Method for detection of drug-induced mutations in the reverse tr

Details Method for detection of drug-induced mutations in the reverse tr Method for detection of drug-induced mutations in the reverse tr

1997-09-15

2000-07-11

Horlick, Kenneth R.

Chemistry: molecular biology and microbiology

Measuring or testing process involving enzymes or...

Involving virus or bacteriophage

435 912, 536 2432, C12Q 170, C12P 1934, C07H 2104

Patent

active

060870931

DESCRIPTION:

BRIEF SUMMARY
BACKGROUND OF THE INVENTION

The present invention relates to the field of HIV diagnosis. More particularly, the present invention relates to the field of diagnosing the susceptibility of an HIV sample to antiviral drugs used to treat HIV infection.
1. Field of the Invention
The present invention relates to a method for the rapid and reliable detection of drug-induced mutations in the HIV reverse transcriptase gene allowing the simultaneous characterization of a range of codons involved in drug resistance using specific sets of probes optimized to function together in a reverse-hybridisation assay.
2. Description of the Related Art
During the treatment of human immunodeficiency virus (HIV) type 1 infected individuals with antiretroviral nucleoside analogs emergence of resistance against these drugs has been observed. The mechanism responsible for the resistance is not fully understood, since the appearance of a resistant virus in not always correlated with clinical deterioration (Boucher et al. 1992). Amongst the reverse transcriptase (RT) inhibitors, the nucleoside analogs 3'-azido-2',3'-dideoxyThymidine (AZT, Zidovudine), 2',3'-dideoxyInosine (ddI), 2',3'-dideoxyCytidine (ddC), (-)-.beta.-L-2',3'-dideoxy-3'-thioCytidine (3TC), 2',3'-didehydro-3'deoxyThymidine (D4T) and (-)-2'3'-dideoxy-5-fluoro-3'-thiacytidine (FTC) are the most important, since they show a favourable ratio of toxicity for the host versus efficacy as antiviral. All these compounds act in a similar way, namely they serve, after intracellular phosphorylation, as chain terminators of the RT reaction. Upon prolonged treatment with these nucleoside analogs, accumulation of mutations in the viral reverse transcriptase gene (RT) occur, thereby escaping the inhibitory effect of the antivirals. The most important mutations induced by the above compounds and leading to gradually increasing resistance were found at amino acid (aa) positions 41 (M to L), 69 (T to D), 70 (K to R), 74 (L to V), 181 (Y to C), 184 (M to V) and 215 (T to Y or F) (Schinazi et al., 1994). Mutations at aa 65, 67, 75 and 219 have also been reported but these were only showing a minor decrease in sensitivity. More recently, multi-drug-resistant HIV-1 strains were described showing aa changes at codon 62, 75, 77, 116, and 151 (Iversen et al., 1996). In general, these aa changes are the consequence of single point mutations at the first or second codon letter, but in the case of T69D (ACT to GAT), T215Y (ACC to TAC) and T215F (ACC to TTC), two nucleotide mutations are necessary. Whether in these cases the single nucleotide mutation intermediates exist, and if they show any importance in the mechanism for acquiring resistance is as yet not reported. Third letter variations are in general not leading to an amino acid change, and are therefore seen as natural polymorphisms.
The regime for an efficient antiviral treatment is not clear at all. The appearance of one or several of these mutations during antiviral treatment need to be interpreted in conjunction with the virus load and the amount of CD4 cells. Indeed, since it has been shown that the effect of AZT resistance mutations can be suppressed after the appearance of the 3TC induced M184V mutation, it is clear that disease progression is multifactorial. The influence of other simultaneous occuring mutations under different combination therapies with respect to the outcome and resistance of the virus has not yet been analysed systematically. In order to get a better insight into the mechanisms of resistance and HIV biology, it is necessary to analyse follow-up plasma samples of antiviral treated patients for these mutational events together with the simultaneous occuring changes of virus titre and CD4 cells.


BRIEF SUMMARY OF THE INVENTION

It is an aim of the present invention to develop a rapid and reliable detection method for determination of the antiviral drug resistance of viruses which contain reverse transcriptase genes such as HIV retroviruses and Hepadnaviridae present in a biological sample.
More particularly it is a

REFERENCES:
patent: 5409810 (1995-04-01), Larder et al.
patent: 5631128 (1997-05-01), Kozal et al.
patent: 5759770 (1998-06-01), Guertler et al.
patent: 5827648 (1998-10-01), Eastman et al.
Mellors et al., "Mutations in HIV-1 Reverse Transcriptase and Protease Associated with Drug Resistance," Int. Antiviral News 3:8-13 (1995).
Schinazzi et al., "Mutations in HIV-1 Reverse Transcriptase and Protease Associated with Drug Resistance," Int. Antiviral News 2:72-74 (1994).
Larder et al., "Multiple Mutations in HIV-1 Reverse Transcriptase Confer High-Level Resistance to Zidovudine (AZT)," Science 246:1155-1158 (1989).
Eron et al., "Susceptibility Testing by Polymerase Chain Reaction DNA Quantitation: A Method to Measure Drug Resistance of Human Immunodeficiency Virus Type 1 Isolates," Proceedings of the National Academy of Sciences of USA 89:3241-3245 (1992).
Gingeras et al., "Use of Self-Sustained Sequence Replication Amplification Reaction to Analyze and Detect Mutations in Zidovudine-Resistant Human Immunodeficiency Virus," The Journal of Infectious Diseases 164:1066-1074 (1991).
Stuyver et al., "Typing of HCV Isolates and Characterisation of New Subtypes Using a Line Probe Assay," J. Gen. Virol. 74:1093-1102 (1993).
Lipshutz et al., "Using Oligonucleotide Probe Arrays to Access Genetic Diversity," Biotechniques 19:442-447 (1995).
Iversen et al., "Multidrug Resistant HIV-1 Strains Resulting form Combination Antiretroviral Therapy," J. Virol. 70:1086-1090 (1996).
Mellors et al., Int. Antiviral News 3(1), 8-13 (1995).
Shafer et al., J. Inf. Diseases 172, 70-78 (1995).
Stuyver et al., Antimicrobial Agents and Chemo. 41, 284-291 (1997).

Affiliated with

Also associated with

Rating

Method for detection of drug-induced mutations in the reverse tr does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Method for detection of drug-induced mutations in the reverse tr, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Method for detection of drug-induced mutations in the reverse tr will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-540701