Hur Jae Young, Lee Jong Sik, Kim In Ae, Kim Hee Joung, Kim Wan Seop, Lee Kye Young
Precision Medicine Lung Cancer Center, Konkuk University Medical Center, Seoul, Republic of Korea.
Department of Pathology, Konkuk University School of Medicine, Seoul, Republic of Korea.
Transl Lung Cancer Res. 2019 Dec;8(6):1051-1060. doi: 10.21037/tlcr.2019.12.16.
Extracellular vesicles (EV) have been proven to contain double-stranded DNA reflecting the mutational status of the parental tumor cells in non-small cell lung cancer (NSCLC), which can be translated into clinically useful EV-based liquid biopsy for Epidermal growth factor receptor (EGFR) genotyping using bronchoalveolar lavage fluid (BALF) obtained from tumor site.
Patients subjected for an initial lung cancer work-up underwent bronchoscopy and BALF was obtained from tumor site. After isolating EVs from BALF by ultracentrifugation, EV-derived DNA (EV DNA) was extracted for subsequent EGFR genotyping performed through peptide nucleic acid (PNA)-mediated Real-Time PCR. The sensitivity, specificity, and concordance rate of BALF EV-based EGFR genotyping were calculated in comparison to tissue genotyping.
The average sensitivity and specificity of BALF EV-based EGFR genotyping were 76% and 87%, respectively, while the sensitivity significantly increased as the stage progressed. Especially, in stage IV, BALF EV-based EGFR typing identified all tissue-proven EGFR mutant cases (n=31) and detected 6 additional mutant cases. The concordance rate was 79% in stage I, 100% in stage II, 74% in stage III, and 92% in stage IV. As TNM stage advanced, especially in the presence of metastasis, concordance rate significantly increased (P<0.05).
The use of BALF for the collection of EV DNA in lung cancer patients resulted in a highly accurate diagnosis. The establishment of a fast and reliable method to identify target genes using EV DNA illustrated that it can overcome the problems of low sensitivity and instability in using cell-free DNA (cfDNA).
细胞外囊泡（EV）已被证明含有反映非小细胞肺癌（NSCLC）亲本肿瘤细胞突变状态的双链DNA，这可以转化为临床上有用的基于EV的液体活检，用于使用从肿瘤部位获得的支气管肺泡灌洗液（BALF）进行表皮生长因子受体（EGFR）基因分型。
接受初始肺癌检查的患者接受支气管镜检查，并从肿瘤部位获取BALF。通过超速离心从BALF中分离出EV后，提取EV衍生的DNA（EV DNA），用于随后通过肽核酸（PNA）介导的实时PCR进行EGFR基因分型。与组织基因分型相比，计算基于BALF EV的EGFR基因分型的敏感性、特异性和符合率。
基于BALF EV的EGFR基因分型的平均敏感性和特异性分别为76%和87%，而敏感性随着分期进展而显著增加。特别是在IV期，基于BALF EV的EGFR分型识别出所有经组织证实的EGFR突变病例（n = 31），并检测到另外6例突变病例。I期的符合率为79%，II期为100%，III期为74%，IV期为92%。随着TNM分期进展，尤其是在存在转移的情况下，符合率显著增加（P<0.05）。
使用BALF收集肺癌患者的EV DNA可实现高度准确的诊断。建立一种使用EV DNA识别靶基因的快速可靠方法表明，它可以克服使用游离DNA（cfDNA）时敏感性低和稳定性差的问题。
