| Part One The relevance of advanced non-small cell lung cancer metastasis sites with plasma EGFR mutant typesObjectiveAnalyze the relevance of advanced non-small cell lung cancer(NSCLC)metastasis sites with plasma EGFR mutant types,to demonstrate whether EGFR mutant types could be biomarkers for the prognoses of advanced NSCLC metastasis.Methods1.One-hundred and five advanced NSCLC patients,be hospitalized in Qilu Hospital of Shandong University or Liaocheng People's Hospital,were enrolled in this study.Fasting blood of these patients were sampled and transferred to anticoagulant tubes,and in 2 hours after sampled,the samples were subjected to 10 minutes,2,00g centrifuge for plasm sepreation.Twenty-four of these 105 patients'postoperative specimens were collected and stored in-80?.2.Plasma DNA were extracted using a plasma DNA extracting kit,and the tissue DNA were extracted using a tissue DNA extrationg kit.Amplification refractory mutation system polymerase chain reaction(ARMS-PCR)was used for EGFR mutant detection.EGFR mutants of EGFR19-Del.21-L858R and 20-T790M in plasma and tumor tissue specimens were comparied.The relevance of metastasis site with EGFR mutant types was analysed.Results1.The mutant rate of EGFR in plasma was 41.6%(10/24),and the mutant rate of EGFR in tissue specimens was 50.0%(12/24).EGFR 19-Del.21-L858R and 20-T790M mutant rates in plasma were]2.5%(3/24),29.17%(7/24)and 0%(0/24);EGFR 19-Del?21-L858R and 20-T790M mutant rates in tissue specimens were 16.67%(4/24),33.33%(8/24)and 0%(0/24).The mutant of EGFR in plasma was consistent with that in tissue specimens.2.Plasma EGFR mutant rate of patients with lymph node metastasis,bone metastasis,pleura metastasis and brain metastasis were 15.15%(5/33),43.59%(17/39),45.00%(9/20),69.23%(9/13)respectively.The rates of each mutant types showed no difference in patients with lymph node metastasis;mutant rate of EGFR 21-L858R was higher than EGFR 19-Del and 20-T790M in patients with bone metastasis;mutant rate of EGFR 21-L858R was higher than EGFR 19-Del and 20-T790M in patients with pleura metastasis;The rates of each mutant types showed no difference in patients with brain metastasis.3.EGFR mutant showed significant relevance with gender and brain metastasis:EGFR mutant rate in female patients(63.64%)were higher than that in male patients(21.31%);EGFR mutant rate in patients with brain metastasis(69.23%)were higher than those without brain metastasis(33.70%).EGFR 19-Del mutant showed significant relevance with clinical stage,brain metastasis:the EGFR 19-Del rate in patients of stage ?(2.41%)was lower than that in stage ?(45.45%).The EGFR 19-Del rate in patients with brain metastasis(38.46%)were higher than those without brain metastasis(7.61%).EGFR 21-L858R mutant showed significant relevance with gender:EGFR 21-L858R mutant of male patients(9.84%)was lower than that of'female patients(47.73%).ConclusionThe consistence of EGFR mutant in plasma and tissue specimens suggested plasma EGFR mutant could be replacement of tissue specimens EGFR as biomarkers of advanced NSCLC.The revelance of EGFR 19-Del,21-L858R mutation with clinicopathologic feature indicated EGFR 19-Del,21-L858R mutation could be biomarkers for predicting advanced NSCLC metastasis and for making individual therapy strategy.Part Two The relevance between EGFR mutants of advanced NSCLC with metastasis sites and serum tumor biomarkersObjectiveAnalyze the relevance of metastasis sites,plasma EGFR mutant types with serum tumor biomarkers(CEA,CA125,CYFRA21-1,NSE and SA)in advanced NSCLC,to ellustrate whether serum tumor biomarkers could be used for predicting the metastasis site and EGFR mutant type in advanced NSCLC patients.Methods1.Sample collection:Five milliliter fasting blood of these patients were sampled and transferred to plain tubes,and after the blood clotting,the samples were subjected to 10 minutes-3,000r/min centrifu ge for serum sepreation.2.Serum concentrations of carcino-embryonic antigen(CEA),carbohydrate antigen 125(CA125),cytokeratin fragment 21-1(CY21-1),neuronspecific enolase(NSE)and sialic acid(SA)were detected using chemiluminescent immunoassay.3.Tumor biomarkers concentrations were presented in Median(Quartile).Mann-Whitney U test was used to compare the defference of serum tumor biomarkers between different EGFR mutants or between different metastasis sites.Results1.CEA concentration of advanced NSCLC with bone metastasis(57.180 ng/ml(11.000,166.590))was higher than that of pleura metastasis(18.560 ng/ml(11.800,71.040))and brain metastasis((8.450 ng/ml(3.510,50.550)).Whereas,other tumor biomarkers including CA125?CY21-1?NSE?SA showed no difference between advanced NSCLCs with bone metastasis,pleura metastasis and brain metastasis.2.CEA of NSCLC with EGFR mutant(29.26 ng/ml(14.98,163.88))was higher than NSCLC without EGFR mutant(16.68 ng/ml(6.48,43.38)).All the tumor biomarkers showed no difference between advanced NSCLC with EGFR 19-Del and advanced NSCLC without EGFR mutant.CEA concentration of advanced NSCLC with EGFR 21-L858Rmutant(62.89 ng/ml(15.64,166.34))was higher than advanced NSCLC without EGFR mutant(16.68 ng/ml(6.48,43.38)),whereas,other tumor biomarkers including CA125?CY21-1?NSE?SA showed no difference between advanced NSCLC with 21-L858R mutant and advanced NSCLC without EGFR mutant.3.Among advanced NSCLC with EGFR mutant,CA125 concentrations of bone metastasis(47.450 U/ml(25.998,103.025))and pleura metastasis(66.300 U/ml(38.250,132.450))were much higher than that of brain metastasis(19.900 U/ml(8.725,30.230)),whereas,other biomarkers including CEA,CY21-I.NSE.SA showed no difference.ConclusionCEA concentration of advanced NSCLC with bone metastasis was higher than that of pleura metastasis and brain metastasis,CEA showed a positive correlation with EGFR 21-L858R mutation.CEA concentration may be prognostic marker of advanced NSCLC with bone metastasis and EGFR 21-L858R mutant.Among advanced NSCLC with EGFR mutant,CA125 concentrations of bone metastasis and pleura metastasis were much higher than that of brain metastasis.The increased CA125 concentration of advanced NSCLC with bone metastasis or pleura metastasis may prognosis EGFR mutation.This study indicated that CEA and CA125 may be used promising markers for advanced NSCLC EGFR mutant type and metastasis sites.however,further studies are needed to confirm the feasibility of clinical use. |
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