LA genética del asma en Hispanos

La Genética del Asma in Hispanos Juan C. Celedón, M.D., Dr.P.H.

Channing Laboratory Division of Pulmonary and Critical Care Medicine Brigham and Women’s Hospital Department of Medicine Harvard Medical School

BRIGHAM AND WOMEN’S HOSPITAL

HARVARD MEDICAL SCHOOL

Outline • • • •

Definition of the term Hispanic Asthma among Hispanics in the United States The Genetics of Asthma in Hispanics Study of Asthma Genetics in Costa Rica – Heritability Analysis – Genome-wide linkage analysis of asthma and airway responsiveness – Future Directions

1.

Definition of the Term “Hispanic”

Introduction • 2000 Census: ~32.8 million Hispanics in the United States (~12% of the population) • “Hispanic”: Common heritage of language and culture

2.

Asthma in Hispanics

Asthma in Hispanics • Asthma is a significant cause of morbidity in certain Hispanic subgroups in the U.S. (e.g., Puerto Ricans) and Hispanic America (e.g., Costa Ricans) • Little is known about genetic and environmental influences on asthma in Hispanics Hunninghake GM, Weiss ST, Celedón JC. State of the Art: Asthma in Hispanics. Am J Respir Crit Care Med 2006; 173:143-163.

3.

The Genetics of Asthma in Hispanics

The Genetics of Asthma in Hispanics • To date, genome-wide analyses for linkage to asthma phenotypes have been conducted in 18 distinct populations – Until recently, only one genome scan included any Hispanic individuals

Results of Selected Candidate-Gene Association Studies for Asthma and/or Asthma-Related Phenotypes in Hispanics Gene

Study Sample Hispanics with (n=112) and without (n=126) asthma

Mexican (n=265) and Puerto Rican ADAM33 (n=318) families of individuals with asthma 47 Hispanic families of children with asthma

Number of SNPs Studied

Results

Reference

8

Two SNPs in ADAM33 were Howard T, associated with asthma and et al1 allergen sensitization.

6

No association between any Lind D, et of the SNPs tested and al2 asthma or asthma-related phenotypes.

17

No association between any Raby B, et of the SNPs tested and al3 asthma or asthma-related phenotypes.

Candidate-Gene Association Studies in Hispanics: Limitations • • • • •

Small sample size Inadequate definition of Hispanic subgroups Heterogeneity of populations studied Lack of correction for multiple comparisons Lack of adequate data on environmental exposures • (For case-control studies): lack of detection and control of potential population stratification

4.

The Genetics of Asthma in Costa Rica

4.a.

Rationale and Study Design

Asthma in Costa Rica • Prosperous Latin American Country – Universal access to health care – High literacy rate (~95%)

• High prevalence of childhood asthma in Costa Rica (~23%)

Costa Rica • 1697 Census – 2,100 Spaniards – 1,400 Amerindians – 300 individuals of African descent – 160 Mestizos

The Central Valley of Costa Rica • Most Costa Ricans live in a Central Valley separated from the coasts by mountain ranges • Minimal migration in and out of the Central Valley until the late 1800s

Family-based Study of Asthma Genetics in Costa Rica • Rationale: – To study “complex” diseases, a useful strategy is to study communities that are relatively isolated genetically

• Study Hypothesis: Hypothesis – A small number of chromosomal regions contribute to the expression of asthma in Costa Ricans

Family-based Study of Asthma Genetics in Costa Rica • Funded by the NHLBI since March of 2001 • Phenotypic and genotypic data collected in 2,471 individuals – 671 members of large families of children with asthma – 1,800 members of nuclear families of children with asthma

Data Collected in Members of Family Pedigrees • Children and adults: – – – – – – –

Questionnaire information Spirometry with B.D.R. Methacholine challenge testing Total and specific serum IgE Eosinophil count Skin test reactivity to allergens Blood sample for DNA extraction

4.b.

Subject Characteristics and Heritability Analysis

Table 1. Characteristics of Family Members (n=687) Female sex, n(%)

368 (52.7)

Age in years, mean (range)

29.5 (6.4-92.6)

Total serum IgE (IU/ml), geometric mean (range)

97.0 (0-5,000)

Eosinophil count (cells/mm3), geometric mean (range)

208.4 (30 to 7,500)

Number of positive skin tests, median (range)

2 (0 to 9)

% Predicted Post-BD FEV1, mean (range)

102.4 (33.6-152.4)

% Predicted Post-BD FEV1/FVC, mean (range)

99.9 (45.8-115.9)

Table 1. Characteristics of Family Members (n=687) Physician-diagnosed asthma, n(%) Age <13 years Age >=13 years Current wheezing, n (%) Age <13 years Age >=13 years

69 (39.7) 128 (25.0) 64 (36.8) 166 (32.4)

Physician-diagnosed asthma and current wheezing, n (%) Age <13 years Age >=13 years

52 (29.9) 87 (17.0)

Ever smoker, n (%)

109 (15.9)

Current smoker, n (%)

47 (6.8)

Table 2. Heritability Analysis of Asthma-Related Traits h2N (standard error) Total serum IgE (IU/ml)&

56.6% (6.5%)

Eosinophil count (cells/mm3)&

29.3% (7.6%)

Airway responsiveness (d.r.s. to methacholine)*

49.9% (9.2%)

Post-BD FEV1 (L)*

23.0% (6.5%)

Post-BD FEV1/FVC*

14.3% (6.3%)

& Variance components analysis adjusted for age and sex. *Variance

components analysis adjusted for age, sex, and height.

Note: All heritability (h2N) values > 0 with p<0.0001

4.c.

Genome-wide Linkage Analysis

Overview of Complex Trait Genetics Demonstrate Familial Aggregation and/or Heritability of Trait Positional Cloning

Candidate Gene Approach

Localize Susceptibility Gene(s)

Identify Disease Susceptibility Gene and Functional Variants

Diagnostics

Determine Fraction of Explained Variation

Treatment

How to find a disease-susceptibility gene?

• Difficult challenge: like finding a misspelled word in a set of encyclopedias!

A 1

Z 24

7

Vol 7

page

‘which chromosome?’ ‘chromosomal region’

Plausible candidate genes >100 !

This is a sentence in a paragraph…

‘gene’

This it a sentence in a paragraph…

‘mutation’

Linkage Analysis â&#x20AC;˘ Definition: A group of methods that analyze the distribution of DNA markers within families to determine if a particular region of the genome contains a gene related to the phenotype of interest. â&#x20AC;˘ LOD Score: Result of a statistical test to determine if genetic loci are linked, expressed as log10 of the odds that the loci are linked. LOD scores of 3-3.6 correspond to highly significant linkage.

Whole Genome Scans â&#x20AC;˘ A genome scan consists of ~400 STR markers spread across all 23 human chromosomes. These markers are like a dragnet, enabling us to pinpoint the location of genomic regions containing disease susceptibility genes. â&#x20AC;˘ Shown on the right is a map of human chromosome 1 with different type of markers (SNPs and STR or microsatellite markers) overlaid on the map.

Genome Scan for Asthma and Asthmarelated Phenotypes in Costa Rica â&#x20AC;˘ Genome scan was completed in November of 2004 at the Genome Quebec Innovation Centre in 671 individuals â&#x20AC;&#x201C; 380 short-tandem repeat (STR) markers, with an average spacing of 8.2 cM (ranging from 6.2 cM on chromosome 22 to 9 cM on chromosome 17) - Marker locations were determined on the basis of the deCODE map

Short-Tandem Repeats: Repetitive elements flanked by unique sequence 5’

CAG

CAG

CAG

CAG

GTC

GTC

GTC

GTC

2

3

1

3’

4

GTC

GTC

GTC

GTC

GTC

GTC

GTC

CAG

CAG

CAG

CAG

CAG

CAG

CAG

1

2

3

4

5

6

7/4

8/5

7

7/5 GF

GM

Mom

Dad

Son

8/2

Ladder 8 7 6 5 4 3 2

7/2

Genome-wide Linkage Analysis of Airway Responsiveness • Phenotype: – Dose-response slope to methacholine, which was logtransformed (residual kurtosis <0.9)

• Variance component linkage analysis conducted with the SOLAR program – Age, sex, height, smoking (ever), pack-years, and quadratic terms were considered for inclusion in final models

• All analyses were repeated in nonsmokers only

Table 1. Genome-wide Linkage Analysis of Airway Responsiveness

Chromosome

cM

Maximum LOD score All subjects (n=488)

6

134

10

147

12

23

15

Nonsmokers (n=423)

1.41 1.75 1.50

146

1.32

2.33

119

1.01

1.04

Staged Genomic Search •

A genome scan (Stage 1 or “Random Genome Scan”) is carried out at a low marker density and then those regions which may be linked to the trait of interest are investigated by typing more markers (Stage 2 or “Fine Mapping Linkage Analysis”)

Discussion â&#x20AC;˘ First genome-wide linkage analysis of airway responsiveness in a Hispanic population â&#x20AC;˘ Chromosome 12q24 likely contains a locus (or loci) that influence airway responsiveness, a critical intermediate phenotype of asthma

Sex Differences in Total Serum IgE in Costa Rican Pedigrees

p=0.003

Comparisons adjusted for age, age2, ever smoking, pack-years (py) and py2.

Narrow-sense heritability of log10IgE=0.556 (S.E=0.065), p=10-35

Genome-wide Linkage Analysis of Total Serum IgE in Costa Rican Pedigrees

Sex-specific Linkage to Total Serum IgE in Costa Rican Pedigrees

Discussion â&#x20AC;˘ First demonstration of a distinct genetic architecture of total serum IgE in males and females â&#x20AC;˘ Chromosome 20p12 likely contains a gene(s) that influence total serum IgE in males

4.d.

Future Directions

Family-based Study of Asthma Genetics in Costa Rica • Fine-mapping linkage analysis to confirm and narrow regions linked to asthma and/or asthma-related traits – Currently on chromosomes 1p, 7q, 20p

• Fine-mapping association analysis to identify gene(s) associated with asthma and/or its intermediate phenotypes (in nuclear families) – First region: chromosome 12q24

Collaborators • Channing Laboratory, Brigham and Women’s Hospital (Boston, MA): Jody Senter, Barbara Klanderman, Craig Hersh, Matt Hunninghake, Ngoc Ly, Ross Lazarus, Steve Lake, Catherine Liang, Dan Laskey, Jessica Su, Ed Silverman, and Scott T. Weiss • Dept. of Biostatistics, Harvard School of Public Health (Boston, MA): Christoph Lange and Nan Laird

Collaborators • Genome Quebec Innovation Centre (Montreal, Canada): Thomas Hudson • Hospital Nacional de Niños (San José, Costa Rica): Manuel Soto-Quiros, Lydiana Avila, Eduardo Fournier, and Mitzi Spesny

Genetic Studies of Asthma and COPD in Hispanics at BWH The Genetics of Asthma in Costa Rica HL66289, HL04370

The Genetics of COPD in Costa Rica HL073373

Genes, Allergens, and Asthma in Puerto Ricans HL079966

Genome Scan/Linkage →Flanking Markers/Linkage →Positional Candidates/Association →Animal Models/Function