the importance of research on environmental factors in a highly heritable disorder

The importance of research on environmental factors in a highly

heritable disorder

Mark A. Corrales, MPPUS Environmental Protection Agency

Statements are those of the author, and do not necessarily represent views or policies of the EPA

Autism One ConferenceMay 22, 2009

Parallel Universes?

• General public• Parents & affected kids• Interest groups:

– environmental health advocacy– industries

• Researchers• Policy/ decision-makers:

Lawyers, economists, policy and risk analysts, staff scientists, etc.

Getting from here to there

• If you believe national-level medical and environmental health interventions are needed…

• the autism research literature is a key bottleneck.

From evidence to action

Guesses, theories

Anecdotes, case studies

Ecologic studies

Retrospective epidemiology

Prospectiveepidemiology

Experimental toxicology, mechanisms

RCTs

Drug development

Behav./Educ. therapy guidelines

Regulations to limit risk factors

More studies

Better studies

More types of studies (multiple lines of evidence)

Public & scientific consensus

Is there sufficient evidence for action?

Published scientific research literature on environmental factors

in autism is still quite limited compared to other diseases or

health effects.

Why so little research on environmental factors in autism?

1. All autism research limited (until recently)

2. Behavioral > molecular level(until recently)

3. Genes > Environment:High heritabilitymisinterpreted as“genes not environment”

4. Toxicology and epidemiology not sufficiently integrated with autism research so far

Why so little research on environmental factors in autism?

1. All autism research limited (until recently)

2. Behavioral > molecular level(until recently)

3. Genes > Environment:High heritabilitymisinterpreted as“genes not environment”

4. Toxicology and epidemiology not sufficiently integrated with autism research so far

Most autism science is extremely recent

As of early 2009,

• Half of all the autism research literature indexed in PubMed was published in the past eight years: 2000-2008.

• One third was published in just the last five years: 2004-2008.

Very RecentExplosion of Interest in Autism

In 2008:

• Newspaper articles: >3,000 headlines/ year

• Scientific journal articles:>1,000 / year

• New books:>150 / year

Journal articles

But so far…

• Treatment -- New ASD treatments approved: 0

• Primary prevention (at national level) -- Environmental standards, regulations, or other actions taken based on any potential ASD risk factors: 0

Autism Literature is Still Relatively Limited

• If you believe national-level medical and environmental health interventions are needed…

• the autism research literature is a key bottleneck.

What has autism literature focused on?

Why so little research on environmental factors in autism?

1. All autism research limited (until recently)

2. Behavioral > molecular level(until recently)

3. Genes > Environment:High heritabilitymisinterpreted as“genes not environment”

4. Toxicology and epidemiology not sufficiently integrated with autism research so far

Autism literature has focused on behavioral, not chemical level

• The PDD literature focused on the behavioral and cognitive levels three times as much as did biomedical literature as a whole (as % of articles).

• 56% vs. only 38% of publications were indexed with chemical substances in PubMed vs. PDD literature, even recently (2000-2006).

Corrales, Ringer, & Herbert 2008 IMFAR poster #2999

Moldin vs. Lathe biological level

Focus on Chemicals & Drugs Level

48%

30%

0%

25%

50%

75%

100%

Rhini

tis Per

enPr

ion

Bipol

ar

Epile

psy

Schi

zoph

reni

a TB

Depre

ssio

nHIV

ADHD

PDD[M

H] CP

% o

f p

ub

licati

on

s 1

98

5-2

00

7 w

ith

M

eS

H

Chem level

Comorbid disorders at elevated prevalence in PDD according to Rzhetsky (2007)

Why so little research on environmental factors in autism?

1. All autism research limited (until recently)

2. Behavioral > molecular level(until recently)

3. Genetic > Environmental studies:High heritabilitymisinterpreted as“genes not environment”

4. Toxicology and epidemiology not sufficiently integrated with autism research so far

Genetic etiology pie

A few molecular substances have dominated the literature (1985-2007)

• Approx. 2,000 unique substance terms in ASD literature, occurring 12k times in 4,000 PMIDs– Half the substance terms appear only once.– Avg. substance is in just 6 PMIDs

• Chemicals in >100 articles each: – MECP2 gene or protein (>400)– Serotonin (251)– Risperidone (112)

Corrales, Ringer, & Herbert 2008 IMFAR poster #2999.

Medications vs. environmental pollutants

• >130 medications in >1200 PMIDs ( ~10% of articles)• 43 environmental pollutant terms (other than Hg)

occur 111 times across 84 PMIDs ( <1% of articles)– Only 36/111 refer to a specific chemical rather than class

(“teratogens” or “environmental pollutants” seen up to 13 times each)

– Lead and cocaine were the most common specific toxins.– No other specific toxin (except Hg) appeared in more than 4

PMIDs. 3 Hg-related terms in 91 PMIDs (<1%)• 41 dietary substances in 189 PMIDs (1-2%)

(e.g., gluten, folic acid) (excluding food additives)• A few potential treatments/ causes

(valproic acid, oxytocin, melatonin, secretin) found in 26-91 PMIDs each.

• ~20 other terms in 2-14 PMIDs each:(food additives, illegal drugs, infectious agents or allergens, nicotine, alcohol, caffeine)

Corrales, Ringer, & Herbert 2008 IMFAR poster #2999. Includes publications 1985-2007.

Environment-relevant topics understudied in ASD literature,

relative to all biomedical literature

Hazardous Substances + Organophosph.

0.01 0.10 1.00 10.00 100.00

CarcinogensEndotoxins

Organophosphorus CompoundsCyclophosphamide

InsecticidesMutagens

Air PollutantsWater Pollutants, Chemical

PesticidesAsbestosCadmium

Street DrugsPhosphoric Acid Esters

Chlorpyrifos

Rate Ratio (Rate in ASD vs all PubMed citations)

Haz subst

Cell Types

0.01 0.10 1.00 10.00 100.00

Macrophages

B-Lymphocytes

Killer Cells, Natural

T-Lymphocytes

Neurons

Rate Ratio (Rate in ASD vs all PubMed citations)

Cell types

Understudied in ASD vs. in 10 comorbid disorders

(1985-2007)

• IL-6: Studied 9x more often in comorbid literature than in ASD

• Glucocorticoids: 52x• Anti-Inflammatory Agents: 36x• Adrenal Cortex Hormones: 17x• Thyroxine: 7x• Steroids: 7x• Herpesviridae Infections: 16x

Corrales, Ringer, & Herbert 2008 IMFAR poster #2999

Why so little research on environmental factors in autism?

Highly-cited statements on autism and environment

• “Among cognitive diseases, ASDs are the most heritable (about 80%), suggesting that they are determined largely by genes and not by the environment.” [emphasis added]Südhof (2008) Nature

• Heritability is at least 90%.Moldin, Rubenstein, and Hyman (2006) Journal of Neuroscience

Genes and Environmentin Autism:

From Pie to Venn

G AND EDO NOT FORM A PIE CHART

GENES

ENVIRONMENT

If one is trying to show population attributable fractions (PAF), the sum is not 100%. If one is trying to partition variance, GxE should be explained.

G AND EFORM A VENN DIAGRAM

G EG & E

• “What % is genetic?”– Variance studies misinterpreted as ruling out E,

or as if heritability were PAF– Sum of PAFs is not 100%– A case is not G or E

• G and E are needed to explain the risk

G AND Enot G vs. E

G E

G&E

Tuberculosis as “a genetic disease”?

• TB was thought/ known to be “a genetic disease,” because it was seen to run in families.

• Later, we realized some people are genetically more susceptible to the environmental cause (infectious agent).

• TB in the population is caused by the combination of G and E (“GxE”).

• Many examples of GxE have been discovered (mostly in cancers & cardiovascular disease, but also PKU, depression, ADHD, etc.)

How can environment be important if heritability is so high?

I. G x E (GE interaction) Most heritability studies assume no GxE.

II. epiG E (epigenetic change)

III. G E (genetic damage)

IV. G E (GE correlation)

I. Gene-environment interaction (GxE)

• TB example• Most heritability estimates ignore GxEShared • Heritability is overestimated where genes

interact with shared environment.– GxE inflates heritability estimates when

environmental risk factors are shared by dizygotic twins. Looks like the impact of their genes alone, but is actually impact of GxE.

• Shared environment would include:– Prenatal risk factors– Widespread risk factors

Haynes 2003 EHP

G x E: One genotype is more sensitive to environment

II. Epigenetic modifications (de novo or inherited)

• Epigenetic tags add to apparent heritability.

• Epigenetic tags may be set by prenatal or other environment.

• May be de novo or possibly inherited• Epigenetic tags may be amenable to change

via environmental interventions (e.g. diet). (e.g., see Corrales 2009 Autism File magazine)

epiG E

III. Genome damaged by environment • Environmental factors can cause DNA damage (point

mutations or structural variations such as copy number variants, CNVs, due to duplications or deletions)

• This initially appears to be a de novo genetic cause of autism.

• If inherited by subsequent generations it appears to be an inherited genetic cause of autism.

• The root cause actually would be environmental and potentially preventable, not genetic, for such cases.

• Unclear if mutation or CNV rates have risen.• Signatures of mechanism may be informative (type of

SNP, or NHEJ vs. NAHR, FoSTeS, etc.). ROS causes DSBs which precede NHEJ and possibly NAHR. (Wenli Gu et al., 2008 PathoGenetics)

G E

IV. G-E correlation:Genes can affect exposure to

environmental factors• Genes that affect non-shared environmental

exposures increase apparent heritability, but prevention through environmental interventions may be possible in such cases.

• Non-shared exposures driven by genes are those where one DZ twin is more exposed than other, due to their genotype.

• G-> Pica (tendency to put soil, etc. in mouth, causing exposure to lead, etc.)

• G-> Dietary preferences (nutritional deficiency?)• G-> Preference for social interaction, eye contact,

etc.

G E

Historical trends in exposure or environmental interventions are hidden in

heritability estimates if GxE exists

• Irony: If GxE is important, growing environmental exposures will increase disease prevalence, but make heritability estimates higher over time. Apparent impact of environment is reported as smaller and smaller, even though in reality it is growing.

• Twin studies only examine differences attributable to environmental conditions that differ between twins, but some environmental exposures are ubiquitous. If some interventions have not been tried in the studied twins, the potential impacts of those may not be seen.

Recap: How can environment be important if heritability is so high?

I. G x E (interaction)

II. epiG E (epigenetic changes)

III. G E (genetic damage)

IV. G E (correlation)

Why so little research on environmental factors in autism?

1. All autism research limited (until recently)

2. Behavioral > molecular level(until recently)

3. Genes > Environment:High heritabilitymisinterpreted as“genes not environment”

4. Toxicology and epidemiology not sufficiently integrated with autism research so far

Toxicology literature and autism literature have almost no overlap

Published 1985-2007, in PubMed:

– Toxicology literature: 1.5 million articles– Autism literature: 10 thousand– Overlap: Just 690 articles…– Mostly on drug toxicity,

not about environmental chemicals– Perhaps 2-4% of ASD literature

is on chemicals as risk factors?

See also: Herbert, Ringer, Corrales IMFAR 2008 poster #2979

ASD - Symptom - Risk Factor

AUTISM PRENATALINFECTION

Prepulse inhibition

deficit

oftenincludes can be

caused by

One can seek potential risk factors using this logic:

Integrating autism with toxicology or epidemiology literature

ASD - Comorbidity - Risk Factor

AUTISM PRENATALETSADHD

is oftencomorbid with...

has asrisk factor...

In this type of analysis, one identifies known risk factors for diseases that are often comorbid with or autism. The reasoning here is that prenatal environmental tobacco smoke, alcohol, lead, or PCBs might be risk factors for autism because they have been associated with ADHD.

ASD studies rarely investigate environmental risk factors

AUTISM

Behavior orNeuropsych

trait

AUTISM

Gene

AUTISM

Brainregion

AUTISM

Toxin

AUTISM

Cell

MOST STUDIES

VERY FEW

VERY FEW

Much literature linking environmental toxins to autism-related

features, just not to autism per se

AUTISM AUTISM

Gene

AUTISM

Brainregion

AUTISMAUTISM

Cell

AUTISM

Biomarkers

Behavior orNeuropsych

trait

Toxins

ASD – Gene – Toxin

AUTISM

GeneExpression Toxins

Search for Chemicals that Interact with

Autism Gene Candidates • Objective, comprehensive search for risk factors?• 142 genes (autism candidates) selected• Toxicogenomics database queried for reports of

chemicals “affecting” any of these genes(e.g., gene expression changes in animal studies)

• Approx. 500 chemicals were identified, for 122 genes:– xenobiotics (pollutants, pesticides, illegal drugs, etc.)– medications– nutrients– endogenous substances

Corrales, IMFAR 2009 poster

Results: Chemicals that Interact with Autism Gene Candidates

Genes reportedly interacting with the most chemicals

• PON ( >120 chemicals)

• MET, PTEN, ADRB2, TH (>30 each)

• MECP2, TSC2, RELN, UBE3A, GABRB3 (4-14 chemicals each)

• Most other genes (1-2 chemicals so far)

Results: Chemicals that Interact with Autism Gene Candidates

Xenobiotics or related chemicalsreportedly interacting with the most candidate genes

• Carbon tetrachloride (33)• t-butyl peroxide (19), hydrogen peroxide (9)• Sodium arsenite (17), arsenic trioxide (8), arsenic (4), nickel sulfate (9)• LPS (11)• Paraquat (10), chlorpyrifos (4)• Benzene (7), B[a]P (7)• Ethanol (7), tobacco smoke (4)• BPA, TCDD

Endogenous chemicals reportedly interacting with the most candidate genes

• Progesterone & estradiol• Corticosterone• Thyroxine

Dietary • Zinc• Flavonoids• Fats

Recap and Next Steps• Education about heritability of autism:

– G x E (interaction)– epiG E (epi-mutation)– G E (genetic damage)– G E (correlation)

• Research on environmental factors• Integrated study of E and G factors in autism

(e.g. what affects MECP2 expression?)• Integration and prioritization across E findings in

autism literature (zinc, hormones, ROS, cytokines, calcium, cholesterol, neurotransmitters, melatonin, vitamin D, etc.)

• More integration with toxicology and epidemiology from outside the autism literature, pulling in the “autism-relevant” literature by linking autism’s features to environment.

Thank you

APPENDIX/ADDITIONAL SLIDES

Population Attributable Fraction (PAF)

0.0

0.5

1.0

1.5

2.0

e (unexposed) E (exposed)

Ris

k Extra risk

Baseline

What fraction of today’s cases would be avoidedif this risk factor were not present?

News articles

Books with autism in title

Autism funding, trials, research

• NIH research funding per DALY (disease adjusted life year) (FY03-08):– $150 for mental health– $1,100 for cancer– $2,800 for infectious diseases in the U.S.

• Federal grants for autism research (as of mid-2007):– behaviors (almost 40% of grants)– genetics (30-40% of grants)

• Autism research published 1985-2007:– 20% on diagnosis– 10% on therapy– 1% on prevention

• 63 autism-related active intervention trials as of early 2008:– Almost 75% (47 clinical trials) on medications– 14% (9) on behavioral therapy– 10% (6) on dietary therapy or supplements