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N Engl J Med. Author manuscript; available in PMC 2020 January 03.
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Published in final edited form as:

N Engl J Med. 2019 January 03; 380(1): 23–32. doi:10.1056/NEJMoa1811403.

Marine Omega-3 Fatty Acids and Prevention of Vascular Disease

and Cancer
JoAnn E. Manson, MD, DrPH,
Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA

Nancy R. Cook, ScD,

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Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA

I-Min Lee, MBBS, ScD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA

William Christen, ScD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Shari S. Bassuk, ScD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA
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Samia Mora, MD, MHS,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Heike Gibson, PhD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Christine M. Albert, MD, MPH,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

David Gordon, MAT,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Trisha Copeland, MS, RD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA
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Denise D’Agostino, BS,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Georgina Friedenberg, MPH,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

*The VITAL investigator group is listed in the Supplementary Appendix, available at NEJM.org.
Corresponding Author: JoAnn E. Manson, MD, DrPH, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical
School, 900 Commonwealth Avenue, 3rd Fl., Boston, MA; phone: 617-278-0871; fax: 617-731-3843; [email protected].
Conflict of Interest Disclosures: All authors have completed and submitted the ICMJE Form for Disclosure of Potential Conflicts of
Interest.
 Manson et al. Page 2

Claire Ridge, MPH,

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Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Vadim Bubes, PhD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Edward L. Giovannucci, MD, ScD,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Department of Nutrition, Harvard T.H. Chan School of Public Health, Boston, MA

Walter C. Willett, MD, DrPH,

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA

Department of Nutrition, Harvard T.H. Chan School of Public Health, Boston, MA

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Julie E. Buring, ScD, and

Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA

Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA

VITAL Investigators*

Abstract
Background—Whether omega-3 fatty acid supplementation reduces risk of cardiovascular
disease or cancer remains unclear.

Methods—The VITamin D and OmegA-3Trial (VITAL) was a randomized, placebo-controlled,

2X2 factorial trial of vitamin D3 (2000IU/day) and marine omega-3 fatty acids (1 g/day) in the
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primary prevention of cardiovascular disease and cancer among 25,871 U.S. men aged ≥50 and
women aged >55, including 5,106 African Americans. Primary endpoints were major
cardiovascular events (myocardial infarction, stroke, and cardiovascular mortality) and total
invasive cancer. Secondary outcomes included individual components of the cardiovascular
composite, the composite plus coronary revascularization, site-specific cancers, and cancer
mortality. This paper reports the results of omega-3 and placebo.

Results—During a median 5.3 years, rates of the primary outcomes did not differ between the
omega-3 and placebo groups -- 805 participants had a major cardiovascular event, hazard ratio
[HR]= 0.92; 95% confidence interval [CI], 0.80–1.06, p= 0.24. Invasive cancer was diagnosed in
1,617 participants, HR 1.03 (0.93-1.13, p=0.56). In the analysis of key secondary endpoints,
hazard ratios and 95% CIs comparing omega-3 to placebo were: expanded cardiovascular events,
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HR 0.93 (0.82-1.04); total myocardial infarction HR 0.72 (0.59-0.90); total stroke, HR 1.04
(0.83-1.31); cardiovascular mortality HR 0.96 (0.76-1.21); and cancer deaths (n=341, HR 0.97
(0.79-1.20). For all-cause mortality (n=978), the HR was 1.02 (0.90-1.15). No excess risks of
bleeding or other serious adverse events were observed.

Conclusions—Omega-3 fatty acid supplementation did not reduce major cardiovascular events
or cancer incidence.

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Marine-derived long-chain omega-3 ( also called n-3) fatty acids have shown promise for the
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primary prevention of cardiovascular disease in animal studies, small randomized trials

designed with intermediate cardiovascular endpoints, and observational epidemiologic
investigations.1 However, mid-sized to large trials testing the effect of n-3 fatty acid
supplements on clinical cardiovascular outcomes in secondary prevention or high-risk
settings have shown inconsistent results.1, 2 Large trials of n-3 supplements for primary
prevention of cardiovascular disease in a general population selected only on age and not on
vascular risk factors such as diabetes or dyslipidemia are lacking. Data on n-3 fatty acids and
cancer risk have also been inconsistent.3 Given the popularity of fish oil as a strategy to
reduce chronic disease,4 clarifying the relation between supplemental n-3 fatty acids and
risks of cardiovascular disease and cancer and obtaining more definitive data on the benefit-
risk balance of these supplements is a high priority. The VITamin D and OmegA-3 TriaL
(VITAL) was conducted to address these knowledge gaps in a diverse U.S. cohort.
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METHODS
Study Design
This randomized, double-blind, placebo-controlled, 2×2 factorial trial tested the benefits and
risks of vitamin D3 (2000 IU/day) and n-3 fatty acids (1 g/day fish-oil capsule containing
840 mg of n-3 fatty acids including eicosapentaenoic acid [EPA, 460 mg] +
docosahexaenoic acid [DHA, 380 mg]) in the primary prevention of cardiovascular disease
and cancer among 25,871 men aged ≥50 and women aged ≥55, including 5,106 African
Americans. The results are presented in two papers, with details of the full design in the
accompanying paper containing the vitamin D3 data, the Supplementary Appendix, and also
published earlier.5, 6 The protocol is posted at NEJM.org. The n-3 fatty acid dose chosen was
that recommended by the American Heart Association for cardioprotection7 and
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demonstrated as beneficial in a secondary prevention population.8 The recruitment flow

diagram is presented in Figure S1 in the Supplementary Appendix. Randomization to n-3
fatty acids, vitamin D, both active agents, or both placebos was completed in March 2014.
Study medication ceased as planned on December 31, 2017, yielding a median intervention
period of 5.3 years (range 3.8-6.1 years).

Baseline questionnaires collected data on clinical and lifestyle risk factors and included a
dietary questionnaire that ascertained self-reported intake of fish and other foods. Annual
questionnaires assessed adherence to and potential side effects of randomized treatments,
incident major illnesses, and risk factor updates. Baseline blood samples were collected
from all willing participants (n=16,956 of 25,871 [66%]) and were assayed for plasma
omega-3 index (EPA+DHA as a percent of total fatty acids9) by Quest Diagnostics using
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liquid chromatography-tandem mass spectrometry.

Study Endpoints
Primary endpoints were major cardiovascular events (composite of myocardial infarction,
stroke, and cardiovascular mortality) and total invasive cancer. Secondary cardiovascular
endpoints were major cardiovascular events plus coronary revascularization [coronary artery
bypass grafting (CABG) or percutaneous coronary intervention (PCI)] and individual

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components of the primary endpoint. Secondary cancer endpoints were incident colorectal,
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breast, and prostate cancers, and total cancer mortality. Medical records of those reaching
endpoints were reviewed by an Endpoints Committee of physicians blinded to treatment
assignment. Myocardial infarction and stroke were confirmed using established criteria.10, 11
Cancer was confirmed with histologic or cytologic data.12 Additional details regarding
endpoint confirmation can be found in the preceding paper and the Supplementary
Appendix.

Statistical Analysis
Treatment-effect analyses were based on the intention-to-treat principle, as described in the
companion paper on vitamin D. Primary analyses were based on Cox proportional hazards
models controlling for age, sex, and randomization to vitamin D.

Possible variations in treatment effect by age, sex, baseline cardiovascular risk factors,
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baseline dietary fish intake and plasma omega-3 index, and concurrent randomization to
vitamin D were specified a priori. Because vitamin D was also studied, treatment effects in
racial/ethnic groups were of interest. Aspirin and statin use were additional stratification
variables. There was no control for multiple hypothesis testing, with no formal adjustment to
the p-values or confidence intervals. Thus, results for exploratory outcomes and subgroups
should be interpreted with caution. Additional details regarding the analyses are in the
Supplementary Appendix.

RESULTS
Baseline characteristics of study participants are in Table 1 (with further details in Table S1
of the Supplementary Appendix). Of the 25,871 participants, 51% were women and the
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mean age was 67.1 years. The cohort was racially diverse, including 20% African
Americans. Randomization balanced characteristics between groups. The questionnaire
response rate averaged 93.1%, and self-reported treatment adherence rates (percent taking
≥2/3 of the study capsules) in the active and placebo groups averaged 81.6% and 81.5%,
respectively, over 5-year follow-up (Table S2, Supplementary Appendix). The prevalence of
outside use of fish-oil supplements was below 3.5% in both groups throughout follow-up.
Among the 15,535 participants with analyzable baseline blood samples (60%), the mean
plasma omega-3 index was 2.67% (SD, 0.9%) in both groups. Among the 1,583 participants
who also provided an analyzable one-year blood sample, the mean omega-3 index rose to
4.13% (54.7% increase) in the active group and changed <2% in the placebo group.

Cardiovascular Disease
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During follow-up, there were 805 major cardiovascular events, including 386 in the n-3 and
419 in the placebo group (comparing omega-3 with placebo, HR=0.92; 95% confidence
interval, 0.80-1.06; p=0.24) (Table 2). Results of analyses of prespecified secondary
cardiovascular outcomes were as follows: total myocardial infarction (HR=0.72 [0.59-0.90];
cardiovascular mortality (HR=0.96 [0.76-1.21]); total stroke (HR=1.04 [0.83-1.31]); and
expanded cardiovascular composite (HR=0.93 [0.82-1.04]). Additional vascular outcomes
included coronary artery bypass graft (HR=0.99 [0.73-1.33]), percutaneous coronary

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intervention (HR=0.78 [0.63-0.95]); fatal myocardial infarction (HR=0.50 [0.26-0.97]); and

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total coronary heart disease (HR=0.83 [0.71-0.97]) (see Table 2, which also shows results
for stroke subtypes and stroke death).

Cumulative incidence rates of major cardiovascular events are shown in Figure 1. For major
cardiovascular events, the curves did not differ significantly between groups. After excluding
the first 2 years of follow-up, the HR for major cardiovascular events was 0.89 (0.76-1.05),
and the reduction in myocardial infarction persisted (Table 2).

Subgroup analyses showed a possible reduction in the primary cardiovascular outcome with
omega-3 supplementation among those with low fish consumption (Figure 2). Additional
subgroup analyses are presented in the Supplementary Appendix, Tables S3-S4 and Figure
S2, with a focus on exploring differences according to race/ethnicity, diabetes status, number
of traditional cardiovascular risk factors, dietary fish intake, and other variables for the
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primary endpoint of major cardiovascular events and the secondary endpoint of total
myocardial infarction. For myocardial infarction, these are presented as explanatory analyses
to assess whether the risk reduction was similar across subgroups. The suggestion of greater
risk reductions for myocardial infarction among African Americans and those with lower
fish intake, comparing the omega-3 group with the placebo group, is discussed in the
Supplementary Appendix. For the other secondary cardiovascular endpoints of stroke,
cardiovascular mortality, and the expanded composite of major cardiovascular events plus
coronary revascularization, no appreciable effect modification was found (data not shown).

Cancer and All-Cause Mortality

During follow-up, 1,617 participants developed invasive cancer, with similar rates in the n-3
and placebo groups (820 vs. 797 cancers; HR=1.03 [0.93-1.13]; p=0.56) (Table 2). No
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significant differences between randomized groups were observed for breast, prostate, or
colorectal cancer; cancer mortality (RR=0.97 [0.79-1.20]); or all-cause mortality (HR=1.02
[0.90-1.15]).

Cumulative incidence curves for cancer and all-cause mortality did not differ significantly
between treatment groups at any year of follow-up (Fig. 2). Tests for proportionality over
time suggested violation for cancer (p=0.075). In analyses excluding the first two years of
follow-up, the HRs were 1.13 (1.00-1.28) for cancer incidence and 0.93([0.0.73-1.19) for
cancer mortality in the n-3 group (Table 2).

In subgroup analyses, sex may have modified the results for cancer incidence (p-
interaction=0.024) (Table S5). Fish intake at baseline may have modified the intervention’s
effects on all-cause mortality, (p-interaction=0.017; Supplementary Appendix, Table S6).
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There were no other significant interactions for cancer endpoints or all-cause mortality.

Adverse Events
The intervention was not associated with significant differences in gastrointestinal
symptoms, major bleeding episodes, or other serious side effects, compared to placebo
(Supplementary Appendix, Table S7).

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DISCUSSION
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In this 5.3-year primary prevention trial, n-3 supplementation (1 g/day) as compared to

placebo did not lead to a significant reduction in the primary endpoints of major
cardiovascular events (a composite of myocardial infarction, stroke and cardiovascular
mortality) or invasive cancer. Analyses of the components of the primary composite
cardiovascular endpoint suggested a reduction in myocardial infarction and no change in
cardiovascular mortality or stroke. Exploratory analyses excluding the first two years of
follow-up suggested a nonsignificant increase in cancer incidence, but no increase in cancer
mortality.

Meta-analyses of n-3 supplementation trials in adults with or at high risk for cardiovascular
disease have concluded that supplementation has no, or at most a weak, preventive effect on
cardiovascular outcomes, including major vascular events, major coronary events,
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myocardial infarction, stroke, and revascularization.13–15 The recently completed ASCEND

trial,16 which tested n-3 supplementation (1 g/day) in U.K. adults with diabetes, also
reported generally null results. Thus, the possible reduction in the secondary outcomes of
myocardial infarction and PCI in our trial, which tested n-3 fatty acids for primary
prevention in a usual-risk population, raises the question of potential differences in results
between primary and secondary prevention settings. Notably, neither our trial nor the
secondary prevention trials indicate benefit of n-3 supplementation on stroke or composite
cardiovascular endpoints. Our finding of a possible reduction with n-3 supplementation for
the primary cardiovascular outcome among those with low fish consumption -- a
characteristic that has rarely been examined as an effect modifier in previous trials—is
hypothesis-generating and requires further study.

Two early large (n≥10,000) open-label trials8, 17 testing doses of 1 g/day or higher found
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significant coronary protection, but all but one18 of the subsequent placebo-controlled
trials16, 18–23 (some with smaller sample sizes18–21 and lower doses19, 21) did not. The
finding of a possible reduction in our trial may be partly attributable to these design
differences. Also, the prevalence of use of cardiovascular medications, including statins, β-
blockers, and anticoagulants, was higher in recent trials, perhaps reducing the opportunity to
detect incremental benefit. Although a recent meta-analysis14 of n-3 trials found no variation
in results by statin use, dilution of a potential n-3 effect by other medications cannot be ruled
out. Such a dilution would likely be greater in secondary prevention settings, in which
medication use is more prevalent. In addition, participants in secondary prevention trials
generally have more advanced atherosclerosis, which may require more powerful
interventions than n-3 fatty acids or higher n-3 doses to avert clinical events. Indeed, a
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greater n-3 benefit on major vascular events was observed among participants without prior
stroke in a recent meta-analysis14 and in those without prior vascular disease in a trial
among patients with macular degeneration.24 Differences in fish consumption across study
populations may have also influenced findings. Finally, there were few black participants in
the secondary prevention trials, and our trial suggest a greater coronary benefit of
supplemental n-3 fatty acids in this racial group.

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The finding in a subgroup analyses of the secondary outcome of MI suggesting possible

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greater cardiovascular benefits of n-3 supplementation for African Americans than for non-
Hispanic whites, was unexpected, especially given that both racial/ethnic groups had similar
baseline EPA+DHA blood levels and fish intake, and may be a chance finding that requires
corroboration in future trials. Recent observational studies find racial variation in
associations of both marine- and plant-derived n-3 biomarkers with incident coronary
disease.25 Gene variants influence metabolism and bioavailability of n-3 fatty acids, as
demonstrated in Greenland Inuits,26 and may influence coronary risk.27 Other racial/ethnic
differences in clinical, dietary, or environmental factors may also account for this finding.28
Finally, African Americans have a higher prevalence of comorbidities such as diabetes and
hypertension. However, treatment-associated HRs for myocardial infarction remained
reduced across cardiovascular-risk strata in African Americans, with greater risk reductions
than in non-Hispanic whites (Table S3).
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That supplemental n-3 fatty acids confer coronary protection is biologically plausible. Data
from laboratory and animal studies, as well as small trials of intermediate cardiovascular
endpoints in humans, support mechanisms including anti-thrombotic, hypotriglyceridemic,
blood pressure lowering, and anti-inflammatory effects; impeded growth of atherosclerotic
plaques; slowing of heart rate; reduced susceptibility to cardiac arrhythmias; and promotion
of nitric-oxide induced endothelial relaxation whereby n-3 fatty acids may reduce risk.1, 5
Experimental studies support relevant molecular and gene-regulatory effects.1 The dose-
response curve for most effects plateaus at 1 g/day or lower.29 Observational studies suggest
significant inverse associations between fish intake or n-3 fatty acid biomarkers and
coronary outcomes—findings compatible with these mechanisms.25, 30–32

With regard to cancer, our findings are consistent with results of secondary cardiovascular
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disease prevention trials, which have mostly reported neutral effects or slight (but
nonsignificant) elevations in cancer incidence with n-3 fatty acids.8, 16, 17, 22, 23, 33, 34 A
2014 meta-analysis of 10 n-3 fatty acid trials found a nonsignificant 10% increase in cancer
risk (p=0.12).35 A 2018 meta-analysis of n-3 trials of cardiovascular disease14 also reported
no significant association between supplementation and incident cancer but did not provide
an effect estimate. Our finding of a more favorable effect on incident cancer in women
contrasts with results of the SU.FOL.OM3 n-3 trial33 of an increased cancer risk in women
but not men. Three trials have reported on cancer mortality, with two finding a neutral
effect16, 18 and one a borderline significant reduction.34 The absence of a significant n-3
treatment effect on all-cause mortality in the present trial is consistent with results of meta-
analyses of earlier trials13, 15 and ASCEND.16
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The strengths of our trial include a large general population sample with racial/ethnic and
geographic diversity; high follow-up rates and pill-taking adherence; high blood-collection
rates; validated biomarkers of adherence; dietary assessments; and rigorously adjudicated
endpoints. Ancillary studies examining diabetes, atrial fibrillation, cognition, autoimmune
disorders, and other outcomes will inform the overall benefit-risk balance of n-3
supplementation.

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Our study also has certain limitations. Median treatment duration was 5.3 years. The trial’s
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single n-3 dose did not permit exploration of dose-response relationships. However, the dose
used has been recommended by the American Heart Association for cardioprotection in
persons with prior coronary disease7, 36 and is at least twice the dose recommended for
cardiovascular protection in healthy populations (1-2 servings of fish per week).30, 36
Results of ongoing trials37, 38 testing higher doses in high-risk populations will be
informative but may not apply to primary prevention. Some of our subgroup analyses are
based on small numbers of events.

In summary, omega-3 fatty acid supplementation did not reduce the primary outcomes of
major cardiovascular events (myocardial infarction, stroke, and cardiovascular mortality)
and total invasive cancer.

Supplementary Material
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Refer to Web version on PubMed Central for supplementary material.

ACKNOWLEDGMENTS
VITAL Investigators, Staff, and Study Participants

The authors thank the VITAL investigators, staff, and the trial participants for their outstanding dedication and
commitment.

Drs. Manson, Cook, Lee, and Buring had full access to all of the data in the study and take responsibility for the
integrity of the data and the accuracy of the data analyses.

Funding/Support: VITAL was supported by grants U01 CA138962 and R01 CA138962, which included support
from the National Cancer Institute, National Heart, Lung and Blood Institute, Office of Dietary Supplements,
National Institute of Neurological Disorders and Stroke, and the National Center for Complementary and
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Integrative Health. The ancillary studies are supported by grants from multiple Institutes, including the National
Heart, Lung and Blood Institute; the National Institute of Diabetes and Digestive and Kidney Diseases; the National
Institute on Aging; the National Institute of Arthritis and Musculoskeletal and Skin Diseases; the National Institute
of Mental Health; and others.

Pharmavite LLC of Northridge, California (vitamin D) and Pronova BioPharma of Norway and BASF (Omacor fish
oil) donated the study agents, matching placebos, and packaging in the form of calendar packs. Quest Diagnostics
(San Juan Capistrano, CA) measured the plasma omega-3 index at no cost to the study.

VITAL was approved by the Institutional Review Board of Partners Healthcare/Brigham and Women’s Hospital,
and the study agents have received Investigational New Drug Approval from the U.S. Food and Drug
Administration.

VITAL is registered at clinicaltrials.gov (NCT01169259). The VITAL website is www.vitalstudy.org.

(Funded by the National Institutes of Health; VITAL ClinicalTrials.gov number: NCT01169259.)

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Role of the Sponsor: The National Institutes of Health sponsors of the trial had a collaborative role in the design
and conduct of the study and interpretation of the data. Final decisions concerning the above, however, as well as
data collection, management, analysis, review or approval of the manuscript, and decision to submit the manuscript
for publication resided with study investigators and the study research group. The opinions expressed in the
manuscript are those of the study authors and do not necessarily represent the views of the Department of Health
and Human Services/National Institutes of Health.

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JAMA Internal Medicine. 2014;174:763–71. [PubMed: 24638908]
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25. Del Gobbo LC, Imamura F, Aslibekyan S, et al.; Cohorts for Heart and Aging Research in
Genomic Epidemiology (CHARGE) Fatty Acids and Outcomes Research Consortium (FORCe).
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cohort studies. JAMA Internal Medicine. 2016;176:1155–66. [PubMed: 27357102]
26. Fumagalli M, Moltke I, Grarup N, et al. Greenlandic Inuit show genetic signatures of diet and
climate adaptation. Science. 2015;349:1343–7. [PubMed: 26383953]
27. Martinelli N, Girelli D, Malerba G, et al. FADS genotypes and desaturase activity estimated by the
ratio of arachidonic acid to linoleic acid are associated with inflammation and coronary artery
disease. Am J Clin Nutr. 2008;88:941–9. [PubMed: 18842780]
28. Dietary Tong H. and pharmacological intervention to mitigate the cardiopulmonary effects of air
pollution toxicity. Biochim Biophys Acta. 2016;1860:2891–8. [PubMed: 27189803]
29. Mozaffarian D, Rimm EB. Fish intake, contaminants, and human health: evaluating the risks and
the benefits. JAMA. 2006;296:1885–99. [PubMed: 17047219]
30. Rimm EB, Appel LJ, Chiuve SE, et al. American Heart Association Nutrition Committee of the
Council on Lifestyle and Cardiometabolic Health, Council on Epidemiology and Prevention,
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Council on Cardiovascular Disease in the Young, Council on Cardiovascular and Stroke Nursing
and Council on Clinical Cardiology. Seafood Long-Chain n-3 Polyunsaturated Fatty Acids and
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31. Chowdhury R, Warnakula S, Kunutsor S, et al. Association of dietary, circulating, and supplement
fatty acids with coronary risk: a systematic review and meta-analysis. Ann Intern Med.
2014;160:398–406. [PubMed: 24723079]
32. Alexander DD, Miller PE, Van Elswyk ME, Kuratko CN, Bylsma LC. A meta-analysis of
randomized controlled trials and prospective cohort studies of eicosapentaenoic and
docosahexaenoic long-chain omega-3 fatty acids and coronary heart disease risk. Mayo Clin Proc.
2017;92:15–29. [PubMed: 28062061]
33. Andreeva VA, Touvier M, Kesse-Guyot E, Julia C, Galan P, Hercberg S. B vitamin and/or omega-3
fatty acid supplementation and cancer: ancillary findings from the supplementation with folate,
vitamins B6 and B12, and/or omega-3 fatty acids (SU.FOL.OM3) randomized trial. Arch Intern
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Med. 2012;172:540–7. [PubMed: 22331983]

34. Bordeleau L, Yakubovich N, Dagenais GR, et al.; ORIGIN Trial Investigators. The association of
basal insulin glargine and/or n-3 fatty acids with incident cancers in patients with dysglycemia.
Diabetes Care. 2014;37:1360–6. [PubMed: 24574355]
35. Zhang YF, Gao HF, Hou AJ, Zhou YH. Effect of omega-3 fatty acid supplementation on cancer
incidence, non-vascular death, and total mortality: a meta-analysis of randomized controlled trials.
BMC Public Health. 2014;14:204. [PubMed: 24568238]
36. Siscovick DS, Barringer TA, Fretts AM, et al.; American Heart Association Nutrition Committee
of the Council on Lifestyle and Cardiometabolic Health, Council on Epidemiology and Prevention,

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Council on Cardiovascular Disease in the Young, Council on Cardiovascular and Stroke Nursing
and Council on Clinical Cardiology. Omega-3 Polyunsaturated Fatty Acid (Fish Oil)
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the American Heart Association. Circulation. 2017;135:e867–e884. [PubMed: 28289069]
37. Bhatt DL, Steg PG, Brinton EA, et al.; REDUCE-IT Investigators. Rationale and design of
REDUCE-IT: Reduction of Cardiovascular Events with Icosapent Ethyl-Intervention Trial. Clin
Cardiol. 2017;40:138–148. [PubMed: 28294373]
38. Nicholls SJ, Lincoff AM, Bash D, et al. Assessment of omega-3 carboxylic acids in Statin Treated
Patients with High Levels of Triglycerides and Low Levels of High Density Lipoprotein
Cholesterol: rationale and design of the STRENGTH Trial. Clin Cardiol. 2018 [e-pub 2018 Aug
19].
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Figure 1.
Cumulative Incidence Rates of A) Major Cardiovascular Events, B) Total Invasive Cancer,
by Year of Follow-up. From Cox regression models controlling for age, sex, and vitamin D
randomization group (intention-to-treat analyses).
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 Manson et al. Page 14
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Figure 2:
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Hazard Ratios of Major Cardiovascular Events by Subgroups, comparing Omega-3 Fatty

Acids (n-3) and Placebo Groups. From Cox regressions models controlling for age, sex, and
vitamin D randomization group (intention-to-treat analyses). Analyses have not been
adjusted for multiple comparisons.

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 Manson et al. Page 15

Table 1.

Baseline Characteristics of the 25,871 VITAL Participants, According to Randomized Assignment to Marine
Author Manuscript

a
Omega-3 Fatty Acids.

Baseline Treatment Group

Characteristic
All Participants Omega-3s Placebo
N 25,871 12,933 12,938

Sex, % female 13085 (50.6) 6547 (50.6) 6538 (50.5)

Mean age ± SD, years 67.1 ± 7.1 67.2 ± 7.1 67.1 ± 7.1

Race/ethnicity, %

Non-Hispanic White 18046 (71.3) 9044 (71.5) 9002 (71.2)

African American 5106 (20.2) 2549 (20.1) 2557 (20.2)

Hispanic (not African American) 1013 (4.0) 491 (3.9) 522 (4.1)
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Asian/Pacific Islander 388 (1.5) 200 (1.6) 188 (1.5)

American Indian/Alaskan Native 228 (0.9) 120 (0.9) 108 (0.9)

Other/unknown 523 (2.1) 249 (2.0) 274 (2.2)

Mean body mass index ± SD, kg/m2 28.1 (5.7) 28.1 ± 5.7 28.1 ± 5.8

Current smoking, % 1836 ( 7.2) 920 ( 7.2) 916 ( 7.2)

Hypertension, treated with medication, %, 12791 (49.8) 6338 (49.3) 6453 (50.2)

Cholesterol-lowering medication (current use), % 9524 (37.5) 4788 (37.7) 4736 (37.2)

Diabetes, % 3549 (13.7) 1799 (13.9) 1750 (13.5)

a
Abbreviations: SD = standard deviation. There were no significant differences in the baseline characteristics between the groups.
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 Manson et al. Page 16

Table 2.

Hazard Ratios (HR) and 95% Confidence Intervals (CI) of the Primary, Secondary, and Other Outcomes by
Author Manuscript

Randomized Assignment to Omega-3 (n-3) Fatty Acids in Intention-to-Treat Analyses

No. of Events

Outcome n-3 Placebo HR 95% CI

(N = 12,933) (N = 12,938)

Cardiovascular disease, 1° and 2° Outcomes

a,b 386 419 0.92 0.80-1.06

Major cardiovascular events

c 527 567 0.93 0.82-1.04

Expanded cardiovascular events
Total myocardial infarction 145 200 0.72 0.59-0.90

Total stroke 148 142 1.04 0.83-1.31

Cardiovascular mortality 142 148 0.96 0.76-1.21

d
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Other vascular outcomes

Coronary artery bypass graft (CABG) 85 86 0.99 0.73-1.33

Percutaneous coronary intervention (PCI) 162 208 0.78 0.63-0.95

e 308 370 0.83 0.71-0.97

Total coronary heart disease

Ischemic stroke 111 116 0.96 0.74-1.24

Hemorrhagic stroke 25 19 1.32 0.72-2.39

Coronary heart disease death 37 49 0.76 0.49-1.16

Fatal myocardial infarction 13 26 0.50 0.26-0.97

Stroke death 22 20 1.10 0.60-2.01

a 820 797 1.03 0.93-1.13

Total invasive cancer

Breast 117 129 0.90 0.70-1.16

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Prostate 219 192 1.15 0.94-1.39

Colorectal 54 44 1.23 0.83-1.83

Cancer death 168 173 0.97 0.79-1.20

All-cause mortality 493 485 1.02 0.90-1.15

Excluding the first two years of follow-up:

Major cardiovascular events 269 301 0.89 0.76-1.05

Total myocardial infarction 94 131 0.72 0.55-0.93

Total invasive cancer 536 476 1.13 1.00-1.28

Cancer death 126 135 0.93 0.73-1.19

All-cause mortality 371 381 0.97 0.84-1.12

a
Primary outcomes
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b
A composite of myocardial infarction, stroke, and cardiovascular mortality
c
A composite of myocardial infarction, stroke, cardiovascular mortality, and coronary revascularization (CABG, PCI)
d
Not prespecified as 1° or 2° outcomes.
e
A composite of myocardial infarction, coronary revascularization (CABG, PCI), and coronary heart disease death.

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 Manson et al. Page 17

From Cox regression models controlling for age, sex, and vitamin D randomization group. The 95% CIs have not been adjusted for multiple
comparisons.
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N Engl J Med. Author manuscript; available in PMC 2020 January 03.

 Supplementary Appendix

This appendix has been provided by the authors to give readers additional information about their work.

Supplement to: Manson JE, Cook NR, Lee I-M, et al. Marine n−3 fatty acids and prevention of cardiovascular
disease and cancer. N Engl J Med 2019;380:23-32. DOI: 10.1056/NEJMoa1811403
 Marine n-3 Fatty Acids and Prevention of Cardiovascular Disease and Cancer

SUPPLEMENTARY APPENDIX

Contents

Title Page and Table of Contents…………………………………………………………………………….. 1-2

Members of the VITAL Research Group
Steering Committee………………………………………………………………………………….… 3
Scientific Consultants.……………………………………………………………………………….… 3
Other Members of the VITAL Research Group……………………………………………………... 3
Data and Safety Monitoring Board……………………………………………………………..…..… 4
Detailed Methods ……………………………………………………………………………………… 5

Supplementary Tables
Table S1. Baseline Characteristics of the 25,871 Participants, According to Randomized Assignment to
Marine Omega-3 Fatty Acids……………………………………………………………………………………... 10

Table S2. Participant-Reported Adherence with the Omega-3 and Placebo Study Capsules at Time Points
over 5 Years…………………………………………………………………………………………………………… 12

Additional Details on Subgroup Analyses……………………………………………………………………….. 13

Table S3. Hazard Ratios (HR) and 95% Confidence Intervals (CI) for Major Cardiovascular Events and Total
Myocardial Infarction According to Diabetes Status and Number of Cardiovascular Risk Factors at Baseline,
Comparing Omega-3 Fatty Acids (n-3) and Placebo in Racial/Ethnic groups ……………………................ 14

Table S4. Hazard Ratios (HR) and 95% Confidence Intervals (CI) for Major Outcomes According to Baseline
Fish Intake and Race/Ethnicity, Comparing Omega-3 Fatty Acids (n-3) and Placebo Groups……….…….....15

Table S5. Hazard Ratios (HR) and 95% Confidence Intervals (CI) of Total Invasive Cancer Comparing Active
Omega-3 Fatty Acids (n-3) and Placebo Groups, According to Baseline Characteristics…………….….. 16

Table S6. Hazard Ratios (HR) and 95% Confidence Intervals (CI) of Total Mortality Comparing Active Omega-
3 Fatty Acids (n-3) and Placebo Groups, According to Baseline Characteristics…………….………….... 17

Table S7. Hazard Ratios (HR) and 95% Confidence Intervals (CI) for Safety and Adverse Events by
Randomized Assignment to Omega-3 Fatty Acids (n-3) …………………………………………………… 19

Supplementary Figures
Figure S1. Flow Diagram of Enrollment in the Omega-3 Fatty Acid Component of the Trial…………… 20

Figure S2. Cumulative Incidence Rates of A) Expanded Cardiovascular Events, B) Total Myocardial
Infarction, C) Total Stroke, and D) Cardiovascular Mortality, By Year of Follow-up in the Omega-3 Fatty Acids
and Placebo Groups……………………………………………………………………………………………………... 21

Figure S3. Hazard Ratios of Total Myocardial Infarction by Subgroups, Comparing Omega-3 Fatty Acids
(n-3) and Placebo groups………………………………………………………………………………………. 22

2
 Members of the VITAL Research Group

VITAL Steering Committee:

JoAnn E. Manson (Chair), Julie E. Buring (Chair), Nancy R. Cook, I-Min Lee, William Christen, Shari S.

Bassuk, Samia Mora, Heike Gibson, David Gordon, Trisha Copeland, Denise D’Agostino, Georgina

Friedenberg, Claire Ridge, Vadim Bubes, Edward L. Giovannucci, Walter C. Willett (all at Brigham and

Women’s Hospital, Harvard Medical School, Boston; Drs. Manson, Buring, Cook, Lee, Giovannucci and Willett

are also at the Harvard T.H. Chan School of Public Health).

Scientific consultants:

John Baron (University of North Carolina, Chapel Hill),

Michael Holick (Boston Medical Center), Bruce Hollis (University of South Carolina).

Other Members of the VITAL Research Group:

(Brigham and Women’s Hospital): Christine M. Albert, Diane Gold, Meryl LeBoff, Olivia Okereke, Aruna

Pradhan, Howard Sesso, Wendy Chen, Paulette Chandler, J. Michael Gaziano, Olga Demler, Kathryn

Rexrode, Karen Costenbader, John Forman, Erik Alexander, Sonia Friedman, Jeffrey Katz, Shumin Zhang,

Jennifer Lin, Joseph Walter, Julie Duszlak, Kate Kalan, Jean MacFadyen, Natalya Gomelskaya, David Bates,

Ara Sarkissian, Mary Breen, Yeulolani Andrade, Manickavasagar Vinayagamoorthy, Chunying Li, Eunjung

Kim, Franco Giulianini, Gregory Kotler, Marty Van Denburgh, Rimma Dushkes, Yanyan Liu, Eduardo Pereira,

Lisa Fields-Johnson, George Menjin, Lucy Liu, Lauren Girard, Scott Zeller, Naomi Riches, Katelyn Hasson,

Ellen Bhang, Maria Revilla, Elena McCarthy, Alex Moran, Kristen Haise, Leah Arsenault, Philomena Quinn,

3
Sancia Grimes, Ivan Fitchorov, Kurt Schwerin, Shamikhah Curry, Annie Murray, Angela Zhang, Diana

Walrond-Williams, Alison Weinberg, Chris Pfeffer, Margarette Haubourg, Viviane Nguyen, Henry Ouellette,

Rolando Rodriguez, Tony Montgomery, Keith Morse, Vincent Guzman, Megan Perry, Sandra Weekes, Doug

Smith, Allison Clar, Sara Curran, Yaneve Fonge, David Hibbert, Louisa Paine, Kelly Royce, Courtney Splaine,

Jennifer McMahon, David Eldridge, Laura Hand, Kay Inandan, Meghan Rieu Werden, Harriet Samuelson,

Andrea Hrbek, Megan Mele, Eileen Bowes, Mary Anne Ryan

(Massachusetts General Hospital, Boston): Carlos Camargo, Jacqueline Danik, Ravi Thadhani

(Vanderbilt University, Nashville): Thomas Wang

(Rush University Medical Center, Chicago): Raj C. Shah

(University of California, San Francisco): Michelle A. Albert

(Emory University): Carlos Kase

(Centers for Disease Control and Prevention, Vitamin D Standardization Program): Hubert Vesper and

Julianne Botelho.

Data and Safety Monitoring Board (Voting Members): Nanette Wenger, MD (Chair); Lawrence S. Cohen,

MD; Theodore Colton, ScD; Mark A. Espeland, PhD; Craig Henderson, MD; Alice H. Lichtenstein, ScD; and

Rebecca A. Silliman, MD, PhD. Ex-officio members include Josephine Boyington, PhD, MPH; Rebecca

Costello, PhD; Cindy Davis, PhD; Peter Greenwald, MD; Gabriela Riscuta, MD; and Harold Seifried, PhD.

4
 SUPPLEMENTARY APPENDIX: DETAILED METHODS, TABLES, AND FIGURES

METHODS (details of study methods, omega-3 component)

Study Design

VITAL was a randomized, double-blind, placebo-controlled, 2x2 factorial trial of the benefits and risks of

vitamin D3 (cholecalciferol, 2000 IU/d) and marine omega-3 fatty acids (1 g/d fish oil capsule [Omacor],

containing 840 mg of n-3 FAs including eicosapentaenoic acid [EPA, 460 mg] + docosahexaenoic acid [DHA,

380 mg]) in the primary prevention of cardiovascular disease and cancer among 25,871 men aged ≥50 and

women aged ≥55. Study details are described elsewhere.1,2 The omega-3 fatty acid dose was chosen because

it was recommended by the American Heart Association for cardioprotection3 and demonstrated to be

beneficial and to have minimal side effects in a secondary prevention population.4 The placebo capsule

contained olive oil. Participants were recruited throughout the U.S., balanced by sex, and with a goal to enroll

at least 5,000 African Americans. Eligible participants had no history of cancer (except non-melanoma skin

cancer), myocardial infarction, stroke, transient ischemic attack, or coronary revascularization at enrollment

and were required to forgo use of fish-oil supplements and to complete a 3-month placebo run-in phase. Safety

exclusions included renal failure or dialysis, cirrhosis, fish allergy, anticoagulant use, or other serious

conditions that would preclude participation. The recruitment flow diagram is presented in Figure S1 in the

Supplementary Appendix. Randomization to n-3 fatty acids, vitamin D, both active agents, or both placebos

took place from November 2011 to March 2014 and was computer generated within sex, race (African

American vs. not) and 5-year age groups in blocks of eight. All participants provided written informed consent.

Study pill-taking ended as planned on December 31, 2017, yielding a median treatment period of 5.3 years

(range 3.8-6.1 years). The trial was approved by the institutional review board of Partners Health Care-

Brigham and Women’s Hospital, Boston, and was monitored by an external Data and Safety Monitoring Board.

Baseline questionnaires collected data on risk factors for cardiovascular disease, cancer, and other

conditions, and included a food frequency questionnaire that ascertained intake of fish and other foods.

5
Participants received follow-up questionnaires at 6 months and 1 year after randomization and annually

thereafter to assess compliance with randomized treatments, use of nonstudy fish-oil supplements,

development of major illnesses, updates on risk factors, and potential side effects of the interventions. Study

capsules were mailed to participants with the questionnaires.

Baseline blood samples were collected during the run-in period from all willing individuals, including

16,956 randomized participants. Assays for plasma omega-3 index (EPA+DHA as a percent of total fatty

acids5) were performed on samples by Quest Diagnostics using liquid chromatography-tandem mass

spectrometry.

Study Endpoints

Primary endpoints were major cardiovascular events (composite of myocardial infarction, stroke, and

cardiovascular mortality) and total invasive cancer. Secondary cardiovascular endpoints were expanded

cardiovascular events (major cardiovascular events plus coronary revascularization [coronary artery bypass

grafting (CABG) or percutaneous coronary intervention (PCI)]) and individual components of major

cardiovascular events. Additional cardiovascular events were total coronary heart disease (a composite of

myocardial infarction, coronary revascularization, and coronary heart disease death), fatal myocardial

infarction, fatal stroke, and stroke subtypes. Secondary cancer endpoints were incident colorectal, breast, and

prostate cancers, and total cancer mortality. Participants reporting a relevant endpoint were asked to sign a

release for medical records, which were then reviewed by an Endpoints Committee of physicians blinded to

treatment assignment to confirm or disconfirm reports. Myocardial infarction and stroke were confirmed using

established criteria.6,7 Coronary revascularization was confirmed by medical record review. Cardiovascular

deaths were confirmed by convincing evidence of a cardiovascular event from available sources, including

death certificates, hospital records, autopsy reports, and, for non-hospital deaths, observer accounts. Cancer

was confirmed with histologic or cytologic data.8 Analyses include confirmed endpoints only.

6
 For deaths reported by family members, the next-of-kin was asked for permission to obtain medical

records and a copy of the death certificate. Alternatively, the latter was obtained from the state vital records

bureau. The Endpoints Committee reviewed records to assign cause of death. If records were unavailable (or

participants lost to follow-up), the National Death Index (NDI) Plus was searched to obtain an International

Classification of Disease-coded cause of death based on death-certificate information. Deaths were defined

using all sources.

Statistical Analysis

Treatment-effect analyses compared randomized interventions and were based on the intention-to-treat

principle (all randomized participants were analyzed). The trial was designed to have >85% power to detect

observed hazard ratios (HR) of 0.80 and 0.85 for the primary cardiovascular and cancer endpoints,

respectively.1 Initial analyses compared baseline characteristics of the study population by randomized

treatment assignment using t-tests or chi-square tests as appropriate. Primary analyses compared the main

effects of n-3 fatty acids on cardiovascular disease and cancer using Cox proportional hazards models

controlling for age, sex, and randomization to vitamin D. Person-time was counted from randomization to the

endpoint, to death, or to end of the trial on December 31, 2017. Cumulative incidence plots and interactions

with time examined whether treatment effects varied over time. Prespecified analyses excluding events

occurring during the first year and first two years of follow-up explored latent treatment effects. Compliance

effects were estimated by censoring follow-up when participants stopped taking study capsules and/or began

taking outside fish-oil supplements.

Possible variations in treatment effect by age, sex, concurrent randomization to vitamin D, baseline

cardiovascular risk factors, and baseline fish intake and plasma omega-3 (EPA+DHA) index were specified a

priori. Because vitamin D was also studied, treatment effects in racial/ethnic groups were of interest. We

additionally collected information on aspirin and statin use as stratification variables. There was no control for

multiple hypothesis testing, with no formal adjustment to the P values or confidence intervals. Thus, results for

7
secondary and exploratory outcomes, and for subgroups, should be interpreted with caution. Finally, potential

side effects, including gastrointestinal symptoms and bleeding risks, in the randomized groups were compared.

8
References:

1. Manson JE, Bassuk SS, Lee IM, Cook NR, Albert MA, Gordon D, Zaharris E, Macfadyen JG, Danielson E, Lin J,
Zhang SM, Buring JE. The VITamin D and OmegA-3 TriaL (VITAL): Rationale and design of a large randomized
controlled trial of vitamin D and marine omega-3 fatty acid supplements for the primary prevention of cancer
and cardiovascular disease. Contemp Clin Trials. 2012;33:159-171
2. Bassuk SS, Manson JE, Lee IM, Cook NR, Christen WG, Bubes VY, Gordon DS, Copeland T, Friedenberg G,
D'Agostino DM, Ridge CY, MacFadyen JG, Kalan K, Buring JE. Baseline characteristics of participants in the
VITamin D and OmegA-3 TriaL (VITAL). Contemp Clin Trials. 2016;47:235-243
3. Kris-Etherton PM, Harris WS, Appel LJ. Fish consumption, fish oil, omega-3 fatty acids, and cardiovascular
disease. Circulation. 2002;106:2747-2757
4. GISSI-Prevenzione Investigators (Gruppo Italiano per lo Studio della Sopravvivenza nell'Infarto miocardico).
Dietary supplementation with n-3 polyunsaturated fatty acids and vitamin E after myocardial infarction: Results
of the GISSI-Prevenzione trial. Lancet. 1999;354:447-455
5. Harris WS, Von Schacky C. The omega-3 index: A new risk factor for death from coronary heart disease?
Preventive Medicine. 2004;39:212-220
6. Thygesen K, Alpert JS, Jaffe AS, Simoons ML, Chaitman BR, White HD, Writing Group on behalf of the Joint
ESC/ACCF/AHA/WHF Task Force for the Universal Definition of Myocardial Infarction. Third universal definition
of myocardial infarction. Circulation. 2012;126:2020-2035
7. Adams HP, Jr., Bendixen BH, Kappelle LJ, Biller J, Love BB, Gordon DL, Marsh EE, 3rd. Classification of subtype of
acute ischemic stroke. Definitions for use in a multicenter clinical trial. TOAST. Trial of Org 10172 in Acute Stroke
Treatment. Stroke. 1993;24:35-41
8. Fritz AG, Percy C, Jack A, Shanmugaratham K, Sobin LH, Parkin MD, Whelan S. International Classification of
Diseases for Oncology (ICD-O), Third Edition. Geneva: World Health Organization; 2000.

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Table S1. Baseline Characteristics of the 25,871 Participants, According to Randomized
Assignment to Marine Omega-3 Fatty Acidsa

Baseline Omega-3 Fatty Acids

Characteristic All Participants Active Placebo

N 25,871 12,933 12,938

Sex, % female 13,085 (50.6) 6,547 (50.6) 6,538 (50.5)

Mean age ± SD, years 67.1 ± 7.1 67.2 ± 7.1 67.1 ± 7.1

Age group, years, %

50-54 977 ( 3.8) 488 ( 3.8) 489 ( 3.8)

55-64 8,871 (34.3) 4,431 (34.3) 4,440 (34.3)

65-74 12,705 (49.1) 6,352 (49.1) 6,353 (49.1)

≥75 3,318 (12.8) 1,662 (12.9) 1,656 (12.8)

Race/ethnicity, %

Non-Hispanic White 18,046/25,304 (71.3) 9,044/12,653 (71.5) 9,002/12,651 (71.2)

African American 5,106/25,304 (20.2) 2,549/12,653 (20.1) 2,557/12,651 (20.2)

Hispanic (not African American) 1,013/25,304 (4.0) 491/12,653 (3.9) 522/12,651 (4.1)

Asian/Pacific Islander 388/25,304 (1.5) 200/12,653 (1.6) 188/12,651 (1.5)

Native American or Alaskan Native 228/25,304 (0.9) 120/12,653 (0.9) 108/12,651 (0.9)

Other 523/25,304 (2.1) 249/12,653 (2.0) 274/12,651 (2.2)

Mean body mass index ± SD, kg/m2 28.1 ± 5.7 28.1 ± 5.7 28.1 ± 5.8

Hypertension, treated with medication, %, 12,791/25,698 (49.8) 6,338/12,853 (49.3) 6,453/12,845 (50.2)

Cholesterol-lowering medication (current 9,524/25,428 (37.5) 4,788/12,707 (37.7) 4,736/12,721 (37.2)

use), %

Diabetes, % 3,549/25,828 (13.7) 1,799/12,912 (13.9) 1,750/12,916 (13.5)

Current smoking, % 1,836/2,5485 ( 7.2) 920/12,739 ( 7.2) 916/12,746 ( 7.2)

Any alcohol use, %b 17,443/25,437 (68.6) 8,759/12,728 (68.8) 8,684/12,709 (68.3)

Current regular aspirin use, %b 11,570/25,497 (45.4) 5,771/12,745 (45.3) 5,799/12,752 (45.5)

Current postmenopausal 1,483/12,811 (11.6) 735/6,400 (11.5) 748/6,411 (11.7)

10
 hormone use, % (women only)

Current use of multivitamins, % 11,406/25,439 (44.8) 5,661/12,726 (44.5) 5,745/12,713 (45.2)

Intake of foods related to omega-3 fatty

acids, mean ± SD

Dark-meat fish, servings/week 1.05 ± 1.84 1.05 ± 1.93 1.04 ± 1.76

Other fish and seafood, servings/week 1.11 ± 2.32 1.11 ± 2.36 1.11 ± 2.29

Plasma EPA, %, median [IQR]c 0.50 [0.40-0.70] 0.50 [0.40-0.70] 0.50 [0.40-0.70]

Plasma DHA, %, median [IQR]c 1.90 [1.50-2.40] 1.90 [1.50-2.40] 1.90 [1.50-2.40]

aAbbreviations: SD = standard deviation; BMI = body mass index; EPA = eicosapentaenoic acid;
DHA = docosahexaenoic acid; IQR = interquartile range.
bAt least monthly.
cN = 15,527 with measured values for EPA and 15,534 for DHA.

For body mass index, data were missing for 2.4% of the participants. For dietary factors, data were missing for 1.7% to
2.7% of participants.
There were no significant differences in the baseline characteristics between the groups.

11
Table S2. Participant-Reported Adherence with the Omega-3 and Placebo Study Capsules (percent of
capsules taken) at Time Points over 5 Years.

A. Among those answering the compliance question by questionnaire:

Omega-3 Placebo
Time N Mean (SD) Mean (SD)
6 Months 24108 92.4 (16.8) 92.5 (16.4)
1 Year 24211 89.7 (21.0) 90.2 (20.1)
2 Years 23432 87.0 (25.5) 87.6 (24.7)
3 Years 22630 86.3 (26.9) 86.4 (26.4)
4 Years 21217 86.4 (26.7) 86.5 (26.4)
5 Years 13962 85.7 (27.9) 86.2 (27.0)

B. Including nonrespondents to questionnaires and assuming noncompliance among

all nonrespondents:

Omega-3 Placebo
Time N Mean (SD) Mean (SD)
6 Months 25830 87.8 (25.9) 87.8 (25.8)
1 Year 25751 86.3 (26.9) 86.6 (26.4)
2 Years 25576 82.7 (31.2) 82.9 (31.1)
3 Years 25373 81.2 (33.0) 81.2 (32.8)
4 Years 24616 79.0 (35.2) 79.4 (34.7)
5 Years 16920 75.8 (37.9) 75.7 (37.9)

12
Additional Details on Subgroup Analyses

In subgroup analyses, baseline dietary fish intake modified the n-3 intervention effect on major cardiovascular

events (Fig. 2) and total myocardial infarction (Supplementary Appendix Fig. S3), with nominally significant

reductions of 19% and 40%, respectively, in participants with below-median intake (<1.5 servings per week)

but no reductions in those with higher intake (nominal p-interaction<0.05 for both). The HR for myocardial

infarction among those who ate fish less than once per month was 0.43 (0.21-0.91). For major cardiovascular

events, no other significant interactions were observed. For myocardial infarction, however, race/ethnicity also

modified the treatment effect, with a 77% reduction among African Americans (HR=0.23 [0.11-0.47]) and

smaller reductions in other racial/ethnic groups (nominal p-interaction by race=0.001) (Fig. S3). African

Americans also had reductions in coronary revascularization (HR=0.51 [0.28-0.92]) and total coronary heart

disease (HR=0.47 [0.29-0.75]) with n-3 fatty acids. For myocardial infarction, treatment-associated reductions

were also observed in participants with comorbidities and a larger number of traditional risk factors (nominal p-

interaction=0.047) (Fig. S3). Treatment-associated benefits were more apparent in African Americans than in

non-Hispanic whites across all cardiovascular risk factor strata, including among those with diabetes (HR=0.06

[0.01-0.49] in African Americans vs. HR=0.88 [0.43-1.80] in non-Hispanic whites, nominal p-interaction=0.018)

(Supplementary Appendix, Table S3). For major cardiovascular events, treatment-associated reductions were

greater in those with low baseline fish intake in both racial/ethnic groups (Table S4). For myocardial infarction,

treatment-associated reductions among African Americans were observed irrespective of baseline fish intake,

but non-Hispanic whites benefited from supplementation only if fish consumption was low (Table S4). The

plasma omega-3 index did not significantly modify the intervention’s effects. Analyses that censored for

nonadherence did not materially change results (HR=0.91 [0.78-1.07] for major cardiovascular events and

HR=0.74 [0.58-0.95] for myocardial infarction in the overall cohort).

13
Table S3. Hazard Ratios (HR) and 95% Confidence Intervals (CI) for Major Cardiovascular Events and Total
Myocardial Infarction According to Diabetes Status and Number of Cardiovascular Risk Factors at Baseline,
Comparing Omega-3 Fatty Acids (n-3) and Placebo in Racial/Ethnic Groups

Major Cardiovascular Events Total Myocardial Infarction

Interaction Interaction
No. of Events P-value No. of Events P-value
Participants with HR
Diabetesa Total n-3 Placebo HR (95%CI) n-3 Placebo (95%CI)
Race 2387 0.17 0.018
Non-Hispanic Whites 1449 33 32 1.04 14 16 0.88
(0.64-1.69) (0.43-1.80)
African Americans 938 17 27 0.60 1 15 0.06
(0.33-1.11) (0.01-0.49)

Total Number of CVD Risk Factorsb

Non-Hispanic Whites 18046 0.88 0.82
No Risk Factors 5896 70 70 0.97 34 35 0.95
(0.69-1.35) (0.59-1.53)
1 Risk Factor 6177 105 102 1.01 48 50 0.93
(0.77-1.33) (0.63-1.39)
2 or More Risk Factors 5973 117 117 1.01 44 50 0.89
(0.78-1.30) (0.59-1.33)

African Americans 5106 0.94 0.17

No Risk Factors 1054 7 11 0.62 2 3 0.65
(0.24-1.59) (0.11-3.89)
1 Risk Factor 1857 20 25 0.83 3 11 0.28
(0.46-1.50) (0.08-0.99)
2 or More Risk Factors 2195 35 47 0.72 4 25 0.16
(0.47-1.12) (0.05-0.45)
aReported diabetes treated with medication.
bNumber of traditional cardiovascular disease risk factors (smoking, diabetes, hypertension, high cholesterol, parental
history of premature myocardial infarction).
From Cox regression models controlling for age, sex, and vitamin D randomization group (intention-to-treat analyses).
Analyses have not been adjusted for multiple comparisons.

14
Table S4. Hazard Ratios (HR) and 95% Confidence Intervals (CI) for Major Outcomes According to Baseline
Fish Intake and Race/Ethnicity, Comparing Omega-3 Fatty Acids (n-3) and Placebo Groups

Major Cardiovascular Events Total Myocardial Infarction

No. of Events No. of Events
Total Fish Consumption HR Interaction HR Interaction
(servings/week) Total N n-3 Placebo (95%CI) P-value n-3 Placebo (95%CI) P-value
Total Cohort 25435 0.045 0.048

<median 13514 189 232 0.81 74 121 0.60

(1.5 servings/wk) (0.67-0.98) (0.45-0.81)
>median 11921 189 176 1.08 67 72 0.94
(1.5 servings/wk) (0.88-1.32) (0.67-1.31)
Non-Hispanic Whites 17851 0.043 0.016
<median 9618 146 168 0.86 64 88 0.71
(1.5 servings/wk) (0.69-1.07) (0.51-0.98)
>median 8233 141 116 1.21 59 45 1.32
(1.5 servings/wk) (0.95-1.55) (0.90-1.95)
African Americans 4939 0.23 0.92
<median 2461 26 42 0.61 5 21 0.23
(1.5 servings/wk) (0.37-0.99) (0.09-0.62)
>median 2478 33 36 0.92 3 14 0.21
(1.5 servings/wk) (0.57-1.48) (0.06-0.74)

Total Invasive Cancer Total Mortality

No. of Events No. of Events
Total Fish Consumption Interaction HR Interaction
(servings/week) Total N n-3 Placebo HR (95%CI) P-value n-3 Placebo (95%CI) P-value
Total Cohort 25435 0.094 0.017
<median 13514 421 435 0.96 237 271 0.87
(1.5 servings/wk) (0.84-1.09) (0.73-1.04)
>median 11921 386 346 1.13 240 202 1.19
(1.5 servings/wk) (0.98-1.31) (0.99-1.44)
Non-Hispanic Whites 17851 0.095 0.031
<median 9618 331 347 0.94 175 187 0.93
(1.5 servings/wk) (0.80-1.09) (0.76-1.14)
>median 8233 299 266 1.13 167 127 1.30
(1.5 servings/wk) (0.96-1.34) (1.04-1.64)
African Americans 4939 0.55 0.104
<median 2461 55 58 0.94 37 57 0.64
(1.5 servings/wk) (0.65-1.36) (0.42-0.96)
>median 2478 53 48 1.10 53 52 1.02
(1.5 servings/wk) (0.75-1.63) (0.70-1.50)
From Cox regression models controlling for age, sex, and vitamin D randomization group (intention-to-treat
analyses). Analyses have not been adjusted for multiple comparisons.

15
Table S5. Hazard Ratios (HR) and 95% Confidence Intervals (CI) of Total Invasive Cancer Comparing Active
Omega-3 Fatty Acids (n-3) and Placebo Groups, According to Baseline Characteristics

Total Invasive Cancer

No. of Events
Interaction
Subgroup Category Total n-3 Placebo HR (95%CI) P-value

Pre-specified
Age 25871 0.50
<median (66.7 years) 12859 323 301 1.07 (0.92-1.25)
>median (66.7 years) 13012 497 496 1.00 (0.89-1.14)
Sex 25871 0.024
Male 12786 498 442 1.13 (1.00-1.29)
Female 13085 322 355 0.90 (0.78-1.05)
Race 25304 0.86
Non-Hispanic White 18046 637 621 1.02 (0.91-1.13)
African American 5106 113 111 1.02 (0.79-1.33)
Other 2152 54 51 1.13 (0.77-1.66)
Current Smoker 25485 0.79
No 23649 740 705 1.05 (0.95-1.16)
Yes 1836 73 75 1.01 (0.73-1.39)
Total Fish Intake 25435 0.09
<median (1.5 servings/wk) 13514 421 435 0.96 (0.84-1.09)
>median (1.5 servings/wk) 11921 386 346 1.13 (0.98-1.31)
Vitamin D randomization 25871 0.56
Placebo group 12944 412 412 1.00 (0.87-1.15)
Active group 12927 408 385 1.06 (0.92-1.22)

Other Subgroup Analyses

Baseline Aspirin Use 25497 0.50
No 13927 432 424 1.01 (0.88-1.16)
Yes 11570 378 352 1.08 (0.94-1.25)
Baseline Statin Use 25447 0.63
No 16557 494 483 1.03 (0.91-1.17)
Yes 8890 318 293 1.08 (0.92-1.27)

From Cox regression models controlling for age, sex, and vitamin D randomization group (intention-to-treat
analyses). Analyses have not been adjusted for multiple comparisons.

16
Table S6. Hazard Ratios (HR) and 95% Confidence Intervals (CI) of Total Mortality Comparing Active Omega-3
Fatty Acids (n-3) and Placebo Groups, According to Baseline Characteristics

Total Mortality
No. of Events

Interaction
Subgroup Category Total n-3 Placebo HR (95%CI) P-value

Pre-specified
Age 25871 0.93
<median (66.7 years) 12859 163 158 1.03 (0.83-1.28)
>median (66.7 years) 13012 330 327 1.01 (0.87-1.18)
Sex 25871 0.15
Male 12786 283 256 1.11 (0.93-1.31)
Female 13085 210 229 0.92 (0.76-1.11)
Race 25304 0.26
Non-Hispanic White 18046 351 320 1.09 (0.93-1.26)
African American 5106 97 114 0.84 (0.64-1.11)
Other 2152 37 38 1.08 (0.69-1.70)
Current Smoker 25485 0.093
No 23649 400 406 0.98 (0.85-1.13)
Yes 1836 84 69 1.28 (0.93-1.76)
Diabetes (medication-treated) 25860 0.29
No 23132 413 394 1.05 (0.91-1.20)
Yes 2728 80 91 0.87 (0.65-1.18)
Hypertension (medication-treated) 25698 0.55
No 12907 190 175 1.06 (0.87-1.30)
Yes 12791 293 304 0.98 (0.84-1.15)
High Cholesterol (medication-treated) 25428 0.69
No 15904 304 293 1.05 (0.89-1.23)
Yes 9524 173 174 0.99 (0.80-1.22)
Parental History of Myocardial 22915 0.53
Infarctiona
No 19262 337 327 1.03 (0.89-1.20)
Yes 3653 76 80 0.92 (0.67-1.26)
Total Fish Intake 25435 0.017
<median (1.5 servings/wk) 13514 237 271 0.87 (0.73-1.04)
>median (1.5 servings/wk) 11921 240 202 1.19 (0.99-1.44)
Vitamin D randomization 25871 0.65
Placebo group 12944 252 241 1.05 (0.88-1.25)
Active group 12927 241 244 0.99 (0.83-1.18)

Other Subgroup Analyses

# of Cardiovascular Risk Factorsb 25871 0.55
No risk factors 7802 114 92 1.21 (0.92-1.59)
1 risk factor 8948 162 181 0.90 (0.73-1.11)
2 or more risk factors 9121 217 212 1.03 (0.86-1.25)
Baseline Aspirin Use 25497 0.58
No 13927 255 253 1.00 (0.84-1.19)
Yes 11570 228 215 1.07 (0.89-1.29)
Baseline Statin Use 25447 0.66
No 16557 316 304 1.05 (0.89-1.22)
Yes 8890 161 163 0.98 (0.79-1.22)

17
a
Premature myocardial infarction in a parent (before age 60 in father and before age 65 in mother).
b
Number of traditional cardiovascular disease risk factors (smoking, diabetes, hypertension, high
cholesterol, parental history of premature myocardial infarction).
From Cox regression models controlling for age, sex, and vitamin D randomization group (intention-to-treat
analyses). Analyses have not been adjusted for multiple comparisons.

18
Table S7. Hazard Ratios (HR) and 95% Confidence Intervals (CI) for Safety and Adverse Events by
Randomized Assignment to Omega-3 Fatty Acids (n-3) compared to Placebo

No. of Events
n-3 Placebo
Outcome (N = 12,933) (N = 12,938) HR 95% CI P-value

Monitored safety conditions

Gastrointestinal bleeding 370 374 0.99 0.86-1.14 0.89
Blood in urine 919 874 1.06 0.96-1.16 0.25
Easy bruising 3443 3399 1.02 0.97-1.07 0.48
Frequent nosebleeds 465 491 0.95 0.83-1.07 0.40
Kidney failure or dialysis 85 88 0.97 0.72-1.30 0.82

Other symptoms and side effects

Stomach upset or pain 4887 4843 1.01 0.97-1.05 0.72
Nausea 3558 3550 1.00 0.96-1.05 0.94
Constipation 5184 5111 1.01 0.97-1.05 0.51
Diarrhea 5599 5580 1.00 0.97-1.04 0.77
Skin rash 3331 3367 0.99 0.94-1.03 0.58
Bad taste in mouth 2240 2245 1.00 0.95-1.06 0.92
Increased burping 2217 2158 1.03 0.97-1.10 0.29

From Cox regression models controlling for age, sex, and vitamin D randomization group (intention-to-treat
analyses). Analyses have not been adjusted for multiple comparisons.

19
Figure S1. Flow Diagram of Enrollment in the Omega-3 Fatty Acid Component of the Trial

401,605 Completed initial screening

questionnaire and were
assessed for eligibility

39,430 Initially willing and eligible;

entered run-in phase

13,559 Excluded because they were

non-compliant, or became
unwilling or ineligible

25,871 Randomized

6463 6470 6464 6474

Active omega-3 Active omega-3 Placebo omega-3 Placebo omega-3
Active vitamin D Placebo vitamin D Active vitamin D Placebo vitamin D

12,933 Allocated to active 12,938 Allocated to placebo

omega-3 fatty acids omega-3 fatty acids

Status at end of intervention: Status at end of intervention:

11,440 Known to be alive 11,423 Known to be alive
972 Alive per NDIa search 1,003 Alive per NDIa search)
521 Died 512 Died

12,933 Analyzed 12,938 Analyzed

aNational Death Index

20
Figure S2. Cumulative Incidence Rates of A) Expanded Cardiovascular Events, B) Total Myocardial Infarction, C) Total Stroke, and D) Cardiovascular Mortality,
By Year of Follow-up. From Cox regression models controlling for age, sex, and vitamin D randomization group (intention-to-treat analyses). The insets show the
data on an enlarged y axis.

21
Figure S3. Hazard Ratios of Total Myocardial Infarction by Subgroups, Comparing Omega-3 Fatty Acids (n-3) and
Placebo Groups. From Cox Regression Models Controlling for Age, Sex, and Vitamin D Randomization group (intention-
to-treat analyses). Analyses were not adjusted for multiple comparisons.

22