Inhibition of HIV-1 reverse transcriptase by pyridinone derivatives. Potency, binding characteristics, and effect of template sequence.
The inhibition of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase by pyridinone compounds has been investigated using as templates synthetic RNA with sequences based on the HIV-1 genome sequence. In reactions catalyzed by the enzyme that incorporated more than one nucleotide per primer, inhibition by a representative pyridinone inhibitor, 3-[2-(1,3-benzoxazol-2-yl)ethyl]-5-ethyl-6-methyl-pyridin-2(1H)one (L-696,229), was noncompetitive against deoxynucleotide triphosphate. For reactions that incorporated one deoxynucleotide per primer, IC50 values ranged from 20 to 200 nM, depending on the position of incorporation of the incoming deoxynucleotide base on the template. Inhibition of synthesis on a set of four templates differing only at the template base complementary to the incoming nucleotide had similar IC50 values. These results demonstrate that inhibitory potency is dependent on the primary structure of the template and that inhibitory potency is largely independent of the identity of the incoming nucleotide base. The inhibition of HIV-1 reverse transcription by L-696,229 also displayed slow-binding characteristics. The slow-binding aspect was exploited to gauge the interaction between inhibitor and enzyme. By titrating the reduction in the extent of the burst of synthesis observed in a reaction incorporating dideoxythymidine monophosphate into poly(rA)-oligo(dT)18, the apparent equilibrium constant for dissociation of the reverse transcriptase-L-696,229 complex was estimated to be 400 nM.
Carroll
S S
SS
Department of Biological Chemistry, Merck Research Laboratories, West Point, Pennsylvania 19486.
Olsen
D B
DB
Bennett
C D
CD
Gotlib
L
L
Graham
D J
DJ
Condra
J H
JH
Stern
A M
AM
Shafer
J A
JA
Kuo
L C
LC
D000998:Antiviral Agents / Q000494:pharmacology*; D001483:Base Sequence; D001583:Benzoxazoles / Q000494:pharmacology*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology*; D007700:Kinetics; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000378:metabolism / Q000494:pharmacology*; D011728:Pyridones / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D018894:Reverse Transcriptase Inhibitors; D013329:Structure-Activity Relationship; D013698:Templates, Genetic
D000998:Antiviral Agents; D001583:Benzoxazoles; D009838:Oligodeoxyribonucleotides; D011728:Pyridones; D011994:Recombinant Proteins; D018894:Reverse Transcriptase Inhibitors; C076250:L 696229; D054303:HIV Reverse Transcriptase