STUDY PROTOCOL

Design and Rationale of the Global Phase 3 NEURO-TTRansform Study of Antisense OligonucleotideAKCEA-TTR-LRx (ION-682884-CS3) in HereditaryTransthyretin-Mediated Amyloid Polyneuropathy

Teresa Coelho . Yukio Ando . Merrill D. Benson . John L. Berk . Marcia Waddington-Cruz . Peter J. Dyck .

Julian D. Gillmore . Sami L. Khella . William J. Litchy . Laura Obici . Cecilia Monteiro . Li-Jung Tai .

Nicholas J. Viney . Gustavo Buchele . Michela Brambatti . Shiangtung W. Jung . Louis St. L. O’Dea .

Sotirios Tsimikas . Eugene Schneider . Richard S. Geary . Brett P. Monia . Morie Gertz

Received: December 2, 2020 / Accepted: February 2, 2021 / Published online: February 26, 2021� The Author(s) 2021, corrected publication 2021

ABSTRACT

Introduction: AKCEA-TTR-LRx is a ligand-con-jugated antisense (LICA) drug in developmentfor the treatment of hereditary transthyretinamyloidosis (hATTR), a fatal disease caused bymutations in the transthyretin (TTR) gene.AKCEA-TTR-LRx shares the same nucleotide

sequence as inotersen, an antisense medicineapproved for use in hATTR polyneuropathy(hATTR-PN). Unlike inotersen, AKCEA-TTR-LRx

is conjugated to a triantennary N-acetylgalac-tosamine moiety that supports receptor-medi-ated uptake by hepatocytes, the primary sourceof circulating TTR. This advanced designincreases drug potency to allow for lower andless frequent dosing. The NEURO-TTRansformstudy will investigate whether AKCEA-TTR-LRx

is safe and efficacious, with the aim of improv-ing neurologic function and quality of life inhATTR-PN patients.Methods/Design: Approximately 140 adultswith stage 1 (independent ambulation) or 2(requires ambulatory support) hATTR-PN are

Supplementary Information The online versioncontains supplementary material available at https://doi.org/10.1007/s40120-021-00235-6.

C. Monteiro and L.-J. Tai were employees of IonisPharmaceuticals Inc. at the time of this work. L. St. L.O’Dea was an employee of Akcea Therapeutics at thetime of this work.

T. CoelhoDepartment of Neuroscience, Centro HospitalarUniversitario do Porto, Porto, Portugal

Y. AndoDepartment of Neurology, Graduate School ofMedical Sciences, Kumamoto University,Kumamoto, Japan

M. D. BensonDepartment of Pathology and Laboratory Medicine,Indiana University School of Medicine, Indianapolis,IN, USA

J. L. BerkAmyloidosis Center, Boston University School ofMedicine, Boston, MA, USA

M. Waddington-CruzNational Amyloidosis Referral Center-CEPARM,University Hospital, Federal University of Rio deJaneiro, Rio de Janeiro, Brazil

P. J. Dyck � W. J. LitchyDepartment of Neurology, Mayo Clinic, Rochester,MN, USA

J. D. GillmoreDivision of Medicine, National Amyloidosis Centre,University College London, London, UK

S. L. KhellaDepartment of Neurology, University ofPennsylvania, Philadelphia, PA, USA

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https://doi.org/10.1007/s40120-021-00235-6

anticipated to enroll in this multicenter, open-label, randomized, phase 3 study. Patients willbe assigned 6:1 to AKCEA-TTR-LRx 45 mg sub-cutaneously every 4 weeks or inotersen 300 mgonce weekly until the prespecified week 35interim efficacy analysis, after which patientsreceiving inotersen will receive AKCEA-TTR-LRx

45 mg subcutaneously every 4 weeks. Allpatients will then receive AKCEA-TTR-LRx

through the remainder of the study treatmentperiod. The final efficacy analysis at week 66will compare the AKCEA-TTR-LRx arm with thehistorical placebo arm from the phase 3 trial ofinotersen (NEURO-TTR). The primary outcomemeasures are between-group differences in thechange from baseline in serum TTR, modifiedNeuropathy Impairment Score ? 7, and NorfolkQuality of Life—Diabetic Neuropathyquestionnaire.Conclusion: NEURO-TTRansform is designed todetermine whether targeted delivery of AKCEA-TTR-LRx to hepatocytes with lower and less fre-quent doses will translate into clinical andquality-of-life benefits for patients with hATTR-PN.Trial Registration: The study is registered atClinicalTrials.gov (NCT04136184) and EudraCT(2019-001698-10).

PLAIN LANGUAGE SUMMARY

• Hereditary transthyretin amyloidosis withperipheral neuropathy (hATTR-PN for short)is a rare inherited condition.

– In hATTR-PN, a protein called transthyr-etin (TTR for short) builds up and dam-ages nerves throughout the body.

– This neuropathy causes symptoms suchas weakness, loss of sensation, and pain.

• Currently available medicines can slow dis-ease progression, but researchers are lookingfor more effective treatments with fewer sideeffects.

• AKCEA-TTR-LRx is an investigational treat-ment for hATTR-PN.

– AKCEA-TTR-LRx prevents the liver frommaking TTR, reducing the amount thatcauses disease progression.

– It is similar to an existing treatmentcalled inotersen, but designed for betterdelivery to the liver and is more potent.

• This article describes the NEURO-TTRans-form study that will evaluate how effectiveAKCEA-TTR-LRx is for treating hATTR-PN.

– Around 140 adults with hATTR-PN fromthe USA, Canada, and Europe will be ableto take part in this study.

– The study treatment period will be85 weeks long. People will receive injec-tions underneath the skin of either:

AKCEA-TTR-LRx every 4 weeks, orInotersen once a week for 35 weeks,followed by a switch to AKCEA-TTR-LRx

every 4 weeks.

– PeoplemaycontinuetoreceiveAKCEA-TTR-LRx after the study treatment period ends.

• In this study, researchers will compareresults from people who received AKCEA-TTR-LRx to results from people who receivedno active ingredients (called placebo) in asimilar study (called NEURO-TTR).

• Researchers will measure the differences inpeoples’:

L. ObiciAmyloidosis Research and Treatment Center,Fondazione IRCCS Policlinico San Matteo, Pavia,Italy

C. Monteiro � L.-J. Tai � N. J. Viney � G. Buchele �M. Brambatti � S. W. Jung � S. Tsimikas �E. Schneider � R. S. Geary � B. P. MoniaIonis Pharmaceuticals, Inc., Carlsbad, CA, USA

L. St. L. O’DeaAkcea Therapeutics, Boston, MA, USA

S. TsimikasVascular Medicine Program, Sulpizio CardiovascularCenter, Division of Cardiology, University ofCalifornia San Diego, La Jolla, CA, USA

M. Gertz (&)Division of Hematology, Department of InternalMedicine, Mayo Clinic, Rochester, MN, USAe-mail: gertz.morie@mayo.edu

376 Neurol Ther (2021) 10:375–389

– Neuropathy symptoms.– Quality of life.– TTR protein levels in the blood.

Keywords: AKCEA-TTR-Lrx; Antisenseoligonucleotide; Clinical trial design;Hereditary transthyretin-mediated amyloidpolyneuropathy; Phase 3 clinical trial

Key Summary Points

Why carry out this study?

Hereditary transthyretin amyloidosis(hATTR) is a fatal disease caused bymutations in the transthyretin (TTR) genethat result in the synthesis ofunstable TTR tetramers and consequentsystemic deposition of insoluble amyloidfibrils, which most often manifest ascardiomyopathy or polyneuropathy(hATTR-PN).

Although existing treatments can slow orhalt neurologic impairment associatedwith hATTR-PN, there remains a need forother disease-modifying agents that offerimproved efficacy, safety, andconvenience of use.

AKCEA-TTR-LRx is a novel antisenseoligonucleotide (ASO) designed forenhanced delivery to hepatocytes, theprimary site of TTR protein production,for increased potency and convenience ofuse to potentially translate into clinicaland quality-of-life benefits for patientswith hATTR-PN.

This paper describes the design andrationale of the NEURO-TTRansform trial,a multicenter, open-label, randomized,phase 3 study that aims to assess theefficacy and safety of AKCEA-TTR-LRx inapproximately 140 adult patients withstage 1 (independent ambulation) or 2(requiring ambulatory support) hATTR-PN.

DIGITAL FEATURES

This article is published with digital features,including a summary slide and a plain languagesummary, to facilitate understanding of thearticle. To view digital features for this article goto https://doi.org/10.6084/m9.figshare.13677805.

INTRODUCTION

Hereditary transthyretin amyloidosis (hATTR) isa progressive, irreversible, and fatal autosomaldominant disorder that manifests most com-monly as peripheral sensorimotor and auto-nomic axonal neuropathy or infiltrativecardiomyopathy [1–4]. hATTR is caused bytransthyretin (TTR) gene mutations that resultin the synthesis of unstable TTR tetramers [5, 6].TTR tetramers consisting of mutant TTRmonomers are more susceptible to tetramerdissociation and consequent misfolding of themonomer, resulting in aggregation into solubleoligomeric structures and insoluble TTR-derivedamyloid deposits [7, 8]. The accumulation ofTTR-derived amyloid fibrils in multiple organsystems results in a wide spectrum of clinicalmanifestations [1, 2]. TTR genotype–phenotypecorrelations have been reported for a predomi-nant clinical phenotype; for example, pV142Imutation carriers manifest preferentially withhATTR cardiomyopathy, whereas pV50Mmutation carriers manifest preferentially withhATTR polyneuropathy (hATTR-PN) [9–12].Additionally, factors that contribute to the rateof progression, such as geographic location, sexof transmitting parent, genetic anticipation,and genetic modifiers, are associated with thevariability of the clinical presentation [11–13].Estimates suggest that the global prevalence ofhATTR-PN is approximately 10,000 patients,although many individuals may be undiag-nosed [14]. The median age of onset of hATTR-PN varies geographically (from 33 years in Japanand Portugal to 56 years in Sweden), andpatients progress to death 6–12 years aftersymptom onset [1, 2, 15, 16].

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Clinical staging of hATTR-PN is often basedon ambulatory status, which assesses the pro-gression of sensorimotor axonal peripheralneuropathy severity. Disease metrics includethe Familial Amyloid Polyneuropathy or Cou-tinho stage scoring system, and the Polyneu-ropathy Disability (PND) Score [1]. For the

Coutinho scoring system, stage 1 is character-ized by independent ambulation, stage 2 ischaracterized by the need for unilateral orbilateral support for ambulation, and stage 3 ischaracterized by being wheelchair-bound orbedridden [1].

Table 1 Treatments for polyneuropathy of hATTR

Inotersen [24, 25] Patisiran [22, 23] Tafamidis[18, 29]

Diflunisal [20, 36]

Agent type ASO siRNA Stabilizer Nonsteroidal anti-

inflammatory drug

Indication Approved for ATTR-PN in

the United States and for

stage 1 or 2 hATTR-PN

in the European Union,

Canada, Brazil, and other

countriesa

Approved for ATTR-PN in

the United States,

Canada, and Japan and

for stage 1 or 2 hATTR-

PN in the European

Union, Brazil, and

Switzerland

Approved in

the European

Union (stage

1) and in over

40 countriesb

Off-label

Dose 284 mg subcutaneously

once weekly

0.3 mg/kg intravenously

every 3 weeks

20 mg orally

once daily

250–1000 mg daily

Premedication None Intravenous corticosteroid

Oral acetaminophen

Intravenous H1 blocker

Intravenous H2 blocker

None None

Precautions Vitamin A

supplementation

Vitamin A

supplementation

None None

Safety

monitoring

Thrombocytopenia

Glomerulonephritis

Liver function

Infusion-related reactions None Serious cardiovascular

thrombotic events,

myocardial infarction,

stroke, and serious

gastrointestinal adverse

events including bleeding,

ulceration, and

perforation of the

stomach or intestines

ASO antisense oligonucleotide, ATTR transthyretin amyloidosis, ATTR-PN transthyretin-mediated amyloid polyneu-ropathy, hATTR hereditary transthyretin amyloidosis, hATTR-PN hereditary transthyretin-mediated amyloid polyneu-ropathy, siRNA small interfering ribonucleic acida Iceland, Liechtenstein, Norway, United Kingdom (license issued based on European Medicines Agency)b Approved for ATTR-PN in over 40 countries, including Japan, European countries, Brazil, Mexico, Argentina, Israel,Russia, and South Korea

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Since the TTR protein is primarily producedby the liver, liver transplant has been the his-torical standard of care, acting to slow or haltdisease progression by replacing a liver thatproduces mutant TTR protein with one thatexpresses only wild-type TTR [1]. Owing todonor shortages and progressive end-organamyloid disease following liver transplantation,medical therapies have been pursued [1]. Overthe past 13 years, several pharmacologic thera-pies have been developed, including TTR-stabi-lizing agents that preserve the tetramericstructure and ribonucleic acid (RNA)-targetingtherapeutics that suppress TTR messenger RNA(mRNA) expression (Table 1). Oral tafamidismeglumine is a TTR-stabilizing agent that wasapproved for the treatment of familial amyloidpolyneuropathy (FAP) stage 1 ATTR-PN in 2011by the European Union, with subsequentapprovals in Japan and other countries [17–19].Diflunisal, a nonsteroidal anti-inflammatorydrug, has been shown to significantly reducethe progression of neurological impairmentamong patients with hATTR-PN but is notapproved for the treatment of hATTR-PN[20, 21]. With regard to RNA-targeting thera-peutics, two treatments have recently beenapproved in the United States, the EuropeanUnion, and other countries for patients withhATTR-PN: inotersen and patisiran [22–25].Inotersen is an antisense oligonucleotide (ASO)inhibitor of TTR production, which is adminis-tered at a dosage of 300 mg by subcutaneous(SC) injection once weekly [24, 25]. Patisiran is asmall interfering RNA (siRNA) inhibitor of TTRproduction, which is formulated in a lipidnanoparticle for administration at a dosage of0.3 mg/kg by intravenous infusion every3 weeks and requires premedication with a cor-ticosteroid [22, 23].

Inotersen is approved for the treatment ofadult patients with hATTR-PN [24, 25]. Inot-ersen binds to and induces ribonuclease H1-mediated degradation of TTR mRNA (wild-typeand all known mutations), thereby loweringTTR protein production [21]. NEURO-TTR was apivotal randomized, double-blind, placebo-controlled phase 2/3 study of inotersen 300 mg/week over 15 months in adult patients withstage 1 or stage 2 hATTR-PN [26]. Compared

with placebo, inotersen significantly improvedoutcomes on the modified Neuropathy Impair-ment Score ? 7 (mNIS ? 7) and patient-re-ported Norfolk Quality of Life—DiabeticNeuropathy (Norfolk QOL-DN) questionnaire[26]. The maximum mean reduction frombaseline in serum TTR was 74% [26]. The safetyprofile of inotersen was considered accept-able with regular safety laboratory assessmentsfor thrombocytopenia and glomerulonephritis[24–26]. Although the approved treatments canslow or halt the neurologic impairment associ-ated with hATTR-PN, there remains a need formore efficacious disease-modifying agents thatoffer a higher degree of TTR silencing, greaterconvenience of use, and a more favorable safetyprofile [26].

AKCEA-TTR-LRx (ION-682884) is a ligand-conjugated antisense (LICA) drug designed forpreferential delivery to hepatocytes, the pri-mary source of systemically circulating TTR(Fig. 1). This advanced antisense design utilizesa triantennary N-acetylgalactosamine (GalNAc)moiety to support productive receptor-medi-ated uptake by the high-capacity asialoglyco-protein receptors (ASGPR) expressed byhepatocytes [27, 28]. An integrated analysis ofeight GalNAc-conjugated ASOs tested in phase 1randomized placebo-controlled studies of heal-thy human volunteers showed a 20–30-foldincrease in potency compared with the parentunconjugated ASOs [27]. Notably, these Gal-NAc-conjugated ASOs were well tolerated, withno discontinuations due to adverse events (AEs)[27].

AKCEA-TTR-LRx is a GalNAc-conjugated ASOwith a nucleotide sequence identical to inot-ersen, but with fewer phosphorothioate-modi-fied internucleotide linkages, which is expectedto improve its safety profile. Results from a first-in-human phase 1 study of AKCEA-TTR-LRx inhealthy volunteers demonstrated that AKCEA-TTR-LRx 45 mg SC once every 4 weeks producedsustained decreases in serum TTR levels within30 days of treatment initiation, achieving amean reduction from baseline of 86% after fourdoses of treatment [29]. No treatment-relatedsafety or tolerability issues were identified underthis dosing regimen. Consequently, AKCEA-TTR-LRx requires a lower dose and less frequent

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administration (45 mg SC every 4 weeks, a27-fold reduction in monthly exposure) thanthe unconjugated ASO, inotersen, to achieve asimilar (if not better) pharmacological effect[26, 29, 30].

NEURO-TTRansform is a global, open-label,randomized phase 3 study that aims to evaluatethe efficacy and safety of AKCEA-TTR-LRx inpatients with hATTR-PN. This paper describesthe rationale and the design of the trial.

METHODS

Study Design

NEURO-TTRansform (EudraCT: 2019-001698-10; ClinicalTrials.gov: NCT04136184; ION-682884-CS3) is a phase 3 multicenter, open-la-bel, randomized study of patients with stage 1or stage 2 hATTR-PN assigned to receive AKCEA-TTR-LRx or an active reference arm (inotersen;Fig. 2). The primary aim is to evaluate the

efficacy of AKCEA-TTR-LRx in patients withhATTR-PN relative to the historical placebocontrol arm of the NEURO-TTR trial [26]. Theprimary endpoints include the percentagechange from baseline in serum TTR, changefrom baseline in mNIS ? 7, and Norfolk QOL-DN score (Fig. 3). Key efficacy endpoints will beassessed at week 35 (interim analysis), week 66(final efficacy analysis), and week 85. The sec-ondary aims are to evaluate the efficacy ofAKCEA-TTR-LRx relative to the historical pla-cebo group with respect to the NeuropathySymptom and Change (NSC) score, PhysicalComponent Summary (PCS) score of the36-Item Short-Form Health Survey (SF-36), PNDscore, and modified body mass index (mBMI;Fig. 3). AE monitoring and safety laboratoryassessment will be performed throughout thetrial. Discontinuation of study drug will occurimmediately if the patient becomes pregnant,receives a major organ transplantation, or pre-sents with predefined laboratory testabnormalities.

Fig. 1 ASGPR mediates the uptake of the GalNAc3-conjugated ASO AKCEA-TTR-LRx by hepatocytes, whereit binds to the TTR mRNA through Watson–Crickhybridization and prevents production of TTR protein viaRNase H1-mediated degradation of the target TTR

mRNA. ASGPR asialoglycoprotein receptor, ASO anti-sense oligonucleotide, DNA deoxyribonucleic acid, mRNAmessenger ribonucleic acid, RNase ribonuclease, TTRtransthyretin

380 Neurol Ther (2021) 10:375–389

Patients will be randomized 6:1 to receive SCinjections of either AKCEA-TTR-LRx (n & 120)45 mg every 4 weeks or inotersen (n& 20)300 mg once per week. As the analyses willutilize comparisons to the external placebocontrol arm of the NEURO-TTR trial, the inot-ersen reference arm is intended to ensure thatno gross differences in patient population andresponse exist between the NEURO-TTR and thecurrent study. Patients in the reference inot-ersen arm will be switched to receive AKCEA-TTR-LRx at week 37. All patients will then con-tinue receiving treatment with AKCEA-TTR-LRx

through week 81, with end-of-treatment (EOT)assessments at week 85, 4 weeks after the lastdose. Following treatment and EOT assess-ments, patients will be eligible to enter an open-label extension study to continue receivingAKCEA-TTR-LRx once every 4 weeks or enter a20-week post-treatment evaluation period.

The distribution of patients according to thedisease stage will mirror the NEURO-TTR trial.Patients will also take supplemental doses of therecommended daily allowance of vitamin A toensure adequate delivery of vitamin A to tissuesin a setting of low serum TTR.

The study protocol and amendments havebeen approved by the relevant local institu-tional review boards (IRBs) or ethics committees(ECs) for currently activated clinical study sites.Activation of additional clinical study sites isongoing, and the study protocol and amend-ments will be approved by the relevant localIRBs/ECs before participants are allowed toenroll in the study. The full list of IRB/EC namesand approval numbers is available uponrequest. The study is being conducted accordingto the International Conference on Harmoni-sation guidelines and relevant country-specificlaws. All participants must provide writteninformed consent prior to study inclusion. Adata and safety monitoring board will periodi-cally review safety, tolerability, and efficacydata relating to AKCEA-TTR-LRx and inotersen,including the results of the prespecified interimanalysis at week 35.

Study Population

NEURO-TTRansform will enroll approximately140 patients from * 70 clinical research sites in15 countries. Adults aged 18–82 years are

Fig. 2 NEURO-TTRansform study design. EOT end oftreatment, hATTR-PN hereditary transthyretin-mediatedamyloid polyneuropathy, SC subcutaneously. aThe screen-ing period is B 6 weeks (or B 10 weeks if genetic testingis required). bConcomitant therapy with tafamidis or off-label use of diflunisal is not allowed. Doxycycline use for

the indication of infection (\ 15 days) is allowed. cPatientsnot participating in the open-label extension will enter a20-week post-treatment evaluation after completing EOTassessments. dPlacebo arm of the NEURO-TTR study(NCT01737398)

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eligible for enrollment if they have stage 1 orstage 2 hATTR-PN, a documented TTR muta-tion, and signs and symptoms consistent withpolyneuropathy (including an NIS C 10 andB 130). Key inclusion and exclusion criteria areshown in Fig. 4, and a complete list can befound in Table S1 in the supplementary

material. Patients who have had a liver or hearttransplant are ineligible. Any medicationsdeemed necessary by the investigator areallowed, although concomitant tafamidis,inotersen, patisiran, or off-label use of diflunisalor doxycycline are not permitted. Patients whohave received prior treatment with inotersen or

Fig. 3 NEURO-TTRansform study endpoints. 10MWT10-min walk test, COMPASS-31 Composite AutonomicSymptom Score-31, ECHO echocardiogram, EQ-5D-5L5-level EuroQol 5-dimension, EOT end of treatment,mBMI modified body mass index, mNIS ? 7 modifiedNeuropathy Impairment Score ? 7, Norfolk QOL-DNNorfolk Quality of Life Questionnaire—Diabetic Neu-ropathy, NSC Neuropathy Symptom and Change, NT-proBNP N-terminal pro-brain natriuretic peptide, PCSPhysical Component Summary, PND PolyneuropathyDisability Score, R-ODS Rasch-built Overall DisabilityScore, SC subcutaneously, SF-36 36-Item Short-Form

Health Survey, TTR transthyretin. aPercentage change.bThe mNIS ? 7 assessment procedure includes: Neuropa-thy Impairment Score, quantitative sensory testing, heartrate response to deep breathing, and predetermined sensoryand nerve conduction testing. Additional clinical evalua-tions done during mNIS ? 7 assessment include: lowerlimbs function test and NSC. cweeks 35 and 66. dweek 65.eweek 37. fweek 35. gweek 81. hVital signs, body weight,physical examination, clinical laboratory tests, electrocar-diogram, use of concomitant medication, thyroid panel,inflammatory panel, coagulation, immunogenicity test

382 Neurol Ther (2021) 10:375–389

patisiran are not eligible; patients previouslytreated with tafamidis, diflunisal, or doxycy-cline are eligible as long as these medicationswere discontinued at least 2 weeks prior to ini-tiation of study treatment (day 1).

Planned Outcomes

Co-primary endpoints at the week 35 interimanalysis include percentage change from base-line to week 35 in serum TTR levels and changefrom baseline in mNIS ? 7 (Fig. 3). Co-primaryendpoints at the week 66 final analysis are thesame as those for the week 35 interim analysis,but also include change from baseline in Nor-folk QOL-DN score.

Secondary efficacy, safety, and exploratoryendpoints measured at weeks 35, 66, and 85(EOT) are shown in Fig. 3. The week 35 interimanalysis key secondary endpoint is change frombaseline to week 35 in the Norfolk QOL-DNscore. The final analysis secondary endpointswill include NSC score at weeks 35 and 66, andSF-36 PCS score, PND score, and mBMI at week65. Non-compartmental pharmacokinetic anal-ysis of AKCEA-TTR-LRx or inotersen will be car-ried out on each individual patient data set inthe pharmacokinetic subgroup, where plasmasamples are collected following drug adminis-tration on days 1, 225, and 449.

All treatment-emergent AEs, serious AEs, andreasons for any withdrawals will be summarizedfor each treatment group using the MedicalDictionary for Regulatory Activities coding

Fig. 4 Key inclusion and exclusion criteria for theNEURO-TTRansform study. ASO antisense oligonu-cleotide, eGFR estimated glomerular filtration rate, FAPfamilial amyloid polyneuropathy, hATTR-PN hereditarytransthyretin-mediated amyloid polyneuropathy, NIS Neu-ropathy Impairment Score, NYHA New York HeartAssociation, siRNA small interfering ribonucleic acid, TTRtransthyretin, UPCR urine protein/creatinine ratio. aStage

1 (ambulatory without assistance) or stage 2 (ambulatorywith assistance). bIn the event of UPCR C 1000 mg/g,eligibility may be confirmed by a repeat random urine testwith UPCR\ 1000 mg/g or a quantitative total urineprotein measurement of\ 1000 mg/24 h. cChronic Kid-ney Disease Epidemiology Collaboration equation 1formula

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system, by system organ class, preferred term,relationship to AKCEA-TTR-LRx or inotersen,and severity. AEs of special interest (includingserious or nonserious events) are severe reduc-tions in platelet count (\50 9 109/L) accom-panied by major bleeding or a clinically relevantnon-major bleeding event, or a platelet countof\ 25 9 109/L independent of such bleedingevents [31].

Statistical Analysis

The primary population for efficacy analyseswill be the full analysis set, defined as all ran-domly assigned patients who receive C 1 doseof AKCEA-TTR-LRx and who have one baselineand at least one post-baseline efficacy assess-ment for mNIS ? 7 score or Norfolk QOL-DNquestionnaire total score.

Power calculations for this study are basedon the assumption that TTR reduction will be80%. Approximately 140 patients (120 admin-istered AKCEA-TTR-LRx) will be enrolled inNEURO-TTRansform to account for a 10%dropout rate. In the NEURO-TTR trial, therewere 52 evaluable completers in the placeboarm [26]. A sample size of 108 evaluable patientsin the AKCEA-TTR-LRx arm will provide C 95%power to detect a 70.3% difference in the per-centage change from baseline in serum TTR,C 90% power to detect a 19.6-point differencein the change from baseline of mNIS ? 7, andC 80% power to detect a 10.7-point differencein the change from baseline of the NorfolkQOL-DN score between AKCEA-TTR-LRx–treatedpatients and the historical NEURO-TTR placeboarm (two-sided alpha level, 0.025).

A sequential multiple testing procedure willbe used to control the overall type 1 error rate at0.05. If both co-primary endpoints of theinterim analysis (TTR and mNIS ? 7) are sig-nificant at an alpha level of 0.025, then thesecondary endpoint (Norfolk QOL-DN score)will be tested at the interim analysis at an alphalevel of 0.025. Regardless of the interim analysisresults, the study will proceed as planned andthe data will be collected from week 66 onwardsat all study endpoints. For endpoints that arestatistically significant at the week 35 interim

analysis, the corresponding tests at the week 66final analysis will not be conducted. The non-significant endpoint(s) (TTR and/or mNIS ? 7)and the co-primary endpoint Norfolk QOL-DNscore will be tested at the final analysis. If all theco-primary endpoints are significant, the sec-ondary endpoints will be tested with the mul-tiplicity controlled by the ranking strategy.Testing of the statistical significance of subse-quent endpoints is contingent upon theattainment of statistical significance by theprevious endpoint analyzed.

The percentage change in TTR will be asses-sed with a mixed-effects model for repeatedmeasures (MMRM) adjusted by propensity scoreweights. The MMRM assumes that missing dataare at random and that patients who discon-tinue prematurely would have behaved simi-larly to other patients in the same treatmentgroup. The MMRM will include the effects oftreatment, time, disease stage, pV50M muta-tion, previous treatment, treatment-by-timeinteraction, baseline value of the endpoint, andbaseline-by-time interaction. The propensityscore will be calculated for each patient in theAKCEA-TTR-LRx or historical placebo arm usinga logistic regression model with baseline valueof the endpoint and covariates including dis-ease stage, pV50M mutation, and previoustreatment. For mNIS ? 7 and Norfolk QOL-DNscore, the treatment comparison at week 35 willbe based on the analysis of covariance modeladjusted by propensity score. In the week 66final analysis, the aforementioned MMRMadjusted by propensity score weights will beused for the treatment comparison for the per-centage change from baseline in serum TTR,change from baseline in mNIS ? 7, and NorfolkQOL-DN score.

DISCUSSION

hATTR-PN is a rare, progressive, and fatal dis-ease caused by mutations in the TTR gene thatresult in the synthesis of a destabilized TTRprotein structure and lead to the formation ofamyloid deposits in multiple organ systems[2, 7]. Although two TTR gene silencing treat-ments for hATTR-PN are available that reduce

384 Neurol Ther (2021) 10:375–389

TTR and slow or halt the progression of thedisease, there is an unmet need for an effectivetreatment with a more favorable risk–benefitprofile and greater convenience of use. Inot-ersen requires weekly SC administration andregular monitoring for thrombocytopenia andglomerulonephritis, while patisiran requiresintravenous infusion every 3 weeks andpremedication with a corticosteroid to preventinfusion-related reactions [26]. The ligand-con-jugated antisense technology is a significantadvancement in the selective delivery to hepa-tocytes, yielding an approximate 20–30-foldincrease in potency that permits substantialreduction in systemic exposure and animproved safety and tolerability profile[27, 32–34]. Similar to other ASO conjugates inthis class, preclinical and phase 1 data onAKCEA-TTR-LRx demonstrated an increase inpharmacological potency compared with inot-ersen, a currently approved antisense medicinefor the treatment of hATTR-PN. On the basis ofthe pathogenesis of ATTR, it is likely that pre-venting TTR from being synthesized wouldeliminate the origin of misfolded protein andsubsequent amyloid fibril deposition, thusleading to the improvement of symptoms ofATTR, including polyneuropathy andcardiomyopathy.

NEURO-TTRansform is a phase 3 multicen-ter, open-label study of patients with stage 1 orstage 2 hATTR-PN randomly assigned to theAKCEA-TTR-LRx arm or an inotersen referencearm. Efficacy and safety data from participantsenrolled in the AKCEA-TTR-LRx treatment armwill be compared with results from the histori-cal placebo group in the NEURO-TTR trial [26].The open-label design was selected because ofthe availability of approved treatments forhATTR-PN in some countries, rendering inclu-sion of a placebo control arm unethical[1, 17, 22–25]. This was not the case while theNEURO-TTR study was conducted, when theonly standard of care was liver transplantationand tafamidis (which was only available at thetime for stage 1 ATTR-PN in some Europeancountries [26]). Patient eligibility criteria andthe distribution of stage 1 and 2 hATTR-PN inthe NEURO-TTRansform study are similar tothose in the NEURO-TTR study, with identical

endpoints and assessment times [26]. This pro-vides an opportunity to adequately compareoutcomes between patients who receivedAKCEA-TTR-LRx in NEURO-TTRansform andthose who received placebo in NEURO-TTR. Thelack of blinding in NEURO-TTRansform meansthat many sources of conscious and uncon-scious bias could be introduced. However, theselimitations can be assessed to an extent bycomparing the performance of inotersen upuntil week 35 in NEURO-TTRansform and theNEURO-TTR study, as inotersen represents acommon denominator between the two studies.

One of the primary endpoints assessed inthis study, change from baseline in mNIS ? 7,was also assessed as a primary endpoint in theNEURO-TTR study [26]. The mNIS ? 7 scoringprocedure utilizes highly standardized, quanti-tative, and referenced assessments that arespecifically designed to assess hATTR-PNimpairment [35]. Assessments include the NIS,sensory and motor nerve conduction testing,quantitative sensory testing, and measurementof heart rate variation with deep breathing. Thenerve conduction test is an objective andquantitative assessment, thereby providing ahighly validated measure of neuropathy [35]. Inthe NEURO-TTR study, the mNIS ? 7 scoringapproach showed highly significant improve-ment in global scores and essentially all sub-scores with inotersen treatment, includingquantitative measures and potentially objectivemeasures in scored neuropathy signs [26, 35].The favorable outcome is a result of utilizingpreviously validated scores, using the samespecially trained mNIS ? 7 evaluator through-out the study for each individual participant,and surveillance by a central expert readingcenter [35].

The AKCEA-TTR-LRx dosage regimen underinvestigation in phase 3 was selected on thebasis of the pharmacodynamics and safetyanalyses of the phase 1 randomized placebo-controlled study in 47 healthy volunteers.AKCEA-TTR-LRx dosed at 45 mg every 4 weeksproduced a mean reduction from baseline inserum TTR of 86% after four doses and anabsence of potential tolerability or safety issues,such as the increases in liver transaminase levelsobserved in the upper dose range of this study

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[29]. The GalNAc-mediated delivery of AKCEA-TTR-LRx to hepatocytes supports low-dose ther-apy and a more convenient dosage regimen(i.e., monthly administration) relative to theunconjugated ASO therapy used for the treat-ment of hATTR-PN [22–25]. Since no placebogroup will be utilized in this trial, enrolledparticipants will receive either an approveddrug, inotersen, or AKCEA-TTR-LRx at a muchlower dose to achieve a similar pharmacologicaleffect. Eligible participants will have the optionto enroll into a long-term extension trial afterthe completion of this study. As a targetedtherapy, AKCEA-TTR-LRx has the potential toreduce the disease burden of hATTR-PN withlower and less frequent dose than the parentcompound. The NEURO-TTRansform trial willdetermine whether a more effective and effi-cient reduction of TTR by AKCEA-TTR-LRx yieldsboth an improved safety and tolerability profileand clinical benefit in terms of disease stabilityor regression.

ACKNOWLEDGEMENTS

Funding. This study and Rapid ServicePublication Fee is supported by Ionis Pharma-ceuticals, Inc. (Carlsbad, CA, USA) and AkceaTherapeutics (Boston, MA, USA).

Medical Writing, Editorial, and OtherAssistance. Medical writing support was pro-vided by Nancy Niguidula and Malcom Darkesof Excel Scientific Solutions (Fairfield, CT, USA)and funded by Ionis Pharmaceuticals. Criticalreview was provided by Sanjay Bhanot, MD,PhD, Brenda F. Baker, PhD, and Lisa Hannan,PhD, of Ionis Pharmaceuticals.

Authorship. All named authors meet theInternational Committee of Medical JournalEditors (ICMJE) criteria for authorship for thisarticle, take responsibility for the integrity ofthe work as a whole, and have given theirapproval for this version to be published.

Disclosures. Teresa Coelho has receivedhonoraria to attend scientific meetings from

Akcea, Alnylam, Biogen, Ionis, and Pfizer andparticipates as a consultant in meetings pro-moted by Akcea, Alnylam, Biogen, Ionis, andPfizer without financial compensation. YukioAndo has received research funding from theJapan Society for the Promotion of ScienceGrants-in-Aid for Scientific Research (KAKENHIgrant numbers 19H03565 and 16H0567). Mer-rill D. Benson has received grant support fromthe Veterans Administration Merit Review Pro-gram. John L. Berk participates as a consultantfor Akcea, Alnylam, Corino, Intellia, and Ionis.Marcia Waddington-Cruz has received hono-raria for consultancy and travel, and has been aprincipal investigator in clinical trials for Alny-lam, FoldRx, Genzyme, Ionis, NHI, Pfizer, Pro-thena, and PTC. Peter J. Dyck has receivedhonoraria for teaching and quality assurance oftherapeutic trials in transthyretin amyloidosisfrom Alnylam and Ionis. Julian D. Gillmore hasreceived honoraria for serving on scientificadvisory boards from Alnylam, Eidos, Ionis/Akcea, and Pfizer. Sami L. Khella has receivedhonoraria for consultancy from Akcea, Alny-lam, Eidos, Ionis, and Pfizer. William J. Litchyhad institutional contracts to train investigatorsand for quality control of data from Alnylamand Ionis. Laura Obici has received honorariafor speaking and consultancy from Akcea,Alnylam, and Pfizer. Nicholas J. Viney, GustavoBuchele, Michela Brambatti, Shiangtung W.Jung, Sotirios Tsimikas, Eugene Schneider,Richard S. Geary, and Brett P. Monia areemployees of and hold stock/stock options inIonis. Cecilia Monteiro and Li-Jung Tai wereemployees of Ionis at the time the analysis wasconducted and holds stock/stock options atIonis. Louis St. L. O’Dea is an employee of andholds stock/stock options in Akcea. Morie Gertzhas received honoraria from Alnylam, Amgen,Annexon, Apellis, Ionis/Akcea, Janssen, Med-scape, Physicians Education Resource, and Pro-thena; workforce training from Sanofi; grantsand personal fees from Spectrum; honoraria forspeaking from DAVA Oncology, Johnson &Johnson, Medscape, and Teva; honoraria forserving on scientific advisory boards fromPharmacyclics and Proclara; honoraria for thedevelopment of educational materials from i3Health; royalties from Springer; grant funding

386 Neurol Ther (2021) 10:375–389

from the Amyloidosis Foundation, Interna-tional Waldenstrom’s MacroglobulinemiaFoundation, and The National Cancer InstituteSpecialized Programs for Research ExcellenceMM SPORE 5P50 CA186781-04; has served onthe data safety monitoring boards for AbbVie,Celgene, and Research to Practice; and holdsstock options from Aurora Bio.

Compliance with Ethics Guidelines. Thestudy protocol and amendments have beenapproved by the relevant local IRBs or ECs forcurrently activated clinical study sites. Activa-tion of additional clinical study sites is ongoing,and the study protocol and amendments will beapproved by the relevant local IRBs/ECs beforeparticipants are allowed to enroll in the study.The full list of IRB/EC names and approvalnumbers is available upon request. The study isbeing conducted according to the InternationalConference on Harmonisation guidelines andrelevant country-specific laws. All participantsmust provide written informed consent prior tostudy inclusion. A data and safety monitoringboard will periodically review safety, tolerabil-ity, and efficacy data relating to AKCEA-TTR-LRx

and inotersen, including the results of the pre-determined interim analysis at week 35.

Data Availability. The study is registered atClinicalTrials.gov (NCT04136184) and EudraCT(2019-001698-10).

Open Access. This article is licensed under aCreative Commons Attribution-NonCommer-cial 4.0 International License, which permitsany non-commercial use, sharing, adaptation,distribution and reproduction in any mediumor format, as long as you give appropriate creditto the original author(s) and the source, providea link to the Creative Commons licence, andindicate if changes were made. The images orother third party material in this article areincluded in the article’s Creative Commonslicence, unless indicated otherwise in a creditline to the material. If material is not includedin the article’s Creative Commons licence andyour intended use is not permitted by statutoryregulation or exceeds the permitted use, youwill need to obtain permission directly from the

copyright holder. To view a copy of this licence,visit http://creativecommons.org/licenses/by-nc/4.0/.

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