Supplementary Appendix

Transcription

1 Supplementary Appendix This appendix has been provided by the authors to give readers additional information about their work. Supplement to: Gaudet D, Alexander VJ, Baker BF, et al. Antisense inhibition of apolipoprotein C-III in patients with hypertriglyceridemia. N Engl J Med 2015;373: DOI: /NEJMoa

2 SUPPLEMENTARY APPENDIX Antisense Inhibition of APOC3 in Patients with Hypertriglyceridemia Gaudet D, et al. Table of Contents Study Investigators. 2 Additional Details on Methods... 3 Figure S1. Flow of study participants... 6 Figure S2. Association between total plasma TG and APOC3 levels for all patients by dose group, and by patients treated with 300 mg ISIS , in the (A, B) monotherapy, (C, D) add-on to fibrate, and (E, F) both cohorts Figure S3. Mean percentage change from baseline over time in the ISIS add-on to fibrate cohort by dose group for (A) APOC3, (B) triglyceride, and (C) HDL-cholesterol Figure S4. Individual TG levels at end of treatment in the ISIS add-on to fibrate cohort by (A) percent change from baseline, and (B) absolute level Table S1. Patient disposition (A) Baseline characteristics, (B) Heterozygous loss-of function LPL mutations, and (C) APOE genotypes Table S2. Correlations between APOC3 and total TG for the absolute and percent change from baseline values over time Table S3. Effect of ISIS treatment on primary and secondary outcomes Table S4. Effect of ISIS treatment on VLDL-apoB and LDL-apoB levels Table S5. Effect of ISIS treatment on (A) Chylo/VLDL, (B) LDL and (C) HDL particles. 18 Table S6. Adverse events in the ISIS (A) monotherapy and (B) add-on to fibrate cohorts Table S7. Percentage of injections with local cutaneous reactions in the ISIS (A) monotherapy and (B) add-on to fibrate cohorts References P a g e

3 Study Investigators (NCT , ISIS CS2) Daniel Gaudet, MD, PhD, study principle investigator of Université de Montréal and Ecogene- 21Clinical Research Center (Chicoutimi Qc, CAN) Business Associate Principal Investigators of CTMG, Inc., a Site-Specific Research Organization (SSRO): Mark R. Cervi, MD of CTMG, Inc. (Greenville NC) Natalie A. Doyle, MD of CTMG, Inc. (Wilson NC) Gary H. Heck, DO of CTMG, Inc. (Kinston NC) Charles E. Jahrsdorfer, MD of CTMG, Inc. (Greenville NC) Enrique V. Marana, MD of CTMG, Inc. (Farmville NC) Richard Z. Shultzaberger, MD of CTMG, Inc. (Greenville NC) 2 P a g e

4 Additional Details on Methods STUDY DRUG ISIS (5 -AGCTTCTTGTCCAGCTTTAT-3 ) is a second-generation antisense inhibitor of APOC3 synthesis that incorporates several chemical modifications to improve potency, duration of action, and tolerability. All of the internucleotide phosphates are chemically modified with a phosphorothioate substitution, in which one of the non-bridging oxygen atoms is substituted with sulfur. Additionally the drug incorporates five 2 -O-(2-methoxyethyl) (2 -MOE) modified ribonucleosides (underlined) at the 3 and 5 ends while retaining ten 2 -O-deoxyribonucleosides within the central portion of the molecule. ISIS was supplied by Isis Pharmaceuticals Inc. (Carlsbad, CA, USA) in 2-mL stoppered glass vials as a 1-mL solution (200 mg/ml) for single use only. Patients, investigator(s), and study staff were blinded to the treatment assignment, except for the pharmacist who prepared the drug. Placebo was 0.9% sterile saline. ISIS , or placebo, was administered as a single subcutaneous injection once a week for 13 weeks at the specified dose. STUDY PARTICIPANTS Study participants were 18 years old, had a body mass index (BMI) 40 kg/m 2, and had no current diagnosis or history of endocrine, hematologic, renal, hepatic, metabolic, psychiatric, neurologic, pulmonary, or cardiovascular disease, including any condition that pre-disposed to secondary hyperlipidemia, such as uncontrolled diabetes mellitus or hypothyroidism, nor were they taking drugs that increase TG levels. After a 6 to 8-week diet run-in period, eligible patients not on a TG-lowering therapy had fasting TG 350 mg/dl (4.0 mmol/l) and 2000 mg/dl (22.6 mmol/l), and those on a stable dose of fibrate had fasting TG 225 mg/dl (2.5 mmol/l) and 2000 mg/dl (22.6 mmol/l). STUDY DESIGN A randomized, double-blind, placebo-controlled, dose-ranging study design was used to evaluate the effects of ISIS in patients with severe or uncontrolled hypertriglyceridemia. Eligible patients not on fibrate therapy were assigned to the ISIS monotherapy cohort, and those on a stable dose of fibrate were assigned to the add-on to fibrate cohort. ISIS monotherapy patients were randomized equally (1:1:1) to receive doses of 100, 200, and or 300 mg in a 3:1 active to placebo ratio. Eligible patients on a fibrate were randomized equally (1:1) to receive 200 or 300 mg ISIS , and in a 2:1 active-to-placebo ratio. Patients received 13 doses of study 3 P a g e

5 drug by SC injection once a week for 13 weeks, and then were followed in post-treatment evaluation for 13 weeks. All enrolled patients received diet/alcohol counseling and monitoring for the duration of study participation. Patients were allowed to continue ongoing stable doses of statin therapy and/or oral antidiabetic medication. Alcohol consumption was not allowed for 48 hours prior to visits where lipids were measured. In addition, patients were provided a standardized pre-cooked meal for the dinner (after which they remained fasted for at least 10 hours) prior to these visits. Samples were collected before dosing during the treatment period. OUTCOMES The primary outcome was the percent change in fasting total APOC3 levels from baseline to the end of treatment (average of Day 85 and Day 92). Secondary outcomes included percent change in fasting TG, total cholesterol (TC), high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, very low-density lipoprotein (VLDL) cholesterol (density gradient <1.006 g/ml), non-hdl cholesterol, total apolipoprotein B (apob), apolipoprotein B-48 (apob-48) and VLDL- APOC3 (density gradient <1.006 g/ml), from baseline to the end of treatment, as well as over time for APOC3 and these other listed parameters. Safety was assessed by determining the incidence, severity, and dose relationship of adverse events (AEs) and changes in laboratory parameters. Safety laboratory evaluations included routine hematology, blood chemistry, urinalysis, and coagulation parameters. Other assessments included a physical examination, a 12 lead electrocardiogram, and vital signs. LIPID MEASUREMENTS Fasted blood samples were collected for measurement of lipid parameters at baseline, on Day 8 and then weekly to every other week during the treatment period (immediately prior to administration of ISIS ), and on Days 92, 99, 127 and 176 during the post-treatment follow-up period. Total plasma APOC3, chylomicron/vldl APOC3 (density gradient <1.006 g/ml), TGs, total cholesterol, HDL cholesterol (precipitated), non-hdl cholesterol (calculated), LDL cholesterol (isolated by ultracentrifugation), chylomicron/vldl cholesterol (density gradient <1.006 g/ml), and apob-48 levels were determined at MedPace Reference Labs (Cincinnati, OH). Triglycerides and cholesterol were measured by standard enzyme-based colorimetric assays. APOC3, total apob and apob-48 were measured by rate nephelometry. Non-HDL cholesterol was calculated. VLDL and LDL 4 P a g e

6 fractions were isolated from samples by ultracentrifugation, and then assayed for apolipoprotein B content (VLDL-apoB and LDL-apoB) at Ecogene-21 (Chicoutimi, QC Canada). GENOTYPING LPL and APOE genotyping were performed at Ecogene-21 (Chicoutimi, QC Canada). STATISTICAL ANALYSIS Primary and secondary pharmacodynamic outcomes were analyzed per protocol. These analyses included all patients who received at least 9 doses of study drug and had no significant protocol deviations that would be expected to affect the outcome measures. Analyses evaluating safety of the intervention included all randomized patients who received at least one dose of study drug. The primary analysis compared the percent change in fasting total APOC3 levels from baseline to the end of treatment (primary outcome) in the 300 mg dose group of the monotherapy cohort, compared to that of the placebo group. Secondary analyses included comparisons of the primary outcome for other dose groups to the placebo group of each cohort. Similar analyses were done for the secondary outcomes. These latter analyses were exploratory in nature and performed without adjustment for multiple comparisons. Least squares means were estimated from a mixed-effect model repeated-measures analysis of a modified intent-to-treat population, which included all dosed patients with at least one post-baseline APOC3 measurement. P-values were determined from the Wilcoxon Rank Sum test or t-test depending on normality of the data. Normality was determined using the Kolmogorov-Smirnov test. All statistical tests were unpaired and two-sided, with a significance level of 0.05 for the primary analysis. Post-hoc correlation coefficients (r) were derived from Pearson product-moment correlations. Software utilized for the analyses was SAS version 9.2 or higher (SAS Institute). Sample size was based on a standard deviation of the percent change in total APOC3 of approximately 46% as estimated from prior clinical experience. Under these assumptions, there was estimated to be at least 80% power to detect a 45% difference in total APOC3 between ISIS and placebo group at an alpha level of 0.05, assuming 50% reduction in patients receiving ISIS and 5% in the placebo group. 5 P a g e

7 Figure S1. Flow of Study Participants Abbreviations: w/c, withdrew consent; AE, adverse event; PD, pharmacodynamics. Analysis of PD outcomes was performed on the per-protocol population as defined in the methods section. * Patients assigned to ISIS monotherapy were randomized equally (1:1:1) to receive doses of 100, 200 or 300 mg, and then 3:1 active to placebo. Patients assigned to ISIS add-on to fibrate were randomized equally (1:1) to receive doses of 200 or 300 mg, and then 2:1 active-to-placebo. Principle reason for dose discontinuation: injection site reaction (moderate); abdominal distension (severe); mononucleosis (moderate); musculoskeletal pain (mild); ** serum sickness-like reaction (moderate); fatigue (mild). 6 P a g e

8 Figure S2. Association between total plasma TG and APOC3 levels for all patients by dose group, and by patients treated with 300 mg ISIS , in the (A, B) monotherapy, (C, D) add-on to fibrate, and (E, F) both cohorts. 7 P a g e

9 Figure S3. Mean percentage change from baseline over time in the ISIS add-on to fibrate cohort by dose group for (A) APOC3, (B) triglyceride, and (C) HDL-cholesterol. Solid blue triangles indicate dosing days. 8 P a g e

10 Figure S4. Individual TG levels at end of treatment in the ISIS add-on to fibrate cohort by (A) percent change from baseline, and (B) absolute level. 9 P a g e

11 Table S1. Patient Disposition. (A) Baseline characteristics, (B) Heterozygous loss-of-function LPL mutations, and (C) APOE genotypes A ISIS Monotherapy ISIS Add-on to Fibrate * Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Characteristic (N=16) (N=13) (N=15) (N=13) (N=8) (N=8) (N=12) Gender, F:M 3:13 5:8 1:14 5:8 4:4 2:6 5:7 Age, years 48.6 ± ± ± ± ± ± ±13.8 BMI, kg/m ± ± ± ± ± ± ±4.6 Statins, n (%) 4 (25.0) 6 (46.2) 4 (26.7) 5 (38.5) 4 (50.0) 4 (50.0) 4 (33.3) Maximal Statin 2 (12.5) 2 (15.4) 2 (13.3) 2 (15.4) 1 (12.5) 4 (50.0) 0 (0.0) Triglycerides, mg/dl ± ± ± ± ± ± ±207.4 Median (IQR) 459 (356, 582) 558 (351, 649) 588 (464, 858) 566 (422, 702) 475 (253, 602) 272 (230, 295) 302 (246, 451) HDL-C, mg/dl 33.0 ± ± ± ± ± ± ±11.5 LDL-C, mg/dl ± ± ± ± ± ± ±27.7 Results presented are based on analysis of the Safety population. Values are the mean ± standard deviation, except where indicated. IQR denotes interquartile range, Q1 and Q3. * 25 of 28 (89%) patients were on stable doses of fenofibrate (median daily dose = 145 mg); 3 of 28 (11%) patients were on stable doses of gemfibrozol (median daily dose = 1200 mg). Maximal statin (total daily dose): 40 mg and above of rosuvastatin, atorvastatin, pravastatin, or simvastatin. 10 P a g e

12 Table S1 (cont). B ISIS Monotherapy ISIS Add-on to Fibrate LPL Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Mutation, n (%) (N=16) (N=13) (N=15)* (N=13) (N=8) (N=8) (N=12) Heterozygous mutation 4 (25.0) 5 (38.5) 1 (6.7) 4 (30.8) 2 (25.0) 2 (25.0) 2 (16.7) P207L 2 (12.5) 4 (30.8) 1 (6.7) 2 (15.4) 1 (12.5) 1 (12.5) 2 (16.7) D9N 3 (18.8) 1 (7.7) 0 (0.0) 1 (7.7) 0 (0.0) 1 (12.5) 1 (8.3) G188E 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) N291S 0 (0.0) 0 (0.0) 0 (0.0) 1 (7.7) 1 (12.5) 0 (0.0) 0 (0.0) Results are based on analysis of the Safety population. *Data not available for 2 patients; Data not available for 3 patients. C ISIS Monotherapy ISIS Add-on to Fibrate APOE Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Genotype, n (%) (N=16) (N=13) (N=15)* (N=13) (N=8) (N=8) (N=12) E2/2 1 (6.3) 1 (7.7) 0 (0.0) 2 (15.4) 3 (37.5) 1 (12.5) 1 (8.3) E2/3 3 (18.8) 2 (15.4) 2 (13.3) 1 (7.7) 2 (25.0) 0 (0.0) 2 (16.7) E2/4 0 (0.0) 0 (0.0) 2 (13.3) 0 (0.0) 0 (0.0) 0 (0.0) 0 (0.0) E3/3 6 (37.5) 3 (23.1) 4 (26.7) 1 (7.7) 1 (12.5) 4 (50.0) 3 (25.0) E3/4 6 (37.5) 7 (53.8) 5 (33.3) 6 (46.2) 1 (12.5) 3 (37.5) 2 (16.7) E4/4 0 (0.0) 0 (0.0) 0 (0.0) 3 (23.1) 1 (12.5) 0 (0.0) 1 (8.3) Results are based on analysis of the Safety population. *Data not available for 2 patients; Data not available for 3 patients. 11 P a g e

13 Table S2. Correlations between APOC3 and total TG for the absolute and percent change from baseline values over time. Monotherapy Add-on to Fibrate Both Cohorts APOC3 vs TG All Patients 300 mg ISIS All Patients 300 mg ISIS All Patients 300 mg ISIS Absolute % Change P a g e

14 Table S3. Effect of ISIS treatment on primary and secondary outcomes ISIS Monotherapy ISIS Add-on to Fibrate Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Parameter (N=16) (N=11) (N=13) (N=11) (N=8) (N=8) (N=10) APOC3, mg/dl Baseline 22.2 ± ± ± ± ± ± ±5.9 Endpoint 21.9 ± ± ± ± ± ± ±2.1 % Change 4.2 ± ±32.0* ± ± ± ± ±13.0 LS Mean ±SE 4.2 ± ± ± ± ± ± ±5.3 Triglycerides, mg/dl Baseline ± ± ± ± ± ± ±227.8 Median (IQR) 459 (356, 582) 558 (351, 825) 588 (464, 717) 566 (355, 702) 475 (253, 602) 272 (230, 295) 271 (241, 459) Endpoint ± ± ± ± ± ± ±60.9 Median (IQR) 456 (343, 640) 277 (226, 394) 180 (157, 262) 120 (115, 185) 338 (290, 394) 120 (103, 172) 116 (91, 191) % Change 20.1 ± ±56.8* ± ± ± ± ±8.9 Median (IQR) 12.1 (-24.6, 28.9) (-71.5, -7.0) (-79.5, -43.4) (-82.8, -67.0) (-29.8, 9.7) (-63.5, -39.8) (-70.7, -54.6) LS Mean ±SE 20.1 ± ±15.1* ± ± ± ± ± P a g e

15 ISIS Monotherapy ISIS Add-on to Fibrate Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Parameter (N=16) (N=11) (N=13) (N=11) (N=8) (N=8) (N=10) Total Cholesterol, mg/dl Baseline ± ± ± ± ± ± ±38.8 Endpoint ± ± ± ± ± ± ±54.7 % Change 4.1 ± ± ± ± ± ± ±23.2 LS Mean ±SE 4.1 ± ± ± ± ± ± ±5.7 HDL-C, mg/dl Baseline 33.0 ± ± ± ± ± ± ±12.3 Endpoint 33.2 ± ± v ± ± ± ±19.9 % Change 0.7 ± ± ± ± ± ±32.0* ±23.7 LS Mean ±SE 0.7 ± ± ± ± ± ± ±8.2 Non-HDL-C, mg/dl Baseline ± ± ± ± ± ± ±44.4 Endpoint ± ± ± ± ± ± ±64.2 % Change 6.2 ± ± ± ± ± ± ±28.8 LS Mean ±SE 6.2 ± ± ± ± ± ± ± P a g e

16 ISIS Monotherapy ISIS Add-on to Fibrate Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Parameter (N=16) (N=11) (N=13) (N=11) (N=8) (N=8) (N=10) LDL-C, mg/dl Baseline ± ± ± ± ± ± ±26.2 Endpoint ± ± ± ± ± ± ±62.1 % Change 10.8 ± ±39.0* 79.4 ± ±177.3* 5.3 ± ± ±49.7 LS Mean ±SE 10.8 ± ± ± ± ± ± ±12.4 VLDL-C, mg/dl Baseline ± ± ± ± ± ± ±48.2 Endpoint 96.5 ± ± ± ± ± ± ±15.3 % Change 6.3 ± ±41.8* ± ± ± ±22.5* ±21.6* LS Mean ±SE 6.3 ± ± ± ± ± ± ±10.1 VLDL-APOC3, mg/dl Baseline 11.6 ± ± ± ± ± ± ±2.6 Endpoint 12.1 ± ± ± ± ± ± ±1.0 % Change 21.0 ± ±65.6* ± ± ± ± ±11.4 LS Mean ±SE 21.0 ± ± ± ± ± ± ± P a g e

17 ISIS Monotherapy ISIS Add-on to Fibrate Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Parameter (N=16) (N=11) (N=13) (N=11) (N=8) (N=8) (N=10) Total ApoB, mg/dl Baseline ± ± ± ± ± ± ±27.0 Endpoint ± ± ± ± ± ± ±45.8 % Change 3.9 ± ± ± ± ± ± ±27.7 LS Mean ±SE 3.9 ± ± ± ± ± ± ±6.9 ApoB-48, mg/dl Baseline 0.7 ± ± ± ± ± ± ±0.2 Endpoint 1.0 ± ± ± ± ± ± ±0.2 % Change 64.5 ± ±45.5* ± ± ± ± ±20.1 Results presented are based on the Per-Protocol population, except for the LS mean percent change from baseline results. Values are the means ± standard deviations in mg/dl, except where indicated. Baseline is defined as the Day -8 value. Endpoint represents the average of the Day 85 and Day 92 values. Percent (%) change represents from Baseline to Endpoint. IQR denotes interquartile range, Q1 and Q3; LS, least squares; SE, standard error. To convert apolipoprotein values (APOC3, VLDL-APOC3, ApoB, and ApoB-48) to g/l multiply by 0.01; to convert triglyceride (TG) values to mmol/l multiply by ; to convert cholesterol (C) values to mmol/l, multiply by P-value * <0.05; <0.01; < P a g e

18 Table S4. Effect of ISIS treatment on VLDL-apoB and LDL-apoB Levels ISIS Monotherapy ISIS Add-on to Fibrate ApoB, mg/dl Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Mean ±SD (N=16) (N=11) (N=13) (N=11) (N=8) (N =8) (N =10) VLDL-apoB Baseline 20.1 ± ± ± ± ± ± ±11.8 Endpoint 16.5 ± ± ± ± ± ± ±4.0 % Change -7.6 ± ± ± ± ± ± ±370.2* Median (IQR) (-44.3, 21.4) (-51.7, 13.8) (-82.4, 3.1) (-77.3, -50.0) (-31.1, -14.8) (-75.0, -30.0) (-80.6, -42.9) LDL-apoB Baseline 95.8 ± ± ± ± ± ± ±23.2 Endpoint 95.6 ± ± ± ± ± ± ±44.0 % Change 8.5 ± ± ± ± ± ± ±32.0 Median (IQR) 4.9 (-13.4, 16.2) 15.7 (8.9, 31.9) 28.9 (15.3, 53.8) 41.8 (-2.7, 81.5) 6.1 (-21.4, 17.6) 2.7 (-15.3, 16.8) -1.7 (-4.5, 30.6) Results presented are based on analysis of the Per-Protocol population. Values are the means ± standard deviations in mg/dl, except for % change which shows both the mean ±standard deviation and median (interquartile range) results. Baseline is defined as the average of Day -8 and Day -1, or Day 1, pre-dose results. Endpoint represents the average of the Day 85 and Day 92 values. To convert apolipoprotein values to g/l multiply by P-values were determined for percent change from baseline at end point, vs placebo (* <0.05; <0.01; <0.001). 17 P a g e

19 Table S5. Effect of ISIS treatment on lipoprotein particle size: Chylomicron/VLDL, LDL and HDL particles ISIS Monotherapy ISIS Add-on to Fibrate Size, nm Placebo 100 mg 200 mg 300 mg Placebo 200 mg 300 mg Median (Q1,Q3) (N=16) (N=11) (N=13) (N=11) (N=8) (N=8) (N=10) Chylo/VLDL-P Baseline 63.4 (58.1, 71.1) 58.1 (51.2, 68.4) 66.8 (62.1, 71.1) 64.2 (61.5, 70.3) 56.9 (54.7, 58.8) 58.2 (52.8, 62.1) 57.7 (49.5, 62.9) Day (56.2, 70.9) 56.0 (48.2, 61.6) 56.7 (49.5, 60.6) 53.8 (48.6, 57.1) 53.8 (50.0, 58.0) 49.7 (45.3, 59.4) 51.5 (48.1, 55.9) Change 1.0 (-3.5, 3.1) -4.0 (-9.1, 2.6) (-15.3, -4.2) (-18.5, -6.3) -3.6 (-5.3, -0.1) -7.1 (-8.7, 0.0) -4.2 (-14.1, -1.2) LDL-P Baseline 19.6 (19.6, 19.7) 19.6 (19.4, 19.6) 19.5 (19.5, 19.6) 19.6 (19.6, 19.6) 19.7 (19.6, 19.8) 19.7 (19.7, 20.0) 19.7 (19.6, 19.8) Day (19.5, 19.6) 19.6 (19.6, 20.0) 20.3 (19.8, 20.6) 20.1 (19.7, 20.5) 19.7 (19.6, 20.1) 20.6 (20.3, 20.6) 20.3 (19.8, 21.1) Change -0.1 (-0.1, 0.0) 0.2 (0.0, 0.3) 0.7 (0.2, 1.1) 0.4 (0.1, 0.8) 0.0 (-0.1, 0.5) 0.6* (0.5, 0.9) 0.7* (0.3, 1.3) HDL-P Baseline 8.9 (8.6, 9.2) 8.9 (8.7, 9.1) 8.8 (8.8, 9.1) 8.8 (8.5, 9.1) 8.7 (8.4, 8.8) 8.6 (8.5, 8.7) 8.8 (8.6, 9.0) Day (8.7, 9.2) 8.8 (8.7, 9.2) 8.5 (8.5, 8.9) 8.7 (8.6, 8.8) 8.7 (8.5, 8.8) 8.5 (8.3, 8.8) 9.1 (8.6, 9.2) Change 0.0 (-0.1, 0.2) 0.1 (-0.4, 0.2) -0.3 (-0.5, -0.1) -0.1 (-0.6, 0.4) 0.0 (0.0, 0.2) 0.0 (-0.1, 0.2) 0.3 (-0.4, 0.4) Particle parameters were derived by NMR spectroscopy analysis (LipoScience Inc., Raleigh NC). 1 Values shown are the median (interquartile range). Change is from baseline to Day 92. Chylo/VLDL-P denotes chylomicron/very-low-density lipoprotein particle; LDL-P, low-density lipoprotein particle; HDL-P, high-density lipoprotein particle. P-values * <0.05; <0.01; <0.001 (vs placebo). 18 P a g e

20 Table S6. Adverse events in the ISIS (A) monotherapy and (B) add-on to fibrate cohorts* A 100 mg 200 mg 300 mg Total Placebo ISIS ISIS ISIS ISIS Preferred Term, n (%) (N=16) (N=13) (N=15) (N=13) (N=41) Fatigue 1 (6.3) 2 (15.4) 2 (13.3) 2 (15.4) 6 (14.6) Musculoskeletal pain 0 (0.0) 0 (0.0) 2 (13.3) 2 (15.4) 4 (9.8) Nausea 0 (0.0) 0 (0.0) 0 (0.0) 4 (30.8) 4 (9.8) Chills 0 (0.0) 1 (7.7) 1 (6.7) 1 (7.7) 3 (7.3) Myalgia 0 (0.0) 1 (7.7) 1 (6.7) 1 (7.7) 3 (7.3) B 200 mg 300 mg Total Placebo ISIS ISIS ISIS Preferred Term, n (%) (N=8) (N=8) (N=12) (N=20) Diarrhoea 1 (12.5) 2 (25.0) 0 (0.0) 2 (10.0) Abdominal pain upper 0 (0.0) 0 (0.0) 2 (16.7) 2 (10.0) Fatigue 0 (0.0) 1 (12.5) 1 (8.3) 2 (10.0) Feeling of Relaxation 0 (0.0) 0 (0.0) 2 (16.7) 2 (10.0) * Adverse events related or possibly related to study drug; events at the injections site are excluded (see Table S7). 19 P a g e

21 Table S7. Percentage of injections with local cutaneous reactions in the ISIS (A) monotherapy and (B) add-on to fibrate cohorts* A 100 mg 200 mg 300 mg Total Placebo ISIS ISIS ISIS ISIS % Injections (N=16) (N=13) (N=15) (N=13) (N=41) Mean (SD) 0.0 (0.0) 11.5 (28.0) 5.4 (7.4) 24.7 (30.5) 13.4 (24.5) Median (Q1, Q3) 0.0 (0.0, 0.0) 0.0 (0.0, 7.7) 0.0 (0.0, 12.5) 15.4 (0.0, 38.5) 0.0 (0.0, 15.4) B 200 mg 300 mg Total Placebo ISIS ISIS ISIS % Injections (N=8) (N=8) (N=12) (N=20) Mean (SD) 0.0 (0.0) 19.2 (31.3) 11.7 (16.4) 14.7 (23.0) Median (Q1, Q3) 0.0 (0.0, 0.0) 3.8 (0.0, 34.6) 3.8 (0.0, 19.2) 3.8 (0.0, 19.2) * Local cutaneous reactions are adverse events at the injection site defined as erythema, swelling, pruritus, pain and/or tenderness, which started on the day of injection and persisted for two days or more. 20 P a g e

22 References 1. Jeyarajah EJ, Cromwell WC, Otvos JD. Lipoprotein particle analysis by nuclear magnetic resonance spectroscopy. Clin Lab Med. 2006;26: P a g e