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Nucleic Acids as Therapeutic Targets and Agents
Published in David E. Thurston, Ilona Pysz, Chemistry and Pharmacology of Anticancer Drugs, 2021
Discovered independently by the groups of Takeshi Imanishi and Jesper Wengel in the late 1990s, the exclusive rights were secured by Exiqon A/S, a Danish biotech company in 1997 marketed long-RNA GapmeR LNATM oligomers designed using advanced computer algorithms to ensure that they act as potent RNase H-activating antisense oligonucleotides both in cell lines and in vivo.
Lipid Nanocarriers for Oligonucleotide Delivery to the Brain
Published in Carla Vitorino, Andreia Jorge, Alberto Pais, Nanoparticles for Brain Drug Delivery, 2021
Andreia F. Jorge, Santiago Grijalvo, Alberto Pais, Ramón Eritja
Once internalised, ASOs modulate target mRNA activity through two main mechanisms: (i) ribonuclease H (RNAse H)-dependent mechanism [22] and (ii) nondegradative steric blocking mechanisms [23]. In the degradative mechanism, RNAse H enzyme binds to the mRNA-ASO heteroduplex through a binding domain inserted on the N terminus of the protein and catalyses the cleavage of the mRNA strand. In humans, the specific enzyme recruited is RNAse H1, and it is ubiquitously found in the nucleus, mitochondria and cytoplasm [22]. The cleaved debris of mRNA is then processed by the typical cellular degradation pathways, while the intact DNA is again available to hybridise with an additional target mRNA [24]. Endonuclease-mediated degradation is particularly attractive for the treatment of neurological diseases in which the aetiology is known to be dependent on protein dysregulation or accumulation. The high number of preclinical and clinical studies which employ ASOs designed for RNAse activation validates the interest in the field [25, 26]. However, since RNAse-H is a required mediator, it should be considered a rate-limiting step for ASO activity. As alternative to the use of the catalytic activity of endonucleases, ASOs with intrinsic enzymatic activity can be synthetically designed, as, for instance, DNAzymes or ribozymes [27]. The number of modifications on the ASO backbone may also interfere in their activity. The triggered mechanism(s) by which ASOs exert their antisense activity are thus dependent on ASO chemistry and design. RNase H-mediated ODNs often include internucleotide linkage modifications such as PS, phosphonoacetate (PACE), and boranophosphate [28–30] (Fig. 8.1). Sugar modifications are generally considered incompatible with RNase H-mediated gene silencing. However, they can be implemented in a ‘gapmer’ design, where the unmodified central DNA core guarantees high affinity to RNAse H, while their overhangs can be successfully protected with 2’-O-Me, 2/-O-(2-methoxyethyl) (2’-MOE), bicyclic locked nucleic acid (LNA) and 2’-deoxy-2’-fluoro-(3-D-arabino (2’-F-ANA) sugar modifications [31–33] (Fig. 8.1). Gapmers modified with 2’-MOE are currently tested clinically for the treatment of amyotrophic lateral sclerosis (ALS) [34].
The Promising Therapeutic Potential of Oligonucleotides for Pulmonary Fibrotic Diseases
Published in Expert Opinion on Drug Discovery, 2023
Divyani Paul, Madelyn H Miller, Josh Born, Shayak Samaddar, Huanzhen Ni, Hugo Avila, Venkata R. Krishnamurthy, Kannan Thirunavukkarasu
Three generations of ASO sequence design have been proposed. The first generation includes all PS DNA ASOs, used in lengths of 14–25 nucleotides (nts) long. The second generation was the MOE-DNA ‘Gapmer,’ where nucleotides 1–5 were 2′-MOE, nucleotides 6–10 were comprised of DNA, and nucleotides 11–15 were 2′-MOE, such that a gap of DNA existed around the scissile bond to induce RNAse H cleavage. This improved potency, stability, and durability over the first generation. Further exploration has led to design choices to arrive at the 2.5/3rd generation cEt/LNA Gapmers (Figure 3C) [41]. This Gapmer design increased the potency by 20× time and has since become a staple in design choice by Ionis Pharmaceuticals. ION-827359 from Ionis is an inhalable Gapmer ASO formulation being developed for CF to assess the hyperactivity of ENaCs, though this trial has been paused due to safety concerns. The ASO sequence has the (S)-cEt modification that provides excellent target affinity and nuclease resistance. It binds to αENaC pre-mRNA via complementarity to the fourth intronic region to recruit RNAse-H1 for cleavage [42]. The ASO was evaluated in a Phase 1/2a study involving healthy volunteers and CF patients. Single (3–100 mg) and multiple dosage (10–75 mg, 5 doses, weekly) of the drug were shown to be well tolerated with low systemic exposure [43]. The inhaled formulation was absorbed rapidly and led to a dose-dependent reduction in ENaC mRNA levels in the airway epithelial cells.
Antisense oligonucleotide therapeutics in clinical trials for the treatment of inherited retinal diseases
Published in Expert Opinion on Investigational Drugs, 2020
Kanmin Xue, Robert E. MacLaren
An ASO therapy targeting the P23 H mutation in RHO-adRP (QR-1123, ProQR Therapeutics) is currently under evaluation through a multi-center phase I/II clinical trial (ClinicalTrials.gov ID: NCT04123626). QR-1123 has been designed to hybridize with the mutant P23 H mRNA and induce allele-selective knockdown via RNase H-mediated cleavage while sparing the normal RHO transcripts. The molecule is a gapmer ASO containing a central sequence of DNA nucleotides flanked by RNA residues on either side, which facilitates RNase H-mediated cleavage of the target mRNA across the DNA gap. It was first developed by Ionis Pharmaceuticals Inc (Carlsbad, California, USA) with proof-of-concept studies performed in a P23 H rat model, demonstrating slowing of photoreceptor degeneration and preservation of visual function [34]. The license was subsequently acquired by ProQR Therapeutics NV, who is conducting the current phase I/II trial. The study design includes an open-label group which will receive single dose-escalation therapy with the ASO, and a double-masked randomized group which will receive 3 monthly intravitreal injections of either ASO or sham. In total, 35 adult patients are being recruited and the planned follow-up is 12 months. The main objectives are to assess dose-related safety, and the effects on visual function (visual acuity, visual field, and FST) as well as retinal structure (OCT). Secondary outcomes also include changes in quality-of-life. Interim results may be expected within the next year.
New pharmacotherapies for genetic neuromuscular disorders: opportunities and challenges
Published in Expert Review of Clinical Pharmacology, 2019
Federica Ricci, Martina Vacchetti, Chiara Brusa, Liliana Vercelli, Chiara Davico, Benedetto Vitiello, Tiziana Mongini
It has been recently confirmed that the reduction of CUGexpRNA improves muscle strength in DM1 mice, suggesting that DM1 patients may benefit from elimination of toxic RNAs [82]. A phase 1/2a blinded, placebo-controlled study to assess safety, tolerability, and dose-range finding of ISIS 598769, an ASO drug administered subcutaneously, was conducted in adult patients and showed good safety profile (NCT02312011). It is a gapmer ASO, a DNA–RNA hybrid with target mRNA, that recruits RNase H and promotes degradation of mRNA [83]. In preclinical studies, systemic administration of ISIS 486178 induced rapid knockdown of CUG-repeat RNAs and corrected pathogenic features of the disease [84]. Unfortunately, after phase 1/2a trial results the company discontinued further development of the drug.