genetic variants and genetic testing in cpvt - sads 2015... · genetic variants and genetic testing...

Genetic Variants and Genetic Testing in CPVT

2015 SADS Foundation International Meeting New York City, NY

May 29, 2015

Michael J. Ackerman, MD, PhD, FACC Windland Smith Rice Cardiovascular Genomics Research Professor Professor of Medicine, Pediatrics, and Pharmacology Director, Long QT Syndrome Clinic and the Mayo Clinic Windland Smith

Rice Sudden Death Genomics Laboratory President, Sudden Arrhythmia Death Syndromes (SADS) Foundation

WINDLAND Smith Rice Sudden Death

Genomics Laboratory

Learning Objectives to Disclose: • To ASSESS clinical genetic testing for CPVT and EVALUATE its role and yield • To EXPOSE the “dark side” (aka, Genetic Purgatory) of the CPVT genetic test

Conflicts of Interest to Disclose: • Consultant – Boston Scientific, Gilead Sciences, Medtronic, St. Jude Medical, and Transgenomic/FAMILION • Royalties – Transgenomic/FAMILION

www.StopSADS.org 1-800-Stop SAD

Cardiac Channelopathies LQTS - RWS - JLNS

CPVT

FAVCB

IVF IER

BrS

ATS

SSS SIDS FAF

DI-TdP

SQTS

SUDS TS

DCM

August 2011 HRS/EHRA Consensus Statement on the

State of Genetic Testing for the Channelopathies and Cardiomyopathies

Ackerman, Priori, et al. Heart Rhythm 8:1308-1339, 2011

Decoding a CPVT Genetic Test Result

- What is the test’s DIAGNOSTIC, PROGNOSTIC, and THERAPEUTIC impact?

- What is the test’s YIELD?

- What is the test’s Achilles’ heel (its “NOISE”)?

- What is my index of suspicion?

Clinical

Catecholaminergic Polymorphic Ventricular Tachycardia (CPVT)

Exertion Induced Syncope or Sudden Cardiac Death No Structural Heart Defect

Phenotypically Mimics Long QT Syndrome

Bi-Directional Ventricular Tachycardia

Hallmark Arrhythmia

Priori et al. J Clin Invest 115:2033-2038, 2005

Clinical


Exertion Induced Syncope or Sudden Cardiac Death No Structural Heart Defect

Phenotypically Mimics Long QT Syndrome

Bi-Directional Ventricular Tachycardia

Hallmark Arrhythmia

Priori et al. J Clin Invest 115:2033-2038, 2005

Exercise-induced PVCs in bigeminy initiating at heart rates > 120 beats per minute – suspicious for CPVT! Horner … Ackerman. Heart Rhythm 2008


HCM

LQTS CPVT BrS

Diagnostic Prognostic Therapeutic Disease

ARVC DCM

CCD SQTS AF

LVNC RCM

+++ + - +++ +++ ++

+ - -

+ + - +/-

+ + - - -

+++ ++ + + +/- -

+/- - - + - - + + +


“CPVT genetic testing is recommended for any patient in whom a cardiologist has established a clinical index of suspicion for CPVT…[and] is recommended for family members and appropriate relatives”

“…genotype influences the management and treatment of a patient with genetically confirmed CPVT“

- “Positive” Test Result



Mutation-specific genetic testing is recommended for family members and other appropriate relatives subsequently following the identification of the disease-causative mutation in an index case.

- “Negative” Test Result - What was my index of suspicion?

- What exons were included/excluded? - Was it a clinical or research test result?


Calcium Induced Calcium Release (CICR) 105 RyR2 (CPVT1, Ch 1q42.1-q43)

1 14 47 49 83 88 90 93 97 100 8 15 44 45 46 102 103 101

CASQ2 (CPVT2, Ch 1p13.1) 1 2 3 4 5 6 7 8 9 10 11

Molecular

105 Exons

4967 amino acids

11 Exons

339 amino acids

Cell membrane

Sarcoplasmic reticulum

RyR2 –

CASQ2


Cellular

RyR2 Targeted Screening

1000 2000 3000 4000 4967 0

Mutations seem to be distributed across three canonical structure-function domains

Yano et. al, Nat Clin Prac Card 3:43-52,2006 George et al. J Mol Cardiol; 42:34-50, 2007

I II III

77-466 2246-2534 3778-4959

The Cardiac Ryanodine Receptor (RyR2)

Regions of Higher

Pathogenicity

Exons 3, 8, 10, 12-15, 17, 19, 21, 26-28 Exons 38, 40-50 Exon 75

Exons 83, 86-91, 93-97, 99-105

RYR2 “Hot Spot

Domains”

Central Domain Channel Region N-terminal Domain

2246-2534 3778-4959 77-466

3564-3575

Targeted Genetic Test

Region

Exons 1 – 28 Exons 37 – 50 Exon 75 Exons 83 – 105

1-1141 1638-2578 3776-4967

Targeted Genetic Test 66/105 exons tested, covering 66% of the protein

RyR2 1 4967

Exon 1 Exon 105

Yano et. al, Nat Clin Prac Card 3:43-52,2006 George et al. J Mol Cardiol; 42:34-50, 2007

Medeiros-Domingo et al. JACC 54:2065-74, 2009


1 14 47 49 83 88 90 93 97 100 8 15 44 45 46 102 103 101

CASQ2 (CPVT2, Ch 1p13.1) 1 2 3 4 5 6 7 8 9 10 11

Molecular

105 Exons

4967 amino acids

11 Exons

339 amino acids

Cell membrane


RyR2 –

CASQ2


Cellular

Medeiros-Domingo…Ackerman. JACC 54:2064-2075, 2009 Medeiros-Domingo…Ackerman. JACC 54:2064-2075, 2009

Yield of Genetic Testing


1 14 47 49 83 88 90 93 97 100 8 15 44 45 46 102 103 101

CASQ2 (CPVT2, Ch 1p13.1) 1 2 3 4 5 6 7 8 9 10 11

Molecular

105 Exons

4967 amino acids

11 Exons

339 amino acids

Cell membrane


RyR2 –

CASQ2


Cellular

Medeiros-Domingo…Ackerman. JACC 54:2064-2075, 2009 Medeiros-Domingo…Ackerman. JACC 54:2064-2075, 2009

Yield of Genetic Testing

CPVT Strong – 60% CPVT Possible – 30% Real World (N = 1200) – 18% Autopsy Negative SUDS – 10%

Total Cases 1200 Avg Age (years) 31 ± 19 % Female 46% Caucasian 78% African 9% Hispanic 7% Asian 3% Other 3%

Test Version

Exons on Panel Exons Sample Count

1 38 3, 8-15, 37, 41, 44-50, 83, 87-105 291 2 66 1-28, 37-50, 75, and 83-105 656 3 105 1-105 253

CPVT Genetic Testing in Real World

Kapplinger … Ackerman. 2015 (submitted)

Is the “X” that marks the spot truly THE disease-causing mutation?

Genetic Testing’s Achilles’ Heel

Non-Synonymous Single Nucleotide Polymorphism • Common/rare SNP w/o functional/clinical significance • Common/rare SNP w/ functional/clinical significance • Pathogenic (“disease-causing”) mutation

“With great power comes great responsibility” Spider Man’s Uncle

Background Noise Issue 2003

RESULTS

2%

1.4%

0.6%

Yield of CPVT Genetic Test in Controls

0.0%

0.5%

1.0%

1.5%

2.0%

All Targeted Region

Outside Targeted Region

Yiel

d N1

C4967

Cytosol

SR lumen

1

11

RyR2 (n = 221) Control Variants > 8000 “controls”

FKBP12.6 Binding Domain

754

40 250

328 507

1136

3152 3510

377

1013 1810

2094 2156 2435

2284

2812

3973 4281 4573

4653 4344


• 3% background rate of rare variants identified in 200 ostensibly healthy controls

• 15/142 (10.5%) of the previously published CPVT variants were found in controls

(Medeiros-Domingo…Ackerman JACC 2009)

• Reconfirmed – 11% (Jabbari…Olesen Circ. Cardiovasc. Genet. 2013)

Background Rate Issue in CPVT

Reported Variants Represented in Controls*

Mutation Reference (PMID) Count (Out of 8075)

R1013Q PMID: 21964171 13 (0.16%)

R4307C PMID: 19926015 10 (0.12%)

V2113M PMID: 19926015 7 (0.09%)

T1107M PMID: 19926015 5 (0.06%)

V919M Kellen et al., HRS 2012 3 (0.04%)

V377M PMID: 19926015 3 (0.04%)

P466A PMID: 19926015 2 (0.02%)

V4010M PMID: 19926015 2 (0.02%)

A4556T PMID: 19926015 2 (0.02%)

*Variants found only once in controls were omitted

- “Maybe” Test Result ?

- What was my index of suspicion? - Have I done my homework?

“Possible Deleterious” “Variant of Uncertain Significance (VUS)”

“Genetic Purgatory”


“Genetic Purgatory is a Real Place and its

Scary!”

MAF < 1:10000 MAF > 1:10000 Case

68 non-synonymous variants with a MAF > 1/10,000 alleles

1053 non-synonymous variants with a MAF < 1/10,000 alleles

Background rate of 3.2%


59.0%

31.2%

18.2%

3.2%

0%

10%

20%

30%

40%

50%

60%

70%

Strong (n = 78) Possible (n = 77) Referral (n = 1,200) ExAC (n = 60,706)

Yiel

d of

Rar

e Va

riant

s (M

AF <

1/1

0,00

0)

Molecular Autopsy

18:1

10:1

5.5:1

Strong = exertional syncope plus documentation of bidirectional or

polymorphic ventricular tachycardia

10.0%

3.2% ExAC Rate

Possible = exertional syncope plus either stress test induced ventricular ectopy

or corrected QT interval values <480 ms

3:1


25.0%

18.0%

11.0% 9.7%

0%

5%

10%

15%

20%

25%

30%

1st 300 2nd 300 3rd 300 4th 300

Yiel

d

27.3 28.6 31.2 33.5

05

10152025303540

1st 300 2nd 300 3rd 300 4th 300

Aver

age

Age

1.1x10-5

0.004

8.7x10-7

0.002 0.003

3.2% ExAC Rate

3:1


28.1%

18.4%

4.7%

0%

5%

10%

15%

20%

25%

30%

<18 (n= 335)

18-35 (n= 244)

>35 (n= 387)

Yiel

d of

Rar

e Va

riant

s

Age at Genetic Testing

9:1

6:1

3.2% ExAC Rate

<18 (n = 335)

18-35 (n = 244)

>35 (n = 387)

1.5:1

161

24

7 2

7

0

20

40

60

80

100

120

140

160

180

1 2 3 4 ≥ 5

# of

Uni

que

Varia

nts

# of Unrelated Cases

36/201 case-derived variants were overrepresented (p<4.8x10-4) in disease and completely absent from the ExAC

exomes

N1

C4967

Cytosol

SR lumen

FKBP12.6 Binding Domain

RyR2 N57_G91del

R169Q E243K F329L

R332W

G357S R420Q

E1724K H2168Q

C2201Y

S2246L

E2296Q

A2387T

R2401H

R4959Q

F4851L (2x)

I4848V

V4771I

N4763S

G4749V G4749E R4497C

L4188P

N4178S

R4157Q

S4124T

A2403T

A2416D

R2420W

A4091T A4091V

E4076K

M3972I

G3946S

C3800F

59.0%

31.2%

18.2%

3.2%

0%

10%

20%

30%

40%

50%

60%

70%

Strong (n = 78) Possible (n = 77) Referrals (n = 1200) ExAC

Yiel

d of

Rar

e Va

riant

s (M

AF <

1/1

0,00

0)

10:1

5.5:1

3.2%

11.2%

16.9%

35.9%

5.3:1

11:1

18:1

3.5:1


0.0%

1.0%

2.0%

3.0%

4.0%

5.0%

1 4 7 10 13 16 19 22 25 28 31 34 37 40 43 46 49 52 55 58 61 64 67 70 73 76 79 82 85 88 91 94 97 100

103

Yiel

d of

Rar

e Va

riant

s

Exon

ReferralsPhenotyped CasesExAC ExomesIdentified ExonsOREAE

35/105 exons carried an overrepresentation of “new” rare variants (exons 1-3, 6, 8, 14, 17, 27, 40, 43-45, 47-50, 54, 80-81, 83, 86, 88-90, 93, 95-101,

103-105)

These 35 exons only account for ~1/3 of the protein (1664/4967 amino acids)

33.3%

15.6%

8.8%

0.8% 2.6% 1.3% 2.3% 2.3%

0%

5%

10%

15%

20%

25%

30%

35%

Strong (n = 78) Possible (n = 77) Referrals (n = 1200) ExAC

Yiel

d of

Rar

e Va

riant

s (M

AF <

1/1

0,00

0)

Identified Exons

Outside Identified Exons

20:1

42:1

11:1

Outside Identified Exons 1:1 Signal-to-noise

Regardless of Phenotype


Take Home Points 1. The yield of CPVT genetic testing is 60%, BUT

2. The signal-to-noise ratio can be up to 43:1, BUT

3. “X” does NOT always mark the spot!

4. Genetic purgatory is a real place and it’s scary! 5. Time to go retro with targeted RYR2 testing (35 exons)?


Dr. Scholl Foundation, CJ Foundation for SIDS Hannah Wernke Memorial Foundation

Sheikh Zayed Saif Mohammed Al Nahyan Fund National Institutes of Health


Genomics Laboratory


Genomics Laboratory

“To heal the sick and advance the science” Dr. Charles W. Mayo

If you feel you have benefitted from this presentation,

please make a donation now.

1-800-STOP SAD(801) 531-0937

www.StopSADS.org

genetic variants and genetic testing in cpvt - sads 2015... · genetic variants and genetic testing...

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