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16th International HIV Drug Resistance Workshop12-16 June 2007, Barbados |
BIOCHEMICAL CHARACTERIZATIONS OF THE EFFECT OF MUTATIONS SELECTED IN HIV-1 INTEGRASE GENE ASSOCIATED WITH FAILURE TO RALTEGRAVIR (MK-0518)
Antivir Ther. 2007; 12:S9 (abstract no. 7)
I Malet1, O Delelis2, MA Valantin3, B Montes4, C Soulie1, M Wirden1, L Tchertanova2, G Peytavin5, J Reynes4, JF Mouscadet2, C Katlama3, AG Marcelin1 and V Calvez1
1Laboratoire de Virologie, Hôpital Pitié-Salpêtrière, Paris, France; 2LBPA, CNRS, Ecole Normale Supérieure de Cachan, France; 3Service des Maladies Infectieuses, Hôpital Pitié-Salpêtrière, Paris, France; 4Département de Maladies Infectieuses, CHU de Montpellier, France; 5Laboratoire de Toxicologie, Hôpital Bichat Claude Bernard, Paris, France
BACKGROUND: Raltegravir (MK-0518) is a potent integrase inhibitor that is highly active in patients harbouring resistance viruses to other antiretroviral classes. It has been recently shown that raltegravir can select mutations in integrase gene in patients failing to raltegravir regimen.
METHODS: Three heavily pretreated patients who received a salvage therapy including raltegravir and who subsequently developed virological failure under raltegravir therapy were studied. To describe the genetic changes in the integrase gene, the complete nucleotide sequence of the 864 nucleotides integrase coding region was amplified and determined at different time points during the follow-up of each patient. The 288 corresponding amino acid were deduced, analysed and compared to the point at the beginning of the treatment. The PCR products were cloned into bacterial expression vectors and the 6 proteins produced and purified were assayed in vitro.
RESULTS: Three different profiles of mutations were identified: E92Q, G140S+Q148H that seemed to appear at the same time and N155H mutations. No other specific change all along the integrase gene was identified. In vitro, the three wild-type recombinant integrases were sensitive to raltegravir showing a strand transfer activity specifically inhibited by raltegravir and an unaffected 3' processing activity. All identified mutations altered the functions of integrase protein, E92Q moderately affecting both activities, G140S+Q148H strongly impairing both activities and N155H markedly affecting strand transfer. For each profile, the wild-type and mutant integrases’ sensitivity to raltegravir were compared. The two first profiles displayed a comparable seven- to eight-fold resistance index while the third was more than 14-fold less sensitive to raltegravir.
CONCLUSION: E92Q, G140S+Q148H and N155H integrase mutations, located near the catalytic site of the enzyme, have been identified in patients failing to raltegravir and led to an in vitro resistance. Finally these mutations led to strongly crippled enzymes when tested in vitro in the absence of raltegravir affecting the strand-transfer activity in all cases and the 3' processing in some cases.
2007-06-12
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