用于检测非小细胞肺癌患者白细胞分离产物中循环肿瘤细胞的微筛

Microsieves for the detection of circulating tumor cells in leukapheresis product in non-small cell lung cancer patients.

作者信息

Tamminga Menno, Oomens Lisa, Hiltermann T Jeroen N, Andree Kiki C, Tibbe Arjan, Broekmaat Joska, Schuuring Ed, Terstappen Leon W M M, Groen Harry J M

机构信息

Department of Pulmonary Diseases, University Medical Center Groningen, University of Groningen, Groningen, The Netherlands.

VyCAP, Enschede, Groningen, The Netherlands.

出版信息

Transl Lung Cancer Res. 2020 Aug;9(4):1093-1100. doi: 10.21037/tlcr-19-413.

DOI:10.21037/tlcr-19-413

PMID:32953488

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC7481649/

Abstract

BACKGROUND

Circulating tumor cells (CTC) in non-small cell lung cancer (NSCLC) patients are a prognostic and possible therapeutic marker, but have a low frequency of appearance. Diagnostic leukapheresis (DLA) concentrates CTC and mononuclear cells from the blood. We evaluated a protocol using two VyCAP microsieves to filter DLA product of NSCLC patients and enumerate CTC, compared with CellSearch as a gold standard.

METHODS

DLA was performed in NSCLC patients before starting treatment. DLA product equaling 2×10 leukocytes was diluted to 9 mL with CellSearch dilution buffer in a Transfix CTC tube. Within 72 hours the sample was filtered with a 7 µm pore microsieve and subsequently over a 5µm pore microsieve. CTC were defined as nucleated cells which stained for cytokeratin, but lacked CD45 and CD16. CellSearch detected CTC in the same volume of DLA.

RESULTS

Of 29 patients a median of 1.4 mL DLA product (range, 0.5-4.1) was filtered (2% of total product) successfully in 93% and 45% of patients using 7 and 5 µm pores, respectively. Two DLA products were unevaluable for CTC detection. Clogging of the 5 µm but not 7 µm microsieves was positively correlated with fixation time (ρ=0.51, P<0.01). VyCAP detected CTC in 44% (12/27) of DLA products. Median CTC count per mL DLA was 0 [interquartile range (IQR): 0-1]. CellSearch detected CTC in 63% of DLA products (median =0.9 CTC per mL DLA, IQR: 0-2.1). CTC counts detected by CellSearch were significantly higher compared with VyCAP (P=0.05).

CONCLUSIONS

VyCAP microsieves can identify CTC in DLA product, but workflows need to be optimized.

摘要

背景

非小细胞肺癌（NSCLC）患者循环肿瘤细胞（CTC）是一种预后指标及可能的治疗标志物，但出现频率较低。诊断性白细胞分离术（DLA）可从血液中富集CTC和单核细胞。我们评估了一种使用两个VyCAP微筛过滤NSCLC患者DLA产物并计数CTC的方案，并与作为金标准的CellSearch进行比较。

方法

在NSCLC患者开始治疗前进行DLA。将相当于2×10白细胞的DLA产物在Transfix CTC管中用CellSearch稀释缓冲液稀释至9 mL。在72小时内，先用7 µm孔径的微筛过滤样品，随后再用5 µm孔径的微筛过滤。CTC定义为细胞角蛋白染色阳性、但缺乏CD45和CD16的有核细胞。CellSearch在相同体积的DLA中检测CTC。

结果

29例患者中，分别有93%和45%的患者使用7 µm和5 µm孔径成功过滤了中位数为1.4 mL的DLA产物（范围为0.5 - 4.1）（占总产物的2%）。有两份DLA产物无法用于CTC检测。5 µm微筛而非7 µm微筛的堵塞与固定时间呈正相关（ρ = 0.51，P < 0.01）。VyCAP在44%（12/27）的DLA产物中检测到CTC。每毫升DLA中CTC计数的中位数为0 [四分位间距（IQR）：0 - 1]。CellSearch在63%的DLA产物中检测到CTC（每毫升DLA中位数 = 0.9个CTC，IQR：0 - 2.1）。与VyCAP相比，CellSearch检测到的CTC计数显著更高（P = 0.05）。