| ObjectiveThe aim of the paper is to explore the expression of PITX2 and evaluate the value of PITX2 as a prognostic marker in ?A-N2 NSCLC.MethodsThe clinical datas and tumor tissue specimens of patients diagnosed with ?A-N2 NSCLC between January 1st,2008 and December 1st,2008 were collected,besides all the patients were treated with radical surgery in tianjin medical university cancer hospital.The expression level of PITX2 was examined using genome sequencing and immunohistochemical method.Tumor tissues from 20 cases of ?A-N2 NSCLC were selected.The signals of genes were collected after a series of steps including extracting RNA,reversing transcription synthesis of cDNA,making gene chip,fluorescence labeling and hybridization of gene chip.The expression level of PITX2 in different patients was also analysed.Paraffin-embedded tissues from 77 cases of ?A-N2 NSCLC were selected(including the 20 patients being genome sequenced).The protein expression of PITX2 was measured by immunohistochemical method,and then sored using the microscope.All statistical analyses were performed using SPSS 20.0 software.The chi square test was used to detect differences between groups,survival was estimated using Kaplan-Meier curves and Cox model,and Log-rank method was used to compare differences of survival rates between groups.P < 0.05 were defined as a statistically significant difference.The correlation between PITX2 and clinicopathological characteristics(age,gender,T stage,pathological type,tumor location,the number of positive N2 stations,the number of positive N2 lymph nodes,positive subcarinal lymph node)was analyzed.The study also elucidate the correlation between PITX2 and OS,LRFS and DMFS in ?A-N2 NSCLC.Further more,the correlation between PITX2 and beta-catenin,HDAC1,Akt and mTOR was also examined using immunohistochemical method,in order to explore the mechanism how PITX2 promotes the progression of NSCLC.Results1.For the 77 patients,the 3-and 5-year OS were 60% and 47%,respectively;DFS were 41% and 36% at 3 years and 5 years;LRFS were 78% and 74% at 3 years and 5 years;DMFS were 43% and 41% at 3 years and 5 years.T stage(P=0.018),positive PITX2(P=0.036),number of positive N2 stations(P=0.042),subcarinal lymph node metastasis(P=0.003)are factors influencing OS;T stage(P = 0.001),number of positive N2 stations(P=0.048)and subcarinal lymph node metastasis(P = 0.001)were factors influencing the DFS of patients.2.Whole genome sequencing were carried in 20 cases of NSCLC,which were divided into two groups,one was defined as distant-metastasis group including patients occurring distant metastasis within 1 year,another group was defined as non-distant-metastasis group including patients without occurring distant-metastasis in three years.Compared with patients without distant metastasis,the topest up-regulated 10 genes in distant-metastasis group were XAGE1 B,HAPLN1,CEBPZOS,KIF1 A,NTRK3,TCAM1 P,PITX2,TUBB2 B,DDX3X and PGC.And PITX2 was up-regulated by 18.7-fold change.3.According to the results of immunohistochemical method,positive rate of PITX2 was 41.6% in ?A-N2 NSCLC,which was higher in patients with adenocarcinoma,positive N2 stations ?3 and positive subcarinal lymph node than patients with squamous cell carcinoma ? positive N2 stations <3 and negative subcarinal lymph node,respectively(57.8% vs 18.8%,P=0.001)(68.2% vs 30.6%,P=0.011;68.2% vs 31.6%,P=0.011)(57.1% vs 28.6%,P=0.012).Positive PITX2 rates of patients with positive-N2 nodes number of 1-3,4-6 and 7 were 28.9%,52.4% and 72.7%,respectively(P = 0.015).4.The 5 years OS and DMFS were superior in PITX2-negative patients to that of PITX2-positive patients(60.0% vs 37.5%,P = 0.036)(51.1% vs 28.1,P = 0.042).No significant difference was seen between the two groups in 5 years LRFS(80% vs 75%,P = 0.576).5.Positive rates of PITX2 were 56.4%(31/55)and 4.5%(1/22)(P=0.000),respectively in patients with positive and negative expression of beta-catenin.In patients with positive expression of PITX2 or not,the high expression rates of HDAC1 were 90.6%(29/32)and 62.2%(28/45)(P= 0.005),Akt-positive rates were 78.1%(25/32)and 55.6%(25/45)(P= 0.041),mTOR-positive rates were 56.3% and 33.3%(P= 0.045),respectively.Conclusions1.Over-expression of PITX2 can be detected in NSCLC,and the positive expression rate of PITX2 was significantly higher in adenocarcinoma than squamous cell carcinoma.There is no correlation between PITX2 expression and gender,age,T stage and tumor location.2.The positive expression of PITX2 was correlated with multi-level stations of positive N2 nodes,the increase of numbers of positive N2 nodes and the positive subcarinal lymph node,indicating that PITX2 was predictive factor for the characteristics of positive N2 nodes.3.PITX2 has value in predicting failure modes for ?A-N2 NSCLC.The expression of PITX2 was correlated with poor OS and DMFS,and has no correlation with the LRFS.Patients with positive PITX2 expression are favorable to receive postoperative chemotherapy.4.Co-expression of PITX2,?-catenin,HDAC1,Akt and mTOR was observed in NSCLC.More researches are needed to explore details. |
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