From Mar 2011 to September 2016, 2213 patients were consecutively screened, among whom 1614 patients who met the inclusion and the exclusion criteria were enrolled. Of the 1614 enrolled patients, 3 were not included in the final analysis due to unsatisfactory blood sample quality. All the remaining patients completed the genotype assessment and 1-year clinical follow-up. Platelet aggregation testing were performed in 1175 patients at baseline and in 624 patients at 1-month follow-up (Figure 1).
Patients’ Characteristics
The baseline characteristics of patients included in this study are summarized in Table 1. Compared with patients who did not experience MACE, those who experienced MACE were older [69.00 (14.50) vs. 64.00 (15.00), P=0.0069], more commonly had reduced LVEF (25.0% vs. 7.66%, P<0.001) and history of ST-segment-elevation myocardial infarction (STEMI) or non-ST-segment elevation acute coronary syndromes (NSTE-ACS) (63.63% vs. 42.44%, P=0.0010).
Table 1
Baseline Characteristics of Patients Grouped by the Occurrence of MACE
Variables | MACE (n=44) | MACE free (n=1, 567) | P value |
Age, median (IQR), years | 69.00 (14.50) | 64.00 (15.00) | 0.0069 |
Sex, No. (%) | | | 0.2966 |
Female | 8(18.18) | 393(25.08) | |
Male | 36(81.82) | 1174(74.92) | |
Previous MI, No. (%) | | | 0.7166 |
No | 42(95.45) | 1499(95.66) | |
Yes | 2(4.55) | 68(4.34) | |
Hypertension, No. (%) | | | 0.4109 |
No | 12(27.27) | 520(33.18) | |
Yes | 32(72.73) | 1047(66.82) | |
Diabetes Mellitus, No. (%) | | | 0.3568 |
No | 30(68.18) | 1165(74.35) | |
Yes | 14(31.82) | 402(25.65) | |
Smoking, No. (%) | | | 0.3503 |
No | 24(54.55) | 743(47.42) | |
Yes | 20(45.45) | 824(52.58) | |
Previous PCI, No. (%) | | | 0.4246 |
No | 42(95.45) | 1424(90.87) | |
Yes | 2(4.55) | 143(9.13) | |
LVEF, No. (%) | | | <0.001 |
≥ 55% | 33(75.00) | 1447(92.34) | |
< 55% | 11(25.00) | 120(7.66) | |
Serum creatinine, No. (%) | | | 0.2168 |
≤ 133µmol/L | 42(95.45) | 1537(98.09) | |
> 133µmol/L | 2(4.55) | 30(1.91) | |
Low density lipoprotein, No. (%) | | | 0.5350 |
≥ 1.8mmol/L | 36(81.82) | 1335(85.19) | |
< 1.8mmol/L | 8(18.18) | 232(14.81) | |
Diagnosis, No. (%) | | | 0.0010 |
SA | 16(36.36) | 902(57.56) | |
NSTE-ACS | 12(27.27) | 412(26.29) | |
STEMI | 16(36.36) | 253(16.15) | |
Values are presented as median (IQR) or number of patients (percentage) as appropriate. P values were calculated with the use of t test or \({\chi }^{2}\) test as appropriate. Abbreviations: LVEF. left ventricular ejection fraction; MACE. major adverse cardiovascular events, including cardiovascular death, myocardial infarction and ischemic stroke; MI. myocardial infarction; NSTE-ACS. non-ST-segment elevation acute coronary syndromes; PCI. percutaneous coronary intervention; SA. stable angina pectoris; STEMI. ST-segment-elevation myocardial infarction. |
On-Treatment Platelet Reactivity and Genotypes.
The baseline and 1-month PLADP were (29.88±14.34)% and (26.27±15.10)%, respectively. There was no significant difference in PLADP among different rs956115 genotypes at the baseline assessment (F=0.20, P=0.8200, Figure 2A). At 1-month follow-up, PLADP was significantly different among the three genotypes (F=3.28, P=0.0381, Figure 2A). CG genotype was associated with a significantly lower PLADP compared with CC genotype (P=0.0158, Figure 2A). Regarding PLAA, there were no significant difference among the three genotypes of rs956115 either at baseline (F=2.73, P=0.0656, Figure S1A) or at 1-month follow-up (F=0.20, P=0.8180, Figure S1A).
For rs4244285, PLADP were significantly different among the three genotypes at baseline (F=53.27, P<0.001, Figure 2B) and 1-month follow-up (F=12.07, P<0.001, Figure 2B). By pairwise comparisons, the platelet reactivities corresponding to different genotypes of rs4244285 were all significantly different except the comparison between GA and AA at 1-month follow-up (P=0.4392, Figure 2B). As shown in Figure 2B, the platelet reactivity increased with the number of the A alleles of rs4244285. Regarding PLAA, there were no significant difference among the three genotypes of rs4244285 either at baseline (F=0.38, P=0.6870, Figure S1B) or at 1-month follow-up (F=0.78, P=0.4590, Figure S1B).
Association between IRS/CYP2C19 Genotypes and Cardiovascular Outcomes.
A total of 44 patients experienced MACE, including 15 cardiac deaths, 16 nonfatal myocardial infarctions, and 13 ischemic strokes.
For rs956115, patients with CG or GG genotypes had a 1.99-fold higher MACE risk than those with CC homozygote (dominant model, adjusted HR=1.99, 95%CI: 1.00-3.98, P=0.0499; additive model, adjusted HR=1.95, 95%CI: 1.05-3.61, P=0.0341; Table 2). When further adjusted for rs4244285 genotypes, patients with CG or GG genotypes had a 2.09-fold higher MACE risk than those with CC homozygote (dominant model, adjusted HR=2.09, 95%CI: 1.04-4.19, P=0.0376; additive model, adjusted HR=2.04, 95%CI: 1.10-4.19, P=0.0244; Table 2 and Figure 3A). There was no significant difference in risk of MACE risk between CG and CC genotypes (adjusted HR=1.91, 95%CI: 0.94-3.88, P=0.0751) or between GG and CC genotypes (adjusted HR=4.23, 95%CI: 0.55-32.29, P= 0.1643) (Table 2).
Table 2
MACE risk by Multi-Cox regression
SNP | Gene | Geno-type | MACE N | Censored N | Comparison | Unadjusted model | Adjusted model a | Adjusted model b |
HR (95%CI) | P value | HR (95%CI) | P value | HR (95%CI) | P value |
rs956115 | IRS1 | CC | 32 | 1245 | | 1.00 (Reference) | 1.00 (Reference) | 1.00 (Reference) |
| | CG | 11 | 305 | CG vs.CC | 1.66(0.82,3.34) | 0.1571 | 1.91(0.94,3.88) | 0.0751 | 1.99(0.98,4.08) | 0.0586 |
| | GG | 1 | 17 | GG vs.CC | 2.65(0.36,19.53) | 0.3377 | 4.23(0.55,32.29) | 0.1643 | 4.70(0.62,35.84) | 0.1351 |
| | | | | Dominant | 1.71(0.87,3.38) | 0.1211 | 1.99(1.00,3.98) | 0.0499 | 2.09(1.04,4.19) | 0.0376 |
| | | | | Recessive | 2.35(0.32,17.11) | 0.3992 | 3.58(0.48,26.96) | 0.2157 | 3.91(0.52,29.33) | 0.1851 |
| | | | | Additive | 1.65(0.91,3.00) | 0.1013 | 1.95(1.05,3.61) | 0.0341 | 2.04(1.10,3.81) | 0.0244 |
rs4244285 | CYP2C19 | GG | 14 | 712 | | 1.00 (Reference) | 1.00 (Reference) | 1.00 (Reference) |
| | GA | 28 | 666 | GA vs.GG | 2.04(1.07,3.90) | 0.0303 | 2.13(1.10,4.12) | 0.0248 | 2.19(1.13,4.24) | 0.0200 |
| | AA | 2 | 189 | AA vs.GG | 0.60(0.14,2.65) | 0.5010 | 0.58(0.13,2.61) | 0.4814 | 0.58(0.13,2.60) | 0.4787 |
| | | | | Dominant | 1.76(0.93,3.33) | 0.0843 | 1.81(0.94,3.49) | 0.0759 | 1.85(0.96,3.56) | 0.0666 |
| | | | | Recessive | 0.40(0.10,1.65) | 0.2049 | 0.37(0.09,1.56) | 0.1767 | 0.37(0.09,1.53) | 0.1702 |
| | | | | Additive | 1.17(0.75,1.82) | 0.4853 | 1.16(0.74,1.81) | 0.5114 | 1.17(0.75,1.81) | 0.4936 |
a Model adjusted for clinical covariates, including age, previous MI, hypertension, diabetes mellitus, LVEF, serum creatinine, diagnosis, low density lipoprotein, smoking status, previous PCI. b Model adjusted for rs4244285/ rs956115 and clinical covariates, including age, previous MI, hypertension, diabetes mellitus, LVEF, serum creatinine, diagnosis, low density lipoprotein, smoking status, previous PCI. Dominant model: CG and GG vs.CC. Recessive model: GG vs.CC and CG. Addictive model: the number of risk alleles is proportional to the risk of MACE. Abbreviations: CI, confidence intervals; HR, hazard ratio; Other abbreviations as in Table 1. |
For rs4244285, patients with GA genotype had a 2.13-fold higher MACE risk than those with GG genotype (adjusted HR=2.13, 95%CI: 1.10-4.12, P=0.0248; Table 2). Patients with AA genotype showed no increased MACE risk to GG genotype (adjusted HR=0.58; 95%CI:0.13-2.60; P=0.4787). When further adjusted for rs956115 genotypes, patients with GA genotype had a 2.19-fold higher risk than those with GG genotype (adjusted HR=2.19, 95%CI: 1.13-4.24, P=0.0200; Table 2 and Figure 3B). No significant difference of MACE risk was found while using either dominant (P=0.0666; Table 2) or additive model (P=0.4936; Table 2).
Interaction Analysis
Among patients with GG genotype of rs4244285, those who had CG or GG genotype of rs956115 presented a 4.85-fold higher MACE risk than those who had CC genotype (adjusted HR=4.85, P=0.0081; Figure 4). By comparison, among patients with non-GG genotype of rs4244285, those who had CG or GG genotype of rs956115 presented a 1.40-fold higher risk than those who had CC genotype (adjusted HR=1.40, P=0.4764; Figure 4). The interaction between rs956115 and rs4244285 was none-statistically significant (P=0.1453; Figure 4).
Association Of Rs956115 With Mace In Subgroup Analysis
We performed multivariable Cox-regression analysis for rs956115 in different patient subgroups (Figure 5). The association between rs956115 genotypes and MACE remained statistically significant in subgroup of normal serum creatinine (adjusted HR=2.09, 95%CI: 1.04-4.18) (Figure 5). Although the adjusted HR between CG or GG and CC genotypes of rs956115 did not reach statistically significant in the diabetes subgroup (Figure 5), the dominant model HR of MACE for patients with CG or GG genotype of rs956115 tended to be similar across subgroups. No significant interactions were observed in any of those subgroups except LVEF subgroup (Interaction P=0.0006) (Figure 5).