SUPPLEMENTAL METHODS

Genotyping Methods and Quality Control

DNA was isolated and purified from whole blood using the Qiagen QIAamp DNA purification kit (Quiagen, Germantown, MD). Genotyping of CYP polymorphisms was performed by pyrosequencing or TaqMan assay. Pyrosequencing was performed as previously described.1, 2 PCR was carried out using Amplitaq Gold PCR master mix (ABI, Foster City, CA), 1 pmole of each primer (IDT, Coralville, IA), and 1ng DNA. Pyrosequencing primers and conditions/TaqMan genotyping assays (available upon request) were performed according to manufacturer's directions (Applied Biosystems, Carlsbad, California). The DNA segments containing the region of interest were amplified with PCR using TaqMan genotyping master mix and 5-10ng DNA. Allelic discrimination was performed using sequence detection software (Applied Biosystems, Carlsbad, California). PCR and allelic discrimination were performed using the ABI 7500real time PCR platform. All genotype data were transferred to a Microsoft Access database for permanent storage and merging with the clinical datasets through SAS v9.1.

For all variants, genotype call rates were greater than 93%. After pyrosequencing, the CYP2C19*2 (rs4244285) variant was found to be out of Hardy Weinberg equilibrium (HWE) in African-Americans (p=0.03). All samples were genotyped again using TaqMan genotyping assay. All but 3 samples matched genotypes. These 3 results were removed but the variant remained out of HWE in African-Americans.

References

1.Marsh S, King CR, Garsa AA, McLeod HL. Pyrosequencing of clinically relevant polymorphisms. Methods Mol Biol 2005; 311: 97-114.

2.Cresci S, Dorn GW, 2nd, Jones PG, Beitelshees AL, Li AY, Lenzini PA, et al. Adrenergic-pathway gene variants influence beta-blocker-related outcomes after acute coronary syndrome in a race-specific manner. J AmColl Cardiol2012; 60: 898-907.

Supplemental Table S1A: Baseline Clinical Characteristics of CYP2C19*1 Homozygotes with and without the use of a Proton Pump Inhibitor

Caucasians
PPI
(n = 122 patients) / No PPI
(n = 537 patients) / p-value
Age / 60.63 (12) / 58.53 (11.9) / 0.08
Male / 80 (65.6) / 412 (76.7) / 0.01
Hypertension / 84 (68.9) / 287 (53.4) / 0.002
Dyslipidemia / 76 (62.3) / 258 (48) / 0.004
Diabetes Mellitus / 41 (33.6) / 105 (19.6) / 0.0007
Smoking History / 91 (74.6) / 393 (73.6) / 0.82
Congestive Heart Failure / 9 (7.4) / 30 (5.6) / 0.45
Chronic Kidney Disease / 10 (8.2) / 21 (3.9) / 0.044
Atrial fibrillation/flutter / 8 (6.6) / 21 (3.9) / 0.20
Peripheral Vascular Disease / 8 (6.6) / 18 (3.4) / 0.10
Prior Myocardial Infarction / 26 (21.3) / 80 (14.9) / 0.08
Prior PCI / 35 (28.7) / 105 (19.6) / 0.03
Prior CABG / 27 (22.1) / 44 (8.2) / 0.000007
Prior Stroke / 7 (5.7) / 19 (3.5) / 0.26
Prior TIA / 7 (5.7) / 12 (2.2) / 0.037
GRACE Risk Score / 99.36 (29.7) / 94.97 (27.8) / 0.12
Clinical Presentation of AMI
STEMI / 54 (44.3) / 301 (56.1) / 0.02
NSTEMI / 66 (54.1) / 234 (43.6) / 0.04
Index hospitalization Revascularization
PCI / 102 (83.6) / 464 (86.4) / 0.42
CABG / 2 (1.6) / 18 (3.4) / 0.56
Discharge Medications
Aspirin / 119 (97.5) / 520 (96.8) / 0.68
Clopidogrel / 122 (100) / 537 (100) / 1.00
Beta-blocker / 112 (91.8) / 497 (92.6) / 0.78
ACEI/ARB / 98 (80.3) / 418 (77.8) / 0.55
Lipid lowering medication / 116 (95.1) / 502 (93.5) / 0.51

Values are shown as absolute numbers (percentages) or mean ± SD.

ACE = angiotensin-converting enzyme; AMI = acute myocardial infarction; ARB = angiotensin receptor blocker; CABG = coronary artery bypass graft; NSTEMI = non-ST-elevation myocardial infarction; PCI = percutaneous coronary intervention; STEMI = ST-elevation myocardial infarction; TIA = Transient ischemic attack.

Supplemental Table S1B: Baseline Clinical Characteristics of CYP2C19*2 Carriers with and without the use of a Proton Pump Inhibitor

Caucasians
PPI
(n = 80 patients) / No PPI
(n = 366 patients) / p-value
Age / 60.6 (11.3) / 59.44 (12.1) / 0.43
Male / 48 (60) / 265 (72.4) / 0.028
Hypertension / 49 (61.3) / 226 (61.7) / 0.93
Dyslipidemia / 50 (62.5) / 180 (49.2) / 0.03
Diabetes Mellitus / 32 (40) / 103 (28.1) / 0.04
Smoking History / 59 (73.8) / 266 (73.5) / 0.96
Congestive Heart Failure / 4 (5) / 17 (4.6) / 0.78
Chronic Kidney Disease / 1 (1.3) / 14 (3.8) / 0.49
Atrial fibrillation/flutter / 5 (6.3) / 18 (4.9) / 0.58
Peripheral Vascular Disease / 5 (6.3) / 13 (3.6) / 0.34
Prior Myocardial Infarction / 11 (13.8) / 71 (19.4) / 0.24
Prior PCI / 17 (21.3) / 82 (22.4) / 0.82
Prior CABG / 12 (15) / 44 (12) / 0.47
Prior Stroke / 2 (2.5) / 13 (3.6) / 1.00
Prior TIA / 1 (1.3) / 7 (1.9) / 1.00
GRACE Risk Score / 98.56 (25.8) / 96.67 (29.3) / 0.59
Clinical Presentation of AMI
STEMI / 41 (51.3) / 207 (56.6) / 0.39
NSTEMI / 39 (48.8) / 158 (43.2) / 0.36
Index hospitalization Revascularization
PCI / 73 (91.3) / 324 (88.5) / 0.48
CABG / 2 (2.5) / 8 (2.2) / 0.70
Discharge Medications
Aspirin / 78 (97.5) / 356 (97.3) / 0.91
Clopidogrel / 80 (100) / 366 (100) / 1.00
Beta-blocker / 78 (97.5) / 344 (94) / 0.21
ACEI/ARB / 57 (71.3) / 290 (79.2) / 0.12
Lipid lowering medication / 70 (87.5) / 339 (92.6) / 0.13

Values are shown as absolute numbers (percentages) or mean ± SD.

All abbreviations as shown in Supplemental Table S1A.

Supplemental Table S1C: Baseline Clinical Characteristics of CYP2C19*17 Carriers with and without the use of a Proton Pump Inhibitor

Caucasians
PPI
(n = 121 patients) / No PPI
(n = 453 patients) / p-value
Age / 59.4 ±11.3 / 58.3 ±12.1 / 0.37
Male / 84 (69.4) / 349 (77.0) / 0.08
Hypertension / 79 (65.3) / 259 (57.2) / 0.11
Dyslipidemia / 68 (56.2) / 217 (47.9) / 0.10
Diabetes Mellitus / 28 (23.1) / 99 (21.9) / 0.76
Smoking History / 85 (70.8) / 335 (74.4) / 0.42
Congestive Heart Failure / 7 (5.8) / 11 (2.4) / 0.06
Chronic Kidney Disease / 6 (5.0) / 14 (3.1) / 0.32
Atrial fibrillation/flutter / 10 (8.3) / 14 (3.1) / 0.01
Peripheral Vascular Disease / 1 (0.8) / 12 (2.6) / 0.32
Prior Myocardial Infarction / 25 (20.7) / 66 (14.6) / 0.10
Prior PCI / 29 (24.0) / 86 (19.9) / 0.22
Prior CABG / 19 (15.7) / 41 (9.1) / 0.03
Prior Stroke / 3 (2.5) / 12 (2.6) / 1.00
Prior TIA / 3 (2.5) / 8 (1.8) / 0.71
GRACE Risk Score / 97.8 ±28.1 / 92.7 ±27.5 / 0.07
Clinical Presentation of AMI
STEMI / 65 (53.7) / 259 (57.2) / 0.50
NSTEMI / 54 (44.6) / 193 (42.6) / 0.69
Index hospitalization Revascularization
PCI / 103 (85.1) / 405 (89.4) / 0.19
CABG / 1 (0.8) / 13 (2.9) / 0.32
Discharge Medications
Aspirin / 116 (95.9) / 438 (96.7) / 0.66
Clopidogrel / 121 (100.0) / 453 (100.0) / 1.00
Beta-blocker / 112 (92.6) / 416 (91.8) / 0.79
ACEI/ARB / 100 (82.6) / 369 (81.5) / 0.76
Lipid lowering medication / 110 (90.9) / 417 (92.1) / 0.68

Values are shown as absolute numbers (percentages) or mean ± SD.

All abbreviations as shown in Supplemental Table S1A.

Supplemental Table S2: Type of proton pump inhibitor used during the study period within each CYP2C19 genotype group

CYP2C19 *1 homozygotes / CYP2C19*2 carriers / CYP2C19*17 carriers / p-value*
Caucasians
Omeprazole / 60% / 63% / 66% / 0.22
Pantoprazole / 28% / 27% / 27% / 0.71
Esomeprazole / 20% / 18% / 24% / 0.17
Lansoprazole / 15% / 17% / 9% / 0.23
Rabeprazole / 5% / 4% / 3% / 0.86
African Americans
Omeprazole / 67% / 56% / 55% / 0.27
Pantoprazole / 25% / 39% / 45% / 0.13
Esomeprazole / 24% / 22% / 28% / 0.48
Lansoprazole / 25% / 6% / 10% / 0.40
Rabeprazole / 0% / 0% / 0% / na

Values are shown as percentages.

*p-values were calculated after excluding individuals who were both *2 and *17 carriers. na = not applicable.

Supplemental Table S3: Causes for Non-elective Cardiac Rehospitalization in Clopidogrel-treated Caucasian Patients with and without PPI use

All Caucasians / CYP2C19*17 Carriers
PPI / No PPI / p-value / PPI / No PPI / p-value
STEMI / 0.4% / 0.3% / 0.58 / 0.0% / 0.0% / na
NSTEMI / 3.2% / 1.4% / 0.04 / 2.0% / 0.3% / 0.12
Unstable Angina – with ischemia / 0.4% / 0.2% / 0.48 / 0.0% / 0.3% / 1.0
Unstable Angina – without ischemia / 9.2% / 4.3% / 0.002 / 10.2% / 2.3% / 0.0004
Stable Angina / 1.2% / 1.0% / 0.73 / 1.0% / 0.8% / 1.0
Restenosis / 1.2% / 0.3% / 0.09 / 2.0% / 0.6% / 0.21
Demand Ischemia / 0.0% / 0.1% / 1.00 / 0.0% / 0.0% / na
Stroke / 0.0% / 0.1% / 1.00 / 0.0% / 0.0% / na
Sudden Cardiac Death / 0.0% / 0.1% / 1.00 / 0.0% / 0.0% / na
Arrhythmia / 1.2% / 0.7% / 0.42 / 3.1% / 0.3% / 0.03
Heart Failure / 4.0% / 2.2% / 0.11 / 5.1% / 2.0% / 0.15
Other Cardiac Cause / 1.6% / 0.4% / 0.05 / 3.1% / 0.3% / 0.03
CYP2C19*2 Carriers / CYP2C19 Wildtypess
PPI / No PPI / p-value / PPI / No PPI / p-value
Non-Elective Rehospitalization:
STEMI / 0 / 0.4 / 1.00 / 1.0 / 0.5 / 0.48
NSTEMI / 4.6 / 2.5 / 0.41 / 3.1 / 1.5 / 0.39
Unstable Angina – with ischemia / 0 / 0.4 / 1.00 / 1.0 / 0.0 / .19
Unstable Angina – without ischemia / 4.6 / 6.5 / 0.78 / 10.3 / 3.9 / 0.01
Stable Angina / 0 / 0.4 / 1.00 / 2.1 / 1.2 / .63
Restenosis / 1.5 / 0 / 0.19 / 0 / 0.2 / 1.00
Demand Ischemia / 0 / 0 / - / 0 / 0.2 / 1.00
Stroke / 0 / 0 / - / 0 / 0.2 / 1.00
Sudden Cardiac Death / 0 / 0 / - / 0 / 0 / -
Arrhythmia / 1.5 / 1.1 / 0.57 / 0 / 0.5 / 1.00
Heart Failure / 3.1 / 2.9 / 1.00 / 4.1 / 2.5 / .32
Other Cardiac Cause / 1.5 / 0.7 / .47 / 0 / 0.2 / 1.00

Values are shown as percentages.

NSTEMI = non-ST-elevation myocardial infarction; STEMI = ST-elevation myocardial infarction.

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