Human immunodeficiency virus (HIV) isolates with reduced sensitivity to zidovudine (3'-azido-3'-deoxythymidine, AZT) from individuals with acquired immunodeficiency syndrome (AIDS) or AIDS-related complex were studied to determine the genetic basis of their resistance. Most were sequential isolates obtained at the initiation of and during therapy. Comparative nucleotide sequence analysis of the reverse transcriptase (RT) coding region from five pairs of sensitive and resistant isolates identified three predicted amino acid substitutions common to all the resistant strains (Asp67----Asn, Lys70----Arg, Thr215----Phe or Tyr) plus a fourth in three isolates (Lys219----Gln). Partially resistant isolates had combinations of these four changes. An infectious molecular clone constructed with these four mutations in RT yielded highly resistant HIV after transfection of T cells. The reproducible nature of these mutations should make it possible to develop rapid assays to predict zidovudine resistance by performing polymerase chain reaction amplification of nucleic acid from peripheral blood lymphocytes, thereby circumventing current lengthy HIV isolation and sensitivity testing.
Resistant variants of human immunodeficiency virus type 1 (HIV-1) have been selected by limited passage in MT4 cells of both wild-type and 3'-azido-3'-deoxythymidine (AZT, zidovudine)-resistant strains with the nucleoside analogues (-)-2'-deoxy-3'-thiacytidine (3TC) and (-)-2'-deoxy-5-fluoro-3'-thiacytidine (FTC). Virus variants selected independently were crossresistant to both inhibitors. This rapid in vitro selection of resistant virus has not previously been seen with nucleoside analogues but is reminiscent of that observed with the nonnucleoside reverse transcriptase inhibitors. However, passage of wild-type virus with a combination of AZT and FTC appreciably delayed emergence of FTCresistant virus. DNA sequence analysis of the reverse transcriptase coding region from FTC-resistant virus revealed changes at codon 184 in the highly conserved Tyr, Met, Asp,
Combinations of antiretroviral drugs that prevent or delay the appearance of drug-resistant human immunodeficiency virus-type 1 (HIV-1) mutants are urgently required. Mutants resistant to 3'-azidothymidine (AZT, zidovudine) became phenotypically sensitive in vitro by mutation of residue 184 of viral reverse transcriptase to valine, which also induced resistance to (-)2'-deoxy-3'-thiacytidine (3TC). Furthermore, AZT-3TC coresistance was not observed during extensive in vitro selection with both drugs. In vivo AZT-3TC combination therapy resulted in a markedly greater decreased in serum HIV-1 RNA concentrations than treatment with AZT alone, even though valine-184 mutants rapidly emerged. Most samples assessed from the combination group remained AZT sensitive at 24 weeks of therapy, consistent with in vitro mutation studies.
SUMMARYA genetically engineered herpes simplex virus type 1 (HSV-1) thymidine kinase (TK) deletion mutant has been constructed and used to investigate the role of this gene in pathogenesis. Inoculation of mice with the HSV TK deletion mutant resulted in the establishment of latent ganglionic infection as demonstrated by superinfection of explanted ganglia with wild-type (wt) virus bat not by routine explant culture suggesting that the virus-encoded TK is not essential for the establishment of latent infection but may be necessary for either reactivation or virus replication following reactivation. In addition, Southern blot hybridization has been used to demonstrate in vivo complementation of this mutant by wt virus in both peripheral and central nervous system tissues of mice during acute infection and to show that such complementation can result in the establishment and reactivation of latent TK-infection.
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