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

Abstract

Secondary damage after spinal cord injury (SCI) occurs due to a sequence of events after the initial injury, including exacerbated inflammation that contributes to increased lesion size and poor locomotor recovery. Thus, mitigating secondary damage is critical to preserve neural tissue and improve neurological outcome. In this work we examined the therapeutic potential of a novel antisense oligonucleotide with special chemical modifications (FANA ASO) for specifically inhibiting an inflammatory molecule in the injured spinal cord. The chemokine CCL3 plays a complex role in the activation and attraction of immune cells and is upregulated in the injured tissue after SCI. We used specific FANA ASO to inhibit CCL3 in a contusive mouse model of murine spinal cord injury. Our results show that self-delivering FANA ASO molecules targeting the chemokine CCL3 penetrate the spinal cord lesion site and suppress the expression of CCL3 transcripts. Furthermore, they reduce other pro-inflammatory cytokines such as TNF and IL-1β after SCI. In summary, we demonstrate for the first time the potential of FANA ASO molecules to penetrate the spinal cord lesion site to specifically inhibit CCL3, reducing pro-inflammatory cytokines 2 and improve functional recovery after SCI. This novel approach may be utilized in new treatment strategies for SCI and other pathological conditions of the central nervous system (CNS). Significance statement Preserving white matter tissue following SCI is an important therapeutic goal to minimize tissue damage and improve neurological outcome. Unresolved inflammation contributes to secondary tissue damage. Specifically targeting pro-inflammatory molecules, such as the chemokine CCL3, in the injured spinal cord can present technical challenges including tissue penetration, stability and efficacy of suppression. Here, we present the use of novel FANA ASO molecules as a feasible approach to specifically target proinflammatory molecules in the injured spinal cord.

2′-Fluoro-arabinonucleic Acid (FANA): A Versatile Tool for Probing Biomolecular Interactions

“…FANA ASOs, in particular, have been receiving increased attention. In the past year, four studies have highlighted the use of FANA ASOs in vivo to target Crohn’s disease, androgen receptor (AR)-RNA complexes implicated in cancer, inflammation after spinal cord injury, and angiogenesis . A fifth study showcased antibacterial FANA ASOs to tackle citrus greening disease, which affects citrus tree groves worldwide .…”

Host microRNA interactions with the SARS-CoV-2 viral genome 3’-untranslated region

“…Nucleic acid analogs have been employed as therapeutic agents in a vast majority of disorders, allowing for sequence specific targeting of genomic or proteomic targets. (42,43,56,57) Of particular interest in this study, we focused on 2'-fluoro-D-arabinonucleic acid analogs (FANAs), as these oligomers have been shown to be self-delivering in vivo and provide effective binding to their targets within both in vitro and in vivo systems. (42)(43)(44) Thus, FANA analogs designed to bind to the miR binding sites could be potentially used as a therapeutic option as a binding inhibitor to block miR binding on the SARS-CoV-2 genome 3'-UTR, relieving the wild-type miRs to perform their normal functions in the host.…”

“…Antisense oligonucleotides, like 2'-fluoro-D-arabinonucleic acids (FANAs), have been employed in several applications as binding inhibitors or in siRNA knockdowns, and have been theorized and developed to be effective oligos due to their ability to self-deliver in vivo and prove effective in many in vitro systems. (42)(43)(44) Often, antisense oligonucleotides have been employed for these purposes, designed as sequence specific targets, such as for siRNA-mediated knockouts or targeted binding. (42)(43)(44)(45) Thus, FANAs could sequencespecific target these binding interactions and allow for a potential mechanism of intervention.…”

“…(42)(43)(44) Often, antisense oligonucleotides have been employed for these purposes, designed as sequence specific targets, such as for siRNA-mediated knockouts or targeted binding. (42)(43)(44)(45) Thus, FANAs could sequencespecific target these binding interactions and allow for a potential mechanism of intervention. In this work, we show that the SARS-CoV-2 viral genome 3'-UTR binds host cellular miR-760-3p, miR-34a-5p, and miR-34b-5p, and that these binding interactions can be inhibited by intervention with antisense FANA oligonucleotides complementary to the genome miR binding sites.…”

Plant biomacromolecule delivery methods in the 21st century

“…FANA ASOs have also proven to be more stable than dsRNAs (Ferrari et al, 2006;Hunter et al, 2021) and have shown activity in both the cytoplasm and the nucleus (Liang X.-H. et al, 2017). Perhaps the most intriguing (and most useful) property of FANA ASOs is their capacity for self-delivery (gymnosis), independent of transfection agents (Soifer et al, 2011;Souleimanian et al, 2012;Pelisch et al, 2021). These properties have spurred great interest in utilizing FANA ASOs in a wide array of fields and are of particular interest in biopesticide applications (Hunter et al, 2021;Sandoval-Mojica et al, 2021), especially given the ability of FANA ASOs to target bacteria.…”