暴露于铟-111标记的二乙三胺五乙酸-人表皮生长因子（¹¹¹In-DTPA-hEGF）的乳腺癌细胞中磷酸化H2AX的诱导与细胞存活之间的关系

Relationship between induction of phosphorylated H2AX and survival in breast cancer cells exposed to 111In-DTPA-hEGF.

作者信息

Cai Zhongli, Chen Zhuo, Bailey Kristy E, Scollard Deborah A, Reilly Raymond M, Vallis Katherine A

机构信息

Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Ontario, Canada.

出版信息

J Nucl Med. 2008 Aug;49(8):1353-61. doi: 10.2967/jnumed.108.051805. Epub 2008 Jul 16.

DOI:10.2967/jnumed.108.051805

PMID:18632822

Abstract

UNLABELLED

The Auger electron-emitting radiopharmaceutical 111In-diethylenetriaminepentaacetic acid human epidermal growth factor (111In-DTPA-hEGF) binds the epidermal growth factor receptor (EGFR), is internalized, and translocates to the nucleus. The purpose of this study was to investigate the relationship between EGFR expression, DNA damage, and cytotoxicity in cells exposed to 111In-DTPA-hEGF.

METHODS

Breast cancer cell lines with a range of EGFR expression levels were exposed to 111In-DTPA-hEGF or gamma-radiation. The cell lines (followed by number of EGFR per cell in parentheses) were MDA-MB-468 (1.3 x 10(6)), MDA-MB-231 (1.3 x 10(5)), and MCF-7 (1.5 x 10(4)). The proportion of radioactivity partitioning into the nucleus was measured by cell fractionation. DNA double-strand breaks were evaluated using the gamma-H2AX assay. Clonogenic survival assays were used to measure cytotoxicity.

RESULTS

All data are presented as mean +/- SD. The amount of 111In-DTPA-hEGF that translocated to the nucleus (in mBq/nucleus) in MDA-MB-468, MDA-MB-231, and MCF-7 cells incubated with 111In-DTPA-hEGF (5.2 MBq/mL, 43 nM) for 20 h was 131 +/- 6, 8.1 +/- 0.1, and 1.1 +/- 0.9, respectively. The number of gamma-H2AX foci per nucleus was 35 +/- 15, 19 +/- 10, and 1.7 +/- 0.3, respectively. A reduction in the surviving fraction (SF) in MDA-MB-468 (0.013 +/- 0.001) and MDA-MB-231 (0.5 +/- 0.1) but not in MCF-7 cells after exposure to 111In-DTPA-hEGF (5.2 MBq/mL, 43 nM) for 20 h has been demonstrated. The SF of MDA-MB-468 cells after exposure to DTPA-EGF (43 nM) and 111In-acetate (5.2 MBq/mL) for 20 h was 0.5 +/- 0.1 and 0.53 +/- 0.05, respectively. MDA-MB-468 was the most sensitive of the cell lines to gamma-irradiation, with an SF after 2 Gy of 0.45 +/- 0.04, compared with 0.7 +/- 0.1 and 0.8 +/- 0.1 for MCF-7 and MDA-MB-231, respectively. The number of gamma-H2AX foci per nucleus in MDA-MB-468 cells correlated with the concentration, specific activity, and incubation time of 111In-DTPA-hEGF.

CONCLUSION

DNA damage caused by 111In-DTPA-hEGF correlates with the EGFR expression level of the exposed cells and with concentration, specific activity, and incubation time of 111In-DTPA-hEGF. The gamma-H2AX assay may be a useful biomarker to predict and monitor the outcome of treatment with 111In-DTPA-hEGF.

摘要

未标记

发射俄歇电子的放射性药物铟-111-二乙烯三胺五乙酸人表皮生长因子（111In-DTPA-hEGF）可结合表皮生长因子受体（EGFR），被内化并转运至细胞核。本研究旨在探讨暴露于111In-DTPA-hEGF的细胞中EGFR表达、DNA损伤与细胞毒性之间的关系。

方法

将一系列EGFR表达水平不同的乳腺癌细胞系暴露于111In-DTPA-hEGF或γ射线。这些细胞系（括号内为每个细胞的EGFR数量）分别为MDA-MB-468（1.3×10⁶）、MDA-MB-231（1.3×10⁵）和MCF-7（1.5×10⁴）。通过细胞分级分离法测量放射性物质在细胞核中的分配比例。使用γ-H2AX检测法评估DNA双链断裂情况。采用克隆形成存活检测法测量细胞毒性。

结果

所有数据均以平均值±标准差表示。将MDA-MB-468、MDA-MB-231和MCF-7细胞与111In-DTPA-hEGF（5.2 MBq/mL，43 nM）孵育20小时后，转运至细胞核的111In-DTPA-hEGF量（以mBq/细胞核计）分别为131±6、8.1±0.1和1.1±0.9。每个细胞核中的γ-H2AX焦点数量分别为35±15、19±10和1.7±0.3。已证明，将MDA-MB-468和MDA-MB-231细胞暴露于111In-DTPA-hEGF（5.2 MBq/mL，43 nM）20小时后，其存活分数（SF）降低，而MCF-7细胞未降低。将MDA-MB-468细胞暴露于DTPA-EGF（43 nM）和铟-111-乙酸盐（5.2 MBq/mL）20小时后的SF分别为0.5±0.1和0.53±0.05。MDA-MB-468是对γ射线最敏感的细胞系，2 Gy照射后的SF为0.45±0.04，而MCF-7和MDA-MB-231分别为0.7±0.1和0.8±0.1。MDA-MB-468细胞中每个细胞核的γ-H2AX焦点数量与111In-DTPA-hEGF的浓度、比活度和孵育时间相关。