Limits...
Nanotherapeutics Using an HIV-1 Poly A and Transactivator of the HIV-1 LTR-(TAR-) Specific siRNA.

Mahajan SD, Aalinkeel R, Reynolds JL, Nair B, Sykes DE, Law WC, Ding H, Bergey EJ, Prasad PN, Schwartz SA - Patholog Res Int (2011)

Bottom Line: We used a well-validated siRNA (si510) which targets the poly A/TAR (transactivator of the HIV-1 LTR) site and suppresses viral replication.Our results suggest that the QR-si510 HIV-1 siRNA nanoplex is not only effective in delivering siRNA, but also in suppressing HIV-1 viral replication for a longer time period.HIV-1 nanotherapeutics can thus enhance systemic bioavailability and offer multifunctionality.

View Article: PubMed Central - PubMed

Affiliation: Division of Allergy, Immunology, and Rheumatology, Department of Medicine, University at Buffalo, The State University of New York, 640 Ellicott Street, Room 444 Innovation Center, Buffalo, NY 14203, USA.

ABSTRACT
HIV-1 replication can be efficiently inhibited by intracellular expression of an siRNA targeting the viral RNA. We used a well-validated siRNA (si510) which targets the poly A/TAR (transactivator of the HIV-1 LTR) site and suppresses viral replication. Nanotechnology holds much potential for impact in the field of HIV-1 therapeutics, and nanoparticles such as quantum rods (QRs) can be easily functionalized to incorporate siRNA forming stable nanoplexes that can be used for gene silencing. We evaluated the efficacy of the QR-si510 HIV-1 siRNA nanoplex in suppressing viral replication in the HIV-1-infected monocytic cell line THP-1 by measuring p24 antigen levels and gene expression levels of HIV-1 LTR. Our results suggest that the QR-si510 HIV-1 siRNA nanoplex is not only effective in delivering siRNA, but also in suppressing HIV-1 viral replication for a longer time period. HIV-1 nanotherapeutics can thus enhance systemic bioavailability and offer multifunctionality.

No MeSH data available.


Related in: MedlinePlus

The dose-dependent kinetics of QR-si510 siRNA nanoplex on HIV-1-infected THP-1 cells. THP-1 monocytic cultures were treated with lipofectamine transfected scrambled siRNA (5–100 nM), Lipofectamine transfected si510 HIV-1 siRNA (5–100 nM) and  the QR-si510 siRNA (5–100 nM) nanoplex for a 48 hr time period and the p24 production was measured in the supernatants at the end of the incubation period using a commercially available p24 ELISA kit. Untransfected HIV-1-infected THP-1 and the non-HIV-1-infected THP-1 cells (nonvirus control) were the experimental controls. The lipofectamine was used as per manufacturer's protocol. Our results show that 10 nM of the QR-si510 HIV-1siRNA nanoplex was optimal in achieving almost 85% suppression of viral replication. The results shown are mean ± SD of 3 separate experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3108557&req=5

fig3: The dose-dependent kinetics of QR-si510 siRNA nanoplex on HIV-1-infected THP-1 cells. THP-1 monocytic cultures were treated with lipofectamine transfected scrambled siRNA (5–100 nM), Lipofectamine transfected si510 HIV-1 siRNA (5–100 nM) and the QR-si510 siRNA (5–100 nM) nanoplex for a 48 hr time period and the p24 production was measured in the supernatants at the end of the incubation period using a commercially available p24 ELISA kit. Untransfected HIV-1-infected THP-1 and the non-HIV-1-infected THP-1 cells (nonvirus control) were the experimental controls. The lipofectamine was used as per manufacturer's protocol. Our results show that 10 nM of the QR-si510 HIV-1siRNA nanoplex was optimal in achieving almost 85% suppression of viral replication. The results shown are mean ± SD of 3 separate experiments.

Mentions: The results of our dose response experiments (Figure 3) show a significant decrease in HIV-1 p24 production in HIV-1-infected THP-1 cells transfected with the QR-si510 HIV-1siRNA nanoplexes as compared to HIV-1-infected THP-1 cells transfected with a commercially available transfection reagent Lipofectamine (Invitrogen, Carlsbad, CA). No significant differences in p24 levels was observed when THP-1 cells were treated with different concentration of si510 HIV-1siRNA which ranged from 5 nM to 100 nM. Comparisons between the untransfected control and the QR-si510 HIV-1 siRNA nanoplex transfected THP-1 cells show a significant decrease in p24 production. The p24 levels in cells treated with 5, 10, 40, 80, and 100 nM concentrations of si510 HIV-1 siRNA were 159 ± 9.6 pg/mL, 111 ± 10.9 pg/mL, 102 ± 11.3 pg/mL, 89 ± 8.3 pg/mL and 91 ± 9.6 pg/mL, respectively, that corresponded to a 79.05% decrease (P < .001), 84.63% decrease (P < .001), 85.65% decrease (P < .001), 88.15% decrease (P < .0001) and 87.69% decrease (P < .0001) in viral replication as measured by p24 production. While the concentrations of the si510 HIV-1 siRNA varies from 5–100 nM, the QR concentration in the nanoplexes was constant and was 200 pg. In our time kinetics experiments we used the optimal 10 nM concentration of si510 HIV-1 siRNA. Comparisons between the untransfected control and the QR-si510 HIV-1 siRNA nanoplex transfected THP-1 cells at 12, 24, 48, 96 and 1 week posttransfection showed a significant decrease in p24 production as early as 12 hrs posttransfection (Figure 4). The p24 levels in THP-1 cells at 12, 24, 48, 96 hr, and 1 week posttransfection were 114 ± 10.5 pg/mL, 122 ± 11.2 pg/mL, 76 ± 9.4 pg/mL, 72 ± 8.7 pg/mL and 64  ±  6.3 pg/mL, respectively, and corresponded to a 85.33% decrease (P < .0001), 83.93% decrease (P < .0001), 90.04% decrease (P < .0001), 89.79% decrease (P < .0001) and 91.00% decrease (P < .0001) in viral replication as measured by p24 production. The QR concentration in the nanoplex was 200 pg. The results of our time kinetics experiments show that 48 hrs posttransfection, a 90% suppression of viral replication was observed (Figure 3) and this suppression was observed even 1 week posttransfection. The 48 hr time point was determined to be the optimal time for transfection with the nanoplex, as a 90% decrease in p24 production was observed at that time point.


Nanotherapeutics Using an HIV-1 Poly A and Transactivator of the HIV-1 LTR-(TAR-) Specific siRNA.

Mahajan SD, Aalinkeel R, Reynolds JL, Nair B, Sykes DE, Law WC, Ding H, Bergey EJ, Prasad PN, Schwartz SA - Patholog Res Int (2011)

The dose-dependent kinetics of QR-si510 siRNA nanoplex on HIV-1-infected THP-1 cells. THP-1 monocytic cultures were treated with lipofectamine transfected scrambled siRNA (5–100 nM), Lipofectamine transfected si510 HIV-1 siRNA (5–100 nM) and  the QR-si510 siRNA (5–100 nM) nanoplex for a 48 hr time period and the p24 production was measured in the supernatants at the end of the incubation period using a commercially available p24 ELISA kit. Untransfected HIV-1-infected THP-1 and the non-HIV-1-infected THP-1 cells (nonvirus control) were the experimental controls. The lipofectamine was used as per manufacturer's protocol. Our results show that 10 nM of the QR-si510 HIV-1siRNA nanoplex was optimal in achieving almost 85% suppression of viral replication. The results shown are mean ± SD of 3 separate experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3108557&req=5

fig3: The dose-dependent kinetics of QR-si510 siRNA nanoplex on HIV-1-infected THP-1 cells. THP-1 monocytic cultures were treated with lipofectamine transfected scrambled siRNA (5–100 nM), Lipofectamine transfected si510 HIV-1 siRNA (5–100 nM) and the QR-si510 siRNA (5–100 nM) nanoplex for a 48 hr time period and the p24 production was measured in the supernatants at the end of the incubation period using a commercially available p24 ELISA kit. Untransfected HIV-1-infected THP-1 and the non-HIV-1-infected THP-1 cells (nonvirus control) were the experimental controls. The lipofectamine was used as per manufacturer's protocol. Our results show that 10 nM of the QR-si510 HIV-1siRNA nanoplex was optimal in achieving almost 85% suppression of viral replication. The results shown are mean ± SD of 3 separate experiments.
Mentions: The results of our dose response experiments (Figure 3) show a significant decrease in HIV-1 p24 production in HIV-1-infected THP-1 cells transfected with the QR-si510 HIV-1siRNA nanoplexes as compared to HIV-1-infected THP-1 cells transfected with a commercially available transfection reagent Lipofectamine (Invitrogen, Carlsbad, CA). No significant differences in p24 levels was observed when THP-1 cells were treated with different concentration of si510 HIV-1siRNA which ranged from 5 nM to 100 nM. Comparisons between the untransfected control and the QR-si510 HIV-1 siRNA nanoplex transfected THP-1 cells show a significant decrease in p24 production. The p24 levels in cells treated with 5, 10, 40, 80, and 100 nM concentrations of si510 HIV-1 siRNA were 159 ± 9.6 pg/mL, 111 ± 10.9 pg/mL, 102 ± 11.3 pg/mL, 89 ± 8.3 pg/mL and 91 ± 9.6 pg/mL, respectively, that corresponded to a 79.05% decrease (P < .001), 84.63% decrease (P < .001), 85.65% decrease (P < .001), 88.15% decrease (P < .0001) and 87.69% decrease (P < .0001) in viral replication as measured by p24 production. While the concentrations of the si510 HIV-1 siRNA varies from 5–100 nM, the QR concentration in the nanoplexes was constant and was 200 pg. In our time kinetics experiments we used the optimal 10 nM concentration of si510 HIV-1 siRNA. Comparisons between the untransfected control and the QR-si510 HIV-1 siRNA nanoplex transfected THP-1 cells at 12, 24, 48, 96 and 1 week posttransfection showed a significant decrease in p24 production as early as 12 hrs posttransfection (Figure 4). The p24 levels in THP-1 cells at 12, 24, 48, 96 hr, and 1 week posttransfection were 114 ± 10.5 pg/mL, 122 ± 11.2 pg/mL, 76 ± 9.4 pg/mL, 72 ± 8.7 pg/mL and 64  ±  6.3 pg/mL, respectively, and corresponded to a 85.33% decrease (P < .0001), 83.93% decrease (P < .0001), 90.04% decrease (P < .0001), 89.79% decrease (P < .0001) and 91.00% decrease (P < .0001) in viral replication as measured by p24 production. The QR concentration in the nanoplex was 200 pg. The results of our time kinetics experiments show that 48 hrs posttransfection, a 90% suppression of viral replication was observed (Figure 3) and this suppression was observed even 1 week posttransfection. The 48 hr time point was determined to be the optimal time for transfection with the nanoplex, as a 90% decrease in p24 production was observed at that time point.

Bottom Line: We used a well-validated siRNA (si510) which targets the poly A/TAR (transactivator of the HIV-1 LTR) site and suppresses viral replication.Our results suggest that the QR-si510 HIV-1 siRNA nanoplex is not only effective in delivering siRNA, but also in suppressing HIV-1 viral replication for a longer time period.HIV-1 nanotherapeutics can thus enhance systemic bioavailability and offer multifunctionality.

View Article: PubMed Central - PubMed

Affiliation: Division of Allergy, Immunology, and Rheumatology, Department of Medicine, University at Buffalo, The State University of New York, 640 Ellicott Street, Room 444 Innovation Center, Buffalo, NY 14203, USA.

ABSTRACT
HIV-1 replication can be efficiently inhibited by intracellular expression of an siRNA targeting the viral RNA. We used a well-validated siRNA (si510) which targets the poly A/TAR (transactivator of the HIV-1 LTR) site and suppresses viral replication. Nanotechnology holds much potential for impact in the field of HIV-1 therapeutics, and nanoparticles such as quantum rods (QRs) can be easily functionalized to incorporate siRNA forming stable nanoplexes that can be used for gene silencing. We evaluated the efficacy of the QR-si510 HIV-1 siRNA nanoplex in suppressing viral replication in the HIV-1-infected monocytic cell line THP-1 by measuring p24 antigen levels and gene expression levels of HIV-1 LTR. Our results suggest that the QR-si510 HIV-1 siRNA nanoplex is not only effective in delivering siRNA, but also in suppressing HIV-1 viral replication for a longer time period. HIV-1 nanotherapeutics can thus enhance systemic bioavailability and offer multifunctionality.

No MeSH data available.


Related in: MedlinePlus