Intratumoral heterogeneity of EGFR-activating mutations in advanced NSCLC patients at the single-cell level
Longhua Guo,
Zhihong Chen,
Chongrui Xu,
Xuchao Zhang,
Honghong Yan,
Jian Su,
Jinji Yang,
Zhi Xie,
Weibang Guo,
Feng Li,
Yilong Wu,
Qing Zhou
Affiliations
Longhua Guo
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Zhihong Chen
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Chongrui Xu
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Xuchao Zhang
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Honghong Yan
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Jian Su
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Jinji Yang
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Zhi Xie
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Weibang Guo
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Feng Li
Department of Medical Oncology, Cancer Center, Meizhou People’s Hospital (Huangtang Hospital), Meizhou Hospital Affiliated to Sun Yat-Sen University
Yilong Wu
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Qing Zhou
Guangdong Lung Cancer Institute, Guangdong Provincial Key Laboratory of Translational Medicine in Lung Cancer, Guangdong Provincial People’s Hospital and Guangdong Academy of Medical Sciences
Abstract Background Intratumoral epidermal growth factor receptor (EGFR) mutational heterogeneity is yet controversial in non-small cell lung cancer (NSCLC) patients. Single-cell analysis provides the genetic profile of single cancer cells and an in-depth understanding of the heterogeneity of a tumor. Methods Firstly, single H1975 cells harboring the EGFR L858R mutation were submitted to flow cytometry isolation, nested polymerase chain reaction (nested-PCR) amplification, and direct DNA sequencing to assess the feasibility of single-cell direct DNA sequencing. Then, the single cells of patients with lung adenocarcinoma receiving gefitinib were captured by laser capture microdissection and analyzed by the above methods to identify the intratumoral heterogeneity of the EGFR L858R mutant. Three patients with progression-free survival (PFS) > 14 months were categorized as the long PFS group, and 3 patients with PFS < 6 months as the short PFS group. The correlation between the abundance of EGFR L858R mutant and PFS was analyzed. Results 104 single H1975 cells were isolated. 100/104 were amplified by nested-PCR and confirmed by direct sequencing. We captured 135 tumor cells from the tissues of six patients. 120 single tumor cells were successfully amplified and sequenced. The rate of EGFR exon 21 mutation was only 77.5% (93/120). Furthermore, the rate of mutation in exon 21 of EGFR was significantly higher in the long PFS group than in the short PFS group (86.4 ± 4.9% vs. 68.9 ± 2.8%, P = 0.021). Conclusion Our study suggested the intratumoral heterogeneity of EGFR-activating mutations in lung adenocarcinoma confirmed on the single-cell level, which might be associated with EGFR-TKIs response in lung adenocarcinoma patients harboring the EGFR L858R mutation.
