Safety and Tolerability of GalNAc3-Conjugated Antisense Drugs Compared to the Same-Sequence 2'-O-Methoxyethyl-Modified Antisense Drugs: Results from an Integrated Assessment of Phase 1 Clinical Trial Data.

The triantennary N-acetylgalactosamine (GalNAc3) cluster has demonstrated the utility of receptor-mediated uptake of ligand-conjugated antisense drugs targeting RNA expressed by hepatocytes. GalNAc3-conjugated 2'-O-methoxyethyl (2'MOE) modified antisense oligonucleotides (ASOs) have demonstrated a higher potency than the unconjugated form to support lower doses for an equivalent pharmacological effect. We utilized the Ionis integrated safety database to compare four GalNAc3-conjugated and four same-sequence unconjugated 2'MOE ASOs. This assessment evaluated data from eight randomized placebo-controlled dose-ranging phase 1 studies involving 195 healthy volunteers (79 GalNAc3 ASO, 24 placebo; 71 ASO, 21 placebo). No safety signals were identified by the incidence of abnormal threshold values in clinical laboratory tests for either ASO group. However, there was a significant increase in mean alanine transaminase levels compared with placebo in the upper dose range of the unconjugated 2'MOE ASO group. The mean percentage of subcutaneous injections leading to local cutaneous reaction was 30-fold lower in the GalNAc3-conjugated ASO group compared with the unconjugated ASO group (0.9% vs. 28.6%), with no incidence of flu-like reactions (0.0% vs. 0.7%). Three subjects (4.2%) in the unconjugated ASO group discontinued dosing. An improvement in the overall safety and tolerability profile of GalNAc3-conjugated 2'MOE ASOs is evident in this comparison of short-term clinical data in healthy volunteers.

Predicting the pharmacokinetics and pharmacodynamics of antisense oligonucleotides: an overview of various approaches and opportunities for PBPK/PD modelling.

INTRODUCTION: Advances in research and development (R&D) have enabled many approvals of antisense oligonucleotides (ASOs). Its administration expanded from systemic to local for treating various diseases, where predicting target tissue exposures and pharmacokinetics (PK) and pharmacodynamics (PD) in human can be critical. AREAS COVERED: A literature search for PBPK/PD models of ASOs was conducted using PubMed and Embase (to 1 April 2023). ASO PK and PD in animals and humans and modeling approaches including physiologically based (PB) are summarized; and relevance and impacts of PBPK/PD modeling are assessed. EXPERT OPINION: Allometric scaling and compartmental PK/PD modeling have been successful to predict human ASO PK/PD, addressing most R&D needs. Understanding tissue distribution of ASOs can be crucial for their efficacy and safety especially for intrathecal (IT), pulmonary, or other local routes. PBPK/PD modeling is expected to improve such understanding, for which, efforts have been sporadic. However, developing a PBPK/PD model requires careful review of known biology/pharmacology and thoughtful experimental designs. Resulting models have the potential to predict target/specified tissue exposures and responses in human adults and pediatrics. Ultimately, a PBPK/PD modeling approach can lead to more efficient and rational clinical development, resulting in well-informed decision making and a shortened timeline.

Integrated Assessment of Phase 2 Data on GalNAc3-Conjugated 2'-O-Methoxyethyl-Modified Antisense Oligonucleotides.

Receptor-mediated delivery of an antisense oligonucleotide (ASO) using the ligand-conjugated antisense technology is establishing a new benchmark for antisense therapeutics. The triantennary N-acetylgalactosamine (GalNAc3) cluster is the first conjugated ligand to yield a marked increase in ASO potency for RNA targets expressed by hepatocytes, compared to the unconjugated form. In this study, we present an integrated safety assessment of data available from randomized, placebo-controlled, phase 2 studies for six GalNAc3-conjugated 2'-O-methoxyethyl (2'MOE)-modified ASOs. The total study population included 642 participants (130 placebo; 512 ASO) with up to 1 year of exposure. The primary measures were the incidence of signals from standardized laboratory tests and the mean test results over time. The GalNAc3-conjugated ASOs were well tolerated with no class effect identified across all doses tested compared to placebo. These results extend prior observations from phase 1 studies, now with treatment up to 1 year.

Assessment of the Effect of Organ Impairment on the Pharmacokinetics of 2'-MOE and Phosphorothioate Modified Antisense Oligonucleotides.

The pharmacokinetics (PK) of 2'-O-methoxyethyl and phosphorothioate antisense oligonucleotides (ASOs), with or without N-acetyl galactosamine conjugation, have been well characterized following subcutaneous or intravenous drug administration. However, the effect of organ impairment on ASO PK, primarily hepatic or renal impairment, has not yet been reported. ASOs distribute extensively to the liver and kidneys, where they are metabolized slowly by endo- and exonucleases, with minimal renal excretion as parent drug (<1%-3%). This short review evaluated the effect of organ impairment on ASO PK using 3 case studies: (1) a phase 1 renal impairment study evaluating a N-acetyl galactosamine-conjugated ASO in healthy study participants and study participants with moderate renal impairment, (2) a phase 2 study evaluating an unconjugated ASO in patients with end-stage renal disease; and (3) a phase 3 study evaluating an unconjugated ASO, which included patients with mild hepatic or renal impairment. Results showed that patients with end-stage renal disease had a mild increase (≈34%) in total plasma exposure, whereas mild or moderate renal impairment showed no effect on plasma PK. The effect of hepatic impairment on ASO PK could not be fully evaluated due to lack of data in moderate and severe hepatic impairment study participants. Nonetheless, available data suggest that mild hepatic impairment had no effect on ASO exposure.

Assessment of efficacy and safety of volanesorsen for treatment of metabolic complications in patients with familial partial lipodystrophy: Results of the BROADEN study: Volanesorsen in FPLD; The BROADEN Study.

BACKGROUND: Volanesorsen, an antisense oligonucleotide, is designed to inhibit hepatic apolipoprotein C-III synthesis and reduce plasma apolipoprotein C-III and triglyceride concentrations. OBJECTIVE: The present study assessed efficacy and safety of volanesorsen in patients with familial partial lipodystrophy (FPLD) and concomitant hypertriglyceridemia and diabetes. METHODS: BROADEN was a randomized, placebo-controlled, phase 2/3, 52-week study with open-label extension and post-treatment follow-up periods. Patients received weekly subcutaneous volanesorsen 300 mg or placebo. The primary endpoint was percent change from baseline in fasting triglycerides at 3 months. Secondary endpoints included relative percent change in hepatic fat fraction (HFF), visceral adiposity, and glycated hemoglobin levels. RESULTS: Forty patients (11 men, 29 women) were enrolled, majority of whom were aged <65 years (mean, 47 years) and White. Least squares mean (LSM) percent change in triglycerides from baseline to 3 months was -88% (95% CI, -134 to -43) in the volanesorsen group versus -22% (95% CI, -61 to 18) in the placebo group, with a difference in LSM of -67% (95% CI, -104 to -30; P=0.0009). Volanesorsen induced a significant LSM relative reduction in HFF of 53% at month 12 versus placebo (observed mean [SD]: 9.7 [7.65] vs. 18.0 [8.89]; P=0.0039). No statistically significant changes were noted in body volume measurements (fat, liver, spleen, visceral/subcutaneous adipose tissue) or glycated hemoglobin. Serious adverse events in patients assigned to volanesorsen included 1 case each of sarcoidosis, anaphylactic reaction, and systemic inflammatory response syndrome. CONCLUSION: In BROADEN, volanesorsen significantly reduced serum triglyceride levels and hepatic steatosis in patients with FPLD.

Population pharmacokinetic/pharmacodynamic modelling of eplontersen, an antisense oligonucleotide in development for transthyretin amyloidosis.

AIMS: Transthyretin-mediated amyloidosis is a progressive and fatal disease caused by the build-up of misfolded transthyretin (TTR) protein. Eplontersen is a triantennary N-acetyl galactosamine (GalNAc3)-conjugated antisense oligonucleotide targeting TTR messenger ribonucleic acid (mRNA) to inhibit production of both variant and wild-type TTR. We aimed to develop a population pharmacokinetic/pharmacodynamic (PK/PD) model for eplontersen and to evaluate the impact of covariates on exposure and response. METHODS: Plasma eplontersen and serum TTR concentration data were obtained from two phase 1 studies in healthy volunteers (ClinicalTrials.gov: NCT03728634, NCT04302064). Model development was conducted using a nonlinear mixed-effects approach. RESULTS: Eplontersen PK was well described by a two-compartment model. Evaluation of demographics identified significant covariates of lean body mass on clearance and body weight on intercompartmental clearance and volumes of distribution. Population PK modelling showed the absorption rate was 29.6% greater with injection into the abdomen versus the arm. The typical population terminal elimination half-life was 25.5 days. Serum TTR was well described by an indirect response model with inhibition of TTR production by eplontersen. Maximum fractional inhibition (Imax ) was 0.970 (0.549%RSE) and the half maximal inhibitory concentration (IC50 ) was 0.0283 ng/ml (13.3%RSE). Simulations showed subjects with lower weight had higher exposure (AUC, Cmax ), while higher Cmax was observed when comparing site of administration (ratio abdomen/arm = 1.18), but differences in exposure did not significantly impact response at evaluated doses. CONCLUSION: The exposure-response relationship of eplontersen was well characterised by the PKPD model. Weight and injection site were found to affect systemic exposure, but this effect does not seem to result in clinically relevant variation in response.

Phase 2 Study of the Factor XI Antisense Inhibitor IONIS-FXIRx in Patients With ESRD.

INTRODUCTION: Patients with end-stage renal disease (ESRD) requiring hemodialysis (HD) have an increased risk of thrombotic events and bleeding. Antisense reduction of factor XI (FXI) with IONIS-FXIRx is a novel strategy that may safely reduce the risk of thrombotic events. METHODS: This multicenter study enrolled 49 patients receiving HD in 2 parts. First, 6 participants (pharmacokinetics [PK] cohort) received 1 open-label 300 mg dose of IONIS-FXIRx both before and after HD. Subsequently, 43 participants were treated in a double-blind, randomized design with 200 mg or 300 mg IONIS-FXIRx or placebo for 12 weeks. The PK, pharmacodynamics (PD), and adverse events of IONIS-FXIRx were evaluated (ClinicalTrials.gov: NCT02553889). RESULTS: The PK of IONIS-FXIRx was consistent with previous studies and similar whether injected before or after HD. No accumulation of IONIS-FXIRx was observed after repeat administration. By day 85, mean levels of FXI activity fell 56.0% in the 200 mg group, 70.7% in the 300 mg group, and 3.9% in the placebo group compared with baseline. FXI antigen levels paralleled FXI activity. Dose-dependent prolongation of activated partial thromboplastin time (aPTT) was observed, with no changes in international normalized ratio (INR). IONIS-FXIRx was not associated with drug-related serious adverse events. In the randomized phase of the study, major bleeding events occurred in 0 (0.0%; 200 mg), 1 (6.7%; 300 mg), and 1 (7.7%; placebo) patients and were not considered related to treatment. CONCLUSION: IONIS-FXIRx reduced FXI activity in patients with ESRD receiving HD. Further studies are needed to determine the benefit-risk profile of FXI as a therapeutic target for patients who require HD.

Apolipoprotein C-III reduction in subjects with moderate hypertriglyceridaemia and at high cardiovascular risk.

AIMS: Hypertriglyceridaemia is associated with increased risk of cardiovascular events. This clinical trial evaluated olezarsen, an N-acetyl-galactosamine-conjugated antisense oligonucleotide targeted to hepatic APOC3 mRNA to inhibit apolipoprotein C-III (apoC-III) production, in lowering triglyceride levels in patients at high risk for or with established cardiovascular disease. METHODS AND RESULTS: A randomized, double-blind, placebo-controlled, dose-ranging study was conducted in 114 patients with fasting serum triglycerides 200-500 mg/dL (2.26-5.65 mmol/L). Patients received olezarsen (10 or 50 mg every 4 weeks, 15 mg every 2 weeks, or 10 mg every week) or saline placebo subcutaneously for 6-12 months. The primary endpoint was the percent change in fasting triglyceride levels from baseline to Month 6 of exposure. Baseline median (interquartile range) fasting triglyceride levels were 262 (222-329) mg/dL [2.96 (2.51-3.71) mmol/L]. Treatment with olezarsen resulted in mean percent triglyceride reductions of 23% with 10 mg every 4 weeks, 56% with 15 mg every 2 weeks, 60% with 10 mg every week, and 60% with 50 mg every 4 weeks, compared with increase by 6% for the pooled placebo group (P-values ranged from 0.0042 to <0.0001 compared with placebo). Significant decreases in apoC-III, very low-density lipoprotein cholesterol, non-high-density lipoprotein cholesterol, and apolipoprotein B were also observed. There were no platelet count, liver, or renal function changes in any of the olezarsen groups. The most common adverse event was mild erythema at the injection site. CONCLUSION: Olezarsen significantly reduced apoC-III, triglycerides, and atherogenic lipoproteins in patients with moderate hypertriglyceridaemia and at high risk for or with established cardiovascular disease. TRIAL REGISTRATION NUMBER: NCT03385239.

Antisense Inhibition of Angiotensinogen With IONIS-AGT-LRx: Results of Phase 1 and Phase 2 Studies.

Targeting angiotensinogen (AGT) may provide a novel approach to more optimally inhibit the renin-angiotensin-aldosterone system pathway. Double-blind, placebo-controlled clinical trials were performed in subjects with hypertension as monotherapy or as an add-on to angiotensin-converting enzyme inhibitors/angiotensin receptor blockers with IONIS-AGT-LRx versus placebo up to 2 months. IONIS-AGT-LRx was well tolerated with no significant changes in platelet count, potassium levels, or liver and renal function. IONIS-AGT-LRx significantly reduced AGT levels compared with placebo in all 3 studies. Although not powered for this endpoint, trends were noted in blood pressure reduction. In conclusion, IONIS-AGT-LRx significantly reduces AGT with a favorable safety, tolerability, and on-target profile. (A Study to Assess the Safety, Tolerability and Efficacy of IONIS-AGT-LRx; NCT04083222; A Study to Assess the Safety, Tolerability and Efficacy of IONIS-AGT-LRx, an Antisense Inhibitor Administered Subcutaneously to Hypertensive Subjects With Controlled Blood Pressure; NCT03714776; Safety, Tolerability, Pharmacokinetics, and Pharmacodynamics of Ionis AGT-LRx in Healthy Volunteers; NCT03101878).

Improvements in the Tolerability Profile of 2'-O-Methoxyethyl Chimeric Antisense Oligonucleotides in Parallel with Advances in Design, Screening, and Other Methods.

The development process of antisense oligonucleotides (ASOs) as therapeutic agents in humans has advanced through the implementation of chemical compound modifications as well as increasingly sophisticated toxicological preclinical screening techniques. The Ionis Integrated Safety Database was utilized to determine if advances in ASO screening and clinical lead identification methods have improved the tolerability profiles of 2'-O-methoxyethyl (2'MOE)-modified ASOs as a class, relative to the first 2'MOE ASO approved for use in humans, mipomersen. Tolerability was assessed by the incidence and percentage of subcutaneous doses leading to adverse events at the injection site or flu-like reactions (FLRs), as well as by the incidence of dose discontinuations due to these events. In randomized placebo-controlled phase 1 and phase 2 trials, the incidence of each measure of tolerability was lower in the test group of 12 ASOs (713 ASO-treated subjects) compared with the reference, mipomersen (266 ASO-treated subjects); with the most marked reduction in the incidence of FLRs (0.6% vs. 9.4%). A similar reduction in the incidence of dose discontinuation due to FLRs was also observed (0.2% vs. 0.9%). When compared with mipomersen, 8 of 12 ASOs showed significant improvements in their respective mean percentage of doses leading to adverse events at the injection site, whereas 7 ASOs showed a significant improvement in mean percentage of doses leading to FLRs. These results support an overall improvement in the tolerability profile in 2'MOE ASOs that entered development after mipomersen, in parallel with advances in the drug discovery screening process as well as the gains in clinical experience during development of each ASO.

PK/PD modeling of FXI antisense oligonucleotides to bridge the dose-FXI activity relation from healthy volunteers to end-stage renal disease patients.

IONIS-FXIRX (BAY2306001) is an antisense oligonucleotide that inhibits the synthesis of coagulation factor XI (FXI) and has been investigated in healthy volunteers and patients with end-stage renal disease (ESRD). FXI-LICA (BAY2976217) shares the same RNA sequence as IONIS-FXIRX but contains a GalNAc-conjugation that facilitates asialoglycoprotein receptor (ASGPR)-mediated uptake into hepatocytes. FXI-LICA has been studied in healthy volunteers and is currently investigated in patients with ESRD on hemodialysis. We present a model-informed bridging approach that facilitates the extrapolation of the dose-exposure-FXI relationship from IONIS-FXIRX to FXI-LICA in patients with ESRD and, thus, supports the selection of FX-LICA doses being investigated in patients with ESRD. A two-compartment pharmacokinetic (PK) model, with mixed first- and zero-order subcutaneous absorption and first-order elimination, was combined with an indirect response model for the inhibitory effect on the FXI synthesis rate via an effect compartment. This PK/pharmacodynamic model adequately described the median trends, as well as the interindividual variabilities for plasma drug concentration and FXI activity in healthy volunteers of IONIS-FXIRX and FXI-LICA, and in patients with ESRD of IONIS-FXIRX . The model was then used to predict dose-dependent steady-state FXI activity following repeat once-monthly doses of FXI-LICA in a virtual ESRD patient population. Under the assumption of similar ASGPR expression in patients with ESRD and healthy volunteers, doses of 40 mg, 80 mg, and 120 mg FXI-LICA are expected to cover the target range of clinical interest for steady-state FXI activity in the phase IIb study of FXI-LICA in patients with ESRD undergoing hemodialysis.

An oral antisense oligonucleotide for PCSK9 inhibition.

Inhibitors of proprotein convertase subtilisin/kexin type 9 (PCSK9) reduce low-density lipoprotein (LDL) cholesterol and are used for treatment of dyslipidemia. Current PCSK9 inhibitors are administered via subcutaneous injection. We present a highly potent, chemically modified PCSK9 antisense oligonucleotide (ASO) with potential for oral delivery. Past attempts at oral delivery using earlier-generation ASO chemistries and transient permeation enhancers provided encouraging data, suggesting that improving potency of the ASO could make oral delivery a reality. The constrained ethyl chemistry and liver targeting enabled by N-acetylgalactosamine conjugation make this ASO highly potent. A single subcutaneous dose of 90 mg reduced PCSK9 by >90% in humans with elevated LDL cholesterol and a monthly subcutaneous dose of around 25 mg is predicted to reduce PCSK9 by 80% at steady state. To investigate the feasibility of oral administration, the ASO was coformulated in a tablet with sodium caprate as permeation enhancer. Repeated oral daily dosing in dogs resulted in a bioavailability of 7% in the liver (target organ), about fivefold greater than the plasma bioavailability. Target engagement after oral administration was confirmed by intrajejunal administration of a rat-specific surrogate ASO in solution with the enhancer to rats and by plasma PCSK9 and LDL cholesterol lowering in cynomolgus monkey after tablet administration. On the basis of an assumption of 5% liver bioavailability after oral administration in humans, a daily dose of 15 mg is predicted to reduce circulating PCSK9 by 80% at steady state, supporting the development of the compound for oral administration to treat dyslipidemia.

Interspecies Scaling of Human Clearance and Plasma Trough Exposure for Antisense Oligonucleotides: A Retrospective Analysis of GalNAc3-Conjugated and Unconjugated-Antisense Oligonucleotides.

It is well documented and generally accepted that human clearance (CL) of unconjugated single-strand antisense oligonucleotides (ASOs) can be directly predicted from monkeys by body weight (BW) on a mg/kg dose basis. However, the scaling for triantennary N-acetyl galactosamine (GalNAc3)-conjugated ASOs has not been fully established. In this study, we retrospectively analyzed pharmacokinetic data from 9 GalNAc3-conjugated and 12 unconjugated single-stranded ASOs (ten 2'-methoxyethyl and two 2', 4'-constrained ethyl ASOs) to identify an appropriate allometric scaling factor between the two species. In addition, we compared the trough plasma concentrations (Ctrough, a surrogate for tissue exposure) between monkeys and humans at comparable dose levels, aiming at predicting tissue distribution in humans from monkeys. Overall, the median plasma CL ratios (monkey CL/human CL) were 1.05 and 0.94 when CL was normalized by BW, as compared with 0.33 and 0.29 when CL was normalized by body surface area (BSA) for the 12 unconjugated and 9 GalNAc3-conjugated ASOs, respectively. Similarly, the median Ctrough ratios (Ctrough in monkeys/Ctrough in humans) were 0.96 and 1.71, respectively, when Ctrough was normalized by mg/kg dose as compared with 3.10 and 5.50 when Ctrough was normalized by mg/m2 dose for the same unconjugated and conjugated ASOs, respectively. Equivalent CL and dose-normalized plasma Ctrough between monkeys and humans suggest similar pharmacokinetic profiles and tissue distribution between the two species on a per kilogram BW basis. In conclusion, human CL and plasma Ctrough (a surrogate of tissue distribution) can be directly predicted (1:1 or within twofold) from monkeys by BW on a mg/kg dose basis but these parameters can be under- or over-predicted by BSA on a mg/m2 dose basis. These results provide evidence for single species scaling from monkeys to humans directly and, thus, they can facilitate early human dose prediction in ASO drug development.

Design and Rationale of the Global Phase 3 NEURO-TTRansform Study of Antisense Oligonucleotide AKCEA-TTR-LRx (ION-682884-CS3) in Hereditary Transthyretin-Mediated Amyloid Polyneuropathy.

INTRODUCTION: AKCEA-TTR-LRx is a ligand-conjugated antisense (LICA) drug in development for the treatment of hereditary transthyretin amyloidosis (hATTR), a fatal disease caused by mutations in the transthyretin (TTR) gene. AKCEA-TTR-LRx shares the same nucleotide sequence as inotersen, an antisense medicine approved for use in hATTR polyneuropathy (hATTR-PN). Unlike inotersen, AKCEA-TTR-LRx is conjugated to a triantennary N-acetylgalactosamine moiety that supports receptor-mediated uptake by hepatocytes, the primary source of circulating TTR. This advanced design increases drug potency to allow for lower and less frequent dosing. The NEURO-TTRansform study will investigate whether AKCEA-TTR-LRx is safe and efficacious, with the aim of improving neurologic function and quality of life in hATTR-PN patients. METHODS/DESIGN: Approximately 140 adults with stage 1 (independent ambulation) or 2 (requires ambulatory support) hATTR-PN are anticipated to enroll in this multicenter, open-label, randomized, phase 3 study. Patients will be assigned 6:1 to AKCEA-TTR-LRx 45 mg subcutaneously every 4 weeks or inotersen 300 mg once weekly until the prespecified week 35 interim efficacy analysis, after which patients receiving inotersen will receive AKCEA-TTR-LRx 45 mg subcutaneously every 4 weeks. All patients will then receive AKCEA-TTR-LRx through the remainder of the study treatment period. The final efficacy analysis at week 66 will compare the AKCEA-TTR-LRx arm with the historical placebo arm from the phase 3 trial of inotersen (NEURO-TTR). The primary outcome measures are between-group differences in the change from baseline in serum TTR, modified Neuropathy Impairment Score + 7, and Norfolk Quality of Life-Diabetic Neuropathy questionnaire. CONCLUSION: NEURO-TTRansform is designed to determine whether targeted delivery of AKCEA-TTR-LRx to hepatocytes with lower and less frequent doses will translate into clinical and quality-of-life benefits for patients with hATTR-PN. TRIAL REGISTRATION: The study is registered at ClinicalTrials.gov (NCT04136184) and EudraCT (2019-001698-10).

Hereditary transthyretin amyloidosis with peripheral neuropathy (hATTR-PN for short) is a rare inherited condition. In hATTR-PN, a protein called transthyretin (TTR for short) builds up and damages nerves throughout the body. This neuropathy causes symptoms such as weakness, loss of sensation, and pain. Currently available medicines can slow disease progression, but researchers are looking for more effective treatments with fewer side effects. AKCEA-TTR-L_Rx is an investigational treatment for hATTR-PN. AKCEA-TTR-L_Rx prevents the liver from making TTR, reducing the amount that causes disease progression. It is similar to an existing treatment called inotersen, but designed for better delivery to the liver and is more potent. This article describes the NEURO-TTRansform study that will evaluate how effective AKCEA-TTR-L_Rx is for treating hATTR-PN. Around 140 adults with hATTR-PN from the USA, Canada, and Europe will be able to take part in this study. The study treatment period will be 85 weeks long. People will receive injections underneath the skin of either: AKCEA-TTR-L_Rx every 4 weeks, or Inotersen once a week for 35 weeks, followed by a switch to AKCEA-TTR-L_Rx every 4 weeks. People may continue to receive AKCEA-TTR-L_Rx after the study treatment period ends. In this study, researchers will compare results from people who received AKCEA-TTR-L_Rx to results from people who received no active ingredients (called placebo) in a similar study (called NEURO-TTR). Researchers will measure the differences in peoples': Neuropathy symptoms. Quality of life. TTR protein levels in the blood.

Antisense drug discovery and development technology considered in a pharmacological context.

When coined, the term "antisense" included oligonucleotides of any structure, with any chemical modification and designed to work through any post-RNA hybridization mechanism. However, in practice the term "antisense" has been used to describe single stranded oligonucleotides (ss ASOs) designed to hybridize to RNAswhile the term "siRNA" has come to mean double stranded oligonucleotides designed to activate Ago2. However, the two approaches share many common features. The medicinal chemistry developed for ASOs greatly facilitated the development of siRNA technology and remains the chemical basis for both approaches. Many of challenges faced and solutions achieved share many common features. In fact, because ss ASOs can be designed to activate Ago2, the two approaches intersect at this remarkably important protein. There are also meaningful differences. The pharmacokinetic properties are quite different and thus potential routes of delivery differ. ASOs may be designedto use a variety of post-RNA binding mechanismswhile siRNAs depend solely on the robust activity of Ago2. However, siRNAs and ASOs are both used for therapeutic purposes and both must be and can be understood in a pharmacological context. Thus, the goals of this review are to put ASOs in pharmacological context and compare their behavior as pharmacological agents to the those of siRNAs.

Ligand conjugated antisense oligonucleotide for the treatment of transthyretin amyloidosis: preclinical and phase 1 data.

AIMS: Amyloidogenic transthyretin (ATTR) amyloidosis is a fatal disease characterized by progressive cardiomyopathy and/or polyneuropathy. AKCEA-TTR-LRx (ION-682884) is a ligand-conjugated antisense drug designed for receptor-mediated uptake by hepatocytes, the primary source of circulating transthyretin (TTR). Enhanced delivery of the antisense pharmacophore is expected to increase drug potency and support lower, less frequent dosing in treatment. METHODS AND RESULTS: AKCEA-TTR-LRx demonstrated an approximate 50-fold and 30-fold increase in potency compared with the unconjugated antisense drug, inotersen, in human hepatocyte cell culture and mice expressing a mutated human genomic TTR sequence, respectively. This increase in potency was supported by a preferential distribution of AKCEA-TTR-LRx to liver hepatocytes in the transgenic hTTR mouse model. A randomized, placebo-controlled, phase 1 study was conducted to evaluate AKCEA-TTR-LRx in healthy volunteers (ClinicalTrials.gov: NCT03728634). Eligible participants were assigned to one of three multiple-dose cohorts (45, 60, and 90 mg) or a single-dose cohort (120 mg), and then randomized 10:2 (active : placebo) to receive a total of 4 SC doses (Day 1, 29, 57, and 85) in the multiple-dose cohorts or 1 SC dose in the single-dose cohort. The primary endpoint was safety and tolerability; pharmacokinetics and pharmacodynamics were secondary endpoints. All randomized participants completed treatment. No serious adverse events were reported. In the multiple-dose cohorts, AKCEA-TTR-LRx reduced TTR levels from baseline to 2 weeks after the last dose of 45, 60, or 90 mg by a mean (SD) of -85.7% (8.0), -90.5% (7.4), and -93.8% (3.4), compared with -5.9% (14.0) for pooled placebo (P < 0.001). A maximum mean (SD) reduction in TTR levels of -86.3% (6.5) from baseline was achieved after a single dose of 120 mg AKCEA-TTR-LRx . CONCLUSIONS: These findings suggest an improved safety and tolerability profile with the increase in potency achieved by productive receptor-mediated uptake of AKCEA-TTR-LRx by hepatocytes and supports further development of AKCEA-TTR-LRx for the treatment of ATTR polyneuropathy and cardiomyopathy.

Antisense Inhibition of Prekallikrein to Control Hereditary Angioedema.

Hereditary angioedema is characterized by recurrent and unpredictable episodes of subcutaneous and mucosal swelling that can be life threatening. IONIS-PKK-LRx is a ligand-conjugated antisense oligonucleotide designed for receptor-mediated delivery to hepatocytes. In a compassionate-use pilot study, two patients with severe bradykinin-mediated angioedema were initially administered weekly subcutaneous injections of the unconjugated parent drug, IONIS-PKKRx, for 12 to 16 weeks, after which they received IONIS-PKK-LRx at a dose of 80 mg every 3 to 4 weeks for 7 to 8 months. Treatment was accompanied by a reduction in the angioedema attack rate. (Funded by Amsterdam UMC.).

Vupanorsen, an N-acetyl galactosamine-conjugated antisense drug to ANGPTL3 mRNA, lowers triglycerides and atherogenic lipoproteins in patients with diabetes, hepatic steatosis, and hypertriglyceridaemia.

AIMS: Loss-of-function mutations in ANGPTL3 are associated with beneficial effects on lipid and glucose metabolism and reduced risk of coronary artery disease. Vupanorsen (AKCEA-ANGPTL3-L Rx ) is an N-acetyl galactosamine-conjugated antisense oligonucleotide targeted to the liver that selectively inhibits angiopoietin-like 3 (ANGPTL3) protein synthesis. METHODS AND RESULTS: This was a double-blind, placebo-controlled, dose-ranging, Phase 2 study. Patients (N =105) with fasting triglycerides >150 mg/dL (>1.7 mmol/L), type 2 diabetes, and hepatic steatosis were treated for 6 months with 40 or 80 mg every 4 weeks (Q4W), or 20 mg every week (QW) of vupanorsen, or placebo given subcutaneously. The primary efficacy endpoint was per cent change in fasting triglycerides from baseline at 6 months. Median baseline triglycerides were 2.84 mmol/L (252 mg/dL). Significant reductions in triglycerides of 36%, 53%, 47%, and in ANGPTL3 of 41%, 59%, 56%, were observed in the 40 mg Q4W, 80 mg Q4W, and 20 mg QW groups, respectively, compared with 16% reduction in triglycerides and 8% increase in ANGPTL3 in placebo. Compared with placebo, vupanorsen 80 mg Q4W reduced apolipoprotein C-III (58%), remnant cholesterol (38%), total cholesterol (19%), non-high-density lipoprotein cholesterol (HDL-C; 18%), HDL-C (24%), and apolipoprotein B (9%). There was no improvement in glycaemic parameters, or hepatic fat fraction. Treatment with vupanorsen was not associated with clinically significant changes in platelet counts, and the most common adverse events were those at the injection site, which were generally mild. CONCLUSION: Vupanorsen results in a favourable lipid/lipoprotein profile and provides a potential strategy for residual cardiovascular risk reduction.

Immunogenicity Assessment of Inotersen, a 2'-O-(2-Methoxyethyl) Antisense Oligonucleotide in Animals and Humans: Effect on Pharmacokinetics, Pharmacodynamics, and Safety.

Inotersen (TEGSEDI™) is a 2'-O-(2-methoxyethyl)-modified antisense oligonucleotide, intended for treating hereditary transthyretin (TTR) amyloidosis with polyneuropathy. The potential immunogenicity (IM) response to inotersen was evaluated in chronic nonclinical safety studies and the pivotal phase 2/3 clinical study. The evaluation was designed to assess the characteristics of antidrug antibodies (ADAs) and their effects on the pharmacokinetics, pharmacodynamics, clinical efficacy, and safety in animals and humans. No immunogenic response was observed after long-term treatment with inotersen in mice. In monkeys, the incidence rate of IM to inotersen appeared to be dose dependent, with 28.6%-50.0% of animals developing ADAs after 36 weeks of treatment. This was characterized as late onset (median onset of 185 days) with low titers (median titer of 8, or 400 if minimum required dilution of 50 is included). The overall incidence rate of patients who developed ADAs was 30% after 65 weeks of treatment with median onset of 203 days and median peak titer of 300. IM had minimal effect on plasma peak (Cmax) and total exposure (i.e. area under curve, AUC) of inotersen, but showed elevated plasma trough levels in both IM-positive animals and humans. However, ADAs had no effect on tissue exposure, TTR messenger RNA, or plasma TTR levels in the long-term monkey study. Similarly, IM showed no effect on plasma TTR levels in clinical studies. Thus, ADAs antibodies were binding antibodies, but not neutralizing antibodies. Finally, no association was observed between IM and toxicity findings (eg, platelet, complement activation, and histopathology findings) in the inotersen 9-month monkey study. In humans, no difference was observed in hematology, including platelets, kidney function tests, or incidence of adverse events between IM-positive and -negative patients. Overall, IM showed no effect on toxicity or safety of inotersen evaluated in both monkeys and humans. ClinicalTrials.gov Identifier: NCT01737398.