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Antimicrobial Agents and Chemotherapy, February 2001, p. 495-501, Vol. 45, No. 2
Experimental Retroviral Section, Department of Medicine
Branch, National Cancer Institute, National Institutes of Health,
Bethesda, Maryland,2 and Departments of
Pathology3 and Veterinary
Science,4 National Institute of Infectious
Diseases, and AIDS Clinical Center, International Medical
Center of Japan,1 Tokyo, Japan
Received 10 February 2000/Returned for modification 11 April
2000/Accepted 7 November 2000
We describe a rapid and simple novel phenotypic assay for drug
susceptibility of human immunodeficiency virus type-1 (HIV-1) using a
CCR5-expressing HeLa/CD4+ cell clone 1-10 (MAGIC-5).
MAGIC-5 cells produced large amounts of HIV-1 in culture supernatants,
which enabled us to perform the phenotypic resistance assay.
Determination of HIV-1 susceptibility to various protease inhibitors
(PI) and nucleoside reverse transcriptase inhibitors was completed
within 15 days in T-cell-tropic (X4) and macrophage-tropic (R5) viruses
using fresh plasma samples containing at least 104
copies/ml. The nucleotide sequence of the envelope V3 region of HIV-1
in plasma was almost identical to that of the virus isolated by MAGIC-5
cells, suggesting a lack of selection bias in our assay. The assay
variability was confined to within five-fold in all drugs examined.
Accordingly, we used a 10-fold increase in the 50% inhibitory
concentration as the cutoff value for viral resistance in the present
assay. HIV-1 resistant to lamivudine, which was not detected by
conventional genotypic assays, was isolated. In HIV-1 with
PI-associated primary amino acid substitutions, our assay showed that
drug resistance profiles correlated well with previously reported
genotypic-assay data. Furthermore, our assay provided comprehensive
results regarding PI resistance in the presence of multiple mutations.
The novel assay successfully quantified the level of resistance of
clinical HIV-1 isolates to a battery of anti-HIV drugs, indicating its
clinical usefulness, particularly in patients who failed to respond to
antiretroviral chemotherapy.
0066-4804/01/$04.00+0 DOI: 10.1128/AAC.45.2.495-501.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Rapid and Simple Phenotypic Assay for Drug
Susceptibility of Human Immunodeficiency Virus Type 1 Using
CCR5-Expressing HeLa/CD4+ Cell Clone 1-10 (MAGIC-5)
*
Corresponding author. Mailing address: AIDS Clinical
Center, International Medical Center of Japan, 1-21-1, Toyama,
Shinjuku-ku, Tokyo 162-8655, Japan. Phone and fax: 81-3-5273-5193. E-mail: oka{at}imcj.hosp.go.jp.
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