Klimkait T, Schaub G, Brennan L; International Conference on AIDS.
Int Conf AIDS. 2000 Jul 9-14; 13: abstract no. TuPeA3018.
T. Klimkait, Basel Center for HIV Research, University of Basel, Institute for Medical Microbiology, Petersplatz 10, 4003 Basel, Switzerland, Tel.: +41 612 673 264, Fax: +41 612 673 298, E-mail: thomas.klimkait@unibas.ch
Background: Even in the era of "HAART" as standard HIV treatment and despite the improved patients' perspectives, the times of failing therapies are not over: viral drug resistance has advanced to become one of the most serious threats to effective clinical management. Rapid detection and characterization of HIV-resist-ance are now being recognized as crucial success factors for therapy. The most advanced approach uses "genotyping" of the virus in patients' blood; however, this method is unable to effectively discriminate viral variants coexisting in the same sample. Results: As "genotyping" alone detects only the most prominent HIV mutants in a given blood sample, a selective sequence assignment to one viral genome is difficult. We have therefore developed a phenotype-based test-system, in which HIV PR- and/or RT-gene sequences from patients are re-inserted into an engineered isogenic HIV-1 cloning cassette: 1. After gene insertion, plasmids from individual bacterial colonies can be sequenced. We could indeed confirm the coexistence of sequence variants in the original clinical sample 2. After transfection of "plasmid pools" from bacterial cultures into human cells subsequent drug selection revealed the presence of more drug resistant HIV variants than the initial "genotype", performed prior to cloning. 3. Using a "rapid virus amplification format" the test system needed 7 days to assess drug susceptibility of viruses or -mixtures with complex mutational patterns. Conclusions: We present a system for 1) potent determination of viral resistance to individual HIV inhibitors or within a full drug-regimen even before comprehensive resistance information is available, which 2) is suitable even for new, previously not yet described mutations. We hope to develop the assay into a cost-effective and routine test which can be used for the direct detection of complex viral mixtures in clinical specimens including even greatly underrepre-sented HIV variants.
Publication Types:
Keywords:
- AIDS Vaccines
- Acquired Immunodeficiency Syndrome
- Antiretroviral Therapy, Highly Active
- Drug Resistance, Viral
- Genotype
- HIV Infections
- HIV Seropositivity
- HIV-1
- Humans
- In Vitro
- genetics
- methods
- reverse transcriptase, Human immunodeficiency virus 1
Other ID:
UI: 102238864
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