Hu Wenwei, Yang Yang, Zhang Longzhen, Yin Jianxin, Huang Jingwei, Huang Lei, Gu Hua, Jiang Gening, Fang Jianmin
School of Life Sciences and Technology, Tongji University, Shanghai, China.
Shanghai Pulmonary Hospital, Shanghai, China.
Cancer Med. 2017 May;6(5):962-974. doi: 10.1002/cam4.980. Epub 2017 Apr 5.
Cancer cells release DNA fragments into plasma as circulating free DNA (cfDNA). However, quantitative measurement of tumor-derived DNA in cfDNA remains challenge. The purpose of this study was to quantitatively assess tumor-derived DNA in lung cancer patients. By optimizing competitive allele-specific TaqMan PCR (CAST-PCR), we assessed the copy number of mutated Kirsten rat sarcoma viral oncogene homolog (KRAS) and epidermal growth factor receptor (EGFR) alleles in the pre/post surgery plasma of 168 lung cancer patients. An absolute quantitative PCR method was developed to assess the number of total cfDNA. All mutations detected in tumors were also found in the plasma after surgery. At the time of 30 days after surgery, EGFR mutation of circulating cell-free DNA was detected only in two patients who recurred in 4 months after surgery. Compared to that of normal control at 30 days after surgery, five patients who recurred in 4 months had significantly higher circulating cell-free DNA (P < 0.001), whereas six patients who recurred after 4 months (P = 0.207) and five patients without recurrence (P = 0.901) demonstrated significantly lower circulating cell-free DNA. Our findings suggest that cfDNA analysis in plasma is an alternative and supplement to tissue analysis and hold promise for clinical application. Stratification of patients according to cfDNA levels at 30 days after surgery might be helpful in selecting lung cancer patients for adjuvant therapy after surgery.
癌细胞会将DNA片段作为循环游离DNA（cfDNA）释放到血浆中。然而，对cfDNA中肿瘤来源DNA进行定量测量仍然具有挑战性。本研究的目的是对肺癌患者的肿瘤来源DNA进行定量评估。通过优化竞争性等位基因特异性TaqMan PCR（CAST-PCR），我们评估了168例肺癌患者手术前后血浆中突变的 Kirsten 大鼠肉瘤病毒癌基因同源物（KRAS）和表皮生长因子受体（EGFR）等位基因的拷贝数。开发了一种绝对定量PCR方法来评估总cfDNA的数量。肿瘤中检测到的所有突变在手术后的血浆中也能找到。在手术后30天时，仅在术后4个月复发的两名患者中检测到循环游离DNA的EGFR突变。与术后30天的正常对照相比，术后4个月复发的5例患者的循环游离DNA显著更高（P < 0.001），而术后4个月后复发的6例患者（P = 0.207）和5例未复发患者（P = 0.901）的循环游离DNA显著更低。我们的研究结果表明，血浆中的cfDNA分析是组织分析的一种替代和补充，具有临床应用前景。根据术后30天的cfDNA水平对患者进行分层，可能有助于选择术后辅助治疗的肺癌患者。
