CI-101检测肺癌患者外周血循环肿瘤细胞水平及其临床意义

国际肿瘤学杂志››2020,Vol. 47››Issue (4): 199-204.doi:10.3760/cma.j.cn371439-20200116-00002

CI-101检测肺癌患者外周血循环肿瘤细胞水平及其临床意义

田翎含¹, 朱玲², 刘馨¹, 叶静雯¹, 何嘉慧¹, 王熙才¹()

¹昆明医科大学第三附属医院云南省肿瘤医院肿瘤研究所 650118
²昆明市第一人民医院肿瘤科 650034

收稿日期:2020-01-16修回日期:2020-03-11出版日期:2020-04-08发布日期:2020-05-26
通讯作者:王熙才 E-mail:wangxc2005323@126.com
基金资助:
国家自然科学基金(81460358);云南省卫生科技计划(2017NS201);云南省卫生科技计划(2018NS0078);云南省科技计划(2017FE467(-189));云南省科技计划(2014FB066)

Level of peripheral blood circulating tumor cells in lung cancer patients detected by CI-101 and its clinical significance

Tian Linghan¹, Zhu Ling², Liu Xin¹, Ye Jingwen¹, He Jiahui¹, Wang Xicai¹()

¹Cancer Research Institute, Third Affiliated Hospital of Kunming Medical University (Yunnan Cancer Hospital), Kunming 650118, China
²Department of Oncology, First People's Hospital of Kunming, Kunming 650034, China

Received:2020-01-16Revised:2020-03-11Online:2020-04-08Published:2020-05-26
Contact:Wang Xicai E-mail:wangxc2005323@126.com
Supported by:
National Natural Science Foundation of China(81460358);Yunnan Provincial Health Technology Plan(2017NS201);Yunnan Provincial Health Technology Plan(2018NS0078);Yunnan Provincial Science and Technology Plan(2017FE467(-189));Yunnan Provincial Science and Technology Plan(2014FB066)

摘要/Abstract

摘要：

目的应用CI-101细胞搜寻免疫磁珠富集技术结合荧光细胞化学染色方法,探讨肺癌患者外周静脉血中循环肿瘤细胞(CTC)水平与肺癌患者临床病理特征及生物标志物之间的相关性。方法采集2014年3月至2014年9月云南省肿瘤医院胸外科和胸心外科收治的100例首诊肺癌患者、40例肺部良性肿瘤患者(均经过病理活检确诊)及体检中心30例健康志愿者的血样。应用CI-101细胞搜寻免疫磁珠富集外周血中的CTC,应用免疫细胞荧光技术对CTC行形态学分析,并用HE细胞染色方法鉴定肿瘤细胞。检测CI-101细胞搜寻仪对CTC的回收率、灵敏度、特异度;比较肺癌患者、肺部良性肿瘤患者及健康志愿者外周血CTC阳性率的区别;分析肺癌患者CTC阳性率与患者临床病理特征之间的关系;采用列联系数分析肺癌患者及肺部良性肿瘤患者CTC与血清肿瘤标志物之间的相关性。结果用CI-101细胞搜寻仪检测CTC回收率为72.0%~89.0%,且回收的细胞数与掺入细胞数有显著线性相关性,相关系数r=0.998(P<0.001),线性回归方程为:y=0.781x+11.307,灵敏度为85.0%,特异度为71.4%。肺癌患者组CTC阳性率(85.0%,85/100)高于肺部良性肿瘤患者组(15.0%,6/40)及健康志愿者组(46.7%,14/30)(χ²=62.798,P<0.001);CTC阳性率与肺癌患者TNM分期(χ²=19.059,P<0.001)、肿瘤大小(χ²=13.830,P<0.001)、有无远处转移(χ²=6.005,P=0.014)均相关。肺癌患者及肺部良性肿瘤患者CTC阳性与血清肿瘤标志物CEA(φ=0.217,P=0.011)、CA125(φ=0.198,P=0.020)、CA199(φ=0.169,P=0.049)、CA742(φ=0.186,P=0.037)、细胞角蛋白19片段(φ=0.461,P<0.001)呈正相关。结论应用CI-101细胞搜寻仪联合免疫磁珠法可成功富集肺癌患者外周静脉血中的CTC,肺癌患者的CTC阳性率与患者的肿瘤大小、TNM分期、有无远处转移、血清肿瘤标志物有明显相关性,可作为肺癌患者病情监测的辅助指标。

关键词:肿瘤细胞,循环,肺肿瘤,液态活检

Abstract:

ObjectiveTo explore the correlations between circulating tumor cells (CTCs) level in peripheral venous blood and clinicopathological characteristics and biomarkers of lung cancer patients using CI-101 cell search immunomagnetic bead enrichment technology combined with fluorescent cytochemical staining.MethodsBlood samples were collected from 100 patients with first-diagnosed lung cancer treated in Department of Thoracic Surgery and Department of Cardiothoracic Surgery of Yunnan Cancer Hospital from March 2014 to September 2014, 40 patients with lung benign tumor (all confirmed by pathological biopsy) and 30 healthy volunteers from the physical examination center. CTCs in peripheral blood were enriched by CI-101 cell search immunomagnetic bead, the morphology of CTCs was analyzed by immunocytofluorescence technique, and tumor cells were identified using HE cell staining method. The recovery rate, sensitivity and specificity of CI-101 cell search instrument for CTCs were detected. The difference of positive rate of CTCs in peripheral blood among lung cancer patients, lung benign tumor patients and healthy volunteers was compared. The relationship between the positive rate of CTCs and the clinicopathological characteristics of patients with lung cancer was analyzed. The correlations between CTCs and serum tumor markers were analyzed by coefficient of contingency in patients with lung cancer and lung benign tumor.ResultsThe recovery rate of CTCs by CI-101 cell search instrument was 72.0%-89.0%, and there was a significant linear correlation between the number of recovered cells and the number of incorporated cells. The correlation coefficientr=0.998 (P<0.001), the linear regression equation wasy=0.781x+11.307, the sensitivity was 85.0%, and the specificity was 71.4%. The positive rate of CTCs in lung cancer patients (85.0%, 85/100) was higher than that in lung benign tumor patients (15.0%, 6/40) and healthy volunteers (46.7%, 14/30) (χ²=62.798,P<0.001). The positive rate of CTCs in lung cancer patients was correlated with TNM stage (χ²=19.059,P<0.001), tumor size (χ²=13.830,P<0.001) and distant metastasis (χ²=6.005,P=0.014). Coefficient of contingency analysis showed that the positive of CTCs was positively correlated with serum tumor markers CEA (φ=0.217,P=0.011), CA125 (φ=0.198,P=0.020), CA199 (φ=0.169,P=0.049), CA742 (φ=0.186,P=0.037) and cytokeratin 19 fragment (φ=0.461,P<0.001) in patients with lung cancer and lung benign tumor.ConclusionThe application of CI-101 cell search instrument combined with immunomagnetic bead method can successfully enrich CTCs in peripheral venous blood of lung cancer patients. The positive rate of CTCs in patients with lung cancer has obvious correlation with tumor size, TNM stage, distant metastasis, serum tumor markers. It can be used as an auxiliary indicator for monitoring the condition of lung cancer patients.

Key words:Neoplastic cells,circulating,Lung neoplasms,Liquid biopsy

田翎含, 朱玲, 刘馨, 叶静雯, 何嘉慧, 王熙才. CI-101检测肺癌患者外周血循环肿瘤细胞水平及其临床意义[J]. 国际肿瘤学杂志, 2020, 47(4): 199-204.

Tian Linghan, Zhu Ling, Liu Xin, Ye Jingwen, He Jiahui, Wang Xicai. Level of peripheral blood circulating tumor cells in lung cancer patients detected by CI-101 and its clinical significance[J]. Journal of International Oncology, 2020, 47(4): 199-204.

图/表6

图1

图2

表1

5例健康志愿者外周血中掺入不同数量的小细胞肺癌NCI-H446细胞后CI-101细胞搜寻仪的细胞回收情况"

掺入细胞数(个)	回收细胞数(个, $x ˉ$ ±s)	回收率(%)
50	36±6	72.0
100	85±12	85.0
200	178±15	89.0
500	420±30	84.0
1 000	782±45	78.0

表1

组别	CTC检出数	H值	P值
肺癌患者组(n=100)	58(42)^ab
肺部良性肿瘤患者组(n=40)	26(8)	51.874	<0.001
健康志愿者组(n=30)	26(23)

表2

表3

100例肺癌患者外周血中CTC阳性率与临床病理特征的关系"

临床病理特征	CTC阳性 (n=85)	CTC阴性 (n=15)	阳性率 (%)	χ²值	P值
年龄(岁)
<50	17	1	94.4	0.765	0.382
≥50	68	14	82.9
性别
男性	59	11	84.2	0.093	0.760
女性	26	4	86.7
TNM分期
Ⅰ~Ⅱ期	14	11	50.0	19.059	<0.001
Ⅲ~Ⅳ期	71	4	94.4
肿瘤大小(cm)
<3	15	10	57.7	13.830	<0.001
≥3	70	5	86.5
病理类型
鳞状细胞癌	14	3	82.3	2.152	0.348
腺癌	44	10	81.5
其他	27	2	93.1
远处转移
有	72	8	77.5	6.005	0.014
无	13	7	45.0

表3

表4

140例肺癌患者及肺部良性肿瘤患者CTC阳性与血清肿瘤标志物的关系(例)"

肿瘤标志物^a	CTC阳性	CTC阴性	φ值	P值
CEA
阳性	40	49	0.217	0.011
阴性	11	37
CA125
阳性	36	53	0.198	0.020
阴性	10	38
CA153
阳性	2	64	-0.117	0.211
阴性	4	44
CA199
阳性	19	69	0.169	0.049
阴性	4	44
CA242
阳性	16	72	0.164	0.055
阴性	3	45
CA742
阳性	20	62	0.186	0.037
阴性	4	40
细胞角蛋白19片段
阳性	55	27	0.461	<0.001
阴性	8	35
铁蛋白
阳性	38	45	0.060	0.503
阴性	17	26

表4

参考文献17

[1]	Balata H, Fong KM, Hendriks LE , et al. Prevention and early detection for non-small cell lung cancer: advances in thoracic oncology 2018[J]. J Thorac Oncol, 2019,14(9):1513-1527. DOI: 10.1016/j.jtho.2019.06.011. doi:10.1016/j.jtho.2019.06.011
[2]	Siegel RL, Miller KD, Jemal A . Cancer statistics, 2018[J]. CA Cancer J Clin, 2018,68(1):7-30. DOI: 10.3322/caac.21442. doi:10.3322/caac.21442
[3]	Bray F, Ferlay J, Soerjomataram I , et al. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin, 2018,68(6):394-424. DOI: 10.3322/caac.21492. doi:10.3322/caac.v68.6
[4]	Perumal V, Corica T, Dharmarajan AM , et al. Circulating tumour cells (CTC), head and neck cancer and radiotherapy; future perspectives[J]. Cancers (Basel), 2019,11(3):367. DOI: 10.3390/cancers11030367. doi:10.3390/cancers11030367
[5]	Lozar T, Gersak K, Cemazar M , et al. The biology and clinical potential of circulating tumor cells[J]. Radiol Oncol, 2019,53(2):131-147. DOI: 10.2478/raon-2019-0024. doi:10.2478/raon-2019-0024
[6]	Bailey PC, Martin SS . Insights on CTC biology and clinical impact emerging from advances in capture technology[J]. Cells, 2019,8(6):553. DOI: 10.3390/cells8060553. doi:10.3390/cells8060553
[7]	Truini A, Alama A, Dal Bello MG , et al. Clinical applications of circulating tumor cells in lung cancer patients by CellSearch system[J]. Front Oncol, 2014,4:242. DOI: 10.3389/fonc.2014.00242.
[8]	Adams DL, Stefansson S, Haudenschild C , et al. Cytometric characterization of circulating tumor cells captured by microfiltration and their correlation to the CellSearch(®) CTC test[J]. Cytometry A, 2015,87(2):137-144. DOI: 10.1002/cyto.a.22613. doi:10.1002/cyto.a.22613
[9]	秦子敏, 杨丁, 何山宏 , 等. 循环肿瘤细胞在肺癌诊断中的应用及临床意义[J]. 基因组学与应用生物学, 2019,38(7):3228-3232. DOI: 10.13417/j.gab.038.003228.
[10]	Myung JH, Park SJ, Wang AZ , et al. Integration of biomimicry and nanotechnology for significantly improved detection of circulating tumor cells (CTCs)[J]. Adv Drug Deliv Rev, 2018,125:36-47. DOI: 10.1016/j.addr.2017.12.005. doi:10.1016/j.addr.2017.12.005
[11]	Huo J, Chung TH, Kim B , et al. Provider-patient discussions about smoking and the impact of lung cancer screening guidelines: NHIS 2011-2015[J]. J Gen Intern Med, 2020,35(1):43-50. DOI: 10.1007/s11606-019-05111-6. doi:10.1007/s11606-019-05111-6
[12]	Li Y, Tian X, Gao L , et al. Clinical significance of circulating tumor cells and tumor markers in the diagnosis of lung cancer[J]. Cancer Med, 2019,8(8):3782-3792. DOI: 10.1002/cam4.2286.
[13]	Sharma P, Mehta M, Dhanjal DS , et al. Emerging trends in the novel drug delivery approaches for the treatment of lung cancer[J]. Chem Biol Interact, 2019,309:108720. DOI: 10.1016/j.cbi.2019.06.033. doi:10.1016/j.cbi.2019.06.033
[14]	吴方, 赵京, 张侠 , 等. 循环肿瘤细胞与肿瘤标志物在肺癌诊断中的价值比较[J]. 国际检验医学杂志, 2020,41(5):527-531. DOI: 10.3969/j.issn.1673-4130.2020.05.004.
[15]	Maly V, Maly O, Kolostova K , et al. Circulating tumor cells in diagnosis and treatment of lung cancer[J]. In Vivo, 2019,33(4):1027-1037. DOI: 10.21873/invivo.11571. doi:10.21873/invivo.11571
[16]	Hanssen A, Wagner J, Gorges TM , et al. Characterization of different CTC subpopulations in non-small cell lung cancer[J]. Sci Rep, 2016,6:28010. DOI: 10.1038/srep28010. doi:10.1038/srep28010
[17]	Tong B, Xu Y, Zhao J , et al. Prognostic significance of circulating tumor cells in non-small cell lung cancer patients undergoing chemotherapy[J]. Oncotarget, 2017,8(49):86615-86624. DOI: 10.18632/oncotarget.21255. doi:10.18632/oncotarget.v8i49