Discovery (NPNPD) prefractionated natural product library. Purification of the active agent from a single fraction of an Aplidium sp. marine tunicate led to the discovery of two unprecedented alkaloids, aplithianines A ( 1 ) and B ( 2 ). Aplithianine A ( 1 ) showed potent inhibition against J-PKAcα with an IC 50 of ∼1 μM in the primary screening assay. In kinome screening, 1 inhibited wild-type PKA with an IC 50 of 84 nM. Further mechanistic studies including cocrystallization and X-ray diffraction experiments revealed that 1 inhibited PKAcα catalytic activity by competitively binding to the ATP pocket. Human kinome profiling of 1 against a panel of 370 kinases revealed potent inhibition of select serine/threonine kinases in the CLK and PKG families with IC 50 values in the range ∼11–90 nM. An efficient, four-step total synthesis of 1 has been accomplished, enabling further evaluation of aplithianines as biologically relevant kinase inhibitors.