Study on the association between PI3K/AKT/mTOR signaling pathway gene polymorphism and susceptibility to gastric

Transcription

1 JBUON 2017; 22(6): ISSN: , online ISSN: E-mil: ORIGINAL ARTICLE Study on the ssocition between PI3K/AKT/mTOR signling pthwy gene polymorphism nd susceptibility to gstric cncer Lei Qi, Kewen Sun, Yun Zhung, Jing Yng, Jinping Chen Deprtment of Gstroenterology, the Third Affilited Hospitl of Soochow University, Chngzhou, Chin Summry Purpose: Excessive ctivtion of PI3K/AKT/mTOR signling pthwy is one of the most common chnges in humn cncers, nd single nucleotide polymorphisms (SNPs) existing in its functionl region cn ffect the occurrence process of vriety of cncers. This study imed to screen out the SNPs ssocited with susceptibility to gstric cncer in the PI3K/AKT/mT0R signling pthwy. Methods: In this cse-control study, the tgging SNPs in the promoter region5 -UTR, exon region or 3 -UTR of PIK3CA, PIK3CB, PIK3R1, PIK3R2, PIK3R3, AKT1, AKT2, AKT3 nd mtor genes were screened out. The reltionship between the genetic vrition of PI3K/AKT/mT0R signling pthwy genes nd the susceptibility to gstric cncer in Chinese Hn popultion ws investigted by this csecontrol study. Results: The results showed tht the polymorphisms of the two loci, PIK3R3 rs (Additive model: OR=1.16, 95% CI= ) nd mtor rs (GG vs TT: OR=0.75, 95% CI= ; Additive model: OR=0.78, 95% CI= ), were ssocited with the risk of gstric cncer in the studied popultion nd there ws combined effect between the two loci (p trend =0.005). Conclusions: In conclusion, the polymorphisms of the two loci, PIK3R3 rs nd mtor rs , on the PI3K/AKT/mTOR signling pthwy genes re ssocited with genetic susceptibility to gstric cncer in Chinese popultion. Key words: gstric cncer, PI3K/AKT/mTOR, polymorphism Introduction Gstric cncer is the fifth mjor tumor with high incidence rte worldwide, rnking third in the common cuses of deth from tumors. According to the sttisticl dt of the World Helth Orgniztion (WHO) in 2012, there were bout 951,600 new cses of gstric cncer, leding to 723,100 deths ech yer [1]. Studies hve confirmed tht the occurrence of gstric cncer is the result of the combined ction of environmentl nd genetic fctors. Helicobcter pylori (Hp) infection [2], poor eting hbits [3,4], smoking nd drinking [5] re the min risk fctors for gstric cncer. Under the exposure to the sme risk fctors, the risk of gstric cncer is not exctly the sme in individuls, suggesting tht the individul susceptibility my ply n importnt role in the occurrence nd development of gstric cncer. Phosphtidylinositol 3-kinse (PI3K)/protein kinse B (PKB or AKT)/mmmlin trget of rpmycin (mtor) signling pthwy is n importnt intrcellulr signling pthwy, which ffects the occurrence, development, tretment nd prognosis of tumors through regulting vriety of cell physiologicl processes [6]. Mny studies hve confirmed the PI3K/AKT/mTOR signling pthwy disorders in tumors, nd prticulrly in the biologicl regultion of gstric cncer [7], liver cncer [8], brest cncer [9], colorectl cncer [10] Correspondence to: Jinping Chen, MD. Deprtment of Gstroenterology, the Third Affilited Hospitl of Soochow University, 185 Juqin Rod, Chngzhou, Jingsu, Chin. Tel: , E-mil: @qq.com Received: 22/06/2017; Accepted: 03/07/2017

2 PI3K/AKT/mTOR polymorphism in gstric cncer 1489 nd prostte cncer [11], plying role s protooncogene, which hs become hotspot of the new moleculr biomrker-bsed nd trgeted therpy of tumors. Up to now, there hve been more studies on the PI3K signling pthwy gene polymorphism. Li et l. found in Chinese popultion tht the polymorphisms of PI3K/AKT/mTOR pthwy genes in ptients with non-smll cell lung cncer cn predict brin metstsis, which is instructive for the prevention of such metstsis in this cncer [12]. Lin et l. performed cse-control study to investigte whether the combined ction of PI3K/ AKT/mTOR pthwy gene polymorphism nd environmentl fctors would ffect the occurrence nd development of bldder cncer, nd they found tht the risk of bldder cncer in individuls with excessive energy intke, little exercise or severl risk lleles of PI3K/AKT/mTOR pthwy is incresed by nerly 21-fold; these results hve implictions for the prevention of bldder cncer [13]. At present, lrge number of studies on the PI3K/AKT/mTOR pthwy gene polymorphism nd tumors hve been conducted, but only few studies explored the PI3K/AKT/mTOR pthwy gene polymorphism nd gstric cncer. Moreover, such studies re limited to the single pthwy gene polymorphism nd gstric cncer, nd the study on the correltion between PI3K/AKT/mTOR pthwy gene polymorphism nd susceptibility to gstric cncer is lcking. In this reserch, the key genes of the whole PI3K/AKT/mTOR pthwy were studied bsed on the Thousnd Tlents Progrm Dtbse, 5 tgging SNPs in the functionl region of PIK3CA, PIK3CB, PIK3R1, PIK3R2, PIK3R3, AKT1, AKT2, AKT3 nd mtor genes were screened out, nd cse-control study ws performed to investigte the ssocition between the selected loci nd gstric cncer risk in 574 ptients with gstric cncer nd 912 control subjects. Methods Study objectives A totl of 574 ptients with new-onset gstric cncer who were treted in the Third Affilited Hospitl of Soochow University from Mrch 2014 to Jnury 2017 were enrolled in this study, while totl of 912 helthy subjects in the sme hospitl nd time period served s controls. All ptients were dignosed with gstric cncer by biopsy nd did not suffer from other cncers or receive ny rdiotherpy or chemotherpy before enrollment. Cses nd controls were mtched ccording to ge nd gender, nd the ge difference ws ±5 yers. Informed consent ws obtined from ll individuls; 5 ml venous blood ws collected using vcuum nticogulnt tube contining ethylene dimine tetrcetic cid (EDTA) nd stored t -20 C. The bsic informtion of ll subjects, such s gender, ge, tumor type nd tumor site, ws registered. This study ws pproved by the Ethics Committee of Soochow University. SNPs selection strtegy According to the UCSC gene browser, the tgging SNPs in the promoter region, 5 -untrnslted region (5 -UTR), exon region or 3 -untrnslted region (3 -UTR) of PIK3CA, PIK3CB, PIK3R1, PIK3R2, PIK3R3, AKT1, AKT2, AKT3 nd mtor were selected nd screened out using Hploview4.0 softwre ccording to Pirwise tgging nd r 2 threshold of 0.8. The minor llele frequency (MAF) of the selected loci ws higher thn Genotyping DNA ws extrcted from ll smples using the trditionl mmoni-chloroform method, nd the DNA genotype ws identified vi ABI7900HT fluorescent quntittive PCR instrument using TqMn genotyping method. Sttistics Chi-squre test ws used for the frequency distribution of genotyping in the control subjects to ensure tht they met the Hrdy-Weinberg equilibrium (HWE). Person s chi-squre test ws used for the demogrphic chrcteristics, ge nd gender mtching involved in the cse-control study. The odds rtio (OR) nd 95% confidence intervl (CI) of different lleles in clinicopthologic stging were nlyzed using the univrite nd multivrite logistic regression nlysis. SAS9.2 softwre ws used for the dt nlysis. All tests were two-sided. P<0.05 suggested tht the difference ws sttisticlly significnt. Results Generl chrcteristics of the study subjects A totl of 1486 cses nd controls (574 ptients with gstric cncer nd 912 helthy controls) were enrolled, nd the min chrcteristics re shown in Tble 1. There were no differences in ge nd gender between cses nd controls (p=0.418; p=0.115, respectively). There were 214 (37.3%) ptients with gstric crdi cncer nd 360 (62.7%) ptients with non-gstric crdi cncer. There were lso 328 (57.1%) ptients with diffuse type (57.1%) nd 246 (42.9%) ptients with intestinl type. Anlysis of the correltion between the SNPs nd the susceptibility to gstric cncer All loci selected met HWE (p>0.05) nd MAF ws higher thn 0.1 (Tble 2). Univrite logistic regression nlysis results re shown in Tble 3. After djustment for ge nd gender, PIK3R3 JBUON 2017; 22(6): 1489

3 1490 PI3K/AKT/mTOR polymorphism in gstric cncer rs ws found to be ssocited with incresed risk of gstric cncer (Additive model: OR=1.16, 95%CI= ), nd mtor rs ws ssocited with decresed risk of gstric cncer (GG vs TT: OR=0.75, 95%CI= ; Additive model: OR=0.78, 95%CI= ). The strtified nlysis results bsed on ge, gender, lesion site nd pthologicl type showed tht rs could reduce the onset risk of the elderly, femle, non-gstric crdi cncer nd intestinl type gstric cncer ptients (Tble 4). The conjoint nlysis showed tht rs nd rs hd combined effect, nd the onset risk of gstric cncer ws significntly incresed with the increse of risk bse (p trend =0.005). Compred with tht in the group crrying 0-2 risk bses, the incidence rte of gstric cncer in the group crrying 3-4 risk bses ws incresed by In ddition, it ws not found tht the three tgging SNPs, rs , rs nd rs , were ssocited with the onset risk of gstric cncer (p>0.05) (Tble 5). Tble 1. Selected chrcteristics in gstric cncer cses nd controls Vribles Cses Controls p vlue n (%) n (%) All subjects 574 (100) 912 (100) Age, yers (58.7) 516 (56..6) (41.3) 396 (43.4) Gender Mle 361 (62.9) 610 (66.9) Femle 213 (37.1) 302 (33.1) Tumor site Crdi 214 (37.3) Other site 360 (62.7) Pthologicl type Diffuse 328 (57.1) Intestinl 246 (42.9) Two-sided chi-squre test Tble 2. Primry informtion nd minor llele frequencies (MAFs) of selected SNPs SNP Gene Position Bse chnge HWE MAF in our controls rs PIK3R1 Exon G >A rs PIK3R1 3 UTR G >A rs PIK3R3 Promoter A>G rs AKT1 Exon A>G rs mtor Promoter T>G HWE: Hrdy-Weinberg equilibrium, MAF: minor llele frequency, SNP: single nucleotide polymorphism Tble 3. Univrite logistic regression nlysis for ssocitions between selected SNPs nd gstric cncer risk SNP Genotype Cses (%) Controls (%) Adjusted OR (95%CI) rs GG 307(53.5) 472(51.8) 1 p vlue AG 211(36.8) 358(39.3) 0.91( ) AA 56(9.8) 82(9.0) 1.03( ) Additive model 0.98( ) rs GG 161(28.0) 304(33.3) 1 AG 279(49.5) 427(46.8) 0.97( ) AA 101(22.5) 181(19.8) 0.94( ) Additive model 0.94( ) rs AA 194(33.8) 297(32.6) 1 GA 279(48.6) 441(48.4) 1.15( ) GG 101(17.6) 174(19.1) 1.26( ) Additive model 1.16( ) rs AA 177(30.8) 293(32.1) 1 GA 284 (49.5) 464(50.9) 1.01( ) GG 113(19.7) 155(17.0) 1.09( ) Additive model 1.08( ) rs TT 194(33.8) 297(32.6) 1 TG 279(48.6) 441(48.4) 0.81( ) GG 101(17.6) 174(19.1) 0.75( ) Additive model 0.78( ) Adjusted by ge nd sex JBUON 2017; 22(6): 1490

4 PI3K/AKT/mTOR polymorphism in gstric cncer 1491 Tble 4. Strtifiction nlysis for ssocition between vrint genotypes nd gstric risk Vribles rs Adjusted OR p rs Adjusted OR p Age, yers Sex Cses Controls (95%CI) Cses Controls (95%CI) AA /AG/GG AA /AG/GG TT/TG/GG TT/TG/GG /156/81 174/239/ ( ) >65 61/128/48 130/188/ ( ) Mle 61/103/49 102/148/ ( ) Femle 104/181/76 202/279/ ( ) Tumor site Crdi 60/106/48 304/427/ ( ) Other site 105/178/77 304/427/ ( ) Pthologicl type Diffuse 94/161/73 304/427/ ( ) Intestinl 71/123/52 304/427/ ( ) Adjusted by ge nd sex /122/20 265/208/ ( ) /9 204/163/ ( ) /251/49 159/120/ ( ) /140/15 119/480/ ( ) /81/13 469/371/ ( ) /126/78 213/131/ ( ) /127/20 469/371/ ( ) /85/9 469/371/ ( ) Tble 5. Combined effects of rs nd rs on gstric cncer risks No. of risk llele Cses Controls Adjusted OR(95%CI) b p b n (%) n (%) (14.8) 178 (19.5) (36.4) 345 (37.8) 1.27 ( ) (35.7) 295 (32.3) 1.47 ( ) (13.1) 94 (10.3) 1.67 ( ) Trend 1.18 ( ) Binry clssifiction (51.2) 523 (57.3) (48.8) 389 (42.7) 1.28 ( ) The rs G nd rs T llele were ssumed s risk lleles bsed on min effect of individul locus; b Adjusted by ge nd sex Discussion In this study, the ssocition of PI3K/AKT/ mt0r signling pthwy gene polymorphism with the risk of gstric cncer in Chinese popultion ws investigted bsed on the Thousnd Tlents Progrm Dtbse using the cndidte genes. The results showed tht PIK3R3 rs nd mtor rs were ssocited with the onset risk of gstric cncer. In the present study reported were the correltions between the loci rs nd rs nd the genetic susceptibility to gstric cncer. There is no report on PIK3R3 rs in cncer, but rs hs been reported in prostte cncer [11] nd colorectl cncer [10]. Hildebrndt et l. [14] reported the mtor rs polymorphism in 2009 for the first time nd discussed the effect of this locus in the prognosis of esophgel cncer ptients in Europe nd the United Sttes who received rdiotherpy nd chemotherpy, but no correltion between rs polymorphism nd the prognosis of ptients with esophgel cncer ws found [14]. Co et l. found in Chinese popultion tht the risk of renl cell crcinom in individuls crrying rs TG/GG genotype ws significntly reduced compred with tht in individuls crrying TT genotype (OR=0.74, 95%CI= ); the mtor mrna of renl cncer tissues crrying G llele ws significntly decresed, nd the G llele could reduce the mtor trnscriptionl ctivity nd ffect the expression JBUON 2017; 22(6): 1491

5 1492 PI3K/AKT/mTOR polymorphism in gstric cncer of mtor [15]. Chen et l. found in Chinese popultion tht the risk of prostte cncer in individuls crrying rs TG/GG genotype ws significntly reduced compred with tht in individuls crrying TT genotype (OR=0.77, 95%CI= ) [16]. In ddition, Xu et l. hve lso found tht the rs polymorphism plys similr role in colorectl cncer [17]. The results of the present study re consistent with those previous reports. In number of cncer studies it hs been shown tht PI3K/AKT/mTOR signling pthwy does not ct independently s bridge molecule connecting extrcellulr signling nd intrcellulr response effects, but ffects the downstrem signling molecules nd the cell growth, prolifertion, poptosis, migrtion nd other physiologicl functions under the combined ction of mny upstrem nd/or bypss signling molecules, thus ffecting the occurrence nd regression of cncer. Tribbles pseudokinse-3 (TRIB3) is tumor suppressor gene tht cn inhibit the expression of AKT. Downregultion of TRIB3 expression cn promote the phosphoryltion of AKT protein, enhnce the ctivity of AKT downstrem trget gene FOXO3 nd Bcl2 ssocited gonist of cell deth (BAD) nd promote the tumor growth [18]. Qiu et l. found tht the silencing of eukryotic elongtion fctor 1 lph2 (eeflα2) cn inhibit the ctivity of PI3K/AKT/mT0R signling pthwy, led to prolifertion, migrtion nd invsion of heptocellulr crcinom cells nd cuse cell cycle rrest, so eeflα2 my ply role s proto-oncogene through the PI3K/AKT/mT0R signling pthwy [19]. AKT is lowly expressed in the AKT-knockout liver cell lines, which cn directly ct on BAD, upregulte the BAD expression nd promote poptosis of heptocellulr crcinom cells [20]. There re lso some shortcomings in this study. First, the smple size ws too smll; considering the power of sttisticl nlysis, some SNPs tht met the screening criteri but with too smll MAF were not included in the study. In ddition, the power of the results might be ffected becuse of the too smll smple size, so the results need to be verified in future lrger popultion-bsed smples. Second, only SNPs in the functionl region were selected in this study, so smll number of potentil ssocitions with gstric cncer might be ignored. In ddition, the study popultion ws from the hospitl, so selective nd informtion bis were inevitble. Conclusions In conclusion, the results of our study showed tht there is significnt correltion between the polymorphisms of the locus rs in the PIK3R3 gene promoter region nd the locus rs in the mtor gene promoter region nd the risk of gstric cncer in Chinese popultion. Therefore, rs nd rs re expected to be useful moleculr biomrkers of gstric cncer, but the possible moleculr mechnism of the SNPs of the two loci in the occurrence nd development of gstric cncer needs further study. Conflict of interests The uthors declre no conflict of interests. References 1. Torre LA, Bry F, Siegel RL, Ferly J, Lortet-Tieulent J, Jeml A. Globl cncer sttistics, CA Cncer J Clin 2015;65: Peek RJ, Blser MJ. Helicobcter pylori nd gstrointestinl trct denocrcinoms. Nt Rev Cncer 2002;2: Tsugne S, Sszuki S. Diet nd the risk of gstric cncer: Review of epidemiologicl evidence. Gstric Cncer 2007;10: Tsugne S. Slt, slted food intke, nd risk of gstric cncer: Epidemiologic evidence. Cncer Sci 2005; 96: Slspuro M. Interctions of lcohol nd tobcco in gstrointestinl cncer. J Gstroenterol Heptol 2012;27 (Suppl 2): Vivnco I, Swyers CL. The phosphtidylinositol 3-Kinse AKT pthwy in humn cncer. Nt Rev Cncer 2002;2: Smuels Y, Wng Z, Brdelli A et l. High frequency of muttions of the PIK3CA gene in humn cncers. Science 2004;304: Grbinski N, Ewld F, Hofmnn BT et l. Combined trgeting of AKT nd mtor synergisticlly inhibits prolifertion of heptocellulr crcinom cells. Mol Cncer 2012;11: Mohn CD, Srinivs V, Rngpp S et l. Trisubstituted-Imidzoles induce poptosis in humn brest cncer cells by trgeting the oncogenic PI3K/Akt/mTOR signling pthwy. PLoS One 2016;11:e Frncipne MG, Lgsse E. MTOR pthwy in colorec- JBUON 2017; 22(6): 1492

6 PI3K/AKT/mTOR polymorphism in gstric cncer 1493 tl cncer: An updte. Oncotrget 2014;5: Chng L, Grhm PH, Ni J et l. Trgeting PI3K/Akt/ mtor signling pthwy in the tretment of prostte cncer rdioresistnce. Crit Rev Oncol Hemtol 2015;96: Li Q, Yng J, Yu Q et l. Associtions between singlenucleotide polymorphisms in the PI3K-PTEN-AKTmTOR pthwy nd incresed risk of brin metstsis in ptients with non-smll cell lung cncer. Clin Cncer Res 2013;19: Lin J, Wng J, Greisinger AJ et l. Energy blnce, the PI3K-AKT-mTOR pthwy genes, nd the risk of bldder cncer. Cncer Prev Res (Phil) 2010;3: Hildebrndt MA, Yng H, Hung MC et l. Genetic vritions in the PI3K/PTEN/AKT/mTOR pthwy re ssocited with clinicl outcomes in esophgel cncer ptients treted with chemordiotherpy. J Clin Oncol 2009;27: Co Q, Ju X, Li P et l. A functionl vrint in the MTOR promoter modultes its expression nd is ssocited with renl cell cncer risk. PLoS One 2012;7: e Chen J, Sho P, Co Q et l. Genetic vritions in PTEN/AKT/mTOR xis nd prostte cncer risk in Chinese popultion. PLoS One 2012;7:e Xu M, Go Y, Yu T et l. Functionl promoter rs T>G vrint in MTOR gene is ssocited with risk of colorectl cncer in Chinese popultion. Biomed Phrmcother 2015;70: Slzr M, Lorente M, Grci-Tbod E et l. Loss of Tribbles pseudokinse-3 promotes Akt-driven tumorigenesis vi FOXO inctivtion. Cell Deth Differ 2015;22: Qiu FN, Hung Y, Chen DY et l. Eukryotic elongtion fctor-1lph 2 knockdown inhibits heptocrcinogenesis by suppressing PI3K/Akt/NF-kppB signling. World J Gstroenterol 2016;22: Hu B, Sun M, Liu J, Hong G, Lin Q. The preventtive effect of Akt knockout on liver cncer through modulting NF-kppB-regulted inflmmtion nd Bd-relted poptosis signling pthwy. Int J Oncol 2016;48: JBUON 2017; 22(6): 1493