3rd International Workshop on HIV Drug Resistance 2-5 August 1994, Kauai, Hawaii, USA

RAPID SCREEN OF CLINICAL SPECIMENS FOR DRUG RESISTANT HIV PHENOTYPES DURING VIRUS ISOLATION

Int Wkshop HIV Drug Res 1994 Aug 2-5;3:26 (abstract no. 25)

Douglas L. Mayers, J. Lane^* and O.S. Weislow^*
Walter Reed Army Institute for Research, and ^*SRA Technologies, Inc. Rockville, Md, 20850, USA

OBJECTIVE: Identification of drug resistant HIV isolates currently requires virus isolation followed by expansion and titration of the isolate. Phenotyping is then accomplished with a drug susceptibility assay. This process takes 4-6 weeks and outcomes are potentially influenced by frequency of virus passage. To reduce the cost, time to phenotype identification and potential for phenotypic selection during viral propagation, we have developed a rapid drug phenotype screening assay (RDPS).

METHODS: PBMC from HIV-infected patients were separated by ficoll/hypaque centrifugation and co-cultured with PHA-stimulated, donor PBMC until supernatants were positive with p24Ag >200 pg/ml. The cultures were split into three subcultures and the media was adjusted to 0, 0.2 and 2.0 µM ZDV. Supernatant p24Ag was quantitated after 7 days of culture in the presence of drug. Screening assay results were compared with those obtained using the ACTG/DoD drug susceptibility assay.

RESULTS: Of 9 isolates studied, 3 were ZDV-sensitive (IC₅₀ <0.2 µM), 1 partially resistant and 5 resistant (IC₅₀ ≥1 µM) by the ACTG/DoD assay. P24Ag levels in the no drug control cultures ranged from 867 to 2046 pg/ml. There was no p24Ag detected in the presence of 0.2 µM ZDV for the ZDV-sensitive HIV isolates. The partially resistant isolate had 3.8% control p24Ag levels in the presence of 0.2 µM ZDV. The ZDV resistant isolates had >25% control p24Ag levels in the presence of 0.2 µ ZDV. 2 µM ZDV totally inhibited the majority of ZDV-resistant clinical HIV isolates using this assay format.

CONCLUSIONS: This initial study suggests the RDPS will permit differentiation of ZDV-sensitive and resistant HIV phenotypes with results in 2-3 weeks. HIV isolates which produce p24Ag levels >25% no drug control in the presence of 0.2 µM ZDV in this assay should have IC₅₀ values ≥1.0 µM using the ACTG/DoD assay.

Download PDF of this abstract.

1994-08-02
25

Copyright © 1994 - International Medical Press Ltd. Reproduction of this abstract (other than one copy for personal reference) must be cleared through the International Medical Press Ltd. 2-4 Idol Lane, London EC3R 5DD UK.