Download which genes

Assessment of genomewide
association studies
Tuan V. Nguyen
Garvan Institute of Medical Research
Sydney, Australia
WHICH GENES ?
Gene variants ?
False positive problem
Candidate gene studies: reproducibility
problem
600 positive associations between common
gene variants and disease reported 1986-2000
166 were studied 3+ times
6 have been consistently replicated
J N Hirschhorn et al. Genetics in Medicine 2002
Introduction to genomewide
association studies
Genomewide association studies
(GWA)
•
•
•
•
Revolution in gene search
Hypothesis-free driven approach
Scan 100,000-500,000 gene variants (SNPs)
Case – control design (>1000 individuals)
Massive number of tests of
hypothesis
Recent GWA studies in osteoporosis
• Styrkarsdottir U, et al (2008) Multiple genetic loci for bone
mineral density and fractures. N Engl J Med 358:23552365.
• van Meurs JB, et al (2008) Large-scale analysis of
association between LRP5 and LRP6 variants and
osteoporosis. JAMA 299:1277-1290.
• Richards JB, et al (2008) Bone mineral density,
osteoporosis, and osteoporotic fractures: a genome-wide
association study. Lancet 371:1505-1512.
Some gene variants from GWA
Gene variant (SNP)
rs3736228
Gene or location
11q13 (LRP5)
rs3736228
11q13 (LRP5)
rs4355801
rs4988321
LRP5
11q13 (LRP5)
rs2302685
12p12 (LRP6)
rs4355801
rs7524102
8q24 (TNFRSF11B)
1p36 (ZBTB40)
rs6696981
1p36 (close to ZBTB40)
rs3130340
6p21 ()
rs9479055
rs4870044
rs1038304
rs6929137
rs1999805
rs6993813
6q25 (1)
6q25 (1)
6q25 (1)
6q25 (1)
6q25 (1)
8q24 (OPG)
rs6469804
8q24 (OPG)
rs9594738
rs9594759
rs11898505
rs3018362
rs2306033
rs7935346
13q14 (RANKL)
13q14 (RANKL)
2p16 (SPTBN1)
18q21 (RANK)
11p11 (LRP4)
11p11 (LRP4)
Trait and P-value
BMD (p = 2.6 × 10-9)
Fracture (p = 0.02)
BMD (p = 6.3 × 10-12)
Fracture (p = 0.002)
BMD (p = 3.3 × 10-8)
Fracture (p = 0.002)
BMD (p = 0.97)
Fracture (p = 0.95)
BMD (p = 7.6 × 10-10)
BMD (p = 9.2 × 10-19)
Fracture (p = 8.4 × 10-4)
BMD (p = 1.7 × 10-7)
Fracture (p = 2.4 × 10-4)
BMD (p = 1.2 × 10-7)
Fracture (p = 0.008)
BMD (p = 6.2 × 10-7)
BMD (p = 1.6 × 10-11)
BMD (p = 4.0 × 10-11)
BMD (p = 2.5 × 10-10)
BMD (p = 2.2 × 10-8)
BMD (p = 1.8 × 10-14)
Fracture (p = 0.04)
BMD (p = 7.4 × 10-15)
Fracture (p = 0.052)
BMD (p = 2.0 × 10-21)
BMD (p = 1.1 × 10-16)
Fracture (p = 1.8 × 10-4)
Fracture (p = 0.005)
Fracture (p = 0.007)
Fracture (p = 0.02)
What is the credibility of a GWA
finding ?
An observed association with
p<0.05 does not necessarily mean
that the association exists.
In 100,000 tests, 5000 positive
findings could be false positive
Diagnostic test and association test
Diseased
NO
YES
+ve
Sensitivity
P(+ve | D)
Association
-ve
+ve
False +ve
False
True
-ve
+ve
Power
-ve
+ve
P-value
P(+ve | False)
-ve
What do want we to know?
• Probability of association given observed data
(eg posterior probability of association)
or
• Probability of observing data if there is no
association (P-value)
Posterior probability of
association
is a function of
• Prior probability of association (p)
• Power = Pr(significance | association)
Sample size
• P-value = Pr(significance | no association)
Effect size
What is the prior probability of
association for a gene variant ?
Gene search = finding small
needles in a VERY large haystack
• Human genome ~3 billion base pairs long
BUT: Most are vanishingly rare
• 99.9% identical between any two individuals
• ~90% differences between any two individuals
is due to common variants
Hypotheses
• Common disease / common variants (CD/CV)
(Reich & Lander 2001, Pritchard et al 2005)
• ~90% differences between any two individuals
is due to common variants
Prior probability of association (p)
• Common variants in the human population: 10
million (Kruglyak and Nickerson Net Gent 2001)
• No. of genetic variants associated with a common
disease ~100 or less (Yang et al, Int J Epidemiol 2005)
Prior probability of association
p = 0.000001
A Bayesian interpretation of association
Power = 95%;
P-value=0.00001
10,000,000
common variants
True association (100)
Significant
(95)
Non-significant
(5)
No association (9,999,900)
Significant
(100)
Non-significant
(9,999,800)
P(True association given a significant result) = 95 / (95+195) = 48%
A Bayesian interpretation of association
Power = 95%;
P-value=0.00000001
10,000,000
common variants
True association (100)
Significant
(95)
Non-significant
(5)
No association (9,999,900)
Significant
(1)
Non-significant
(9,999,800)
P(True association given a significant result) = 95 / (95+1) = 99%
P-value and “true” association
P-value in the range of 5% - 0.1% will virtually
be false positives even in large scale studies
P-value for a reliable association
P < 5 x 10-5
or
P < 5 x 10-8
For 1000 cases and 1000 controls,
p< 10-8 are more likely to be true than false
Some gene variants from GWA
Gene variant (SNP)
rs3736228
Gene or location
11q13 (LRP5)
rs3736228
11q13 (LRP5)
rs4355801
rs4988321
LRP5
11q13 (LRP5)
rs4355801
rs7524102
8q24 (TNFRSF11B)
1p36 (ZBTB40)
rs9479055
rs4870044
rs1038304
rs6929137
rs1999805
rs6993813
6q25 (1)
6q25 (1)
6q25 (1)
6q25 (1)
6q25 (1)
8q24 (OPG)
rs6469804
8q24 (OPG)
rs9594738
rs9594759
rs11898505
rs3018362
rs2306033
rs7935346
13q14 (RANKL)
13q14 (RANKL)
2p16 (SPTBN1)
18q21 (RANK)
11p11 (LRP4)
11p11 (LRP4)
Trait and P-value
BMD (p = 2.6 × 10-9)
Fracture (p = 0.02)
BMD (p = 6.3 × 10-12)
Fracture (p = 0.002)
BMD (p = 3.3 × 10-8)
Fracture (p = 0.002)
BMD (p = 7.6 × 10-10)
BMD (p = 9.2 × 10-19)
Fracture (p = 8.4 × 10-4)
BMD (p = 6.2 × 10-7)
BMD (p = 1.6 × 10-11)
BMD (p = 4.0 × 10-11)
BMD (p = 2.5 × 10-10)
BMD (p = 2.2 × 10-8)
BMD (p = 1.8 × 10-14)
Fracture (p = 0.04)
BMD (p = 7.4 × 10-15)
Fracture (p = 0.052)
BMD (p = 2.0 × 10-21)
BMD (p = 1.1 × 10-16)
Fracture (p = 1.8 × 10-4)
Fracture (p = 0.005)
Fracture (p = 0.007)
Fracture (p = 0.02)
Number of individuals needed to screen
in population and family
Hypothetical gene
Fracture risk in
Population
Family
5
10
Cumulative risk
40%
80%
Cumulative risk after Rx
20%
40%
Number needed to treat
5
2.5
Frequency of risk
“genotype”
0.2%
50%
Number needed to screen
2500
5
Relative risk
How many genes are required for a
“good” fracture prognosis ?
Odds
ratio
Genotype
frequency
Number of genes needed for AUC of
0.70
0.80
0.90
0.95
1.1
5%
>400
>400
>400
>400
10%
330
>400
>400
>400
30%
150
>400
>400
>400
5%
33
100
280
>400
10%
19
50
150
330
30%
9
23
70
160
1.5
Assessment of GWA finding
• Genetic components of BMD and fracture
• Finding genes of osteoporosis: a
challenge
• Genes can help improve the prognosis of
fracture