Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 https://doi.org/10.1186/s13023-020-01399-4 RESEARCH Open Access A phase II, open-label, extension study of long-term patisiran treatment in patients with hereditary transthyretin-mediated (hATTR) amyloidosis Teresa Coelho1*, David Adams2, Isabel Conceição3, Márcia Waddington-Cruz4, Hartmut H. Schmidt5, Juan Buades6, Josep Campistol7, John L. Berk8, Michael Polydefkis9, Jing Jing Wang10, Jihong Chen10, Marianne T. Sweetser10, Jared Gollob10 and Ole B. Suhr11 Abstract Background: Patisiran, an RNA interference therapeutic, has demonstrated robust reduction of wild-type and mutant transthyretin protein and was able to improve polyneuropathy and quality of life following 18 months of treatment in patients with hereditary transthyretin-mediated (hATTR) amyloidosis. In this 24-month Phase II open- label extension study, we evaluated the effects of patisiran treatment (0.3 mg/kg intravenously every 3 weeks) on safety, serum transthyretin levels, and clinical parameters. Efficacy assessments included modified Neuropathy Impairment Score +7 (mNIS+7) and multiple disease-relevant measures. Cardiac assessments were performed in a pre-specified cardiac subgroup. Results: Twenty-seven patients entered this study, including 12 (44%) with ambulation difficulties due to their neuropathy and 11 (41%) who met criteria for the cardiac subgroup. During treatment, the majority of adverse events were mild/moderate in severity; there were no drug-related adverse events leading to treatment discontinuation. The most common drug-related adverse events were flushing and infusion-related reactions (22% each). Patisiran resulted in rapid, robust (~ 82%), and sustained reduction of mean transthyretin levels over 24 months. A mean 6.95-point decrease (improvement) in mNIS+7 from baseline was observed at 24 months. Patisiran’s impact on mNIS+7 was irrespective of concomitant tafamidis or diflunisal use, sex, or age. Clinical assessments of motor function, autonomic symptoms, disease stage, and quality of life remained stable over 24 months. No significant changes were observed for echocardiographic measures or cardiac biomarkers in the cardiac subgroup. Exploratory analyses demonstrated improvements in nerve-fiber density with corresponding reductions in amyloid burden observed in skin biopsies over 24 months. Conclusions: Long-term treatment with patisiran had an acceptable safety profile and was associated with halting/ improvement of polyneuropathy progression in patients with hATTR amyloidosis. Trial registration: The study was registered at ClinicalTrials.gov (identifier: NCT01961921) on October 14, 2013. Keywords: ATTR amyloidosis, Cardiomyopathy, Patisiran, Polyneuropathy, RNA interference * Correspondence: tcoelho@netcabo.pt 1 Hospital de Santo António, Centro Hospitalar do Porto, 4099-001 Porto, Portugal Full list of author information is available at the end of the article © The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 2 of 10 Background Patisiran is an RNA interference (RNAi) therapeutic Hereditary transthyretin-mediated (hATTR) amyloidosis is that targets both wt and mutant TTR mRNA, thereby re- a progressive and life-threatening disease caused by muta- ducing expression of all forms of TTR protein. Patisiran tions in the gene encoding the transthyretin (TTR) protein is formulated in a lipid nanoparticle facilitating delivery [1, 2]. TTR circulates as a tetrameric protein produced pre- to the liver, which is the primary site of TTR production dominantly by the liver [3], with its primary role transport- [3, 20]. Across the clinical development program, pati- ing vitamin A complexed with retinol-binding protein siran has demonstrated robust, rapid, and sustained re- (RBP), with a minor role as a thyroxine carrier [4]. Over duction of TTR in patients with hATTR amyloidosis 120 TTR mutations confer amyloidogenicity [5], resulting [10, 20, 21]. In the recent Phase III APOLLO study, in the accumulation of misfolded TTR protein as amyloid 18 months of patisiran dosing demonstrated the po- fibrils in multiple sites including nerves, heart, gastrointes- tential to halt or reverse polyneuropathy and improve tinal tract, eye, and central nervous system [6, 7]. This QOL from baseline in the majority of patients [10], pathophysiology leads to a heterogeneous clinical presenta- leading to the approval of patisiran in the USA and tion that includes sensory and motor, autonomic, and car- several other countries for the treatment of hATTR diac symptoms, with the majority of patients displaying a amyloidosis with polyneuropathy [22, 23]. Patisiran mixed phenotype of both polyneuropathy and cardiomyop- also improved exploratory measures of cardiac structure athy [8–11]. hATTR amyloidosis is associated with signifi- and function compared with placebo in a pre-specified cant morbidity and mortality, with a median survival analysis of patients with cardiac involvement in the of 4.7 years following diagnosis [12]. The presence of car- APOLLO study [24]. Here, we report the results from an diac involvement is associated with a worse prognosis with open-label extension (OLE) of a Phase II study of patisiran a reduced survival (3.4 years) reported in these patients [13]. in patients with hATTR amyloidosis with polyneuropathy. A substantial decline in quality of life (QOL) and physical Safety, efficacy, and pharmacodynamic (PD) data are de- functioning is observed as hATTR amyloidosis progresses, scribed over 24 months, which is, to date, the longest reflecting disease involvement across multiple systems. completed clinical trial with patisiran. Thus, there is a need for treatments that target the patho- genic TTR protein, rather than the symptoms associated with specific tissues. Current treatment strategies for Methods hATTR amyloidosis directed at disease pathophysiology in- Study design and participants clude prevention of TTR protein production or stabilization This was a multicenter, international, Phase II OLE study of the TTR protein complex. Orthotopic liver transplant- of patisiran conducted across seven countries: USA, Brazil, ation (OLT), which replaces the source of mutant TTR, has France, Germany, Portugal, Spain, and Sweden. Eligible been mainly used in patients with early-stage polyneurop- patients had biopsy-proven hATTR amyloidosis with evi- athy without cardiac involvement [14]; however, this strat- dence of mild-to-moderate neuropathy, and had previ- egy has been utilized much less in recent years after the ously received and tolerated patisiran in the Phase II study introduction of medical therapy [14]. OLT also does not (NCT01617967) [21]. Patients had Karnofsky Performance prevent deposition of wild-type (wt) TTR protein, which Status ≥60%; adequate liver and renal function; absolute contributes to subsequent disease progression in post-OLT neutrophil count ≥1500 cells/mm3; platelet count ≥100,000 patients [15]. Treatment with TTR tetramer stabilizers cells/mm3; and hemoglobin ≥100 g/l. Key exclusion criteria (tafamidis: approved in EU and select regions for treatment included: prior OLT; unstable angina or uncontrolled car- of the polyneuropathy of hATTR amyloidosis [16]; and diac arrhythmia; and New York Heart Association heart diflunisal: used off-label for hATTR amyloidosis [6]) has failure class > II. Patients who received an investigational been shown to slow progression of neurologic impairment drug other than tafamidis or diflunisal within 30 days of [17, 18]. Additionally, a recent Phase III study of tafamidis the patisiran first dose were also excluded. Further details showing reduced cardiovascular mortality and hospitaliza- on inclusion/exclusion criteria are in Additional file 1: tions in patients with ATTR amyloidosis with cardiomyop- Supplementary methods. athy resulted in USA and EU approval for the treatment of This study was conducted according to the guidelines the cardiomyopathy of ATTR amyloidosis derived from wt of the International Conference on Harmonisation, the or mutant TTR for early-stage disease [19]. Inotersen, an World Health Organization Declaration of Helsinki, and antisense oligonucleotide which reduces circulating TTR the Health Insurance Portability and Accountability Act levels, is approved in the USA and EU for the treatment of of 1996. Written informed consent was obtained from the polyneuropathy associated with hATTR amyloidosis all participants in the study. The study protocol was ap- [11]. While these treatments may slow the natural progres- proved by local Institutional Review Boards and Ethics sion of hATTR amyloidosis, worsening of neurologic and/or Committees, and all subsequent protocol amendments cardiac function is observed in the majority of patients. underwent the same approval procedure. Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 3 of 10 Procedures Outcomes Patients received intravenous infusions of patisiran The primary objective was to evaluate the safety and tol- 0.3 mg/kg (over 70 min) every 3 weeks for approximately erability of long-term dosing with patisiran. Secondary 2 years. Patients received a premedication (dexametha- objectives were to assess the PD effect of long-term pati- sone, paracetamol/acetaminophen, H1 and H2 blockers, or siran dosing on serum TTR levels, and to monitor equivalent) to reduce the likelihood of an infusion-related changes from baseline in the following measures [25]: reaction (IRR). An oral daily supplemental dose of vitamin mNIS+7; QOL (EuroQoL 5-dimensions questionnaire); A was also given. Patients were monitored before the first disability (Rasch-built Overall Disability Scale [R-ODS]); dose (Day − 28 to Day 0), during dosing (Day 0 to Day motor function (10-meter [m] walk test and dynamometric 735), and at follow-up (Day 756 and 791). Safety was eval- grip strength); and nutritional status (modified body mass uated throughout the study, and included vital signs, phys- index). Exploratory endpoints included evaluation of ical and ophthalmologic examinations, clinical laboratory changes in: NIS, NIS+7, familial amyloid polyneuropathy tests, electrocardiograms, and adverse event (AE) moni- (FAP) stage, polyneuropathy disability (PND) score; patient- toring. AEs were categorized by the Investigators as either reported autonomic symptoms (Composite Autonomic mild, moderate or severe, and coded according to the Symptom Score [COMPASS]-31 questionnaire); sensory Medical Dictionary for Regulatory Activities (MedDRA®) and autonomic innervation of the skin; dermal amyloid version 18.0. burden [25]; and cardiac parameters (measured by echo- Serum TTR concentration was assessed using both cardiogram and circulating cardiac biomarkers) in patients enzyme-linked immunosorbent assays and turbidimetric as- in the cardiac subgroup. says. Serum RBP was quantified using nephelometry and serum vitamin A was quantified using a high-performance Statistical analysis liquid chromatography assay. Details of assessment timings For each patient population analyzed in this OLE study, are in Additional file 1: Supplementary methods. categorical variables were calculated as the number and Neurologic function was assessed using the modified percentages of patients; for continuous variables, mean, Neuropathy Impairment Score +7 (mNIS+7), Neuropathy median, standard deviation, and range were calculated. Impairment Score +7 (NIS+7), and Neuropathy Impair- Descriptive statistics provided for clinical laboratory tests, ment Score (NIS), at baseline and every 6 months thereafter vital signs, all clinical activity parameters, and echocardio- [25]. To standardize the efficacy assessment and minimize grams were presented as actual values and changes from variability across the multiple study centers, neurologists baseline relative to each on-study observation. were trained to perform the mNIS+7 evaluation at a central This trial is registered with ClinicalTrials.gov, number center (Dyck Peripheral Nerve Research Laboratory, Mayo NCT01961921. Clinic, Rochester, MN, USA). QOL, disability, motor func- tion, nutritional status, autonomic symptoms, and disease Results stage (outlined in Outcomes below) were assessed at base- A total of 27 of 29 patients from the Phase II study [21] line and every 6 months thereafter (details in Additional file were enrolled in this OLE study (1 patient withdrew for 1: Supplementary methods). Measurements were based on personal reasons, the other discontinued the Phase II two independent readings taken at least 24 h (but no study due to AEs). The time between last patisiran dose greater than 7 days) apart and performed by the same in- in the Phase II study and first dose in this OLE study vestigator when possible. Tandem skin punch biopsies of ranged from 169 to 512 days. Of the 27 patients enrolled, the distal thigh and distal leg were obtained at baseline and 25 completed the OLE study (completion date August every 6 months thereafter (patient status allowing). These 31, 2016); 1 patient withdrew after approximately biopsies were used to quantify intra-epidermal nerve-fiber 19 months due to gastro-esophageal cancer (unlikely re- density (IENFD; sensory innervation) and sweat gland lated to patisiran), which proved fatal, and 1 patient died nerve-fiber density (SGNFD; autonomic innervation) based of a myocardial infarction (not related to patisiran) after on blinded central analysis. Congo Red staining was used to completing all dosing but before the end of the study. serially assess dermal amyloid burden in the biopsies. Median treatment duration was 25 months (range, In a subgroup of patients with baseline cardiac involve- 19–25 months), with a median of 35 doses (range, ment (cardiac subgroup; defined as left ventricular [LV] 27–36 doses) administered per patient. wall thickness of ≥13 mm on echocardiogram in the ab- Baseline clinical characteristics of the OLE population are sence of hypertension, controlled hypertension, or aortic shown in Table 1. At study entry, 12 patients (44%) had valve disease), serial echocardiograms were performed every ambulatory difficulties due to neuropathy (PND score > I), 6 months and blood samples were taken every 3 months for and 20 (74%) and 14 (52%) had a medical history of gastro- central assessment of the cardiac biomarkers troponin I intestinal or renal and urinary disorders, respectively. and N-terminal pro-brain natriuretic peptide. Twenty patients (74%) were receiving TTR tetramer Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 4 of 10 Table 1 Patient baseline demographics and disease Table 2 Summary of safety data characteristics AE Total population (n = 27) n (%) Characteristic Total population (n = 27) Summary of AEs Age, years, median (range) 64 (29 to 77) Any AE 26 (96) Sex, n Any AE related to study drug 16 (59) Male 18 (67%) Any serious AE 7 (26) Female 9 (33%) Any study drug-related serious AE 0 Genotype Death 2 (7) Val30Met 20 (74%) Any AE leading to discontinuation 2 (7) a Other 7 (26%) Common AEs (occurring in > 15% of patients) FAP stage Flushing 7 (26) 1 24 (89%) Diarrhea 6 (22) 2 3 (11%) Infusion-related reaction 6 (22) PND score Nasopharyngitis 6 (22) I 15 (56%) Urinary tract infection 6 (22) II 9 (33%) Vomiting 6 (22) IIIa 2 (7%) Wound 6 (22) IIIb 1 (4%) Nausea 5 (19) On TTR stabilizer at study entry AEs related to study drug in > 2 patients Tafamidis 13 (48%) Infusion-related reaction 6 (22) Diflunisal 7 (26%) Flushing 6 (22) None 7 (26%) Diarrhea 3 (11) mNIS+7 (max. impairment: 304) 53.0 (2.0 to 122.5) Abbreviation: AE adverse event NIS (max. impairment: 244) 34.8 (4.0 to 93.4) AEs potentially related to patisiran occurred in 16 (59%) mBMI, (kg/m2 × g/L) 1030.5 (728.6 to 1379.6) patients. No patient discontinued the study due to a EQ-5D (max. impairment: 0) 0.8 (0.3 to 1.0) drug-related AE. The most common drug-related AEs Cardiac subgroup 11 (41%) were mild flushing and IRRs (6 patients [22%] each). Val30Met/non-Val30Metb, n 8/3 The incidence and number of IRRs decreased over time; NT-proBNP (pg/mL)c 809.8 (105.0 to 2070.0) all IRRs were mild in severity. All patients received their d complete dose of patisiran, but 2 patients had temporary Troponin I (ng/mL) 0.14 (0.03 to 0.69) infusion interruptions due to IRRs. In 1 patient, the infu- LV wall thickness (cm) 1.6 (1.3 to 1.9) sion rate was slowed with no subsequent IRRs; the other Abbreviations: EQ-5D EuroQoL 5-dimensions questionnaire, FAP familial amyloid polyneuropathy, LV left ventricular, mBMI modified body mass index, patient had mild local intravenous site irritation. mNIS+7 modified Neuropathy Impairment Score + 7, NIS Neuropathy Serious AEs were reported in 7 patients (26%), none of Impairment Score, NT-proBNP N-terminal pro-brain natriuretic peptide, PND which were considered related to patisiran (Table 2). Of polyneuropathy disability, QOL quality of life, TTR transthyretin. All data are mean (range) or n (%), unless otherwise stated the serious AEs, only osteonecrosis (n = 2) was reported in a Non-Val30Met mutations: Ser77Tyr (n = 2), Ser77Phe (n = 2), Tyr116Ser (n = 1), more than 1 patient (Additional file 2: Table S1). Severe Phe64Leu (n = 1), Arg54Thr (n = 1) b Non-Val30Met mutations in cardiac subgroup: Arg54Thr (n = 1), Ser77Phe AEs were reported in 5 patients, with none considered re- (n = 1), Ser77Tyr (n = 1) lated to patisiran; all severe events were also serious AEs. c NT-proBNP normal range is: ≤ 97 pg/mL (age 18–45); ≤ 121 pg/mL (age 45–55); Two patients died during the study, following cessation of ≤ 198 pg/mL (age 55–65); ≤ 285 pg/mL (age 65–75); ≤ 526 pg/mL (age ≥ 75) d Troponin I normal range is: < 0.03 ng/mL patisiran treatment, as detailed above. There were no clin- ically significant changes related to patisiran in liver func- stabilizers at baseline; 6 of these patients discontinued tion tests, renal function, thyroid function, or hematologic stabilizer use (diflunisal [n = 5]; tafamidis [n = 1]) be- parameters (including platelet count), and no clinical tween 1 and 18 months after entering the study. Eleven manifestations of vitamin A deficiency. There were no patients were included in the cardiac subgroup, for whom clinically relevant changes in vital signs, electrocardiogram the mean LV wall thickness was 1.6 cm (range, 1.3–1.9 parameters, or ophthalmology tests. The overall safety cm) (Table 1). profile of patisiran was similar in the cardiac subgroup Overall, 26 patients (96%) reported AEs (Table 2), the (Additional file 3: Table S2) compared with the overall majority of which were mild or moderate in intensity. study population, and similar irrespective of concomitant Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 5 of 10 TTR tetramer stabilizer therapy (tafamidis and/or difluni- patients < 65 or ≥ 65 years old (Table 3). The mean change sal) (data not shown). Furthermore, the overall safety pro- (standard error of the mean) in NIS and NIS +7 from files of concomitant treatment with either patisiran and baseline at 24 months was + 1.92 (1.84) and + 2.54 (1.84) tafamidis (median exposure 736 days [range: 19–747 days]) points, respectively (Additional file 5: Fig. S1a and b). or patisiran and diflunisal (median exposure 421 days Multiple additional clinical parameters, including [range: 139–736 days]) in small cohorts of patients EuroQoL 5-dimensions questionnaire, R-ODS, 10-m were consistent with the safety profiles of the respective walk distance, hand grip strength, and COMPASS-31 therapies as monotherapies [17, 18]. score, showed stability throughout the 24-month treat- Serum TTR reduction was rapid and robust following ment period (Table 4). Within the cardiac subgroup, the first dose of patisiran, with mean reduction from echocardiographic assessments and serum biomarker baseline nearing 80% by Day 18 (Fig. 1). A sustained levels also remained stable (Table 4). The majority of pa- mean reduction of 82% was achieved over 24 months, tients showed stabilization of FAP stage (20 [74%] pa- with mean maximal reduction of 93%. The magnitude tients with no change in FAP stage at 24 months) or and dynamics of TTR reduction were similar between PND score (23 [85%] patients with no change in PND patients who received patisiran and TTR stabilizers com- score at 24 months) (Additional file 6: Table S4). pared with patisiran alone (Additional file 4: Table S3). Exploratory analyses evaluating SGNFD of the lower limb Similarly, TTR genotype (Val30Met versus non- demonstrated increased fiber density in the distal thigh at Val30Met), sex, and age (< 65 versus ≥65 years) did not 6, 12, 18, and 24 months, and in the distal leg at 24 months affect the PD activity of patisiran (Additional file 4: (p < 0.05) (Fig. 3a and b). Conversely, a decrease (p < 0.05) Table S3). Overall, serum vitamin A and RBP levels were in dermal amyloid burden was demonstrated in both distal reduced by > 65% after 24 months of treatment, as ex- thigh and distal leg over 24 months (Fig. 3c). IENFD pected based on the known role of TTR in binding and remained stable over time (data not shown). transport of the RBP–vitamin A complex [26]. At 24 months, there was a mean mNIS+7 decrease (improvement) from baseline of 6.95 points (Table 3; Discussion Fig. 2); 19 patients (70.4%) had an improvement (de- This OLE study evaluated the safety, PD, and clinical ef- crease from baseline at 24 months) in mNIS+7 score. Al- fect of patisiran on patients with hATTR amyloidosis though patisiran appeared to affect all components of with polyneuropathy over 24 months. Patisiran, dosed mNIS+7 in a similar manner directionally, sensory mea- once every 3 weeks at 0.3 mg/kg, induced rapid and ro- sures exhibited the greatest decrease from baseline (im- bust TTR reduction in serum. A mean reduction of ap- provement) at 24 months by quantitative sensory testing proximately 82% was sustained across the 24-month (Table 3). The mean change in mNIS+7 was similar ac- treatment period, consistent with the median 81% TTR cording to TTR tetramer stabilizer use, sex, and in reduction reported at 18 months in the Phase III Fig. 1 Serum TTR reduction. Percentage change in serum TTR from baseline over time. Pre- and post-dose (0.3 mg/kg Q3W patisiran) values are indicated on the graph with an x. BSL, baseline; ELISA, enzyme-linked immunosorbent assay; Q3W, every 3 weeks; TTR, transthyretin Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 6 of 10 Table 3 Change in mNIS+7 by components and subgroups at 24 months of patisiran treatment Change from baseline to Month 24 n Mean (SEM) Median (range) mNIS+7 component Total 26 −6.95 (2.03) −7.00 (−34.63 to 15.38) NIS-Weakness 26 1.23 (1.43) 0.00 (−13.50 to 24.38) NIS-Reflexes 26 −0.48 (0.53) 0.00 (−6.00 to 7.00) QST 26 −7.4 (2.04) −6.0 (−40.0 to 16.0) NCS ∑5 26 −0.19 (0.18) −0.25 (−2.00 to 2.50) Postural BP 26 −0.10 (0.06) 0.00 (−1.00 to 0.50) mNIS+7 by TTR tetramer stabilizer use Patisiran alone 7 −6.75 (5.24) −8.50 (−28.50 to 15.38) Patisiran + TTR tetramer stabilizer 19 −7.03 (2.11) −6.63 (−34.63 to 3.88) Abbreviations: BP blood pressure, mNIS+7 modified Neuropathy Impairment Score +7, NCS nerve conduction studies, NIS Neuropathy Impairment Score, QST quantitative sensory testing, SEM standard error of the mean, TTR transthyretin APOLLO study. No evidence was found for tachyphyl- levels that accompanied TTR suppression was not associ- axis of any PD effect with chronic dosing. ated with any clinical manifestations of vitamin A defi- In this study, patisiran was generally well tolerated, with ciency (e.g. ocular events), which is possibly due to the majority of AEs mild to moderate in intensity, and no redundant mechanisms for its transport and tissue uptake patient discontinued treatment due to a patisiran-related [28]. The overall safety profile is consistent with that ob- AE. Mild IRRs were among the most common drug- served in the APOLLO study [10], with no new safety sig- related AEs (23%), yet they did not interfere with the com- nals reported with 24 months of patisiran dosing. pletion of dosing, and decreased in incidence over time. Furthermore, the safety data in the cardiac subgroup in No clinically relevant changes in laboratory values related this study were similar to that observed in the overall to patisiran, including indicators of liver or kidney func- population, and with that observed in patients in the car- tion, were observed during the study. In addition, there diac subpopulation in the APOLLO study. Patisiran also were no safety signals regarding thrombocytopenia with had an acceptable safety profile in patients with renal patisiran [27]. The expected reduction in serum vitamin A amyloid involvement at baseline, and in patients taking Fig. 2 Mean change from baseline (SEM) in mNIS+7 in the all-treated population over 24 months. Error bars represent the SEM. mNIS+7, modified Neuropathy Impairment Score +7; SEM, standard error of the mean Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 7 of 10 Table 4 Changes in clinical assessments at 24 months with These data are consistent with those reported in APOLLO patisiran treatment and compare favorably to those reported for other TTR sta- Assessment Change from baseline to bilizing or reduction therapies [11, 17, 18, 32, 33] in patients Month 24 with hATTR amyloidosis with polyneuropathy. It should be n Mean (SEM) noted that measures of neurologic impairment have varied EQ-5D (max. impairment: 0) 26 −0.01 (0.02) across studies of different therapeutic agents. The mNIS+7 R-ODS (no limitations: 48) 25 −1.8 (0.8) scale was used in this study and APOLLO [25] to fully assess 10-m walk (m/s) 21 0.03 (0.04) the progression of this polyneuropathy. The mNIS+7 scale includes measures of autonomic function and nerve conduc- Hand grip strength (kg) 26 1.5 (1.2) tion that are not captured in the NIS score [34]. Notably, mBMI (kg/m2 × g/L) 22 −60.8 (34.9) the mNIS+7 utilizes smart somatotopic quantitative sensa- COMPASS-31 (max. impairment: 100) 26 1.3 (1.8) tion testing to provide a better balance between measure- Cardiac subgroup (n = 11) ment of large and small sensory fibers, and to measure NT-proBNP (pg/mL) 8 −49.6 (170.8) sensation loss across the body [34]. The mNIS+7 scale thus Troponin I (ng/mL) 8 −0.1 (0.1) represents a disease-specific measure of polyneuropathy in hATTR amyloidosis, supporting its use as the primary end- LV mass (g) 10 −16.7 (11.7) point in recent pivotal studies [10, 11]. In the current study, LV wall thickness (cm) 10 −0.08 (0.1) a consistent, favorable effect on mNIS+7 was observed irre- Ejection fraction (%) 10 −0.6 (1.5) spective of age at onset or mutation status, as was also ob- Peak longitudinal strain (%) 10 0.9 (0.9) served in APOLLO [10]. 10-m walk (m/s) 7 0.03 (0.05) It should be noted that there was concomitant use of Abbreviations: EQ-5D EuroQoL 5-dimensions questionnaire, COMPASS either tafamidis or diflunisal in 20 of 27 patients at base- Composite Autonomic Symptom Score, LV left ventricular, m meter, mBMI line. Irrespective of this concomitant TTR tetramer modified body mass index, NT-proBNP N-terminal pro-brain natriuretic peptide, QOL quality of life, R-ODS Rasch-built Overall Disability Scale stabilizer use, patisiran was shown to have a similar ef- fect on mNIS+7; notably, the 6 patients who discontin- ued their stabilizer within 18 months did not exhibit TTR tetramer stabilizers, supporting the potential for pati- neurologic disease progression. siran to treat a diverse patient population. The decrease in mNIS+7 over 24 months supports the The polyneuropathy seen in symptomatic patients with hypothesis that patisiran has the potential to alter the hATTR amyloidosis is rapidly progressive [29]. While pa- course of hATTR amyloidosis with polyneuropathy. The tients with diabetic polyneuropathy exhibit a < 1-point/year exploratory findings in autonomic nerve-fiber regener- increase in NIS-Lower Limbs [30], patients with hATTR ation and amyloid regression in this study may also amyloidosis and a baseline NIS similar to that in the highlight a potential effect of patisiran on disease patho- current study have an estimated 26-point increase in physiology. However, it should be noted that these were mNIS+7 after 24 months [31]. Indeed, patients in the exploratory analyses, and that there are currently no nat- APOLLO placebo arm demonstrated a 28-point increase in ural history data to contextualize the impact of patisiran mNIS+7 over 18 months, highlighting the urgency to treat on nerve-fiber regeneration. patients with hATTR amyloidosis as early in the disease Multiple additional clinical measures stabilized over course as possible [10]. Although the current study did not 24 months, and indeed all measures, whether based have a comparator arm, the mean 6.95-point improvement on neurologists’ assessments (mNIS+7), patient-based in mNIS+7 at 24 months contrasts markedly from the performance (timed 10-m walk), or blinded assessment of expected worsening without active treatment. Notably, pathologic samples (skin biopsy), were similar or im- the magnitude of mNIS+7 score improvement in this proved at 24 months compared with baseline. Addition- study is similar to that observed following 18 months ally, stable COMPASS-31 autonomic symptom scores of patisiran treatment in APOLLO (6.0-point im- over the course of the OLE support a stabilizing effect of provement), supporting the consistent effects of pati- patisiran on hATTR amyloidosis-related autonomic dys- siran on polyneuropathy. In this study, these data function, which is known to impart a substantial disease indicate that patisiran can halt or reverse neuropathy, burden in hATTR amyloidosis. Notably, measures of as 19 of 27 patients showed a decrease from baseline cardiac structure and function were generally unchanged in mNIS+7 at 24 months, demonstrating a reversal in after 24 months of patisiran dosing in patients with car- polyneuropathy manifestations in the majority of pa- diac involvement, contrasting to the worsening of these tients. Similarly, in the APOLLO study the majority parameters observed without active treatment. The of patients demonstrated an improvement in mNIS+7 improvements to neuropathy measures in the cardiac score compared with baseline [10]. subgroup observed in this study further support the Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 8 of 10 Fig. 3 Change in sweat gland nerve-fiber density and dermal amyloid burden of the lower limb. a Change in sweat gland nerve-fiber density in distal thigh and distal leg to 24 months. b Distal thigh sweat gland innervation at baseline and at Month 24 in an individual patient, with nerve fibers immunostained for PGP 9·5 (green), blood vessels immunostained for CD31 (red), and cell nuclei labeled with DAPI (blue). c Change from baseline to 24 months in amyloid burden for the lower limb. The statistical significance (performed as a post hoc analysis) of the change from baseline is shown for each time-point where: *p = 0.01–0.05, **p = 0.001–0.01. BSL, baseline; CD31, cluster of differentiation 31 protein (or platelet endothelial cell adhesion molecule [PECAM1]); DAPI, 4′,6-diamidino-2-phenylindole; PGP, protein gene product positive effect of patisiran in patients with a mixed require premedication. Expanding the potential range of phenotype, which was also observed in the APOLLO pharmacotherapies for ATTR amyloidosis will provide study [10, 24]. greater treatment choice, allowing physicans and patients The beneficial effect and tolerability of patisiran treat- to select the option most appropriate for this debilitating ment, reported here and in the APOLLO study, demon- disease. strate the clinical impact of reducing TTR in patients with hATTR amyloidosis. This extended experience, coupled Conclusions with secondary analyses of the APOLLO data suggesting In summary, these results demonstrate that long-term benefits in a cardiac subpopulation [24], support examin- treatment with patisiran had an acceptable benefit:risk ing the effect of patisiran on patients with ATTR amyloid- profile, and provide further data that it may halt or re- osis with cardiomyopathy (APOLLO-B trial [NCT039973 verse neuropathy progression as well as have beneficial 83]) and the development of the investigational RNAi effects on QOL, physical functioning, autonomic symp- therapeutic vutrisiran. The vutrisiran siRNA utilizes next- toms, and activities of daily living by reducing TTR generation enhanced stabilization chemistry (ESC), and is levels. The safety and efficacy of patisiran is consistent conjugated to a trivalent GalNAc ligand to ensure hepatic with that reported in APOLLO (NCT01960348), the lar- delivery [35]. The ESC platform is designed for increased gest Phase III clinical study to enroll patients with potency and high metabolic stability, allowing for a quar- hATTR amyloidosis with polyneuropathy [10]. These terly subcutaneously administered injection that does not data resulted in the approval of patisiran in select Coelho et al. Orphanet Journal of Rare Diseases (2020) 15:179 Page 9 of 10 countries globally for the treatment of hATTR amyloid- Availability of data and materials osis with polyneuropathy. All 25 eligible patients from Authors can confirm that all relevant data are included in the article and its supplementary information files. this Phase II OLE study have since enrolled in the pati- siran Global OLE study (NCT02510261), which will con- Ethics approval and consent to participate tinue to collect long-term safety and efficacy data for an This study was conducted according to the guidelines of the International additional 5 years. Conference on Harmonisation, the World Health Organization Declaration of Helsinki, and the Health Insurance Portability and Accountability Act of 1996. Written informed consent was obtained from all participants in the study. Supplementary information The study protocol was approved by local Institutional Review Boards and Supplementary information accompanies this paper at https://doi.org/10. Ethics Committees, and all subsequent protocol amendments underwent the 1186/s13023-020-01399-4. same approval procedure. Additional file 1. Supplementary methods Consent for publication Not applicable. Additional file 2: Table S1. Summary of serious adverse events. Seven patients reported a total of 18 serious adverse events. Additional file 3: Table S2. Summary of safety in cardiac subgroup. Competing interests TC reports compensation for training from Pfizer Pharmaceuticals and Additional file 4: Table S3. Summary of mean serum TTR percent Alnylam Pharmaceuticals Inc., and compensation for travel from Pfizer reduction by subgroup analysis. Pharmaceuticals, Alnylam Pharmaceuticals Inc., and IONIS Pharmaceuticals. Additional file 5: Fig. S1. Mean change from baseline (SEM) in NIS (a) DA reports grants from Alnylam Pharmaceuticals Inc., and personal fees from and NIS+7 (b) scores from the all-treated population over 24 months. Pfizer Pharmaceuticals and Alnylam Pharmaceuticals Inc. outside the Additional file 6: Table S4. Change from baseline in PND score and submitted work. IC acknowledges financial support as a primary investigator FAP stage at 24 months. for FoldRx Pharmaceuticals/Pfizer Inc., Alnylam Pharmaceuticals Inc., and IONIS Pharmaceuticals, and serves on the Transthyretin Amyloidosis Outcome Survey (THAOS) scientific advisory board. MWC received honorarium from NHI, Prothena, FoldRx, Akcea Therapeutics, Pfizer Abbreviations Pharmaceuticals, Alnylam Pharmaceuticals Inc., PTC, and Genzyme for travel AE: Adverse event; ATTR: Transthyretin-mediated amyloidosis; BP: Blood expenses related to presentations at medical meetings, and for acting as a pressure; BSL: Baseline; CD31: Cluster of differentiation 31 protein (or platelet consultant and as a principal investigator in clinical trials. HHS reports grants endothelial cell adhesion molecule [PECAM1]); COMPASS-31: Composite and consulting fees from Alnylam Pharmaceuticals Inc. outside the Autonomic Symptom Score-31; DAPI: 4′;6-diamidino-2-phenylindole; EQ- submitted work. JBu reports personal fees from Alnylam Pharmaceuticals Inc. 5D: EuroQoL 5-dimensions questionnaire; FAP: Familial amyloid during the conduct of the study for serving as a principal investigator. JCo polyneuropathy; hATTR: Hereditary transthyretin-mediated; IENFD: Intra- has nothing to disclose. JBe reports serving as a study investigator for IONIS epidermal nerve-fiber density; IRR: Infusion-related reaction; LV: Left Pharmaceuticals and Pfizer Pharmaceuticals, and consultant for Intellia ventricular; mBMI: Modified body mass index; mNIS: Modified Neuropathy Therapeutics Inc. and Corino Therapeutics Inc. outside the submitted work. Impairment Score; mNIS+7: Modified Neuropathy Impairment Score +7; MP reports grants from Alnylam Pharmaceuticals Inc. during the conduct of NCS: Nerve conduction studies; NIS: Neuropathy Impairment Score; the study and honorarium following completion of the study. JJW, JCh, MTS, NIS+7: Neuropathy Impairment Score +7; NT-proBNP: N-terminal pro-brain and JG are all Alnylam employees. OBS reports personal fees from Alnylam natriuretic peptide; OLE: Open-label extension; OLT: Orthotopic liver Pharmaceuticals Inc. during the conduct of the study, personal fees and non- transplantation; PD: Pharmacodynamic; PGP: Protein gene product; financial support from Pfizer Pharmaceuticals, and personal fees from PND: Polyneuropathy disability; Q3W: Every 3 weeks; QOL: Quality of life; Prothena Pharmaceuticals and Intellia Therapeutics, Inc. outside the submit- QST: Quantitative sensory testing; RBP: Retinol-binding protein; R-ODS: Rasch- ted work. built Overall Disability Scale; RNAi: RNA interference; SEM: Standard error of the mean; SGNFD: Sweat gland nerve-fiber density; TTR: Transthyretin; Author details wt: Wild-type 1 Hospital de Santo António, Centro Hospitalar do Porto, 4099-001 Porto, Portugal. 2National Reference Centre for Familial Amyloidotic Polyneuropathy Acknowledgements (NNERF)/APHP/INSERM U 1195/CHU Bicêtre, 78 rue du Général Leclerc, 94270 The authors would like to thank investigators Jean Pouget, Cecile Cauquil, Le Kremlin-Bicêtre, France. 3Hospital de Santa Maria-CHULN, and IMM, Céline Labeyrie, and Marie Théaudin for their valued contribution to this Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal. 4Hospital study. The authors would also like to thank Jennifer Willoughby for editorial Universitário Clementino Fraga Filho, Federal University of Rio de Janeiro, Ilha assistance. This study was funded by Alnylam Pharmaceuticals Inc., USA. The do Fundao, Rio de Janeiro CEP21941-913, Brazil. 5Universitätsklinikum authors acknowledge the medical writing services provided by Neil Harrison, Münster, Waldeyerstr. 1, 48149 Munster, Germany. 6Fundació Institut PhD, and Kristen Brown, PhD, of Adelphi Communications Ltd., Macclesfield, d’Investigació Sanitària Illes Balears (IdISBa), Carretera de Valldemossa, 79, UK, in accordance with the Good Publication Practice (GPP3) guidelines, Palma de Mallorca 07120, Balearic Islands, Spain; Servicio de Medicina funded by Alnylam Pharmaceuticals Inc. Interna, Hospital Universitario Son Llàtzer, Carretera Manacor KM, 7198 Palma de Mallorca, Balearic Islands, Spain. 7Hospital Clinic, University of Barcelona, C/ Villarroel, 170, 8036 Barcelona, Spain. 8Boston University, 72 East Concord Authors’ contributions Street, K-504, Boston 02118, USA. 9Johns Hopkins University, 855 North Wolfe TC, DA, IC, MWC, HHS, JBu, JCo, JBe, MP, and OBS contributed to the Street, Rangos 435, Baltimore, MD 21205, USA. 10Alnylam Pharmaceuticals, conception and design of the study and/or acquired the data. JCh provided 300 Third Street, Cambridge, MA 02142, USA. 11Umeå University, statistical analysis of the data. All authors contributed to the interpretation of Universitetstorget 16, 901 87 Umeå, Sweden. the data and the critical revision of the publication, are accountable for the accuracy and integrity of the publication, and provided final approval to Received: 30 September 2019 Accepted: 4 May 2020 submit for publication. 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