Structure-based design of lipophilic quinazoline inhibitors of thymidylate synthase.

To develop novel lipophilic thymidylate synthase (TS) inhibitors, the X-ray structure of Escherichia coli TS in ternary complex with FdUMP and the inhibitor 10-propargyl-5,8-dideazafolic acid (CB3717) was used as a basis for structure-based design. A total of 31 novel lipophilic TS inhibitors, lacking a glutamate residue, were synthesized; 26 of them had in common a N-((3,4-dihydro-2-methyl-6-quinazolinyl)methyl)-N-prop-2-ynylaniline+ ++ structure in which the aniline was appropriately substituted with simple lipophilic substituents either in position 3 or 4, or in both. Compounds were tested for their inhibition of E. coli TS and human TS and also for their inhibition of the growth in tissue culture of a murine leukemia, a human leukemia, and a thymidine kinase-deficient human adenocarcinoma. The crystal structures of five inhibitors complexed with E. coli TS were determined. Five main conclusions are drawn from this study. (i) A 3-substituent such as CF(3), iodo, or ethynyl enhances binding by up to 1 order of magnitude and in the case of CF(3) was proven to fill a nearby pocket in the enzyme. (ii) A simple strongly electron-withdrawing substituent such as NO(2) or CF(3)SO(2) in the 4-position enhances binding by 2 orders of magnitude; it is hypothesized that the transannular dipole so induced interacts favorably with the protein. (iii) Attempts to combine the enhancements of i and ii in the same molecule were generally unsuccessful (iv) A 4-C(6)H(5)SO(2) substituent provided both electron withdrawal and a van der Waal's interaction of the phenyl group with a hydrophobic surface at the mouth of the active site. The inhibition (K(is) = 12 nM) of human TS by this compound, 7n, showed that C(6)H(5)SO(2) provided virtually as much binding affinity as the CO-glutamate which it had replaced. (v) The series of compounds were poorly water soluble, and also the potent TS inhibition shown by several of them did not translate into good cytotoxicity. Compounds with large cyclic groups linked to position 4 by an SO or SO(2) group did, however, have IC(50)'s in the range 1-5 microM. Of these, 4-(N-((3,4-dihydro-2-methyl-6-quinazolinyl)methyl)-N-prop-2-ynylamino )phenyl phenyl sulfone, 7n, had IC(50)'s of about 1 microM and was chosen for further elaboration.

Crystal-structure-based design and synthesis of benz[cd]indole-containing inhibitors of thymidylate synthase.

The X-ray crystal-structure-based design, synthesis, and biological activity of a novel family of benz[cd]indole-containing inhibitors of thymidylate synthase (TS) are described. The structure-activity of the lead compound was studied by conceptually dividing the molecule into four regions and independently optimizing the substituents for each region. Combination of favored substituents for each region led to inhibitors with Ki's against the human enzyme in the range of 10-20 nM. Thymidine shift experiments suggested that the cytotoxic properties of the best enzyme inhibitors were due to TS targeting in cells. The inhibitors were synthesized from substituted 6-aminobenz[cd]indol-2(1H)-ones by alkylation with both a simple alkyl group and a substituted benzylic portion. The 2,6-diaminobenz[cd]indoles were prepared from the corresponding lactams by conversion to the thiolactam, alkylation to the methylated thiolactam, and then displacement with a substituted or unsubstituted amine.

Pharmacological activity of the isomers of LY53857, potent and selective 5-HT2 receptor antagonists.

LY53857 is an ergoline derivative that is a potent and selective antagonist at 5-HT2 receptors. The present study investigated the activities of the diastereomers of LY53857. The SS- and RR-isomers and the meso mixture of isomers of LY53857 all showed affinity for 5-HT2 receptors approximately 100-fold higher than for 5-HT1 receptors based on radiolabeled ligand binding studies in rat cortical membranes. Based on these binding data, the meso mixture of isomers was one-half as potent as the other two isomers. Consistent with the binding data, the SS- and RR-isomers and the meso mixture of isomers all showed high affinity as antagonists of 5-HT2 receptors in the rat jugular vein, with the meso mixture of isomers being slightly less potent than the other two isomers. Affinity of the isomers at alpha-1 and alpha-2 receptors was low. Approximately 100,000- and 2000-fold higher concentrations were required for antagonism of alpha-1 and alpha-2 receptors, respectively, relative to 5-HT2 receptors. A similar profile of selectivity was demonstrated for each of the isomers studied. These in vitro data were consistent with in vivo data showing the high i.v. and p.o. potency of all the isomers to antagonize the pressor response to i.v. administration of serotonin in pithed spontaneously hypertensive rats. As seen in vitro, the meso mixture of isomers was slightly less potent than the SS- and RR-isomers after p.o. administration in vivo.(ABSTRACT TRUNCATED AT 250 WORDS)