Study On Predictive Peptide Biomarkers And Models Of Pemetrexed Plus Platinum In Lung Adenocarcinoma

Background:Pemetrexed is a new synthetic third-generation multi-target folic acid analog anti-metabolic drug,it can be converted into active polyglutamic acid salts by transferred into cells via folate carrier and binding protein,which plays an anti-tumor effect by blocking the activity of several key enzymes.The results of JMDB showed that pemetrexed plus cisplatin is comparable to gemcitabine plus cisplatin in NSCLC patients,while the effects of PC is superior to GC in non-squamous NSCLC,and the median OS of patients with lung adenocarcinoma is more than 12 months.Based on the results of this study,pemetrexed plus platinum regimen was approved by US FDA as first-line treatment for patients with non-squamous NSCLC.But,the ORR and m PFS of this regimen were only 37.8% and 5.7 months,indicating that most patients didn't benefit from this kind of regimen,predictive biomarkers are desperately need to enhance the chemotherapy efficacy.Several studies showed that the expression of TS and ERCC1 may relate to the efficacy of pemetrexed and platinum,but all of these studies were retrospective ones and neither of the two biomarker were recommend by guidelines.In clinical practice,there are no biomarkers available to select chemo sensitive patients.With the development of Proteomics and Peptidomics,a lot of biomarkers related to early diagnosis,therapeutic prediction and prognosis were discovered in patients with lung cacner.Several studies showed that serum peptide predictive classification model can predict the effects of EGFR-TKIs,but this methods were never used to predict the outcomes of chemotherapy before.Chapter 1Observation of effects and safety of pemetrexed plus platinum as first-linetreatment in patients with advanced lung adenocarcinoma Objective:To observe the clinical effects and safety of pemetrexed plus platinum as first-line treatment in patients with advanced lung adenocarcinoma,and analysis the relationship between clinical characters,treatment and therapeutic effects.Methods:We retrspectively searched all clinical,imaging and follow-up data of lung cancer patients who were admitted to the Department of Lung Cancer Medcine,Affiliated Hospital of PLA Academy of Military Medical Sciences from January 2012 to July 2015.All patients were diagnosed with advanced lung adenocarcinoma with good PS(0~2),at least one lesion was measurable,radiotherapy of the brain were also acceptable.All patients received pemetrexed(500mg/m2)combined with cisplatin(75mg/m2)or carboplatin(AUC=5)after treatment with folic acid tablet,vitamin B12 and hexadecadrol pretreatment.Each patient received at least two cycles(1 cycle = 3weeks)of chemotherapy and radiographic evaluation was done every two cycles,all patients received up to 4-6 cycles of thermotherapy(unless progression and intolerable toxities).The main objectives of this study were to investigate the objective response rate,disease control rate,progression-free survival,overall survival and related side effects of pemetrexed combined with platinum regimen in patients with lung adenocarcinoma.The effects of clinical factors and treatment on efficacy and survival were also analyzed.Results:1.236 patients were enrolled according the criteria.There were 142 males and 94 females;the median age is 58 years(33~78);22 and 214 patients had stage of ?B or ?;109 patients had a history of smoking and 127 had no or light smoking history;all patients had good PS scores.192 patients had mutation tests and 24 and 8 patients had EGFR or ALK mutations,respectively.2.All patients received 2 cycles' chemotherapy and evaluation.There were no complete remission patients,87 patients had PR,101 patients had SD,and 48 patients had PD.The ORR was 36.9 %(87/236),the DCR was 89.7 %(188/236),the m PFS was 6.0 months and the OS was 13.7 months.3.Subgroup stratification analysis were performed according to age,sex,history of smoking,the kind of platinum,maintenance therapy,anti-angiogenesis therapy.There were no difference with age and kind of platinum.We found that there was a significant difference of ORR between non-smoking and smoking patients(42.5% vs.28.4%,P=0.025),and OS between male and female patients(14.5 months vs.13.4 months,P=0.043)was also different,and patients received endostatin had relatively longer PFS(7.0 months vs.6.0 months,P=0.006),and there were significant difference of PFS(8.5 months vs.6.3 months,P=0.046)and m OS(16.7 months vs.13.7 months,P=0.027)between patients did or did't receive pemetrexed maintenance therapy.Patients with ALK+ had relatively higher efficacy and the DCR and median PFS were 87.5% and 8.3 months.The COX regression analysis showed that only maintenance therapy is associated with longer PFS and OS.Common adverse events were neutropenia(48.3%),leukpenia(38.6%),anemia(22%),thrombcytopenia(9.3%),nausea(62.3%),vomitting 31.8%),fatigue(44%),hair loss(14%).Conclusion:PC regimen is effective as first-line treatment for lung adenocarcinoma with good tolerance and safety.Age and the kind of platinum had similar results.Non-smokers and combination of endostatin had the tendency for better response and logner PFS,women had the tendency to obtain longer OS,both the univariate and COX analysis showed that patients received pemetrexed maintenance therapy had longer PFS and OS.Patients with ALK+ had relatively higher DCR and longer PFS,the ALK mutation status may related to the efficacy of pemetrexed plus platinum.Chapter 2Peptidome study on predictive peptide biomarkers and models of pemetrexed plus platinum in patients with lung adenocarcinoma Objective:The objective of this study was to search potential peptide biomarkers of pemetrexed plus platinum regimen in patients with lung adenocarcinoma using matrix-assisted laser desorption/ ionization time of flight mass spectrometry(MALDI-TOF MS),and construct a serum prediction model.Then the predictive model was tested to evaluated and confirm its capability of discriminating patients with different clinical outcomes and eventually help clinicians to select appropriate chemosnsitive patients to this regimen.Methods:Patients visited and treated at oncology department of Affiliated Hospital of PLA Academy of Military Medical Sciences from Dec.2012 to Nov.2014 were screened and enrolled in this study.All patients were diagnosed with advanced lung adenocarcinoma and received first-line pemetrexed plus cisplatin or carboplatin with definite treatment outcome evaluation.Patients were randomized into training and validation set according to treatment outcome and gender.All serum samples were collected before treatment.In the trainsing set,samples were subjected to MALDI-TOF MS analysis,and then the peptodome profiles were used for searching biomarkers and model construction.In validation set,each sample was classified into “good” or “poor” outcome group using the predictive peptide model.The clinical outcomes of ORR,DCR,PFS and OS were analyzed based on classification results.The selected peptide peaks were identified by LTQ-Orbitrap.The matched peptides were identified according to the results of protein screening.Results:1.183 patients were enrolled.Most of the patients had stage IV disease and an Eastern Cooperative Oncology Group performance status of 1.More men smokerswere enrolled.61,79 and 43 had PR,SD and PD,respectively.2.According to response and sex,patients were randomized into training set and validation set.The basic characteristics were in balance.The peptidome MS data were compared between the good and poor outcome groups.A total of 136 peptide peaks were detected,and eight peaks were significantly different between good and poor outcome groups.Eight peaks are highly expressed(m/z 3316.19,6624.02,2142.12,4281.94,3773.02,3029.28,3955.87 and 3323.95 Da)in good group,which can potentially be biomarkers.3.The Clin Prot Tools v3.0 was used for model construction and a predictive classification model was constructed using quick classifier.The model consists of four pepetide peaks(m/z:2142.12,3316.19,4281.94,6624.02 Da),the CVV and recognition capability of this model were 91.74% and 94.74%?4.Then the model was then tested in the validation set.All the samples were successfully classified as “good” 55(60.4%)or “poor” 36(39.6%).As a result,in 30 patients with PR,27 were classified as “good” and 3 “poor”,in 40 patients with SD,26 were classified as “good” and 14 as “poor”.In 21 patients with PD,19 and 3 patents were classified as “poor” or “good”,respectively.We found that the chemo good group exhibited significantly higher ORR(49.1% vs.8.3%,P < 0.001),DCR(96.4% vs.47.2%,P < 0.001)and better PFS(7.3 months [95% CI: 6.735–7.865 months] vs.2.7 months [95% CI: 0.939–4.461 months],P < 0.001).However,the peptide model did not predict OS(13.6 months [95% CI: 11.109–16.091 months] vs.12.7 months [95% CI: 10.201–15.199 months],P = 0.0675).5.We used LTQ-Orbitrap to identify the 8 peptides and only two of the them(2142.12 Da and 3316.19 Da)were successfully revealed as the sequences of K.AVEYYFASDASAVIEHTNR.V and K.NGVDGVYSADPNKDASAVKFDTLTHLDIINK.G,respectively.These sequences correspond to fragments of glucosamine-fructose-6-phosphate aminotransferase and uridylate kinase,respectively.Conclusion:1?The peptidome profiles were significantly different in adenocarcinoma patients with different outcome to pemetrexed plus platinum regimen,all of the 8 peaks(m/z: 3316.19,6624.02,2142.12,4281.94,3773.02,3029.28,3,955.87 ? 3323.95Da)were elevated in patients with good clinical outcome,these peptides may be potential predictive biomarkers for pemetrexed plus platinum regimen.2?Clin Prot Tools software were used to construct a predictive peptide model based on four peaks(m/z:2142.12,3316.19,4281.94,6624.02 Da),the validation results showed that this model can accurately discriminate chemo sensitive patients,further study is needed to confirm this findings.3 ? Two of the eight peaks were successfully identified as fragments of glucosamine-fructose-6-phosphate aminotransferase and uridylate kinase,they probably are potential predictive biomarkers of pemetrexed plus platinum regimen..