Assessment of an anti-HIV-1 combination gene therapy strategy using the antisense RNA and multimeric hammerhead ribozymes.
A combination gene therapy strategy using an ASPsi-gag antisense RNA (targeted against the packaging signal and the gag-coding region) and a multimeric hammerhead ribozyme Rz1-9 (targeted against nine sites within the env-coding region) or Rz1-14 (targeted against 14 sites within the 5' leader and the pro-, pol-, vif- and env-coding regions) was assessed for inhibiting HIV-1 replication. A murine stem cell virus (MSCV)-based MGIN vector was used to express Rz1-9, Rz1-14, ASPsi-gag, Rz1-9ASPsi-gag, or Rz1-14ASPsi-gag RNA in a CD4+ T lymphoid cell line. Stable transductants were shown to express similar levels of interfering RNA. HIV-1 replication was inhibited in cells expressing Rz1-9 and Rz1-14. Little inhibition of HIV-1 replication was observed in cells expressing ASPsi-gag RNA. Thus, the multimeric hammerhead ribozymes inhibit HIV-1 replication better than the antisense RNA. Inhibition of HIV-1 replication in cells expressing Rz1-9ASPsi-gag or Rz1-14ASPsi-gag RNA was worse than that obtained with the multimeric ribozymes alone. This result suggests that co-expression of antisense RNA decreases the anti-HIV potential of ribozymes. The multimeric ribozymes and the antisense RNA were designed to target different sites within the HIV-1 RNA. They are not expected to interact with each other. Neither are they expected to compete with each other for binding to the HIV-1 RNA. Instead, the antisense RNA binding to its (1553 nt-long) target site may have resulted in a decreased ribozyme turn over. Furthermore, since the antisense RNA/HIV-1 RNA hybrids are degraded by the cells, the co-expressed antisense RNA may have led to ribozyme degradation.
Ramezani
Ali
A
Department of Medical Genetics and Microbiology, Faculty of Medicine, University of Toronto, Toronto, Ontario M5S 3E2, Canada.
Ma
Xue Zhong
XZ
Ameli
Masoud
M
Arora
Alka
A
Joshi
Sadhna
S
D015496:CD4-Positive T-Lymphocytes / Q000821:virology; D015316:Genetic Therapy; D005822:Genetic Vectors; D015658:HIV Infections / Q000628:therapy; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology*; D008979:Moloney murine leukemia virus / Q000235:genetics; D016372:RNA, Antisense / Q000235:genetics / Q000378:metabolism*; D016337:RNA, Catalytic / Q000235:genetics* / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014161:Transduction, Genetic; D014779:Virus Replication
D016372:RNA, Antisense; D016337:RNA, Catalytic