CLINICAL AND METABOLIC EFFECTS OF ALTERING OMEGA-3 AND OMEGA-6 FATTY ACIDS IN MIGRAINE February 21, 2014 Doug Mann, MD, Professor of Neurology University.

CLINICAL AND METABOLIC EFFECTS OF ALTERING

OMEGA-3 AND OMEGA-6 FATTY ACIDS IN MIGRAINE

February 21, 2014

Doug Mann, MD, Professor of Neurology

University of North Carolina, Chapel Hill

Outline

• Specific Aims

• Rationale for testing dietary modification for chronic pain

• Preliminary studies: Basic hypotheses, Chronic Daily Headache study, results from animal studies

• Molecular mechanisms linking endogenously-produced lipid mediators to physical pain

• Current R01 : study design, questions.

Studies of Diet and Chronic Pain at UNC, Program on Integrative Medicine

1.) Closed to enrollment – Chronic Daily Headache (CDH) funded by

Mayday Fund 2011- 2013. Ongoing data analysis outcomes. 2.) Starting - RO1 funded Sept 2013.

Episodic Migraine (EM) and diet --- 3 arms. 153 subjects.3.) Pending Review - Post traumatic headache in soldiers --- 3 arms.

SPECIFIC AIMS: R01 MIGRAINE STUDY

Clinical and metabolic effects of altering

omega-3 and omega-6 fatty acids in migraine

Study Purpose

To assess whether:

targeted PUFA modifications designed to increase dietary n-3 EPA and DHA, with or without concurrent reduction in n-6 LA, can increase analgesic derivatives of n-3 EPA and DHA, and improve headache-related clinical outcomes.

Specific Aim 1

• To assess the efficacy of the dietary interventions in inducing the predicted changes in circulating PUFA endovanilloid derivatives. • (1a) Compared to Diet C, Diet A will produce significant

increases in analgesic derivatives of n-3 EPA and DHA and reductions in pro-nociceptive n-6 AA-derived and LA-derived endovanilloids.

• (1b) Diet B (High n-3, High n-6 LA) will result in values for analgesic derivatives of n-3 EPA and DHA intermediate between Diet A and Diet C.

Specific Aim 2

• To compare the clinical efficacy of the dietary interventions in adults with migraine.

• Compared to Diet C, Diet A will produce significant improvement in: • (2a) the Headache Impact Test—a headache-specific

quality of life measure, and• (2b) a significantly steeper rate of decrease in

headache hours per day as compared with Diet C. • (2c) Diet B will result in changes in clinical outcomes

intermediate between Diet A and Diet C.

Specific Aim 3• To test our model of the proposed causal chain linking

changes in n-3 and n-6 PUFAs, endovanilloid derivatives, and HA endpoints.

RATIONALEClinical and metabolic

effects of altering omega-3 and omega-6 fatty acids

in migraine

Rationale

• Migraine common (16% women, 7% men)• Debilitating, painful, costly, life-impacting• Good abortive therapies (cost & side effects)• Preventive therapies overall somewhat disappointing

• Timing ideal for a new strategy

Rationale: food and headache

• Traditional consensus with limited evidence beyond patient reports. • Neurotransmitter precursors in red wine, aged cheese,

pickled foods, processed meats, other.• Allergies – gluten, dairy.• Allergy testing for specific foods. • Patient reports of specific food triggers: citrus,

aspartame, caffeine, eggs, breads, other. • Elimination diet (chicken and rice) with add-backs

Rationale: other potential dietary influences

• Magnesium intake as a daily supplement is effective in some with EM. No predictive elements except menses. No dietary studies.

• Riboflavin as a daily supplement is effective – replicated outcomes in EM. No dietary studies.

• Alpha-lipoic acid – weak clinical evidence of effectiveness in EM. No dietary studies.

Migraine Co-Morbidities• Obesity --- headache (inflammation) • Crohn’s disease --- migraine • Ulcerative colitis --- migraine• Irritable bowel syndrome (Aydinlar, 2013)• Omega-6 FA consumption• Gastrointestinal symptoms

• Nausea, diarrhea, cramping, bloating, pain

Headache and Fatty Acids

• One prior study of Omega-3 FA supplements in migraine (Pradlier, 2001)• Negative clinical, migraine-related outcomes. • Minimally affected group (average < 3 migraines/month)• No confirmation of compliance, no biomarkers• No biochemical outcomes• No consideration of diet and ratios of omega-3 to

omega-6 FA intake.• Active intervention for 16 weeks• 196 subjects (96 intervention, 87 placebo) • Strong placebo effect

Omega-6 Consumption 1950 to 2000

Polyunsaturated Fats in U.S. Diets 2008

Linoleic AcidAlpha Linolenic AcidArachidonic AcidEPA+DHA

n-3 ALA

Estimated from Day 1 of 24-hour dietary recall interviews conducted in What We Eat In America, NHANES 2007-8

n-6 LA

n-6 AAn-3 EPA+DHA

7 % of energy

US per capita consumption of vegetable oils in the 20th century

Blasbalg et al AJCN 2011

kg/

p/y

0

2

4

6

8

10

12

14

Canola0.9

Year

Palm

Peanut

Safflower

SesameSunflower

0.10.20.30.40.50.60.70.8

1909

1919

1929

1939

1949

1959

1969

1979

1989

1999

Coconut

CornCottonseed

Olive

SoybeanLinoleic Acid

US per capita apparent consumption of n-6 LA and n-3 ALA in the 20th century

Blasbalg et al AJCN 2011

LA 2.2

ALA 0.35

n-6 LA

n-3 ALA

Overview: biochemistry of n-3 and n-6 fatty acids

Linoleic Acid

9-HODE

Arachidonic Acid

Prostaglandin E2

Arachidonoyl Ethanolamide

15-HETE

α-Linolenic Acid

9-HOTrE

Eicosapentaenoic Acid

Docosahexaenoic Acid

Resolvin E1

18-HEPE

Docosahexaenoyl Ethanolamide

Protectin D1

EndoVanilloids

Eicosanoids

EndoCannabinoids

EndoVanilloids

Docosanoids/ Protectins

Synaptamide

Essential Dietary Fats and their Bioactive Metabolites

Eicosanoids

13-HODE

EndoCannabinoids

EndoVanilloids

Omega-6 Omega-3

Diet and Physical Pain: Hypotheses

Targeted changes in dietary fatty acids alter tissue fatty acids.

Alters the endogenous production of bioactive lipid mediators

(e.g. eicosanoids, endovanilloids, endocannabinoids, resolvins).

Alters the neurochemical milieu in a manner that mayattenuate physical pain.

General model

Mechanisms linking n-3 & n-6 fatty acids to physical pain

Rationale for targeted

dietary intervention

PRELIMINARY STUDIESClinical and metabolic

effects of altering omega-3 and omega-6 fatty acids

in migraine

Targeted alteration

of dietary n-3 and n-6 fatty acids

for treatment of chronic headaches:

a randomized trial

Ramsden CE, Faurot K, Mann JD et al., Trials 2011, BJN 2012, PAIN 2013

Chronic Daily Headache

• Up to 5% of adults. Migraine - 16 and 6 % • 15 or more headache days per month. • HA for 4 hours or more per headache day.• Six months or more duration. • With or without migraine features. • Excluded “organic causes”. • Medication responses considered in the dx.

Dietary Interventions

• H3-L6 intervention• Increase n-3 EPA and DHA • Reduce n-6 LA

• L6 intervention • Maintain low n-3 EPA and DHA intakes (typical of US)• Reduce n-6 LA and n-6 AA

MacIntosh BA, Ramsden CE et al. BJN 2012

‘Chronic Daily Headache’

Headache characteristics

Chronic migraine

Chronic tension-type headache

15 headache days per month

4 headache hours per day

Ramsden CE, Faurot K, Mann JD et al., Trials 2011

bumpybrains.com

Patient population

Methods

• Adults meeting Inclusions/Exclusions• Provided 2 meals and two snacks per day• We provided all oils and fats• All provided food - biochemical analyses at NIH• Intensive dietary counseling by study dietitian • Guidelines for cooking, shopping, dining out• Baseline phase - 4 weeks; intervention 12 weeks• Visits every 2 weeks during the intervention for dietary

counseling and food pick up, diary review.

Methods• No supplements: requested they not start on them during

the study. • Not given other dietary suggestions relating to

traditional consensus advice re: diet. • If they were on fish oil or PUFAs for headache,

they were excluded. • Web based headache diary

• Continue care with neurologist or other MD.

Methods

• Arm 1. Low omega-6 (L6)• Arm 2. Low omega-6, high omega-3 (L6 H3)

• Both possibly anti-nociceptive• Clinical and biochemical primary endpoints• Multiple secondary outcome measures

Overview of Trial Design

• Randomized, parallel-group clinical trial

L6 intervention

H3-L6 intervention

• Dietitian counseling and food provision every 2 weeks

• Patients continued usual headache care throughout trial

• Headache-related quality-of-life (HIT-6)• Headache days per month • Headache hours per day• Headache medication use • Psychological distress (BSI-18)• Physical and mental function (SF-12)

Clinical Outcomes

• **Erythrocytes (n-6 LA, AA; n-3 EPA, DHA)

Ramsden CE, Mann JD et al., Trials 2011, PAIN 2013

Biochemical Outcomes

• Eicosapentaenoic acid (e.g. 18-HEPE)• Docosahexaenoic acid (e.g. 17-HDHA)• Linoleic acid (e.g. 9- and 13-HODE and -oxoODEs)• Arachidonic acid (e.g. 5-, 8-, 9-, 11-, 12-, 15-HETE)

Circulating fatty acid biosynthetic precursor pools

Anti- and pro-nociceptive n-3 and n-6 metabolites

Trial ProfileAssessed for eligibilityTotal Screened: n=211

EnrollmentIneligible(n=144)

Randomized(n=67)

Allocated to L6 intervention(n=34)

Allocated to H3-L6 intervention(n=33)

Discontinued intervention(n=6)

Intention-to-treat AnalysisHIT-6 and Headache Days

(n=34)

Allocation

Follow-Up

Analysis

Discontinued intervention(n=5)

Intention-to-treat AnalysisHIT-6 and Headache Days

(n=33)

Chronic Daily Headache Patient Characteristics

(g)

MacIntosh BA, Ramsden CE, Mann JD et al. BJN 2012

LA, EPA and DHA Consumption in Chronic Daily Headache Trial

*LA intake is expressed as a percentage of daily food energy (%E). Median intakes assessed via six 24-hour dietary recalls administered on non-consecutive days.

Diets altered erythrocyte fatty acid content in a manner predicted to reduce physical pain

Ramsden CE, Mann JD et al., Trials 2011, PAIN 2013

% c

han

ge

(12

wee

ks)

L6 H3-L6

H3-L6 intervention produced greater pain reduction

HIT-6Headache days/

month

-50

-40

-30

-20

-10

0

Headache hours/day

Severe Headache

days

Between-group comparisons p<.001 p<.02 p<.01 p<.02

P-diff = 0.01

Headache hours per day by diet group

Ramsden CE, Mann JD et al., Trials 2011, PAIN 2013

Was clinical improvement due to increased medication use?

Change in medication use by diet group

Clinical effects of the H3-L6 and L6 interventions on SF 12

H3-L6 Phys-ical

H3-L6 Men-tal

L6 Physical L6 Mental

Base-line

43 47.5 44.8 43.8

12 weeks

49.7 51.7 45.9 47.2

27.532.537.542.547.552.557.562.5

Ramsden, Faurot, Mann et al, unpublished

self-

repo

rted

hea

lth*

0 20 40 60 80

2.4

2.6

2.8

3.0

days since randomization

Daily ratings of general health

Group difference p-value = 0.03**

H3L6

L6

* 1= poor; 4= excellent ** Proportional odds longitudinal model, group x time interaction

Clinical effects of the H3-L6 and L6 interventionson psychological distress (BSI-18)

Ramsden, Faurot, Mann et al, unpublished

Ge

ne

ral

So

ma

tiza

tio

n

An

xie

ty

De

pre

ss

ion

Ge

ne

ral

So

ma

tiza

tio

n

An

xie

ty

De

pre

ss

ion

H3-L6 L6

59

60

61

62

63

64

65

66

67

68

69

Baseline12 weeks

Diet-induced changes in anti- and pro-nociceptive lipid mediators

Results Summary

The H3-L6 intervention:

Produced marked alterations in circulating n-3 and n-6 derived: Endovanilloids

EicosanoidsResolvin pathway precursors – to be analyzedEndocannabinoids** to be analyzed

Produced statistically significant, clinically relevant improvements in:

Headache hours per daySevere headache days

Quality of lifePhysical functionEmotional distress

Limitations

• Small trial• Changes in fatty acids not independent• No true control group• Mediators still unclear• Used only blood tissues—unable to determine if other

pertinent tissues changed

Potential mechanisms responsible for pain reduction

Attenuation of

TRPV1 hyper-activation

Anti-nociception

TRPV1-mediated neurogenic inflammatory cascade

Substance P

CGRP

TRPV1

Physical Pain Anxiety

Alcohol Craving

NK1 Recepto

r

Physical Pain

VasodilationTolerance

Dependence Withdrawal

CGRP Recept

or

pH

Heat

+

-

Endogenous vanilloid agonists

Endogenous vanilloid inhibitors

Linoleic Acid

9-HODE

Oxidized Linoleic Acid Metabolites are TRPV1 Agonists

13-HODE

9-oxo-ODE 13-oxo-ODE

D-series resolvins and NPD1 inhibit TRPV1 activation

Docosahexaenoic Acid

Resolvin D2

Protectin D1

17-hydroxy-DHA

7 (S) Maresin 1

14-hydroxy-DHA

Endovanilloids and the neurochemistry of pain

Can diet alter endovanilloids in key ‘pain tissues’?

Diet-induced changes in anti- and pro-nociceptive lipid mediators in circulation

Summary & Conclusions

Targeted dietary manipulation of n-3 and n-6 fatty acids: • Reduced pain and improved quality of life in

chronic headache

• Could represent a novel strategy for treating chronic pain in general

• Future trials evaluating clinical efficacy and elucidating biochemical mechanisms in chronic pain are warranted

Future Directions

9-HODE 13-HODE

CO

X/L

OX

15-L

OX

-1

MaR1

14-HDoHE

12

-LO

X

15-LOX

NPD1

17-HDoHE

RvD1

RvD2RvD3

RvD4

LA

PGE2

COX1/2

5-LO

X

5-HETE

LTB4

12-LOX

15-LOX-2

15-HETE

LXA4 & LXB4

5-L

OX

TXB2

AA

DHA

EPA

RvE1

18-HEPECOX2/CYP

5-

LO

X

RvE2

12-HETE

5-L

OX

EH

5-LOX

12-LOX

15

-LO

X-1

9-OxoODE 13-OxoODE

PGD2

PGF2α

20-OH-LTB4

20-COOH-LTB4

5,15-DiHETE

12-L

OX

5-HEPE12-HEPE

5-L

OX

15-HEPE

15-

LOX

RvD5RvD6

5-LOX4-HDoHE

7-HDoHE

PGE3

COX1/2

TXB3

PGD3

PGF3α

CYP

LXA5 & LXB5

5,15-DiHEPE

RvE3

12/1

5-

LOX

8-HETE9-HETE11-HETE

Ramsden, Rapoport, Yuan, RemaleyIndicates mediators assayed for Chronic Daily Headache trial

NIH Clinical Center LC/MS/MS assay for lipid mediators of nociception

DESIGN OF R01 TRIAL: DIET AND MIGRAINE

Clinical and metabolic effects of altering

omega-3 and omega-6 fatty acids in migraine

Three arm trial• ARM 1: Diet A—High n-3 EPA+DHA, Low n-6 LA

• ARM 2: Diet B—High n-3 EPA+DHA, High n-6 LA

• ARM 3: Diet C—Low n-3 EPA+DHA, High n-6 LA (average U.S. intake; control group)

Overview of Trial Design

• Randomized, parallel-group clinical trial

• Dietitian counseling and food provision every 2-3 weeks

• Patients continue usual headache care throughout trial

Randomization,Blood Collection

Study End,Blood Collection

6 wk f/u,

Blood Collection

H3 – L6 intervention

Baseline Phase

H3 – H6 intervention

L3 – H6 control

4 weeks 16 weeks 6 weeks

Fatty Acid Targets

Main diet changes to achieve nutrient targets:• Replace all oils, spreads and salad dressings with those provided by the study• Avoid processed foods that contain oils. Replace with foods provided by the

study• Consume study provided fish daily (or consume low fat meat, fish and poultry on

Control)

Fat Low n-6, high n-3

Ave n-6, high n-3

Ave n-6, Ave n-3 (Control)

Total Fat (%en) 30% 30% 30%

Monounsaturated (%en) 15% 12% 12%

Polyunsaturated (%en) 4% 7.5% 7.5%

Saturated Fat (%en) 11% 11% 11%

Linoleic acid (n-6) (%en) 2% 7.2% 7.2%

EPA + DHA (mg) 1500 1500 150

16-Wk Diet Intervention, 6-Wk f/u• Personalized nutrition counseling–9 sessions• Food provision -prepared and

unprepared foods• Diet education materials• Diet website• Seven day meal plan• Self-monitoring• 24-hr dietary recalls

• baseline, intervention, post-intervention

Diet Methods Paper: MacIntosh BA, Ramsden CE, Raurot KR, Zamora D, Mangan M, Hibbeln JR, Mann JD. Low n-6 and low n-6 plus high n-3 diets for use in clinical research. Br J Nutr. 2013 Jan 18:1-10

Meal Plan Comparisons

Measures-Clinical (in addition to daily diary)

Baseline

Wk 0

Wk 4

Wk 10

Wk 16

Wk 22

Demographics X

Headache History/Physical Exam X x

HIT-6 X X X X X X

PROMIS-29 Profile X X X X X X

Comorbid pain assessment X x x x x x

MIDAS X X

Satisfaction with care/diet X X X X X

Periodic Assessment of Care for Headaches and Pregnancy Status

X X X X X

Expectation of Benefit X

Food Preference / Dispensing X X X

Blood draw X X X X X

Measures-BiochemicalAnalyte Formula Method

n-6 fatty acidsLA 18:2 n-6 GCAA 20:4 n-6 GCDTA 22:4 n-6 GCDPA n-6 22:5 n-6 GC n-3 fatty acidsALA 18:3 n-3 GCEPA 20:5 n-3 GCDPA (n-3) 22:5 n-3 GCDHA 22:6 n-3 GCOmega-3 Index EPA + DHA GC Other fatty acidsPalmitic acid 16:0 GCOleic acid 18:1 n-9 GC  Precursor  

Endovanilloids (LA and AA-derived TRPV1 agonists)9-HODE LA LC/MS/MS13-HODE LA LC/MS/MS9-oxoODE LA LC/MS/MS13-oxoODE LA LC/MS/MS12-HETE AA LC/MS/MS15-HETE AA LC/MS/MS

Measures-Biochemical Endovanilloids (EPA and DHA-derived TRPV1 antagonists)17-hydroxy-DHA (Primary Outcome) -precursor to D-series resolvins

DHA LC/MS/MS

18-HEPE -precursor to E-series resolvins

EPA LC/MS/MS

Endocannabinoids2-AG AA LC/MS/MSAEA AA LC/MS/MS2-DHG DHA LC/MS/MSDHEA DHA LC/MS/MSOEA Oleic acid LC/MS/MSPEA Palmitic acid LC/MS/MS Eicosanoids    PGE2 AA LC/MS/MSLTB4 AA LC/MS/MSTXB2 AA LC/MS/MSLipoxin A4 AA LC/MS/MS Other analytesSubstance P N/A ELISACalcitonin gene-related peptide N/A ELISACytokines (IL-1, Il-6, IL-10, TNF, MCP-1) N/A Multiplex-ELISA

References• Ramsden CE, Mann JD, Faurot KR, Lynch C, Imam ST, MacIntosh

BA, Hibbeln JR, Loewke J, Smith S, Coble R, Suchindran C and Gaylord SA. Low omega-6 vs. low omega-6 plus high omega-3 dietary intervention for Chronic Daily Headache:  Protocol for a randomized clinical trial.  Trials Journal 12: 97-105, 2011.

• MacIntosh BA, Ramsden CE, Faurot KR, Zamora D, Mangan M, Hibbeln JR, Mann JD. Low n-6 and low n-6 plus high n-3 diets for use in clinical research. Br J Nutr. 18: 1-10, 2013.

• Ramsden CE, Ringel A, Feldstein AE,  Taha AY, Macintosh BA, Hibbeln JR, Majchrzak- Hong SF, Faurot KR, Rapoport SI, Cheon Y, Chung Y, Berk M, Mann, JD. Lowering dietary linoleic acid reduces bioactive oxidized linoleic acid metabolites in humans. Prostaglandins Leukot Essential Fatty Acids 87: 135-141. 2012.

References continued

• Finkel AG, Yerry, JA, Mann JD. Dietary considerations in migraine management: Does a consistent diet improve migraine? Curr Pain Headache Rep 1: 373-376, 2013.

• Ramsden CE, Faurot KR, Zamora D, Suchindran CM, Macintosh BA, Gaylord S, Ringel A, Hibbeln JR, Feldstein AE, Mori TA, Barden A, Lynch C, Coble R, Mas E, Palsson O, Barrow DA, Mann JD. Targeted alteration of dietary n-3 and n-6 fatty acids for the treatment of chronic headaches: a randomized trial. Pain, 154: 2441-51, 2013.

Acknowledgements

• UNC Program on Integrative Medicine• UNC Dept of Physical Medicine and Rehabilitation• UNC Department of Neurology. • UNC NCCAM T-32 Integrative Medicine Fellowship• Mayday Fund• UNC TraCS• Intramural Program of NIAAA• UNC-Chapel Hill CTSA • UNC NORC• UNC CHAI Core• John M. Davis

AcknowledgementsChris RamsdenDouglas MannKim FaurotBeth MacIntoshC. SuchindranSusan GaylordDaisy ZamoraChanee LynchAngela JohnstonBecky CobleAmit RingelDavid Barrow

Marjorie BusbyOlafur PalssonBeth FowlerCarol CarrTim McCaskillMerit McMannisRegina McCoyGus SwensonMeg Mangan

Joseph R Hibbeln

Sharon Majchrzak-Hong

Jim Loewke

Ariel Feldstein

Alexandros Makriyannis

Jodi Wood

Trevor Mori

Anne Barden

Departments of Physical Medicine & Rehabilitationand Neurology.