title: Rational design and synthesis of a novel anti-leukemic agent targeting Bruton's tyrosine kinase (BTK), LFM-A13 [alpha-cyano-beta-hydroxy-beta-methyl-N-(2, 5-dibromophenyl)propenamide].
abstract: In a systematic effort to design potent inhibitors of the anti-apoptotic tyrosine kinase BTK (Bruton's tyrosine kinase) as anti-leukemic agents with apoptosis-promoting and chemosensitizing properties, we have constructed a three-dimensional homology model of the BTK kinase domain. Our modeling studies revealed a distinct rectangular binding pocket near the hinge region of the BTK kinase domain with Leu460, Tyr476, Arg525, and Asp539 residues occupying the corners of the rectangle. The dimensions of this rectangle are approximately 18 x 8 x 9 x 17 A, and the thickness of the pocket is approximately 7 A. Advanced docking procedures were employed for the rational design of leflunomide metabolite (LFM) analogs with a high likelihood to bind favorably to the catalytic site within the kinase domain of BTK. The lead compound LFM-A13, for which we calculated a Ki value of 1.4 microM, inhibited human BTK in vitro with an IC50 value of 17.2 +/- 0.8 microM. Similarly, LFM-A13 inhibited recombinant BTK expressed in a baculovirus expression vector system with an IC50 value of 2.5 microM. The energetically favorable position of LFM-A13 in the binding pocket is such that its aromatic ring is close to Tyr476, and its substituent group is sandwiched between residues Arg525 and Asp539. In addition, LFM-A13 is capable of favorable hydrogen bonding interactions with BTK via Asp539 and Arg525 residues. Besides its remarkable potency in BTK kinase assays, LFM-A13 was also discovered to be a highly specific inhibitor of BTK. Even at concentrations as high as 100 micrograms/ml (approximately 278 microM), this novel inhibitor did not affect the enzymatic activity of other protein tyrosine kinases, including JAK1, JAK3, HCK, epidermal growth factor receptor kinase, and insulin receptor kinase. In accordance with the anti-apoptotic function of BTK, treatment of BTK+ B-lineage leukemic cells with LFM-A13 enhanced their sensitivity to ceramide- or vincristine-induced apoptosis. To our knowledge, LFM-A13 is the first BTK-specific tyrosine kinase inhibitor and the first anti-leukemic agent targeting BTK.
authors: Parker Hughes Cancer Center, Hughes Institute, St. Paul, Minnesota 55113, USA.
authors: S
authors: S
authors: Mahajan
authors: S
authors: S
authors: Ghosh
authors: E A
authors: EA
authors: Sudbeck
authors: Y
authors: Y
authors: Zheng
authors: S
authors: S
authors: Downs
authors: M
authors: M
authors: Hupke
authors: F M
authors: FM
authors: Uckun
mesh_terms: D000077329:Agammaglobulinaemia Tyrosine Kinase; D000577:Amides / Q000138:chemical synthesis* / Q000494:pharmacology; D000970:Antineoplastic Agents / Q000138:chemical synthesis* / Q000494:pharmacology; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D015195:Drug Design; D004791:Enzyme Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D053613:Janus Kinase 1; D053614:Janus Kinase 2; D008958:Models, Molecular; D009570:Nitriles / Q000138:chemical synthesis* / Q000494:pharmacology; D011487:Protein Conformation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011518:Proto-Oncogene Proteins; D011972:Receptor, Insulin / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives / Q000494:pharmacology; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured; D014750:Vincristine / Q000494:pharmacology; D019061:src-Family Kinases / Q000378:metabolism
chemical_list: D000577:Amides; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; C118451:LFM A13; C064769:N-acetylsphingosine; D009570:Nitriles; D011518:Proto-Oncogene Proteins; D014750:Vincristine; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D000077329:Agammaglobulinaemia Tyrosine Kinase; C000625949:BTK protein, human; C507904:JAK1 protein, human; C507924:JAK2 protein, human; D053613:Janus Kinase 1; D053614:Janus Kinase 2; D019061:src-Family Kinases; D013110:Sphingosine