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Gapmer

A gapmer is a chimeric antisense oligonucleotide (ASO) consisting of a central DNA segment flanked by chemically modified RNA-like wings, engineered to hybridize with target messenger RNA (mRNA) and induce its degradation via RNase H-mediated cleavage for selective gene silencing.^[1] This design leverages the DNA gap—typically 7–12 deoxynucleotides long—to form a substrate for the RNase H enzyme, while the flanking wings (usually 2–7 nucleotides each) incorporate modifications such as 2'-O-methyl (2'-OMe), 2'-O-methoxyethyl (2'-MOE), or locked nucleic acids (LNAs) to confer resistance to nuclease degradation and enhance binding specificity and affinity.^[2] The gapmer approach originated in the 1980s as an evolution of early antisense technologies, with foundational work by researchers like Inoue et al. demonstrating improved efficacy through 2'-O-modified phosphorothioate ASOs.^[3] Gapmers function by binding complementary mRNA sequences in a sequence-specific manner, recruiting endogenous RNase H1 to cleave the RNA strand within the DNA-RNA hybrid, leading to mRNA degradation and reduced protein expression; this mechanism is particularly effective for targeting gain-of-function mutations in genetic disorders.^[1] Compared to other ASO classes like steric blockers or splice modulators, gapmers offer potent and durable knockdown (often lasting weeks after a single dose) due to their reliance on cellular RNase H machinery, though they can pose risks of off-target effects from non-specific hybridization or immune activation if not optimized.^[1] Chemical backbone modifications, such as phosphorothioate linkages throughout the molecule, further improve pharmacokinetics by increasing plasma protein binding and tissue distribution, particularly to the liver and kidney.^[2] Therapeutically, gapmers have advanced from research tools for target validation to approved drugs, including mipomersen (Kynamro), approved in 2013 but discontinued in 2019 for familial hypercholesterolemia, which targets apolipoprotein B mRNA,^[4] and inotersen (Tegsedi) for hereditary transthyretin amyloidosis, both utilizing 2'-MOE gapmer designs.^[5] More recent approvals, such as tofersen (Qalsody) for SOD1-related amyotrophic lateral sclerosis (ALS) in 2023^[1] and olezarsen (Tryngolza) for familial chylomicronemia syndrome in 2025,^[6] highlight their potential in monogenic diseases by reducing toxic protein levels. Ongoing innovations address design challenges like allele-specific silencing for heterozygous mutations, with mixmer variants (alternating RNA-DNA segments) showing up to 10-fold improved specificity over classical gapmers in models of muscular dystrophy.^[2] Despite successes, limitations including delivery barriers beyond the liver and potential hepatotoxicity necessitate continued refinements in chemistry and conjugation strategies.^[1]

Structure and Design

Core Architecture

A gapmer is a chimeric antisense oligonucleotide designed as a single-stranded molecule comprising a central DNA segment, known as the gap, typically consisting of 8-10 unmodified deoxynucleotides, flanked on both sides by shorter segments of modified nucleotides referred to as wings, usually 5-7 nucleotides each.^[7] This architecture allows the gapmer to hybridize to complementary target RNA while incorporating elements that enhance its therapeutic potential.^[2] The canonical gapmer motif is often represented in a 5-10-5 configuration, denoting five modified nucleotides in the 5' wing, a ten-nucleotide DNA gap, and five modified nucleotides in the 3' wing, though variations like 5-8-5 or 7-10-7 exist depending on the target and optimization needs.^[8] Throughout the entire structure, a phosphorothioate (PS) backbone modification replaces the non-bridging oxygen in the phosphate groups with sulfur, conferring resistance to nuclease degradation and improving pharmacokinetic properties such as cellular uptake and tissue distribution.^[7] The basic structural notation is thus 5'-wing-DNA gap-wing-3', where the wings are chemically altered to boost hybridization stability without interfering with the gap's function.^[2] The central DNA gap provides a deoxyribonucleotide stretch that permits access by RNase H1 enzyme, while the flanking wings, often incorporating modifications such as locked nucleic acid (LNA) or 2'-O-methoxyethyl (2'-MOE) residues, increase the binding affinity to the target mRNA through enhanced thermodynamic stability of the duplex.^[7] This balanced design ensures effective target engagement in vivo, distinguishing gapmers from other antisense formats like fully modified oligonucleotides.^[8]

Chemical Modifications

Gapmers are chimeric antisense oligonucleotides (ASOs) featuring specific chemical modifications to enhance binding affinity, nuclease resistance, and compatibility with RNase H1 enzymes. Early ASOs relied solely on phosphorothioate (PS) backbone modifications for basic stability, but these uniform designs suffered from limited target affinity and off-target effects. The evolution to modern chimeric gapmers incorporated sugar and base modifications in the flanking regions, balancing high-affinity binding with a central DNA-like gap for enzymatic cleavage, as exemplified in designs like the 5-10-5 motif where wings flank a 10-nucleotide DNA gap.^[9] The backbone of gapmers typically employs full PS linkages, where non-bridging oxygen atoms in the phosphodiester bonds are replaced by sulfur atoms. This modification imparts resistance to degradation by extracellular and intracellular nucleases, extending the half-life of the ASO in biological fluids compared to unmodified phosphodiester backbones. PS linkages also facilitate cellular uptake via protein binding but can contribute to immune activation if not optimized.^[10]^[11] In the wing regions, sugar modifications such as locked nucleic acid (LNA), 2'-O-methoxyethyl (2'-MOE), or 2'-fluoro (2'-F) are introduced to increase the thermal stability of the RNA-DNA hybrid duplex. LNA constrains the ribose sugar in a C3'-endo conformation via a methylene bridge between the 2'-O and 4'-C atoms, boosting the melting temperature (Tm) by 4-8°C per substitution in DNA-RNA duplexes and enabling shorter ASOs with high specificity. 2'-MOE replaces the 2'-OH with a methoxyethyl group, enhancing Tm by approximately 2°C per modification while improving nuclease resistance and reducing toxicity relative to unmodified RNA. Similarly, 2'-F substitutions, where fluorine replaces the 2'-OH, provide a Tm increase of about 2.5°C per nucleotide and confer substantial stability against exonucleases, though they require careful positioning to avoid hepatotoxicity.^[12]^[13]^[14] The central gap region consists of unmodified or minimally modified DNA nucleotides, typically with only PS backbone alterations, to maintain a deoxyribose-like conformation essential for recruitment and activation of RNase H1. This DNA segment, often 8-12 nucleotides long, ensures substrate recognition by the enzyme while avoiding steric hindrance from sugar modifications that could inhibit cleavage.^[15] Representative hybrid designs include LNA-DNA-LNA gapmers, where LNA wings flank a PS-DNA gap to achieve potent hybridization with minimal length, and MOE-DNA-MOE constructs, such as those in approved therapeutics, which combine moderate affinity gains with favorable pharmacokinetic profiles. These chimeric architectures represent a significant advancement over early PS-only ASOs, allowing tailored optimization of stability and target engagement.^[9]^[16]

Mechanism of Action

Target Recognition

Gapmers recognize their target mRNA through sequence-specific hybridization, primarily via Watson-Crick base pairing between the antisense oligonucleotide and complementary RNA sequences. The central DNA gap region forms the core of this duplex, while the flanking modified wings provide high-affinity anchoring that stabilizes the overall structure and facilitates initial binding. This design ensures that the gapmer can effectively invade secondary structures in the target mRNA, positioning the DNA-RNA hybrid for subsequent enzymatic recruitment.^[17]^[18] Sequence design principles for gapmers emphasize lengths of 18-25 nucleotides to balance potency, specificity, and manufacturability, typically comprising a central DNA gap of 8-10 nucleotides flanked by 3-5 modified nucleotides on each wing. Targeting strategies prioritize regions such as exon-intron junctions or the 3' untranslated region (3' UTR) to enhance specificity, as these sites often minimize cross-reactivity with alternative isoforms or non-target transcripts while maintaining accessibility for hybridization. For instance, designs avoiding highly conserved coding sequences in the 3' UTR can reduce unintended binding to homologous mRNAs.^[19]^[18] Chemical modifications, particularly locked nucleic acid (LNA) in the wings, play a crucial role in mitigating off-target binding by improving mismatch discrimination. LNA incorporation increases the thermal stability of perfectly matched duplexes while disproportionately destabilizing those with single nucleotide mismatches, often by 3-4 kcal/mol depending on the mismatch type and context (e.g., enhanced discrimination for C·C or A·G mismatches). This selective affinity allows gapmers to distinguish subtle sequence variations, such as single nucleotide polymorphisms, thereby reducing non-specific interactions across the transcriptome.^[20]^[18] Thermodynamic considerations guide gapmer optimization through free energy (ΔG°) calculations for duplex formation, often using nearest-neighbor parameters adapted from RNA:RNA models to predict hybridization stability at physiological temperatures. The wings contribute significantly to the overall ΔG°, with LNA modifications raising the melting temperature (Tm) by approximately 3-8°C per residue, thereby lowering the free energy of binding for matched targets while amplifying destabilization for mismatches (ΔΔG° up to 3.4 kcal/mol). This wing-driven stability ensures efficient target capture without excessive off-target hybridization, as mismatches in wing regions are less tolerated than in the central gap.^[21]^[20]

RNase H-Mediated Degradation

Upon hybridization of the gapmer antisense oligonucleotide (ASO) to its complementary target mRNA, the central DNA gap region forms an RNA-DNA heteroduplex that serves as a substrate for endogenous RNase H enzymes, primarily recruiting RNase H1 to initiate degradation.^[22] This recruitment occurs in both the nucleus and cytoplasm, where RNase H1 binds to the hybrid with high specificity, recognizing the DNA-like phosphodiester backbone in the gap while the flanking modified wings protect the ASO from nucleases.^[22] The process is rate-limited by RNase H1 availability, typically taking around 40 minutes after hybridization to achieve cleavage.^[22] RNase H1 catalyzes endonucleolytic hydrolysis of the RNA strand within the DNA-RNA hybrid, cleaving the phosphodiester bonds via a divalent metal ion-dependent mechanism that activates a water nucleophile.^[23] This results in RNA fragments bearing 5'-phosphate and 3'-hydroxyl termini, leaving the DNA strand of the gapmer intact for potential multiple rounds of activity or recycling.^[23] The enzyme exhibits strict substrate specificity for RNA-DNA hybrids, with cleavage efficiency influenced by hybrid length (optimal at 8-12 base pairs in the gap) and minimal activity on RNA-RNA duplexes or single-stranded RNA.^[22] The cleaved mRNA fragments are rapidly processed and degraded by cellular exonucleases through intrinsic RNA turnover pathways, leading to substantial reduction in target mRNA levels and subsequent protein knockdown, often achieving 70-90% efficiency in therapeutic contexts.^[24] This degradation is 2- to 4-fold faster than the natural mRNA decay rate, ensuring potent gene silencing.^[22] In contrast to RNase H2, which is chromatin-associated and primarily functions in DNA replication and repair without participating in ASO-mediated RNA cleavage, RNase H1 is the key enzyme for cytoplasmic and nuclear actions in gapmer therapeutics due to its localization and hybrid-cleavage proficiency.^[25]

Advantages

Potency and Specificity

Gapmers demonstrate high potency in silencing target genes, frequently achieving subnanomolar IC₅₀ values in cell culture assays, such as 0.4 nM for locked nucleic acid (LNA)-modified gapmers targeting VR1 mRNA, owing to their enhanced binding affinity and efficient RNase H-mediated cleavage of the RNA-DNA hybrid.^[26] This allows for substantially lower dosing compared to first-generation antisense oligonucleotides, as the catalytic nature of RNase H degradation enables multiple turnover events per gapmer molecule. The specificity of gapmers is bolstered by their flanking wing regions, which incorporate modifications like 2'-O-methoxyethyl or LNA that stabilize the duplex with perfect-match targets while destabilizing mismatched hybrids, resulting in over 100-fold discrimination between fully complementary sequences and those with a single nucleotide mismatch.^[27] For instance, rationally designed gapmers targeting single-nucleotide polymorphisms exhibit this pronounced selectivity, minimizing off-target effects in allele-specific applications.^[27] In comparison to phosphorothioate-only ASOs, which typically require higher concentrations for modest knockdown (e.g., IC₅₀ values around 70 nM), gapmers achieve 80-95% reduction in target protein expression at 10-50 nM doses in cellular models.^[26]^[28] This superior efficiency, coupled with low immune activation due to reduced protein binding from the modified wings, supports robust in vivo gene knockdown in diverse tissues including the liver and central nervous system.^[8]^[29]

Stability Enhancements

The phosphorothioate (PS) backbone modification in gapmers replaces the non-bridging oxygen in the phosphate backbone with sulfur, conferring substantial resistance to degradation by exonucleases and endonucleases that rapidly break down unmodified DNA oligonucleotides. This enhancement significantly prolongs the stability of gapmers in biological environments, with intracellular and tissue half-lives typically ranging from 2 to 4 weeks, compared to hours for unmodified counterparts.^[10]^[30] The flanking wing regions of gapmers, often incorporating locked nucleic acid (LNA) or 2'-O-methoxyethyl (MOE) modifications, provide additional protection against serum nucleases, further bolstering overall durability. LNA-modified wings, in particular, create a rigid bicyclic structure that shields the oligonucleotide from enzymatic attack, resulting in up to a 10-fold increase in serum half-life relative to standard DNA oligonucleotides. MOE wings similarly contribute by sterically hindering nuclease access while maintaining compatibility with the central DNA gap for RNase H recruitment.^[31]^[32] Pharmacokinetic profiles of gapmers reflect these stability gains, with rapid distribution from plasma to tissues leading to high accumulation in the liver and kidney, where concentrations can persist for weeks. Clearance occurs slowly via urinary excretion of short-chain metabolites and proteolytic/nucleolytic degradation within tissues, minimizing rapid elimination and supporting sustained exposure. This pharmacokinetic behavior underpins the feasibility of infrequent dosing, such as weekly or monthly subcutaneous administrations, as demonstrated in approved gapmer therapeutics like mipomersen and volanesorsen.^[33]^[34]^[35]

Therapeutics

Mipomersen (Kynamro)

Mipomersen, marketed as Kynamro, represents the first gapmer antisense oligonucleotide approved for clinical use, targeting apolipoprotein B (ApoB) mRNA to address severe hypercholesterolemia. Developed by Isis Pharmaceuticals (now Ionis Pharmaceuticals) in collaboration with Genzyme, it received FDA approval on January 29, 2013, as an adjunct to maximally tolerated lipid-lowering therapy and diet for reducing low-density lipoprotein cholesterol (LDL-C), ApoB, total cholesterol, and non-HDL cholesterol in adults with homozygous familial hypercholesterolemia (HoFH), a rare genetic disorder characterized by extremely high LDL-C levels due to mutations in the LDL receptor gene.^[36]^[37]^[38] Structurally, mipomersen is a 20-nucleotide (20-mer) second-generation gapmer with a 5-10-5 design, featuring 2'-O-methoxyethyl (2'-MOE) modifications in the flanking wings for enhanced stability and a central deoxyribonucleotide gap to recruit RNase H for target degradation, all linked by a phosphorothioate backbone. It specifically binds to exon 20-21 of human ApoB mRNA in the liver, inducing RNase H-mediated cleavage and reducing ApoB-100 protein synthesis, which in turn decreases hepatic production of very low-density lipoprotein (VLDL) and circulating LDL particles. This mechanism results in LDL-C reductions of 25-40% in HoFH patients, with the recommended dosing regimen being 200 mg administered subcutaneously once weekly.^[15]^[39]^[40] In pivotal phase 3 clinical trials, such as the randomized, double-blind, placebo-controlled study involving 34 HoFH patients, mipomersen demonstrated a mean 36% reduction in LDL-C from baseline after 26 weeks, compared to a 13% increase in the placebo group, alongside significant decreases in ApoB (38%) and non-HDL-C (35%). However, the therapy carries a black box warning for hepatotoxicity, with elevations in alanine aminotransferase (ALT) ≥3 times the upper limit of normal (ULN) occurring in up to 12% of patients and increases in hepatic fat content observed via magnetic resonance imaging. Due to these safety concerns, including injection-site reactions and flu-like symptoms leading to high discontinuation rates, mipomersen has seen limited clinical adoption; marketing was discontinued worldwide, with the FDA withdrawing approval in 2019 as the product was no longer marketed, and it is no longer available as of 2025.^[41]^[42]^[4]^[43]

Volanesorsen (Waylivra)

Volanesorsen, marketed as Waylivra, is a second-generation antisense oligonucleotide (ASO) developed by Ionis Pharmaceuticals and its affiliate Akcea Therapeutics as the first targeted therapy for familial chylomicronemia syndrome (FCS), a rare genetic disorder characterized by severe hypertriglyceridemia and recurrent pancreatitis due to impaired lipoprotein lipase activity.^[44]^[45] As a 20-nucleotide gapmer with 2'-O-methoxyethyl (2'-MOE) modifications on the wings and phosphorothioate backbone linkages, it specifically binds to apolipoprotein C-III (ApoC-III) mRNA in the liver, recruiting RNase H enzymes to cleave the target mRNA and thereby reducing ApoC-III protein production, which inhibits triglyceride clearance.^[45]^[46] Prior to volanesorsen, no approved pharmacologic treatments existed for FCS, leaving patients reliant on strict dietary management to mitigate life-threatening complications.^[44] The European Medicines Agency (EMA) granted conditional marketing authorization for volanesorsen on May 3, 2019, as an adjunct to diet for adult patients with genetically confirmed FCS at high risk of pancreatitis who are unresponsive to standard triglyceride-lowering therapies.^[47] The recommended regimen involves 300 mg subcutaneous injections once weekly for the first three months, followed by every two weeks if triglyceride levels are adequately reduced, with further adjustments at six and nine months based on response and safety monitoring.^[47] In the pivotal phase 3 APPROACH trial, involving 66 adults with FCS, volanesorsen achieved a mean 77% reduction in plasma triglyceride levels from baseline at three months (from 2202 mg/dL to 485 mg/dL), compared to an 18% increase with placebo, while also lowering ApoC-III by 84%.^[48] Additionally, 77% of treated patients reached triglyceride levels below 750 mg/dL, versus 10% on placebo, demonstrating substantial efficacy in this ultra-rare population with historically untreatable severe hypertriglyceridemia exceeding 1000 mg/dL.^[48] A key challenge with volanesorsen is its association with thrombocytopenia, observed in up to 48% of patients in clinical trials, where platelet counts fell below 100,000 per microliter, though severe cases (<25,000 per microliter) were rare and reversible upon discontinuation.^[48]^[47] This class effect of 2'-MOE ASOs necessitates mandatory baseline and ongoing platelet monitoring every two to four weeks, with dose interruptions or reductions if counts drop below 75,000 per microliter, and permanent discontinuation if below 50,000 per microliter, to prevent bleeding risks.^[47] Other common adverse effects include injection-site reactions in over 60% of patients, but the overall risk-benefit profile supports its use in this high-need population under a conditional approval requiring post-marketing studies to confirm long-term safety and efficacy.^[48]^[47]

Inotersen (Tegsedi)

Inotersen, marketed as Tegsedi, is a 20-mer antisense oligonucleotide gapmer modified with phosphorothioate linkages and 2'-O-methoxyethyl (2'-MOE) wings flanking a central DNA gap, designed to target transthyretin (TTR) mRNA in the liver.^[15] By binding to complementary sequences on TTR mRNA, inotersen recruits RNase H to cleave the target RNA, thereby reducing TTR protein production and limiting the formation of amyloid deposits associated with hereditary transthyretin-mediated (hATTR) amyloidosis.^[49] This approach addresses the polyneuropathy manifestation of hATTR, a progressive neurodegenerative condition caused by mutant TTR misfolding and extracellular deposition.^[50] The U.S. Food and Drug Administration (FDA) approved inotersen in October 2018 for the treatment of polyneuropathy in adults with hATTR amyloidosis, marking it as the second approved therapy for this indication following patisiran.^[50] Administered as a 300 mg subcutaneous injection once weekly, inotersen's formulation supports stable delivery via this route, enabling outpatient use.^[50] In the pivotal phase 3 NEURO-TTR study, a randomized, double-blind, placebo-controlled trial involving 172 patients with stage 1 or 2 hATTR polyneuropathy, inotersen demonstrated a 50% slower progression of neurologic impairment compared to placebo, as measured by the modified Neuropathy Impairment Score plus 7 (mNIS+7) over 66 weeks.^[49] The study also showed significant improvements in quality of life, with serum TTR levels reduced by approximately 80% from baseline, correlating with decreased amyloid deposition potential.^[49] Post-approval, inotersen is distributed exclusively through the Tegsedi Risk Evaluation and Mitigation Strategy (REMS) program to mitigate serious risks, including thrombocytopenia and glomerulonephritis.^[50] The REMS requires prescriber certification, patient enrollment, and mandatory laboratory monitoring—such as weekly platelet counts and monthly serum creatinine with urinalysis—to detect early signs of these adverse events, which occurred in 20% and 3% of trial participants, respectively.^[50] This structured oversight ensures safe use while preserving access for eligible patients with hATTR polyneuropathy.

Tofersen (Qalsody)

Tofersen, marketed as Qalsody, is a 20-mer antisense oligonucleotide with a 5-10-5 phosphorothioate backbone modification, designed as a gapmer to bind SOD1 mRNA and induce its RNase H-mediated degradation in patients with amyotrophic lateral sclerosis (ALS) caused by superoxide dismutase 1 (SOD1) gene mutations.^[51] The U.S. Food and Drug Administration granted accelerated approval for tofersen on April 25, 2023, based on its reduction of neurofilament light chain (NfL) levels as a surrogate biomarker for neurodegeneration in SOD1-ALS adults.^[52] This approval marks the first targeted therapy for a specific genetic form of ALS, which accounts for about 2% of cases.^[53] Clinical data from the phase 1/2 trial demonstrated that intrathecal administration of tofersen at the 100 mg dose resulted in a mean 37% reduction in cerebrospinal fluid (CSF) SOD1 protein concentrations from baseline at day 85, compared to minimal change with placebo.^[54] The subsequent phase 3 VALOR trial, involving 137 participants, showed further biomarker reductions, including up to 40% decrease in CSF SOD1 in slower-progressing subgroups and a 60% reduction in plasma NfL levels versus 20% with placebo over 28 weeks, supporting target engagement despite no significant improvement in primary clinical endpoints like ALS Functional Rating Scale scores.^[55] Tofersen is dosed intrathecally at 100 mg (15 mL) with three loading doses every 14 days, followed by maintenance every 28 days, allowing direct central nervous system delivery to motor neurons.^[51] As of 2025, confirmation of clinical benefit is under evaluation in ongoing phase 3 trials, including the ATLAS study assessing tofersen in presymptomatic SOD1 mutation carriers to determine if early intervention delays ALS onset.^[56] Expanded access programs have been extended to provide tofersen to a broader cohort of early-stage SOD1-ALS patients outside clinical trials, enhancing availability in regions permitting such initiatives.^[57] By reducing toxic SOD1 protein accumulation, tofersen offers a pioneering disease-modifying approach for this monogenic ALS subset, potentially slowing neurodegeneration.^[58]

Safety and Toxicity

Common Adverse Effects

Gapmer antisense oligonucleotides, particularly those administered subcutaneously, frequently cause local injection site reactions including erythema, pain, tenderness, pruritus, and swelling, affecting 20-84% of patients.^[59] These reactions are generally mild to moderate, dose-dependent, and resolve without intervention, though they contribute to treatment discontinuation in a small subset of cases.^[60] Flu-like symptoms, such as pyrexia, chills, fatigue, arthralgia, and myalgia, arise early in therapy due to immune activation via Toll-like receptor (TLR) stimulation by the phosphorothioate backbone of gapmers, with incidences ranging from 10-30% across clinical evaluations.^[60] The phosphorothioate modifications enhance stability but can trigger innate immune responses contributing to these transient effects.^[61] Hematological adverse effects, notably thrombocytopenia, occur in TTR-targeting gapmers such as inotersen, where severe reductions in platelet count (grade 3 or higher) affect approximately 3% of patients and necessitate close monitoring.^[49] These events typically manifest within the first few months and may resolve with dose adjustment or supportive care.^[60] Hepatic transaminase elevations, including increases in alanine aminotransferase (ALT) and aspartate aminotransferase (AST), are observed in 12-16% of patients receiving liver-targeted gapmers like mipomersen, often correlating with hepatic fat accumulation but rarely progressing to severe injury.^[60] Routine liver function monitoring is recommended to manage these reversible changes.^[62]

Long-Term Risks and Mitigation

One of the primary long-term risks associated with gapmer antisense oligonucleotides stems from the accumulation of their phosphorothioate (PS) backbones in the kidney and reticuloendothelial system, which can lead to chronic nephrotoxicity, including glomerulonephritis and renal tubular degeneration.^[34] This buildup occurs due to the chemical stability of PS modifications, resulting in prolonged tissue retention and potential for sustained exposure.^[63] Clinical observations in patients receiving long-term gapmer therapy have reported glomerulonephritis as a rare but serious adverse event, often linked to immune complex deposition in renal tissues.^[64] Off-target effects represent another concern in extended gapmer use, where unintended interactions may alter alternative splicing patterns or trigger immune modulation, such as activation of the interferon response pathway.^[65] These effects arise from partial complementarity between the gapmer and non-target RNAs, potentially leading to widespread changes in gene expression.^[17] In central nervous system-targeted gapmers like tofersen, recent 2025 analyses have noted elevations in neuroinflammatory markers during prolonged treatment, suggesting possible late-onset contributions to neurotoxicity, though direct causation remains under investigation.^[66] To mitigate these risks, strategies include dose adjustments to minimize cumulative exposure while preserving efficacy, as higher doses exacerbate PS accumulation.^[67] N-acetylgalactosamine (GalNAc) conjugation enhances liver-specific targeting, reducing off-target renal uptake and attenuating nephrotoxicity in preclinical models by promoting clearance through productive endocytic pathways.^[68] Regular monitoring of renal function, platelet counts, and inflammatory markers is recommended for long-term users to enable early detection and intervention.^[30] Regulatory bodies have issued class-wide warnings for hepatotoxicity in gapmer-based therapies, with approximately half of FDA-approved antisense oligonucleotides carrying precautions or black-box alerts for liver enzyme elevations and steatosis due to prolonged PS exposure.^[69] Ongoing research focuses on developing safer backbone modifications, such as phosphorodithioate linkages or methylphosphonate alternatives, and on understanding sequence-chemistry interplay in toxicity (as of October 2025), to further reduce accumulation and off-target liabilities without compromising potency.^[11]^[70]

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Mipomersen (Kynamro) Approved by FDA - National Lipid Association
Jan 30, 2013 · On January 29, the US FDA approved mipomersen sodium injection as an adjunct to treat LDL-C, Apo B, total cholesterol, and non HDL-C in patients with ...
[36]
Kynamro wins FDA approval for homozygous familial ... - Healio
Jan 30, 2013 · The FDA has approved mipomersen sodium as an adjunct to lipid-lowering medications and a healthy diet to reduce LDL, apolipoprotein B, total ...
[37]
Emerging LDL therapies: Mipomersen—antisense oligonucleotide ...
Mipomersen was recently approved by the Food and Drug Administration for the treatment of homozygous familial FH. FH is caused by inherited genetic defects ...<|control11|><|separator|>
[38]
Mipomersen Dosage Guide + Max Dose, Adjustments - Drugs.com
Jun 30, 2025 · Usual Adult Dose for Hyperlipidemia: 200 mg subcutaneously once weekly on the same day each week.Dose Adjustments · Precautions · Other Comments
[39]
Mipomersen in Familial Hypercholesterolemia: An Update on Health ...
Feb 21, 2022 · Findings in the mipomersen arm demonstrated reductions in LDL-C of 36% from baseline compared to a 13% increase in LDL-C levels in the placebo ...
[40]
[PDF] This label may not be the latest approved by FDA. For current ...
In a subcutaneous carcinogenicity study in mice, mipomersen sodium was administered for up to 104 weeks at doses of 5, 20, 60 mg/kg/week. There were ...
[41]
Kynamro (mipomersen) FDA Approval History - Drugs.com
FDA approval history for Kynamro (mipomersen) used to treat High Cholesterol, Familial Homozygous. Supplied by Kastle Therapeutics.
[42]
Akcea and Ionis announce approval of WAYLIVRA® (volanesorsen ...
May 7, 2019 · WAYLIVRA is an antisense oligonucleotide drug designed by Ionis and co-developed by Akcea and Ionis to reduce the production of ApoC-III, a ...
[43]
[PDF] Assessment report - WAYLIVRA - European Medicines Agency
Feb 28, 2019 · ISIS 304801 (volanesorsen) was identified as the optimal 2′-MOE antisense inhibitor of human apoC-. III on the basis of its consistent and ...
[44]
Treatment with Volanesorsen, a 2 - PubMed Central
Volanesorsen is a second-generation antisense oligonucleotide (ASO) drug targeted to human APOC3 mRNA. The hybridization of volanesorsen to the cognate mRNA ...Aso Volanesorsen · Results · Concentration/effect...
[45]
Waylivra | European Medicines Agency (EMA)
Waylivra is indicated as an adjunct to diet in adult patients with genetically confirmed familial chylomicronemia syndrome (FCS) and at high risk for ...Missing: 2018 | Show results with:2018
[46]
Volanesorsen and Triglyceride Levels in Familial Chylomicronemia ...
Aug 7, 2019 · Another side effect was thrombocytopenia, which was defined as a platelet count below 140,000 per microliter. A recently reported natural ...
[47]
Inotersen Treatment for Patients with Hereditary Transthyretin ...
Jul 4, 2018 · Inotersen improved the course of neurologic disease and quality of life in patients with hereditary transthyretin amyloidosis.
[48]
[PDF] 1 FULL PRESCRIBING INFORMATION - accessdata.fda.gov
A total of 112 adult patients with polyneuropathy caused by hereditary transthyretin-mediated amyloidosis (hATTR) received TEGSEDI in Study 1 and 60 patients ...
[49]
[PDF] SPINRAZA (nusinersen) injection, for intrathecal use
See full prescribing information for. SPINRAZA. SPINRAZA (nusinersen) injection, for intrathecal use. Initial U.S. Approval: 2016. INDICATIONS AND USAGE.
[50]
Fluoroscopic guidance for intrathecal delivery of nusinersen in ...
Because antisense oligonucleotides do not cross the intact blood-brain barrier, nusinersen is administered by intrathecal injection. Treatment is comprised ...
[51]
[PDF] Spinraza – Prescribing Information
1 Loading doses followed by 12 mg (5 mL) once every 6 months. Post-lumbar puncture syndrome has also been observed after administration of SPINRAZA. 6.2.
[52]
Nusinersen versus Sham Control in Later-Onset Spinal Muscular ...
Feb 14, 2018 · The ENDEAR trial also showed greater improvements in infants who received treatment with nusinersen earlier in their disease course than in ...
[53]
Results from a phase 1 study of nusinersen (ISIS-SMNRx) in ...
In mouse models of SMA, nusinersen enhanced exon 7 inclusion, increased SMN protein production, and improved function. ... fold over a ∼3-fold increase in total ...
[54]
[PDF] This label may not be the latest approved by FDA. For current ...
Tofersen, an antisense oligonucleotide, is a 20-base residue (20-mer) 5-10-5 MOE gapmer mixed backbone oligonucleotide. Of the nineteen internucleotide ...
[55]
FDA approves treatment of ALS associated with a mutation in the ...
Apr 25, 2023 · FDA approved Qalsody (tofersen) to treat patients with amyotrophic lateral sclerosis (ALS) associated with a mutation in the superoxide dismutase 1 (SOD1) gene ...
[56]
Tofersen (Qalsody®) - Amyotrophic Lateral Sclerosis
Jul 31, 2025 · In April 2023, tofersen, marketed in the U.S. as Qalsody® (Biogen), was approved by the FDA for the treatment of SOD1-linked ALS. The SOD1 gene ...
[57]
Phase 1–2 Trial of Antisense Oligonucleotide Tofersen for SOD1 ALS
Jul 8, 2020 · A 25% reduction in the CSF SOD1 protein concentration was estimated to exceed both the SOD1 assay variability and the longitudinal ...
[58]
Trial of Antisense Oligonucleotide Tofersen for SOD1 ALS
Sep 21, 2022 · Tofersen reduced concentrations of SOD1 in CSF and of neurofilament light chains in plasma over 28 weeks but did not improve clinical end points and was ...
[59]
NCT04856982 | A Study of BIIB067 (Tofersen) Initiated in Clinically ...
The primary objective of this study is to evaluate the efficacy of tofersen in presymptomatic adult carriers of a superoxide dismutase 1 (SOD1) mutation ...
[60]
Tofersen/Qalsody - International Alliance of ALS/MND Associations
Since 2021, Biogen has “expanded eligibility for its ongoing early access program to all people with SOD1-ALS, in countries where such programs are permitted by ...
[61]
FDA approves QALSODY™ (tofersen) as the first treatment targeting ...
Accelerated approval marks a scientific advancement in treatment of superoxide dismutase 1 (SOD1)-amyotrophic lateral sclerosis (ALS); Approval based on ...
[62]
Injection site reactions after subcutaneous oligonucleotide therapy
When administered subcutaneously (sc), ONs cause a specific local reaction originating around the injection site, such as erythema, itching, discomfort and pain ...
[63]
Adverse Drug Reactions and Toxicity of the Food and Drug ... - NIH
The major safety concerns for the FDA-approved ASO drugs are severe hepatotoxicity, renal toxicity, and hypersensitivity reactions, which are commonly ...
[64]
Considerations in the Preclinical Assessment of the Safety of ...
... effects that can manifest in different ways in the clinic, including injection site reactions, flu-like symptoms, and thrombocytopenia [72–75]. Therefore ...
[65]
[PDF] lik - accessdata.fda.gov
KYNAMRO can cause elevations in transaminases (5.1). • Measure alanine aminotransferase (ALT), aspartate aminotransferase.
[66]
Off-target effects of oligonucleotides and approaches of preclinical ...
... Flu-like symptoms, such as fatigue, flu-like illness, chills, fever, joint pain, and myalgia, Liver injury, liver steatosis. Inotersen, SC, Vacuolation and ...
[67]
Nephrotoxicity of marketed antisense oligonucleotide drugs
It has also been observed that ONDs cause adverse effects on liver and kidney characterised by glomerulonephritis, tubular degeneration and increased protein ...
[68]
Block or degrade? Balancing on- and off-target effects of antisense ...
RNA-seq reveals activation of the interferon response by all ASOs and additional off-target effects on gene expression by the repeat gapmer. After having ...
[69]
Neurodegenerative and neuroinflammatory changes in SOD1-ALS ...
Apr 1, 2025 · We observed a significant decrease in neurofilament levels during Tofersen treatment, alongside an increase in neuroinflammatory markers.
[70]
Considerations in the Preclinical Assessment of the Safety of ...
Importantly, gapmers using both moderate-affinity MOE and high-affinity LNA have caused unexpected renal toxicity in humans that had not been detected or ...
[71]
GalNAc Conjugation Attenuates the Cytotoxicity of Antisense ...
These data further suggest that GalNAc conjugation can mitigate the nephrotoxic potential of AON sequences by direct protection of renal tubular cells. (A) ...Original Article · Results · Materials And Methods
[72]
Toxicity of Antisense Oligonucleotides is Determined by the ...
Oct 13, 2025 · In fact, about half of approved ASO drugs carry warnings or precautions for toxicities such as hypersensitivity reactions, liver damage, or ...<|separator|>