AEGiS-14IAC: A glycine to serine mutation at position 140 of integrase confers resistance to integrase inhibitors and defective integration.

14th International AIDS Conference

Barcelona, Spain - July 7-12, 2002

A glycine to serine mutation at position 140 of integrase confers resistance to integrase inhibitors and defective integration.

Int Conf AIDS 2002 Jul 7-12; 14:(abstract no. ThOrA1432)

Robinson WE, King PJ, Lee DJ, Reinke RA, Victoria JG
University of California, Irvine, United States

BACKGROUND: Culture of HIV with increasing concentrations of L-chicoric acid (L-CA), an inhibitor of integrase (IN), resulted in a single point mutation within IN that conferred resistance of the virus to L-CA. The effects of this mutation on integration and susceptibility to other IN inhibitors were unknown. It was hypothesized that the mutation would cause resistance of the protein to L-CA and cross-resistance to L-731,988, a diketoacid.

METHODS: IN containing the G140S mutation (ING140S) was purified. Susceptibility to both L-CA and L-731,988 was determined. The enzymatic activities of the IN proteins were measured under steady-state and non-steady-state conditions. Binding of the mutant protein to LTR substrate was measured. The effect of the mutation on HIV replication kinetics was measured using reverse transcriptase release, immunofluorescence, and quantitative real-time polymerase chain reaction (PCR). The ratio of two LTR circle DNA to cDNA in HIV- infected cells was used to determine effects of the mutation on integration.

RESULTS: Using real-time PCR, a growth defect secondary to failure of integration was demonstrated. The ING140S protein was attenuated approximately 4-fold for catalysis under equilibrium conditions compared to wildtype IN (INWT) and attenuated 5-fold in steady-state kinetic analysis of disintegration. Fifty-percent inhibitory concentration assays were performed with IN inhibitors against both ING140S and INWT in disintegration and strand transfer reactions. ING140S was resistant to both L-CA and L-731,988. HIV containing the mutation was resistant to L-731,988 and L-CA.

CONCLUSIONS: Glycine 140 is important for susceptibility of IN to both L-CA and a diketoacid. Mutation of this residue to serine confers cross-resistance to both inhibitors and a defect in catalysis mediated by IN. Therefore, both L-731,988 and L-CA are inhibitors of IN and may interact with the same drug binding pocket on IN.

Keywords: AEGIS, Integrase Inhibitors, Integrases, Serine, Virus Replication, Glycine, Mutation, Succinates, Caffeic Acids, HIV Infections, chicoric acid, virology, genetics

020707
ThOrA1432