Inhibition of Human Immunodeficiency Virus Type 1 by Packageable, Multigenic Antisense RNA

Abstract

Viral-based vectors can provide an efficient delivery mechanism for stable expression of antisense RNA. To enhance and propagate the antiviral effect of antisense RNA, two novel human immunodeficiency virus type 1 (HIV-1)-based vector DNAs, designated as pMAG7 and pMAG19, were constructed which contained HIV-1 cis-acting packaging elements and produced multigenic HIV-1 antisense RNA that could target the entire pol, env, vif, vpu, vpr, rev, and tat and portions of gag and nef. The two DNAs were identical except that pMAG19 had additional gag coding sequences. Cotransfection of pMAG DNA and infectious, cloned HIV-1 DNA in 293 cells inhibited virus production (81%-98% reduction in reverse transcriptase activity) of various T cell-tropic and macrophage-tropic clade B isolates, such as NL4-3, YU-2, and JR-CSF. In addition, virion-associated pMAG antisense RNA was detected in residual virus particles produced by pNL4-3 in the presence of pMAG7 DNA, and the antisense sequences were stably transferred by infection of 174 x CEM cells. The results suggest that pMAG DNA may confer broad protection against HIV-1 by reducing initial virus burden due to antisense RNA and subsequent virus spread by propagation of antisense sequences along with wild-type virus.

Antisense-Mediated Inhibition of Human Immunodeficiency Virus (HIV) Replication by Use of an HIV Type 1-Based Vector Results in Severely Attenuated Mutants Incapable of Developing Resistance

“…Several strategies have been employed, including targeting of the HIV coreceptor to prevent infection and spread (21), inhibition of regulatory genes like the packaging sequence, tar, and primer binding site (6), or targeting of structural or nonstructural viral gene products like rev, envelope, virulence factors, etc. (6,10,19,26,27,40,44,47). Importantly, the ability of antisense to inhibit HIV replication in vivo was demonstrated in a monkey model given autologous T cells modified with a vector containing antisense against the tat/rev gene prior to challenge (10).…”

Inducible Expression of the ΔNGFr/F12Nef Fusion Protein as a New Tool for Anti-Human Immunodeficiency Virus Type 1 Gene Therapy

“…These include the expression of either HIV-1 trans-dominant proteins (i.e., Rev, Tat) (Liu et al, 1994;Fox et al, 1995;Plavec et al, 1997;Rossi et al, 1997;Fraiser et al, 1998;Hamm et al, 1999;Mautino et al, 2001), HIV-1 antisense sequences (Lavigne and Thierry, 1997;Chadwick and Lever, 2000;Shahabuddin and Khan, 2000), ribozymes targeting HIV-1 sequences (for a review, see Rossi, 2000), or intrabodies against HIV-1 proteins (Mhashilkar et al, 1995(Mhashilkar et al, , 1999. We isolated and characterized an HIV-1 nef allele (F12nef) whose expression in trans (D'Aloja et al, 1998) or in cis potently inhibits HIV-1 replication.…”

Selection and Characterization of Human Immunodeficiency Virus Type 1 Variants Resistant to Compounds that Inhibit Rev-RRE Function

“…Similar peptides are currently being studied in clinical trials. More novel approaches use either anti-sense RNA or miRNA to target the mRNA cargo of the Rev-RRE pathway(149). The anti-sense approach, although not fully understood, is exciting and involves the intracelluar "immunization" of HIV target cells with packagable inhibitor RNAs that can be transported to other host cells.…”