| Background and objectives:Recently, gefitinib and erlotinib have been successfully used in clinical therapy for non-small cell lung cancer (NSCLC) as the representative of the epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), and become one of the best specific anti-tumor therapies because of their remarkable efficacy and minimal side effects. Choice of targeted drug depends on the detection and confirmation of molecular targets in tumors. It has been confirmed that clinical sensitivity of NSCLC patients applying with molecular targeted therapy such as TKIs, is related closely to EGFR gene mutations. And so, it was necessary to detection for the EGFR mutations before targeted therapy of EGFR-TKIs. To some extent, it promoted cancer therapeutics from the traditional "treatment response mode" to "individual forecasting model". Gene sequence analysis is a classical technique for EGFR gene mutations detection, and has been widespread recognized by clinical ace demists. More and more molecular diagnostic techniques had emerged currently, with increased sensitivity and specificity for gene mutations. However, most of them require tumor specimens from surgical resection or biopsy, even the amount of tumor cells. Much to regret, most of lung cancer patients lost the chance of operation in diagnosis duration, even difficultly to obtain tissue specimens by needle biopsy. In addition, tomor tissue specimens have great limitations of reflecting EGFR mutations because of the dynamic of tumor cell activities, especially after the impact of radiotherapy and chemotherapy.It reported that there were little free DNA in serum of cancer patients, which mainly from apoptosis and necrosis tumor of cells shed from the primary site and enter the blood circulation in solid tumors. More than 90% of apoptosis cells released genetic material. Therefore, it has become a mainstream idea that to forecast the sensitivity and efficacy of TKIs non-invasive and dynamically, such as test EGFR gene mutations from the peripheral blood of lung cancer patients. As we all know, EGFR gene mutations were found only in somatic cells of tumor-specific genetic, especially in NSCLC, and there is no such mutations in normal cells. It is clear that EGFR mutations might be detected from the free DNA in peripheral blood, if these mutations existed in tumor sites of primary or metastatic NSCLC patients. However, such relative research reports are still rare at present. In 2009, Rosell reported that he and his partners first detected EGFR gene mutations from peripheral blood of NSCLC patients by using of real-time fluorescence quantitive polymerase chain reaction (FQ-PCR) method. They found 97 cases (59%) positive mutations in peripheral blood within 164 NSCLC patients with EGFR mutations in tumor tissues. So far, it has not yet reported that the application researches of FQ-PCR for detecting EGFR mutations from peripheral blood of NSCLC patients in our country, even more, lack of evidence-based medical information from clinical paired sample studies. Based of aboved all, the purpose of this study is to test EGFR gene mutations from tumor tissues and peripheral blood specimens in NSCLC patients by using of FQ-PCR, and to investigate the availability of FQ-PCR technique for forecasting the sensitivity and monitoring therapeutic effects of EGFR-TKIs in NSCLC. Furthermore, provide some value evidences for clinical evaluation of EGFR mutation status in molecular targeted therapy.Method:1. 112 peripheral blood and 87 tumor tissue of NSCLC were detected by using of FQ-PCR, 45 cases in which matched peripheral blood and tumor tissue. 29 patients EGFR mutations that were detected by FQ-PCR were sequenced. Meanwhile, we analyzed the relationship between EGFR gene mutation and clinical feature in peripheral blood and tumor tissue of NSCLC patients, furthermore to compare the consistency of EGFR gene mutations in peripheral blood and tumor tissue.2. 61 patients of this group received gefitinib or erlotinib treatment, 24 patients were also detected in peripheral blood and tumor tissue EGFR mutations, 21 cases were detected only in peripheral blood and 16 cases detected in tumor tissue alone. We evaluated the relationship between EGFR mutations and clinical of gefitinib or erlotinib.Results:1. The characteristics of NSCLC patients EGFR mutations in tumor tissues and the characteristics and the relationship with clinical features(1). EGFR mutations of peripheral blood was significantly higher in adenocarcinoma than non-adenocarcinoma (38.2% vs8.3%, P <0.01), significantly higher non-smoking than in smoking patients (41.0% vs 13.7%, P <0.01). More than 60 years of age and less than 60 years, EGFR mutations had no significant difference (22.6% vs33.9%, P> 0.05), and no significant difference between groups in men and women (22.0% vs35.8% , P> 0.05).(2). EGFR mutations of tumor tissue was significantly higher in adenocarcinoma than non-adenocarcinoma (45.0% vs. 19.1%,P<0.01), significantly higher non-smoking than in smoking patients (41.0% vs. 13.7%, P <0.01). More than 60 years of age and less than 60 years, EGFR mutations had no significant difference (26.3% vs 34.7%,P>0.05), and no significant difference between groups in men and women (24.5% vs39.5%,P>0.05).2. The consistency of NSCLC patients with EGFR mutations in peripheral blood and tumor tissue 10 cases were detected EGFR mutations at the same time, in which 45 cases matched peripheal blood and tumor tissue, and the type of EGFR mutations were identical completely. The consistency of EGFR mutations with tumor tissue and peripheal blood reached 71.4%.3. The relationship between clinical efficacy and EGFR mutations of tumor tissue and peripheral blood in NSCLC.61 patients of this study received EGFR-TKI treatment. The ORR and DCR of NSCLC with EGFR mutations in peripheral blood were 52.0% and 78.1% respectively, and effective was statistically significant difference between EGFR mutations and wild-type(P <0.001). The ORR and DCR of NSCLC with EGFR mutations in tumor tissue were 55.6% and 81.5% respectively, and effective was statistically significant difference between EGFR mutations and wild-type(P <0.01). The DCR of patients with EGFR mutations in peripheral blood and tumor tissue receiving TKI treatment was significantly higher than EGFR wild-type patients.Conclusions:1. It was suggested that EGFR gene mutations were detected sensitively from peripheral blood by using of FQ-PCR, which have high consistency to the positive mutation rate in tumor tissues in NSCLC patients. With feature of highly specific and sensitivity, non-invasive, real-time, dynamic and easily accepted, and so on, this technology is suitable for widely development in clinical laboratory, and might be used as an effective choice to forecast the sensitivity and monitor therapeutic effects of EGFR-TKIs in NSCLC.2. EGFR gene mutations tested either from peripheral blood or from tumor tissues were closely related to clinical features, such as adenocarcinoma and non-smoking in NSCLC, further confirmed the view that "Asian, adenocarcinoma and non-smoking patients are more advantage groups from TKIs targeted therapy".3. The DCR of NSCLC patients with EGFR gene mutations in peripheral blood and tumor tissues was significantly higher than that one with EGFR wild-type after TKIs treatment, suggested again that EGFR gene mutations might be effective predict factor of therapeutic effects of EGFR-TKIs molecular target therapy in NSCLC. |
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