Structure-Based Design of Lipophilic Quinazoline Inhibitors of Thymidylate Synthase

J. Med. Chem., 39 (4), 904 -917, 1996. jm9502652

Structure-Based Design of Lipophilic Quinazoline Inhibitors of Thymidylate Synthase

Terence R. Jones,* Michael D. Varney, Stephen E. Webber, Kathleen K. Lewis, Gifford P. Marzoni, Cindy L. Palmer, Vinit Kathardekar, Katharine M. Welsh, Stephanie Webber, David A. Matthews, Krzysztof Appelt, Ward W. Smith, Cheryl A. Janson, J. E. Villafranca, Russell J. Bacquet, Eleanor F. Howland, Carol L. J. Booth, Steven M. Herrmann, Robert W. Ward, Jennifer White, Ellen W. Moomaw, Charlotte A. Bartlett, and Cathy A. Morse

Agouron Pharmaceuticals, Inc., 3565 General Atomics Court, San Diego, California 92121

Received April 10, 1995

Abstract:

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 CF₃, iodo, or ethynyl enhances binding by up to 1 order of magnitude and in the case of CF₃ was proven to fill a nearby pocket in the enzyme. (ii) A simple strongly electron-withdrawing substituent such as NO₂ or CF₃SO₂ 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-C₆H₅SO₂ 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 C₆H₅SO₂ 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 SO₂ group did, however, have IC₅₀'s in the range 1-5 M. Of these, 4-(N-((3,4-dihydro-2-methyl-6-quinazolinyl)methyl)-N-prop-2-ynylamino)phenyl phenyl sulfone, 7n, had IC₅₀'s of about 1 M and was chosen for further elaboration.

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