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| Title: | Proviral HIV-genome-wide and pol-gene specific Zinc Finger Nucleases: Usability for targeted HIV gene therapy |
| Authors: | Wayengera, Misaki |
| Keywords: | Zinc finger nuclease
HIV/AIDS
DNA
Lentiviral vectors |
| Issue Date: | 22-Jul-2011 |
| Publisher: | BioMed Central |
| Citation: | Wayengera, M., (2011). Proviral HIV-genome-wide and pol-gene specific Zinc Finger Nucleases: Usability for targeted HIV gene therapy. Theoretical Biology and Medical Modelling, 8(26) |
| Abstract: | Background: Infection with HIV, which culminates in the establishment of a latent
proviral reservoir, presents formidable challenges for ultimate cure. Building on the
hypothesis that ex-vivo or even in-vivo abolition or disruption of HIV-gene/genomeaction
by target mutagenesis or excision can irreversibly abrogate HIV’s innate fitness
to replicate and survive, we previously identified the isoschizomeric bacteria
restriction enzymes (REases) AcsI and ApoI as potent cleavers of the HIV-pol gene (11
and 9 times in HIV-1 and 2, respectively). However, both enzymes, along with others
found to cleave across the entire HIV-1 genome, slice (SX) at palindromic sequences
that are prevalent within the human genome and thereby pose the risk of host
genome toxicity. A long-term goal in the field of R-M enzymatic therapeutics has
thus been to generate synthetic restriction endonucleases with longer recognition
sites limited in specificity to HIV. We aimed (i) to assemble and construct zinc finger
arrays and nucleases (ZFN) with either proviral-HIV-pol gene or proviral-HIV-1 wholegenome
specificity respectively, and (ii) to advance a model for pre-clinically testing
lentiviral vectors (LV) that deliver and transduce either ZFN genotype.
Methods and Results: First, we computationally generated the consensus sequences
of (a) 114 dsDNA-binding zinc finger (Zif) arrays (ZFAs or ZifHIV-pol) and (b) two zincfinger
nucleases (ZFNs) which, unlike the AcsI and ApoI homeodomains, possess
specificity to >18 base-pair sequences uniquely present within the HIV-pol gene
(ZifHIV-polFN). Another 15 ZFNs targeting >18 bp sequences within the complete HIV-1
proviral genome were constructed (ZifHIV-1FN). Second, a model for constructing
lentiviral vectors (LVs) that deliver and transduce a diploid copy of either ZifHIV-polFN
or ZifHIV-1FN chimeric genes (termed LV- 2xZifHIV-polFN and LV- 2xZifHIV-1FN,
respectively) is proposed. Third, two preclinical models for controlled testing of the
safety and efficacy of either of these LVs are described using active HIV-infected
TZM-bl reporter cells (HeLa-derived JC53-BL cells) and latent HIV-infected cell lines.
Conclusion: LV-2xZifHIV-polFN and LV- 2xZifHIV-1FN may offer the ex-vivo or even invivo
experimental opportunity to halt HIV replication functionally by directly
abrogating HIV-pol-gene-action or disrupting/excising over 80% of the proviral HIV DNA from latently infected cells. |
| URI: | http://www.tbiomed.com/content/8/1/26
http://hdl.handle.net/123456789/1915 |
| ISSN: | 1742-4682 |
| Appears in Collections: | Research Articles (Bio-Medical)
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| wayengera-chs-res.pdf | | 371Kb | Adobe PDF | View/Open |
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