Polymerase chain reaction assays for monitoring antiviral therapy and making therapeutic decisions in the treatment of acquired immunodeficiency syndrome

The present invention relates to methods of monitoring, via polymerase chain reaction, the clinical progression of human immunodeficiency virus infection and its response to antiretroviral therapy. According to the invention, polymerase chain reaction assays may be used to predict immunological decline and to identify, at an early stage, patients whose infection has become resistant to a particular antiretroviral drug regimen.

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Claims

1. A method of evaluating the effectiveness of antiretroviral therapy of an HIV-infected patient comprising:

(i) collecting a plasma sample from an HIV-infected patient; and
(ii) determining whether the plasma sample comprises nucleic acid encoding HIV reverse transcriptase having a mutation at codons 215 or 74, or codons 215 and 74, in which the presence of the mutations correlates positively with an accelerated immunologic decline of said patient compared to patients who do not have the mutations.

2. The method of claim 1 in which the mutation at codon 215, 74, or 215 and 74 are determined by a method comprising polymerase chain reaction (PCR).

3. The method of claim 2 which comprises a "nested" polymerase chain reaction.

4. The method of claim 2 which utilizes primer B (5'-GGATGGAAAGGATCACC-3')(SEQ ID NO:9).

5. The method of claim 2 which utilizes primer 3M (3'-AAGTGTGGTCTGTTTTTTGTA-5') (SEQ ID NO:11).

6. The method of claim 2 which utilizes primer 74WT (5'-AAGTTCTCTGAAATCTACTTA-3') (SEQ ID NO:5).

7. The method of claim 2 which utilizes primer 74MUT (5'-AAGTTCTCTGAAATCTACTTC-3') (SEQ ID NO:6).

8. The method of claim 2 which utilizes primer X2(5'-AACAATGGCCATTGACAGA-3') (SEQ ID NO:4).

9. The method of claim 1 in which the antiretroviral agent comprises zidovudine.

10. The method of claim 1 in which the antiretroviral agent comprises didanosine.

11. The method of claim 2 wherein PCR is repeated for 30 to 40 cycles.

12. The method of claim 11 wherein PCR is repeated for 30-35 cycles.

13. A method of evaluating the effectiveness of antiretroviral therapy of an HIV-infected patient comprising:

(i) collecting PBMC from an HIV-infected patient; and
(ii) determining whether the PBMC comprise proviral HIV DNA which comprises a mutation at codons 74 or 215, or codons 215 and 74, of the HIV reverse transcriptase gene, in which the presence of the mutations correlates positively with an accelerated immunologic decline of said patient compared with patients who do not have the mutations.

14. The method of claim 13 in which the mutations at codons 74 or 215, or 215 and 74 are determined by a method comprising polymerase chain reaction.

15. The method of claim 14 which comprises a nested polymerase chain reaction.

Referenced Cited
U.S. Patent Documents
4683195 July 28, 1987 Mullis et al.
5008182 April 16, 1991 Sninsky et al.
5409810 April 25, 1995 Larder et al.
5631128 May 20, 1997 Kozal et al.
5650268 July 22, 1997 Kozal et al.
Foreign Patent Documents
0229709 July 1987 EPX
0-422-762-A2 August 1990 EPX
A-60828/90 July 1987 GBX
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Patent History
Patent number: 5856086
Type: Grant
Filed: Jan 15, 1997
Date of Patent: Jan 5, 1999
Assignee: Leland Stanford Junior University (Stanford, CA)
Inventors: Michael J. Kozal (Menlo Park, CA), Thomas C.. Merigan (Portola Valley, CA)
Primary Examiner: Jeffrey Fredman
Law Firm: Pennie & Edmunds LLP
Application Number: 8/783,786
Classifications
Current U.S. Class: Involving Virus Or Bacteriophage (435/5); 435/6; Using Fungi (435/911); Absidia (435/912); 435/9151; 536/231; 536/243; 536/2431; 536/2452; 536/2433; Biological Cellular Material Tested (436/63); Biospecific Ligand Binding Assay (436/501); 436/601
International Classification: C12Q 170; C12Q 168; C12P 1934; C07H 2104;