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  • Research article
  • Open Access
  • Open Peer Review

Genetic variations in PRKAA1 predict the risk and progression of gastric Cancer

Contributed equally
BMC Cancer201818:923

https://doi.org/10.1186/s12885-018-4818-3

  • Received: 8 September 2017
  • Accepted: 13 September 2018
  • Published:
Open Peer Review reports

Abstract

Background

PRKAA1 encodes α-subunit of 5-AMP-activated protein kinase (AMPK), which has been implicated in the pathogenesis of carcinoma of the stomach. Previous works have suggested that polymorphisms in the PRKAA1 may be associated with the risk of non-cardiac gastric cancer (NCGC), but whether PRKAA1 polymorphisms are related to clinical pathologic characteristics of gastric cancer and its clinical outcome is largely unknown.

Methods

We carried out a case-control study including a total of 481 gastric cancer patients and 490 healthy controls. The genotypes of enrolled polymorphisms were identified with Sequenom MassARRAY platform.

Results

This study showed that rs10074991 GG genotype (adjusted OR = 1.44, 95%CI:0.99–2.09, p = 0.056) has a borderline significantly increased risk for gastric cancer, which was consistent with the result of additive model (adjusted OR = 1.21, 95%CI:1.01–1.46, p = 0.042). In similar, an increased risk of gastric cancer was also observed for rs13361707 TC genotype (adjusted OR = 1.47, 95%CI: 1.01–2.14, p = 0.043; additive model: adjusted OR = 1.22, 95%CI: 1.02–1.47, p = 0.033). Furthermore, the rs154268 and rs461404 were also found associated with increased gastric cancer risk, which may be influenced by age, tumor type and differentiation, and tumor stage. Haplotype analysis indicated A-G-C-T-C-G haplotype (rs6882903, rs10074991, rs13361707, rs3805490, rs154268 and rs461404) is associated with increased risk of gastric cancer (OR = 1.29, 95%CI: 1.02–1.62, p = 0.035). The univariate analysis for overall survival (OS) revealed that both of rs10074991 and rs13361707 variants are associated with poor OS in patients with NCGC.

Conclusion

This case-control study provided the evidence thatrs13361707CC, rs10074991GG, rs461404GG, and rs154268CC are associated with increased gastric cancer risk, especially for NCGC, and that patients with rs10074991 G or rs13361707 C allele have a poor OS.

Keywords

  • Gastric cancer
  • Polymorphism
  • PRKAA1
  • Prognosis

Background

Gastric cancer (GC) is one of the most common cancers worldwide and remains the leading cause of cancer related death [1]. The incidence of this disease varies with the geographical region and patient ethnicity. About 70% cases in the world were reported from developing countries, and Eastern Asian countries have the highest GC incidence and mortality [1, 2]. Although mechanism of gastric carcinogenesis is still not fully understood, environmental factors, such as high intake of salt, tobacco smoking, and particularly Helicobacter pylori(H. pylori) infection have been regarded as the risk factors for the disease [3].Genetic factors have also been found to contribute to the risk of GC, with the first-degree relatives of the GC patients tending to have about 1.3 to 3.0 fold higher relative risk for GC than those without relatives with GC [4].

To date, genetic variations have widely been shown to be associated with GC risk [5], with particular importance on the polymorphisms involved in the signal transduction pathways [6, 7]. The 5-AMP-activated protein kinase (AMPK) pathway has been implicated in a series of tumors including GC. This is a heterotrimeric protein that consists of an α-catalytic subunit and 2 regulatory subunits (β and γ), and the α-subunit is encoded either by PRKAA1 or PRKAA2 gene. Previous genome-wide association study (GWAS) identified the PRKAA1 polymorphism rs13361707 as a risk factor for non-cardiac GC (NCGC) in a Chinese population [8]; however, these results were not successfully duplicated [9], which may be due to the different characteristics of enrolled participants, population stratification, and clinical pathologic characteristics of GC. It is also not known whether the polymorphisms in the PRKAA1 gene are related to clinical pathological characteristics of GC and clinical outcome of the patients. We carried out this case-control study on a Chinese population to investigate the susceptibility of six polymorphisms in the PRKAA1 gene (see Additional file 1.) to the risk of GC and their associations with the clinical pathological characteristics, and evaluated the predictive value of these polymorphisms to the clinical outcome of GC patients.

Methods

Study subjects

A total of 481 GC patients, and age- and gender-matched 490 healthy individuals were enrolled in this study. The patients were histologically diagnosed as GC from Nanjing First Hospital, Nanjing Medical University, and the healthy controls were individuals who came to the hospital for routine physical examinations and were confirmed be healthy. All the participants were the heritably unrelated ethnic Han Chinese from the same geographic region of Nanjing City, Jiangsu, China. The whole blood of all enrolled participants were collected before operation and then stored at − 80 °C before genotyping. The clinical features of patients, including tumor size, distant metastasis, and depth of invasion, were collected from the patients’ medical records provided by Department of pathology, and the tumor TNM stages were examined and evaluated using the TNM classification according to American Joint Commission for Cancer Staging in 2002, sixth edition. The clinical outcomes of patients were found through on-site interview, direct calling, or medical chart review.

The characteristics of healthy controls, including age, gender, smoking and drinking, were collected via a questionnaire. Individuals who had smoked daily for more than 1 year were considered smokers, and those who consumed one or more alcoholic drinks per week for at least one year were considered drinkers. The protocol of this study was in accordance with the Declaration of Helsinkiand approved by the Institutional Review Board of the Nanjing First Hospital, and written informed consent was obtained from all the participants.

DNA extraction and genotyping

The genotypes of all polymorphisms were detected with the SequenomMassARRAY platform, as previously described [10, 11]. First, DNA was extracted from whole-blood samples and concentrated by using GoldMag-Mini Whole Blood Genomic DNA Purification Kit according to the manufacture’s protocol (GoldMag Co. Ltd. Xi’an, China), and then DNA purity was measured by spectrometry (DU530 UV/VIS spectrophotometer, Beckman Instruments, Fullerton, CA, US). The qualified DNA samples were genotyped using the SequenomMassARRAY platform followed the standard protocol recommended by the manufacturer of a Sequenom Mass-ARRAY®RS1000(Sequenom, Inc.). Multiplexed SNP MassEXTENDED assay was designed by SequenomMassARRAY Assay Design 3.0 Software [12]. Finally, data management and analysis were performed using SequenomTyper 4.0 Software [12, 13].

H. pylori infection detection

H. pylori infection status of enrolled participants has been determined by serology using a commercial H. pylori Immunogold Testing Kit (KangmeiTianhong Biotech (Beijing) Co., Ltd., Beijing, China), which has been validated in the Chinese population with sensitivity of 98.29% and specificity of 98.51% for the detection of H. pylori infection.

Statistical analysis

The Hardy-Weinberg equilibrium in the healthy control group was tested by using a goodness of fit chi-square test. The statistical analysis for genotype distribution was performed by the χ2 test, and odds ratios (OR) and 95% confidence intervals (CIs) were calculated using logistic regression model. The dominant model, co-dominant model, and additive model were the test for all polymorphisms, with the dominant and co-dominant models being used only if the additive model is significant or there is a previous hypothesis to do this.

Survival curves were analyzed by the Kaplan-Meier method, and the Hazard ration (HR) and 95% CIs were calculated using Cox proportional hazards regression model. The P value < 0.05 was considered statistically significant difference. The haplotype analysis was performed using online software SHEsis (analysis.bio-x.cn/myAnalysis.php).

Results

Characteristics of the participants

There was no significant difference in age (cases: 65.55 ± 11.92 years, healthy controls: 64.85 ± 11.83; p = 0.694), gender (cases: male73.60%, healthy controls: male73.27%; p = 0.782), smoking (cases: 23.08%, healthy controls: 24.29%; p = 0.658), and drinking (cases: 11.02%, healthy controls: 9.59%; p = 0.465) between cases and controls. For H. pylori infection status, the ratio of H. pylori infection in cases (54.47%) was higher than that in healthy controls (49.18%), however there was no significant difference between the two groups (p = 0.099), as presented in Table 1.
Table 1

Clinical characteristics of the participants

Variables

Cases, n (%)

Controls, n (%)

p-Value

Total

481

490

 

Age (mean ± SD)

65.55 ± 11.92

64.85 ± 11.83

0.694a

 >60

168

167

0.782b

  ≤ 60

313

323

 

Gender

 Male

354(73.60)

359(73.27)

0.907 b

 Female

127(26.40)

131(26.73)

 

Drinking

 Yes

53(11.02)

47(9.59)

0.465 b

 No

428(88.98)

443(90.41)

 

Smoking

 Yes

111(23.08)

119(24.29)

0.658 b

 No

370(76.92)

371(75.71)

 

Helicobacter pylori infection status

 Positive

262(54.47)

241(49.18)

0.099 b

 Negative

219(45.53)

249(50.81)

 

Differentiation

 Low

195(40.54)

  

 Med and high

286(59,46)

  

Clinical stages

 T1-T2

159(33.06)

  

 T3-T4

322(66.94)

  

Tumor Site

 GCA

140(29.11)

  

 NGCA

341(70.89)

  

GCA gastric cardiac adenocarcinoma, NGCA non-gastric cardiac adenocarcinoma

aIndependent t-test. bTwo-sided χ2 test for distributions between cases and controls

For the clinical pathological characteristics, a total of 195 (40.54%) and 286 (59.46%) patients had low and median to high pathological differentiation, respectively. For the tumor site classification, a total of 159 (33.06%) and 322 (66.94%) patients were classified to TNM stage T1-T2 and T3-T4, respectively. For the tumor location, a total of 140 (29.11%) and 341(70.89%) patients were diagnosed as gastric cardiac adenocarcinoma (GCA) and non-cardiatic GC (NCGC), respectively.

Association between polymorphisms and risk of GC

The genotype distributions of the selected polymorphisms in cases and controls are presented in Table 2. The observed frequencies of all tested genotypes in controls did not deviate from Hardy-Weinberg equilibrium (HWE) (rs10074991: p = 0.129; rs13361707: p = 0.152; rs1044129: p = 0.368; rs154268: p = 0.140; rs6882903: p = 0.842; rs3805490: p = 0.929; rs461404: p = 0.155).
Table 2

Distribution of the genotypes in all participants

Genotype

Controls, n (%)

Patients, n (%)

OR (95% CI)a

p-Value

rs10074991

 AA

128(26.12)

104(21.62)

Reference

 

 AG

261(53.27)

255(53.01)

1.19(0.87,1.62)

0.283

 GG

101(20.61)

122(25.36)

1.44(0.99,2.09)

0.056

 AG/GG

362(73.88)

377(78.38)

1.26(0.94,1.70)

0.127

 Additive model

  

1.21(1.01, 1.46)

0.042

rs13361707

 TT

129(26.33)

103(21.41)

Reference

 

 TC

260(53.06)

256(53.22)

1.22(0.89,1.66)

0.219

 CC

101(20.61)

122(25.36)

1.47(1.01,2.14)

0.043

 TC/CC

361(73.67)

365(75.88)

1.29(0.96,1.74)

0.093

 Addictive model

  

1.22(1.02,1.47)

0.033

rs154268

 TT

297(60.61)

271(56.34)

Reference

 

 TC

176(35.92)

179(37.21)

1.13(0.86,1.47)

0.388

 CC

17(3.47)

31(6.44)

1.96(1.06,3.63)

0.033

 TC/CC

193(39.39)

210(43.66)

1.20(0.93,1.56)

0.158

 Additive model

  

1.24(1.00,1.53)

0.053

rs6882903

 CC

342(69.80)

312(64.86)

Reference

 

 CA

134(27.35)

149(30.98)

0.86(0.41,1.80)

0.687

 AA

14(2.86)

20(4.16)

1.56(0.77,3.15)

0.217

 CA/AA

148(30.20)

169(35.14)

1.26(0.96,1.65)

0.097

 Additive model

  

1.23(0.98,1.55)

0.078

rs3805490

 TT

279(56.94)

280(58.21)

Reference

 

 TA

181(36.94)

170(35.34)

0.93(0.71,1.21)

0.567

 AA

30(6.12)

31(6.44)

1.02(0.60,1.73)

0.953

 TA/AA

211(43.06)

201(41.79)

0.94(0.73,1.21)

0.627

 Additive model

  

0.97(0.79,1.19)

0.756

rs461404

 AA

298(60.82)

270(56.13)

Reference

 

 GA

175(35.71)

179(37.21)

1.14(0.87,1.49)

0.341

 GG

17(3.47)

32(6.65)

2.05(1.11,3.78)

0.022

 GA/GG

192(39.18)

211(43.87)

1.22(0.95,1.58)

0.125

 Additive model

  

1.26(1.01,1.56)

0.037

aAdjusted for age, gender, smoking, drinking, and Helicobacter pylori infection

Rs10074991 GG genotype had a borderline significantly increased risk of GC (adjusted OR = 1.44, 95%CI: 0.99–2.09, p = 0.056), and the additive model shows rs10074991 is an increased risk factor for GC (adjusted OR = 1.21, 95%CI: 1.01–1.46, p = 0.042). In similar, an increased risk of rs13361707 was also observed for GC (TC vs. GG: adjusted OR = 1.47, 95%CI: 1.01–2.14, p = 0.043; additive model: adjusted OR = 1.22, 95%CI: 1.02–1.47, p = 0.033). Besides, the results have also revealed that rs154268 and rs461404 are associated with increased GC risk (rs154268 TC: adjusted OR = 1.96, 95%CI: 1.06–3.63, p = 0.033; rs154268 additive model: adjusted OR = 1.24, 95%CI: 1.00–1.53, p = 0.053; rs461404 GA: adjusted OR = 2.05, 95%CI: 1.11–3.78, p = 0.022; rs461404 additive model: adjusted OR = 1.26, 95%CI: 1.01–1.56, p = 0.037). However, there was no significant association between rs6882903 and rs3805490 and risk of GC, as summarized in Table 2.

Stratification analysis

To further assess the four potential susceptible polymorphisms (rs10074991, rs13361707, rs154268 and rs461404) to the risk of GC, a stratified analysis was performed by subgroups of participants’ clinical characteristics (age, gender, H. pylori infection status), and tumor pathological characteristics (tumor site, tumor differentiation, and clinical stage).

In China, men usually retire at age of 60, which means they retain a stable and sustainable life style (the environmental factors), so we choose 60 years as the cut-off value for the subgroup analysis. In the subgroup of age ≤ 60, rs10074991GG (adjusted OR = 1.93, 95%CI: 1.00–3.73, p = 0.050), rs13361707CC (adjusted OR = 2.00, 95%CI: 1.04–3.84, p = 0.039) and rs461404GG (adjusted OR = 3.12, 95%CI: 1.05–9.28, p = 0.040) were associated with increased GC risk. However, in the group of age>60, there was no significant association of these four polymorphisms with the risk of GC. For the subgroup of gender, in the male group, rs10074991 (additive model: adjusted OR = 1.25, 95%CI: 1.01–1.56, p = 0.046) and rs13361707 (CC: adjusted OR = 1.44, 95%CI: 1.01–2.06, p = 0.044; additive model: adjusted OR = 1.27, 95%CI: 1.02–1.58, p = 0.034) contributed to increased risk of GC. In similar, in the subgroup of positive H. pylori infection, a borderline significantly increased risk of rs10074991 (AG: adjusted OR = 1.68, 95%CI: 0.98–2.88, p = 0.060; additive model: adjusted OR = 1.30, 95%CI: 0.99–1.69, p = 0.057) and rs13361707 (TC: adjusted OR = 1.75, 95%CI: 1.02–3.00, p = 0.042; additive model: adjusted OR = 1.32, 95%CI: 1.01–1.73, p = 0.041) was observed for GC, as shown in Table 3. For the subgroup of pathological characteristics of tumor, the four polymorphisms were significant associated with increased risk of NCGC, but not GCA. Moreover, the significant associations of these four polymorphisms were observed in the subgroup of patients with tumor in median or high differentiation or T3-T4, but not for low differentiation or T1-T2, as shown in Table 4.
Table 3

PRKAA1 Polymorphisms with Gastric Cancer Risk by Clinical Characteristics of Participants

Genotype

Age

Sex

Helicobacter pylori infection.

≤60

>60

Male

Female

Positive

Negative

Ca/Co

OR (95% CI)

P

Ca/Co

OR (95% CI)a

P

Ca/Co

OR (95% CI)a

P

Ca/Co

OR (95% CI)a

P

Ca/Co

OR (95% CI)a

P

Ca/Co

OR (95% CI)a

P

rs10074991

 AA

33/45

Reference

 

71/83

Reference

 

69/62

Reference

 

35/36

Reference

 

52/57

Reference

 

52/71

Reference

 

 AG

90/90

1.31(0.76,2.25)

0.331

165/171

1.10(0.75,1.62)

0.630

197/193

1.35(0.93,1.95)

0.117

58/68

0.77(0.42,1.41)

0.399

142/139

1.12(0.72,1.75)

0.613

113/122

1.26(0.81,1.96)

0.302

 GG

45/32

1.93(1.00,3.73)

0.050

77/69

1.20(0.76,1.91)

0.436

88/74

1.51(0.97,2.37)

0.070

34/27

1.27(0.63,2.58)

0.503

68/45

1.68(0.98,2.88)

0.060

54/56

1.31(0.77,2.22)

0.318

 AG/GG

135/122

1.46(0.87,2.45)

0.154

242/240

1.15(0.80,1.66)

0.461

285/267

1.40(0.98,1.99)

0.067

92/95

0.91(0.52,1.59)

0.734

210/184

1.26(0.82,1.93)

0.292

167/178

1.27(0.84,1.94)

0.255

 Additive model

 

1.40(1.01,1.93)

0.043

 

1.12(0.89,1.41)

0.332

 

1.25(1.01,1.56)

0.046

 

1.10(0.77,1.55)

0.611

 

1.30(0.99,1.69)

0.057

 

1.14(0.88,1.48)

0.313

rs13361707

 TT

33/46

Reference

 

70/83

Reference

 

68/93

Reference

 

36/35

Reference

 

51/58

Reference

 

52/71

Reference

 

 TC

90/89

1.35(0.79,2.33)

0.273

166/171

1.12(0.76,1.65)

0.563

198/192

1.40(0.96,2.03)

0.080

58/68

0.77(0.42,1.41)

0.399

143/138

1.18(0.76,1.84)

0.465

113/122

1.26(0.81,1.96)

0.302

 CC

45/32

2.00(1.04,3.84)

0.039

77/69

1.22(0.77,1.94)

0.402

88/74

1.56(1.00,2.44)

0.052

34/27

1.27(0.63,2.58)

0.503

68/45

1.75(1.02,3.00)

0.042

54/56

1.31(0.77,2.22)

0.318

 TC/CC

135/121

1.51(0.90,2.53)

0.119

242/240

1.17(0.81,1.69)

0.408

286/266

1.44(1.01,2.06)

0.044

92/95

0.91(0.52,1.59)

0.734

211/183

1.32(0.86,2.03)

0.200

167/178

1.27(0.84,1.94)

0.255

 Additive model

 

1.41(1.02,1.95)

 

0.036

1.13(0.90,1.42)

0.305

 

1.27(1.02,1.58)

0.034

 

1.10(0.77,1.55)

0.611

 

1.32(1.01,1.73)

0.041

 

1.14(0.88,1.48)

0.313

rs154268

 TT

92/106

Reference

 

179/191

Reference

 

195/213

Reference

 

76/84

Reference

 

151/146

Reference

 

120/151

Reference

 

 TC

64/56

1.32(0.83,2.10)

0.241

115/120

1.03(0.74,1.44)

0.844

140/134

0.81(0.60,1.10)

0.357

39/42

0.95(0.55,1.65)

0.866

91/86

1.03(0.71,1.49)

0.891

88/90

1.21(0.83,1.78)

0.315

 CC

12/5

2.77(0.92,8.33)

0.069

19/12

1.63(0.76,3.51)

0.210

19/12

1,73(0.81,3.67)

0.153

12/5

2.44(0.82,7.29)

0.111

20/9

2.16(0.95,4.91)

0.068

11/8

1.77(0.69,4.55)

0.238

 TC/CC

76/61

1.45(0.92,2.26)

0.107

134/132

1.09(0.80,1.50)

0.581

159/146

1.20(0.89,1.62)

0.230

51/47

1.12(0.67,1.87)

0.668

111/95

1.13(0.79,1.62)

0.496

99/98

1.27(0.88,1.84)

0.208

Additive model

 

1.46(1.00,2.12)

0.048

 

1.13(0.87,1.48)

0.355

 

1.21(0.94,1.56)

0.142

 

1.23(0.82,1.85)

0.308

 

1.21(0.90,1.62)

0.201

 

1.26(0.92,1.73)

0.153

rs461404

 AA

91/106

Reference

 

179/192

Reference

 

195/214

Reference

 

75/84

Reference

 

151/146

Reference

 

119/152

Reference

 

 GA

64/56

1.34(0.84,2.13)

0.218

115/119

1.05(0.75,1.45)

0.792

139/133

1.16(0.85,1.58)

0.347

40/42

1.00(0.58,1.72)

0.991

91/86

1.03(0.71,1.49)

0.891

88/89

1.25(0.85,1.83)

0.255

 GG

13/5

3.12(1.05,9.27)

0.040

19/12

1.64(0.76,3.53)

0.207

20/12

1.84(0.87,3.87)

0.109

12/5

2.46(0.82,7.36)

0.108

20/9

2.16(0.95,4.91)

0.068

12/8

1.96(0.77,4.97)

0.156

 GA/GG

77/61

1.49(0.95,2.32)

0.082

134/131

1.10(0.81,1.52)

0.537

159/145

1.21(0.90,1.64)

0.205

52/47

1.16(0.69,1.93)

0.573

111/95

1.13(0.79,1.62)

0.496

100/97

1.31(0.91,1.90)

0.150

 Additive model

 

1.51(1.04,2.19)

0.030

 

1.14(0.88,1.49)

0.327

 

1.23(0.95,1.59)

0.113

 

1.26(0.84,1.89)

0.261

 

1.21(0.90,1.62)

0.201

 

1.31(0.95,1.79)

0.098

aAdjusted for age, gender, smoking, drinking, and Helicobacter pylori infection; Ca, case; Co, control

Table 4

PRKAA1 Polymorphisms with Gastric Cancer Risk by Tumor Classification

Genotype

Co

Site

DIF

TNM

GCA

NGCA

Low

Med-high

T1-T2

T3-T4

Ca

OR (95% CI)a

P

Ca

OR (95% CI)a

P

Ca

OR (95% CI)a

P

Ca

OR (95% CI)a

P

Ca

OR (95% CI)a

P

Ca

OR (95% CI)a

P

rs10074991

 AA

128

43

Reference

 

61

Reference

 

47

Reference

 

57

Reference

 

35

Reference

 

69

Reference

 

 AG

261

73

0.80(0.51,1.24)

0.312

182

1.45(1.01,2.09)

0.043

104

1.09(0.73,1.63)

0.683

151

1.28(0.88,1.86)

0.203

88

1.23(0.79,1.94)

0.362

167

1.16(0.82,1.66)

0.409

 GG

101

24

0.66(0.37,1.16)

0.150

98

2.02(1.32,3.07)

0.001

44

1.17(0.71,1.93)

0.528

77

1.66(1.08,2.57)

0.022

36

1.33(0.77,2.29))

0.308

86

1.51(0.99,2.28)

0.054

 AG/GG

362

97

0.77(0.50,1.16)

0.210

280

1.16(1.14,2.28)

0.007

148

1.20(0.76,1.65)

0.570

229

1.38(0.97,1.98)

0.074

124

1.27(0.82,1.95)

0.283

253

1.26(0.90,1.77)

0.171

 Additive model

  

0.82(2.62,1.08)

0.163

 

1.43(1.16,1.76)

0.001

 

1.09(0.86,1.40)

0.475

 

1.31(1.05,1.62)

0.016

 

1.17(0.89,1.53)

0.255

 

1.24(1.01,1.53)

0.039

rs13361707

 TT

129

43

Reference

 

60

Reference

 

47

Reference

 

56

Reference

 

35

Reference

 

68

Reference

 

 TC

260

73

0.80(0.52,1.25)

0.330

183

1.51(1.05,2.17)

0.027

104

1.10(0.73,1.65)

0.641

152

1.32(0.91,1.93)

0.141

88

1.25(0.80,1.97)

0.331

168

1.20(0.84,1.71)

0.320

 CC

101

24

0.66(0.37,1.17)

0.157

98

2.08(1.36,3.16)

0.001

44

1.19(0.72,1.95)

0.495

78

1.71(1.11,2.65)

0.015

36

1.34(0.78,2.31)

0.290

86

1.55(1.02,2.34)

0.040

 TC/CC

361

97

0.77(0.51,1.17)

0.225

281

1.67(1.18,2.36)

0.004

148

1.13(0.77,1.67)

0.529

230

1.42(1.00,2.05)

0.050

124

1.28(0.83,1.98)

0.257

254

1.30(0.93,1.82)

0.125

 Additive model

  

0.82(0.62,1.09)

0.171

 

1.45(1.18,1.78)

0.001

 

1.10(0.86,1.40)

0.451

 

1.32(1.07,1.64)

0.012

 

1.18(0.90,1.54)

0.240

 

1.26(1.02,1.55)

0.030

rs154268

 TT

297

88

Reference

 

183

Reference

 

115

Reference

 

156

Reference

 

90

Reference

 

181

Reference

 

 TC

176

48

0.89(0.59,1.33)

0.567

131

1.24(0.92,1.66)

0.153

70

1.05(0.74,1.49)

0.794

109

1.18(0.86,1.61)

0.299

63

1.12(0.85,1.82)

0.258

116

1.46(0.85,1.82)

0.704

 CC

17

4

0.77(0.25.2.39)

0.657

27

2.54(1.34,4.81)

0.004

10

1.46(0.65,3.31)

0.364

21

2.38(1.20,4.58)

0.010

6

1.14(0.43,3.02)

0.790

25

2.46(1.28,4.70)

0.007

 TC/CC

193

52

0.88(0.60,1.31)

0.535

158

1.36(1.03,1.81)

0.032

80

1.09(0.78,1.53)

0.614

130

1.28(0.95,1.73)

0.099

69

1.24(0.86,1.79)

0.253

141

1.19(0.89,1.58)

0.240

 Additive model

  

0.89(0.63,1.26)

0.508

 

1.40(1.10,1.76)

0.005

 

1.12(0.84,1.49)

0.449

 

1.33(1.04,1.70)

0.025

 

1.19(0.87,1.63)

0.289

 

1.27(1.00,1.61)

0.046

rs461404

 AA

298

88

Reference

 

182

Reference

 

114

Reference

 

156

Reference

 

90

Reference

 

180

Reference

 

 GA

175

48

0.90(0.60,1.34)

0.596

131

1.25(0.93,1.68)

0.132

71

1.08(0.76,1.54)

0.668

108

1.18(0.86,1.61)

0.383

63

1.25(0.86,1.83)

0.243

116

1.08(0.80,1.46)

0.628

 GG

17

4

0.78(0.25,2.40)

0.659

28

2.67(1.42,5.04)

0.002

10

1.48(0.65,3.34)

0.349

22

2.47(1.27,4.79)

0.007

6

1.23(0.58,2.61)

0.787

26

2.59(1.36,4.93)

0.004

 GA/GG

192

52

0.89(0.60,1.32)

0.563

159

1.38(1.05,1.84)

0.023

81

1.12(0.80,1.58)

0.509

130

1.29(0.96,1.74)

0.116

69

1.25(0.86,1.80)

0.240

142

1.21(0.91,1.62)

0.187

 Additive model

  

0.90(0.63,1.27)

0.532

 

1.42(1.13,1.79)

0.003

 

1.14(0.85,1.52)

0.373

 

1.34(1.05,1.72)

0.019

 

1.19(0.87,1.64)

0.276

 

1.30(1.03,1.65)

0.029

aAdjusted for age, gender, smoking, drinking, and Helicobacter pylori infection; GCA gastric cardia adenocarcinoma, NGCA non-gastric cardia adenocarcinoma, Ca, case Co control

Haplotype analysis of polymorphisms in PRKAA1

The enrolled six polymorphisms locate in the intron or upstream of PRKAA1, so these sites may be in linkage disequilibrium with each other. Therefore, the combined susceptibility of these six polymorphisms to GC risk was calculated by haplotype analysis. The results indicated that the haplotype A-G-C-T-C-G (rs6882903, rs10074991, rs13361707, rs3805490, rs154268, rs461404) is associated with the increased risk of GC (OR = 1.29, 95%CI: 1.02–1.62, p = 0.035), as compared with other haplotypes (Fig. 1).
Fig. 1
Fig. 1

Haplotype analysis of polymorphisms indicating the susceptibility to gastric cancer risk. The linkage disequilibrium (LD) map according to the genotype data, the color and figure show the linkage disequilibrium coefficient with D’ values The prevalence of haplotype A-G-C-T-C-G (rs6882903, rs10074991, rs13361707, rs3805490, rs154268, rs461404) was significantly higher among cases (19.6%) compared to controls (16.2%) (haplotype-specific p = 0.035), and those with this haplotype have 1.29 times higher risk of gastric cancer (OR = 1.29, 95%CI: 1.02–1.62, p = 0.035) compared to noncarriers

Association between polymorphisms and clinical outcome of patients

A total 481 patients were followed up for the survival state. The association of polymorphisms with the overall survival (OS) of patients was assessed for their predictive value for patients with heterozygous and homozygous genotype, or their combination, compared to the wild genotype. The results revealed that rs10074991 (AG: adjusted HR = 1.80, 95%CI:1.21–2.67, p = 0.004; GG: adjusted HR = 1.75, 95%CI: 1.13–2.70, p = 0.012; AG/GG: HR = 1.78, 95%CI: 1.21–2.61, p = 0.003) and rs13361707 (TC: adjusted HR = 1.85, 95%CI: 1.24–2.77, p = 0.003; CC: adjusted HR = 1.79, 95%CI: 1.16–2.78, p = 0.009; TC/CC: adjusted HR = 1.83, 95%CI: 1.24–2.70, p = 0.002) were associated with poor OS of patients with NCGC, indicating these two polymorphisms have a significant prediction value for the patients with NCGC, as shown in Table 5.
Table 5

PRKAA1 Polymorphisms with clinical outcome of patients with NGCA

Genotype

All patients

NGCA

HR (95% CI)

p-Value

HR (95% CI)

p-Value

HR (95% CI)a

p-Value

HR (95% CI)b

p-Value

rs10074991

 AA

Reference

 

Reference

 

Reference

 

Reference

 

 AG

1.16(0.87,1.55)

0.300

1.62(1.10,2.38)

0.015

1.63(1.10,2.41)

0.015

1.80(1.21,2.67)

0.004

 GG

1.15(0.83,1.59)

0.401

1.64(1.08,2.49)

0.020

1.71(1.12,2.60)

0.012

1.75(1.13,2.70)

0.012

 AG/GG

1.16(0.88,1.52)

0.290

1.62(1.12,2.35)

0.011

1.66(1.14,2.41)

0.008

1.78(1.21,2.61)

0.003

rs13361707

 TT

Reference

 

Reference

 

Reference

 

Reference

 

 TC

1.18(0.89,1.57)

0.253

1.67(1.13,2.47)

0.010

1.68(1.13,2.50)

0.010

1.85(1.24,2.77)

0.003

 CC

1.16(0.84,1.61)

0.364

1.68(1.10,2.56)

0.016

1.76(1.15,2.68)

0.009

1.79(1.16,2.78)

0.009

 TC/CC

1.18(0.90,1.54)

0.246

1.67(1.15,2.44)

0.007

1.71(1.17,2.50)

0.006

1.83(1.24,2.70)

0.002

rs154268

 TT

Reference

 

Reference

     

 TC

1.08(0.85,1.36)

0.525

1.18(0.90,1.56)

0.242

    

 CC

1.23(0.78,1.94)

0.367

1.29(0.78,2.12)

0.322

    

 TC/CC

1.10(0.88,1.17)

0.405

1.20(0.92,1.56)

0.183

    

rs6882903

 CC

Reference

 

Reference

     

 CA

0.97(0.76,1.24)

0.796

1.12(0.84,1.48)

0.443

    

 AA

1.48(0.89,2.47)

0.130

1.56(0.90,2.72)

0.113

    

 CA/AA

1.02(0.81,1.29)

0.869

1.17(0.89,1.53)

0.252

    

rs3805490

 TT

Reference

 

Reference

     

 TA

1.03(0.82,1.30)

0.807

1.07(0.81,1.41)

0.626

    

 AA

0.82(0.51,1.31)

0.404

0.97(0.55,1.68)

0.898

    

 TA/AA

0.99(0.80,1.24)

0.953

1.06(0.81,1.37)

0.691

    

rs461404

 AA

Reference

 

Reference

     

 GA

1.08(0.86,1.37)

0.506

1.19(0.90,1.56)

0.352

    

 GG

1.25(0.80,1.94)

0.333

1.31(0.80,2.13)

0.228

    

 GA/GG

1.11(0.89,1.38)

0.379

1.21(0.93,1.57)

0.165

    

Discussion

This study revealed that PRKAA1 genetic polymorphismsrs13361707CC, rs10074991GG, rs461404GG, and rs154268CC were associated with increased risk of GC. The susceptibility of these four polymorphisms to the risk of GC were here observed in the subgroup of age ≤ 60, male, NCGC, median to high differentiation and T3-T4 subgroup. Polymorphisms rs13361707 and rs10074991 were associated with poor survival of patients with NCGC.

Variant rs13361707 is located in the first intron of PRKAA1 at 5p13.1, which was primarily found to be associated with NCGC risk by a GWAS in a Chinese population(1006 non-cardia gastric cancer and 2273 controls, and confirmed with 3288 with non-cardia gastric cancer and 3609 controls) [8], and the significant association was duplicated by other studies on Chinese population(1124 cases and 1,194controls) [14] and on Korean population (Kim et al.: 477 case-control pairs; Song et al.: 3245 cases and 1700 controls) [15, 16]. This study observed that rs13361707 CC genotype was associated with increased risk of GC, and C allele carriers had a higher risk of NCGC, but not of GCA, indicating the association of rs13361707with the increased GC risk is specific to NCGC. Etiological studies have found differences between GCA and NCGC, concerning e.g. H. pylori infection [17, 18], or body mass index [19], and which was confirmed by epidemiological study that also suggested the susceptibility of genetic polymorphism to GC is different for NCGC and GCA [20]. Moreover, in the subgroup of positive H. pylori infection, our study showed rs13361707CC genotype is associated with increased risk of GC, indicating the interaction of rs13361707 and H. pylori can enhance the GC risk, which is consistent with the results of previous study [21]. The polymorphism rs13361707 is located in the first intron of PRKAA1 gene, which is a cellular energy sensor maintaining energy homeostasis, and contributes to cancer development by regulating mRNA translation and protein synthesis [22, 23]. Although the function of rs13361707 is largely unknown, several published studies and the current work indicated that risk of rs13361707 for GC was associated with the type of GC, and its susceptibility may be influenced by H. pylori infection [5].

This study also showed that rs10074991GG genotype is borderline significantly associated with increased risk of GC, and stratification analysis revealed the genotype to be associated with increased risk of NCGC, which is consistent with the reports of Hu et al. [20] that rs10074991 G allele linked with rs13361707 C allele (these two polymorphisms locate in the intron of PRKAA1 with the distance of 1333 bp) was a risk factor of NCGC. Moreover, such an association was also reported by Kim et al. [15] in a Korean population [15]. However, the function of these two sites remains unclear and the mechanism has yet to be established.

In this study, rs154268 CC genotype was also found to be associated with increased risk of GC for all participants and especially for the subgroup of NCGC, tumor with median to high differentiation, and T3-T4, suggesting rs154268 could be associated with pathological characteristics of GC. Consistent with this, the rs154268 TC genotype was also previously reported to be associated with the risk of GC [15], indicating that the C allele is a risk factor for GC. Actually, this study revealed the linkage disequilibrium (LD) between rs154268 and rs461404 (D′ = 1.0), which means the result of rs461404 is in accord with that of rs154268. However, to date, there is no functional study regarding the potential functional role of these two polymorphisms in carcinogenesis. In general, in this study, the result of rs461404 was inconsistent with that of rs154268.

The present work showed that rs10074991 G and rs13361707 C allele carriers with NCGC have poor OS, and this association was still observed after being adjusted by basic clinical characteristics (age, gender, H. pylori infection, drinking, and smoking) or pathological characteristics (tumor differentiation, tumor stage), indicating these two polymorphisms were independent factors for predicting the clinical outcome for NCGC. To our knowledge, this is the first report to discuss the role of these two polymorphisms in prognosis for patients with NCGC, which however should be verified by a further research with larger samples.

There are some limitations of this study. First, the sample size is relatively small, which may limit the statistical power, especially for the multiple stratified analyses. Second, the polymorphisms discussed in this study were limited in number and based on previous knowledge of potential functional significance of polymorphisms that have been found to be related to GC risk. Thus, a more comprehensive tagging SNP-based approach and a haplotype block analysis would better assesses the association and provides more complete information regarding the associations of AMPK pathway genes and GC risk.

Conclusions

This case-control study provided the evidence that rs13361707CC, rs10074991GG, rs461404GG, and rs154268CC are associated with increased GC risk, especially for NCGC, and that rs10074991 G and rs13361707 C alleles are independent prognostic factors for NCGC.

Notes

Abbreviations

AMPK: 

5’-AMP-activated protein kinase

CI: 

confidence intervals

GCA: 

gastric cardiac adenocarcinoma

GWAS: 

genome-wide association study

H. pylori

Helicobacter pylori

HR: 

hazard ration

HWE: 

Hardy-Weinberg Equilibrium

NCGC: 

non-cardiac gastric cancer

OR: 

odds ratios

OS: 

overall survival

Declarations

Acknowledgements

This work is supported by the National Natural Science Foundation (Grant numbers: 81472786, 81472305, 81773192); Suzhou Municipal Health Bureau projects (Grant number: LCZX201318); The Foundation of tumor clinical and basic research team (KYC005); The Six Talents Peak Project of Jiangsu Province (2014-WSW-061).

Availability of data and materials

Data supporting our findings are presented in the “Results” section. Researchers interested in source data are invited to write to the corresponding author.

Authors’ contributions

MC, BJ and PL designed the study. B.H collected the sample and information. M.T and PW performed the statistical analysis and drafted and revised the manuscript. LC and JL participated with the H. pylori detection and data collection. All authors read and approved the final version of the manuscript.

Ethics approval and consent to participate

The protocol of this study was in accordance with the Declaration of Helsinkiand approved by the Institutional Review Board of the Nanjing First Hospital, and written informed consents were obtained from all participants.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

Authors’ Affiliations

(1)
Department of Radiotherapy & Oncology, Kunshan First People’s Hospital Affiliated to Jiangsu University, Kunshan, Jiangsu Province, China
(2)
Department of Critical Care Medicine, The affiliated Yixing Hospital of Jiangsu University, Yixing, 214200, Jiangsu Province, China
(3)
General Clinical Research center, Nanjing First Hospital, Nanjing Medical University, Nanjing, 220006, Jiangsu Province, China
(4)
Departments of Medical biology, Wannan Medical College, Wuhu, Anhui Province, China
(5)
Department of Gastroenterology, Xuzhou Hospital of Traditional Chinese Medicine Affiliated to Nanjing University of Chinese Medicine, Xuzhou, China
(6)
Departments of Medical Oncology, Jiangsu Cancer Hospital Affiliated to Nanjing Medical University, Jiangsu Province Institute of Cancer, Nanjing, Jiangsu Province, China
(7)
Department of Medical Oncology, Wuxi People’s Hospital of Nanjing Medical University, No. 299, Qingyang Road, Wuxi, 214023, Jiangsu Province, China

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Copyright

© The Author(s). 2018

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