Steven Harrison June 8 2017 sharrisonbwhharvardedu RASopathies Developmental Delay Cardiovascular Metabolism Hereditary Cancer Sequence Variant Interpretation WG Harmonize recommendations for modifying ACMG guidelines ID: 915221
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Slide1
BioCuration WGVariant Curation
Steven Harrison
June 8 2017
sharrison@bwh.harvard.edu
Slide2Slide3RASopathies
Developmental
Delay
Cardiovascular
Metabolism
Hereditary Cancer
Sequence Variant Interpretation
WG
Harmonize recommendations for modifying ACMG guidelines
Gene/Disease Specific ACMG Guidelines
General recommendations to ACMG Guidelines
ClinGen Disease
WGs
ACMG/AMP Guidelines
Slide4“…To provide critical flexibility to variant classification, some criteria listed as one weight can be moved to another weight using professional judgment, depending on the evidence collected…”
No direction was provided regarding what criteria code to use in these instances
Slide5To document strength-modified evidence, SVI recommends using the original criteria code followed by an underscore and new level of strength
Pathogenic
Supporting
Moderate
Strong
Segregation
Data
Co-segregation with disease in multiple affected family
members
PP1
Increased segregation data
Slide6To document strength-modified evidence, SVI recommends
using the original criteria code followed by an underscore and new level of strength
Pathogenic
Supporting
Moderate
Strong
Segregation
Data
Co-segregation with disease in multiple affected family
members
PP1
# Co-segregation with disease in multiple affected family
members
PP1_Moderate
## Co-segregation with disease in multiple affected family
members
PP1_Strong
Slide7Population
Data
Segregation
Data
Prevalence in affecteds statistically increased over
controls (OR approach)
Variant identified in
###
probands with consistent phenotypes
PS4
MAF is too high for disorder BA1/BS1
Observation in controls inconsistent with disease
penetrance BS2
Predicted null variant in a gene where LOF is a known mechanism of
diseasePVS1
De novo (paternity & maternity confirmed)
PS2
Well-established functional studies show a deleterious effect PS3
Novel missense change at an AA residue where a different pathogenic missense change has been seen
before PM5
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
De novo
(without paternity & maternity confirmed
)
PM6
Non-segregation with
disease
BS4
Patient’s
phenotype
or FH highly specific for gene
PP4
For
AR
disorders, detected in
trans
with a
path variant
PM3
Found in case with an alternate
cause
BP5
Multiple lines of
comp
evidence
suggest
no impact
BP4Missense when only truncating cause disease BP1Silent variant w/ non predicted splice impact BP7In-frame indels in repeat w/out known function BP3
Well-established functional studies show no deleterious effect BS3
Mutational hot spot or well-studied functional domain PM1
Same AA change as an established path variant PS1
Protein length changing variant PM4
Observed in trans with a dominant variant or Observed in cis with a pathogenic variant BP2
Functional Data
# Co-seg w/ disease in multiple affected family members PP1
De novo Data
Allelic Data
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Missense in gene with low rate of benign missense
variants and path. missenses common PP2
Other Data
Benign
Pathogenic
## Co-
seg w/ disease in multiple affected family members PP1_Moderate
### Co-seg w/ disease in multiple affected family members PP1_Strong
# De novo (without paternity & maternity confirmed) PM6_Strong
# For AR disorders, detected in trans with a path variant PM3_Strong
# Found in case with an alternate cause BP5_Strong
Functional
studies show a deleterious effect
PS3_Moderate
Variant identified in
##
probands with consistent phenotypes
PS4_Moderate
Variant identified in
#
probands with consistent phenotypes
PS4_Supporting
Slide8MAF is too high for disorder
BA1/BS1
Observation in controls inconsistent with disease penetrance
BS2
Absent in population databases
PM2
Prevalence in
affecteds
statistically increased over controls
PS4
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
POPULATION DATA (BA1, BS1, BS2, PM2, PS4
)
Slide9MAF is too high for disorder
BA1/BS1
Observation in controls inconsistent with disease penetrance
BS2
Absent in population databases
PM2
Prevalence in
affecteds
statistically increased over controls
PS4
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
POPULATION DATA (BA1, BS1, BS2, PM2, PS4
)
Slide10Proposed BA1: Allele frequency is >5% in any general continental population dataset of at least 2,000 alleles for a gene without a gene- or variant-specific recommendation
Slide11Proposed BA1: Allele frequency is >5% in any general continental population dataset of at least 2,000 alleles
for a gene without a gene- or variant-specific recommendation
Can compare to individual, continental populations (>2,000 alleles)
Tested individual does not need to match ethnic origin of population dataset used
Slide12Proposed BA1: Allele frequency is >5% in any general continental population dataset of at least 2,000 alleles for a gene without a gene- or variant-specific recommendation
Slide13Proposed BA1: Allele frequency is >5% in any general continental population dataset of at least 2,000 alleles for a gene without a gene- or variant-specific recommendation
Gene
Variant
Max MAF
GJB2
NM_004004.5: c.109G>A (V37I)
0.072
EAS-ExAC
HFE
NM_000410.3: c.845G>A (C282Y)
0.051
NFE-ExAC
BTD
NM_000060.4:
c.1330G>C (D444H)0.054 FIN-
ExAC
Clinical domain working groups can define BA1 thresholds <5% (based on prevalence and penetrance)
Exception list for non-benign alleles >5% in continental populations
Slide14Determine Max Credible Population AFPrevalence X
Heterogeneity
Penetrance
BA1
(stand alone criteria for Benign)
Use MOST conservative estimates
BS1
(Benign strong criteria)
Use less conservative estimates
Slide15Determine Max Credible Population AF
BA1 Max Credible AF: 0.00487 (~0.5%)
Assuming…
1/40,000 prevalence
95% of Pompe due to
GAA
All of
GAA
-Pompe caused by single variant
Penetrance 100%
Slide16Determine Max Credible Population AF
BS1 Max Credible AF: 0.00096 (~0.1%)
Assuming…
1/300,000 prevalence
95% of Pompe due to
GAA
One
GAA
variant account for 50%
Penetrance 85%
Slide17Great tool to help determine AF thresholds:https://cardiodb.org/allelefrequencyapp/
Slide18Filtering Allele Freq = highest AF that would be compatible with the observed allele count (95% CI Poisson)
Slide19PM2 – “Absent”Dominant versus
Recessive
However, dominant does
not
need to be 100% absent, depending on penetrance / age of onset
Hypertrophic cardiomyopathy: <0.00004 (~5 alleles max)
RASopathy: strictly absent
Slide20PM2 – “Absent”PM2 threshold could be
inverse of BS1
BA1: ≥0.1%
BS1: ≥0.05%
PM2: <0.05%
IF disorder is recessive or reduced penetrant dominant
Slide21MAF is too high for disorder
BA1/BS1
Observation in controls inconsistent with disease penetrance
BS2
Absent in population databases
PM2
Prevalence in
affecteds
statistically increased over controls
PS4
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
POPULATION DATA (BA1, BS1, BS2, PM2, PS4
)
Generally, should not be used based on occurrences in ExAC/
gnomAD
as phenotypes are unknown
If disorder IS NOT penetrant at early age, consider removing this criteria
Can also add occurrence thresholds
RASopathy:
variant identified in ≥3 well
phenotyped
individuals
MAF is too high for disorder
BA1/BS1
Observation in controls inconsistent with disease penetrance
BS2
Absent in population databases
PM2
Prevalence in
affecteds
statistically increased over controls
PS4
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
POPULATION DATA (BA1, BS1, BS2, PM2, PS4
)
Because the denominator for cases/probands is not always known (and thus cannot calculate an OR), guidelines state PS4 can also be used to count probands w/ consistent phenotypes
Slide23MAF is too high for disorder
BA1/BS1
Observation in controls inconsistent with disease penetrance
BS2
Variant identified in
#
probands with consistent phenotypes
PS4_Supporting
Absent in population databases
PM2
Variant identified in
##
probands with consistent phenotypes PS4_Moderate
Prevalence in
affecteds statistically increased over controls (OR method)
Variant identified in #
## probands with consistent phenotypes PS4
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
POPULATION DATA (BA1, BS1, BS2, PM2, PS4
)
PS4 could be used for typical case-control studies with an OR
OR
PS4 could be used for multiple unrelated probands with consistent phenotypes
Slide24Comparison of PS4 Specification
PS4_Supporting
PS4_Moderate
PS4 (Strong)
PM2 threshold
MYH7
≥2 probands
≥6 probands
≥15 probands
<0.004%
RASopathy
≥1 probands
≥3 probands
≥5 probands
Strictly absent
Slide25Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Novel missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
Slide26Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Novel missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
BP1 could also be used for:
LOF variants in a gene where the disease:
is caused by
GOF
variants
i
s caused by dominant/negative LOF variants
Slide27Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Novel missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
Slide28Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
Slide29Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
General modifications for PM5 and PS1:
Allow criteria to be used for analogous genes
[HRAS, NRAS, KRAS] ; [MAP2K1, MAP2K2] ; [HBA1, HBA2]
Assessing variant c.37G>A (p.Gly13Ser) in HRAS; variant c.38G>A (p.Gly13Asp) in KRAS is established Pathogenic
Important to define mapping between genes!
Does Gly13 in HRAS = Gly13 in KRAS?
Slide30Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
≥2 different missense changes at AA residue have been seen before
PM5_Strong
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
General modifications for PM5:
Allow PM5 to be upgraded to Strong (PM5_Strong) if ≥2 different pathogenic missense changes at the residue have been seen before
Slide31Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
≥2 different missense changes at AA residue have been seen before
PM5_Strong
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
General modifications for PS1:
Can also be used for splice site variants
Slide32PS1: Splice Site VariantsExample: You are assessing IVS3+5G>T
and IVS3+5G>
A
meets criteria for established pathogenic and has been showed to impact splicing in a functional assay. IVS3+5G>
T
has not been studied functionally but in
silico
splicing predictions show IVS3+5G>
T is equally or more damaging than IVS3+5G>A =
PS1 metIf in the same scenario (with IVS3+5G>A meeting criteria for established pathogenic and has been showed to impact splicing in a functional assay)
silico splicing predictions suggest IVS3+5G>T is not as damaging as
IVS3+5G>A = PS1 not metIf PS1 is met in these instances, PP3 (in silico prediction) would NOT be applied
Slide33Multiple lines of comp evidence suggest no impact
BP4
Missense when only truncating cause disease
BP1
Silent variant w/ non predicted splice impact
BP7
In-frame indels in repeat w/out known function
BP3
Multiple lines of computational evidence support a deleterious effect on the gene /gene product
PP3
Predicted
null variant in a gene with moderate level of evidence for LOF as a disease mechanism
PVS1_Moderate
Missense change at an AA residue where a different pathogenic missense change has been seen before
PM5
Protein length changing variant
PM4
≥2 different missense changes at AA residue have been seen before
PM5_Strong
Same AA change as an established path variant
PS1
Predicted null variant in a gene where LOF is a known mechanism of disease
PVS1
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
COMPUTATIONAL AND PREDICTIVE DATA
General modifications for PVS1:
Can be downgraded to Moderate if:
LOF is not an established disease mechanism
Only moderate level of evidence for gene/disease pair, but all the evidence supports LOF as the disease mechanism
Slide34De novo
(without paternity & maternity confirmed)
PM6
De novo
(paternity & maternity confirmed)
PS2
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
DE NOVO DATA
Slide35DE NOVO DATA
Example:
Patient had a de novo missense variant in MECP2 but only presented with ID – no Rett phenotypes
Slide36De novo
(without paternity & maternity confirmed)
PM6
De novo
(paternity & maternity confirmed)
PS2
≥2 De novo occurrences
(without paternity & maternity confirmed)
PM6_Strong
≥2 De novo
occurrences (paternity & maternity confirmed)
PS2
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
DE NOVO DATA
For PS2 and PM6, can increase strength with increasing independent de novo occurrences
Slide37Observed in
trans
with a dominant variant
or
Observed in
cis
with a pathogenic variant
BP2
For recessive
disorders, detected
in trans with a
pathogenic variant
PM3
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
ALLELIC DATA
For PM3, shift weight according to variant strength and number of occurrences
Slide38Observed in
trans
with a dominant variant
or
Observed in
cis
with a pathogenic variant
BP2
For recessive
disorder,
b
iallelic variants but variant on the other allele does
not meet pathogenic criteriaPM3_Supporting
For recessive
disorders, detectedin trans with a
pathogenic variantPM3
For recessive
disorders, ≥2 occurrences when detectedin trans with a
pathogenic variantPM3_Strong
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
ALLELIC DATA
Supporting:
Variant on the other allele “suspicious” and absent from control
dbs
OR
Proband homozygous for the variant and variant absent from control
dbs
Strong:
≥2 unrelated probands compound het with different pathogenic
variants in trans
Proband 1:
p.H308fs
;p.Y3955X
Proband 2:
p.H308fs
;p.N405fs
Proband 3:
p.H308fs
;p.N405fs
Slide39Reputable source = benign
BP6
Reputable source = pathogenic
PP5
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
OTHER DATABASES
Slide40Reputable source = benign
BP6
Reputable source = pathogenic
PP5
Strong
Strong
Very Strong
Moderate
Supporting
Supporting
Benign
Pathogenic
OTHER DATABASES
CRITERIA SHOULD NOT BE USED
Slide41Move to Bayes ApproachACMG/AMP criteria are compatible with this quantitative approach
Slide42Allow us to make better definitions for VUS and when Pathogenic and Benign data conflict(PVS# + PM# + PM#) + BP#
=
Pathogenic
(PS# + PS#) + BS#
=
Uncertain significance
Move to Bayes Approach