Inhibition of human immunodeficiency virus type 1 replication in vitro by a novel combination of anti-Tat single-chain intrabodies and NF-kappa B antagonists.
Human immunodeficiency virus type 1 (HIV-1) Tat, an early regulatory protein that is critical for viral gene expression and replication, transactivates the HIV-1 long terminal repeat (LTR) via its binding to the transactivation response element (TAR) and, along with other cellular factors, increases viral transcription initiation and elongation. Tat also superactivates the HIV-1 promoter through a TAR-independent mechanism, including tumor necrosis factor alpha-induced and protein kinase C (PKC)-dependent activation of NF-kappa B, and inhibitors of Tat and NF-kappa B cooperatively down-regulate this Tat-mediated LTR superactivation. In this study, a combined pharmacologic and genetic strategy using two PKC (NF-kappa B) inhibitors, pentoxifylline (PTX) and Gö-6976, and a stably expressed anti-Tat single-chain intracellular antibody (sFv intrabody) was employed to obtain cooperative inhibition of both HIV-1 LTR-driven gene expression and HIV-1 replication. Treatment of cells with PTX and Gö-6976 resulted in cooperative inhibition of both HIV-1 LTR-driven gene expression and HIV-1 replication. In addition, the combined use of anti-Tat sFv intrabodies and the two NF-kappa B inhibitors retained the virus in the latent state for as long as 45 days. The combined treatment resulted in more durable inhibition of HIV-1 replication than was seen with the NF-kappa B inhibitors alone or the anti-Tat sFv intrabodies alone. Together, these results suggest that in future clinical gene therapy trials, a combined pharmacologic and genetic strategy like the one reported here may improve the survival of transduced cells and prolong clinical benefit.
Mhashilkar
A M
AM
Division of Human Retrovirology, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA.
Biswas
D K
DK
LaVecchio
J
J
Pardee
A B
AB
Marasco
W A
WA
D019380:Anti-HIV Agents / Q000494:pharmacology*; D001569:Benzodiazepines / Q000494:pharmacology; D015496:CD4-Positive T-Lymphocytes; D002227:Carbazoles / Q000494:pharmacology*; D002460:Cell Line; D015696:Gene Products, tat / Q000276:immunology* / Q000378:metabolism; D015483:HIV Antibodies / Q000235:genetics / Q000276:immunology*; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000187:drug effects* / Q000502:physiology; D006801:Humans; D007211:Indoles / Q000494:pharmacology*; D019169:Jurkat Cells; D007963:Leukocytes, Mononuclear; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D010431:Pentoxifylline / Q000494:pharmacology*; D010835:Phytohemagglutinins / Q000494:pharmacology; D011758:Pyrroles; D013997:Time Factors; D015533:Transcriptional Activation; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D014779:Virus Replication / Q000187:drug effects; D054322:tat Gene Products, Human Immunodeficiency Virus
D019380:Anti-HIV Agents; D002227:Carbazoles; D015696:Gene Products, tat; D015483:HIV Antibodies; D007211:Indoles; D016328:NF-kappa B; D010835:Phytohemagglutinins; D011758:Pyrroles; D014409:Tumor Necrosis Factor-alpha; D054322:tat Gene Products, Human Immunodeficiency Virus; D001569:Benzodiazepines; C081021:Go 6976; C070024:Ro 24-7429; D010431:Pentoxifylline