Dual Mechanisms of Action of Self-Delivering, Anti-HIV-1 FANA Oligonucleotides as a Potential New Approach to HIV Therapy

Abstract

Currently, the most effective and durable therapeutic option for HIV-1 infection is combination antiretroviral therapy (cART). Although cART is powerful and can delay viral evolution of drug resistance for decades, it is associated with limitations, including an inability to eradicate the virus and a potential for adverse effects. Therefore, it is imperative to discover new HIV therapeutic modalities. In this study, we designed, characterized, and evaluated the in vitro potency of 2′-deoxy-2′-fluoroarabinonucleotide (FANA) modified antisense oligonucleotides (ASOs) targeting highly conserved regions in the HIV-1 genome. Carrier-free cellular internalization of FANA ASOs resulted in strong suppression of HIV-1 replication in HIV-1-infected human primary cells. In vitro mechanistic studies suggested that the inhibitory effect of FANA ASOs can be attributed to RNase H1 activation and steric hindrance of dimerization. Using 5′-RACE PCR and sequencing analysis, we confirmed the presence of human RNase H1-mediated target RNA cleavage products in cells treated with FANA ASOs. We observed no overt cytotoxicity or immune responses upon FANA ASO treatment. Together, our results strongly suggest that FANA ASOs hold great promise for antiretroviral therapy. The dual ability of FANA ASOs to target RNA by recruiting RNase H1 and/or sterically blocking RNA dimerization further enhances their therapeutic potential.

KCTD15 is overexpressed in human childhood B-cell acute lymphoid leukemia

“…According to the findings of Mayumi Takahashi et al . 46 , we confirmed that FANA oligos penetrate into the cells very easily. Then, for our experiments, a concentration of 8 µM of 2′F-ANA was chosen.…”

Use of a Self-Delivering Anti-CCL3 FANA Oligonucleotide as an Innovative Approach to Target Inflammation after Spinal Cord Injury

“…One of the most important modification to the biological function of FANA ASO molecules is their ability to penetrate cells without the necessity of delivery vehicles required in transfection approaches. Our results revealed that FANA ASO molecules targeting CCL3 can indeed penetrate BMDMs in gymnotic fashion, as previously described for other cell types in vitro (Baby et al, 2020;Margiotta and Howard, 2020;Schmidt et al, 2019;Smaldone et al, 2019;Suwanmanee et al, 2019;Takahashi et al, 2019). We also demonstrated the successful suppression of CCL3 expression in BMDMs by CCL3 specific FANA ASOs.…”

“…FANA ASO mediated mRNA suppression was observed to the same extent in a model of Huntington's disease, where the suppression persisted for 12 weeks after continuous infusion for 2 weeks (Kordasiewicz et al, 2012). In vitro, sustained antisense activity has been described for other FANA ASO molecules for four to 13 days (Ferrari et al, 2006;Takahashi et al, 2019).…”

“…FANA ASO molecules targeting a variety of genes have been utilized in various models. For instance, FANA ASOs targeting the HIV-1 genome successfully suppressed viral replication in CD4+ T-cells in culture (Takahashi et al, 2019). In other work, FANA ASO molecules were used to confirm a direct link between Abi-1 loss and cell cycle activity in hematopoietic stem cells.…”

Battle for supremacy: nucleic acid interactions between viruses and cells

“…Lipid nanoparticles and viral vectors are being studied to ensure intracellular delivery. There has been some success in vitro using ASOs targeting HIV by targeting both viral and host factors and delivering them using a self-inactivating lentiviral vector system to HIV-infected human primary cells and producing strong suppression of HIV replication (103). HITS-CLIP found miR-122 binding directly to the 5'UTR of HCV protecting HCV RNA from degradation and promoting viral replication.…”