²¹³Bi-anti-EGFR radioimmunoconjugates and X-ray irradiation trigger different cell death pathways in squamous cell carcinoma cells

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• 作者：Anja Pickhard ; Guido Piontek ; Christof Seidl ; Samuel Kopping ; Birgit Blechert ; Martin Mi脽lbeck ; Gero Brockhoff ; Frank Bruchertseifer ; Alfred Morgenstern ; Markus Essler
• 关键词：Head and neck squamous cell carcinoma ; 伪-emitter 213Bi ; Radioimmunotherapy ; UD-SCC5 cells ; Cytotoxicity ; Cell-cycle arrest ; Apoptosis
• 刊名：Nuclear Medicine and Biology
• 出版年：January, 2014
• 年：2014
• 卷：41
• 期：1
• 页码：68-76
• 全文大小：1376 K

文摘

Introduction

Treatment of patients with squamous cell carcinoma of head and neck is hampered by resistance of tumor cells to irradiation. Additional therapies enhancing the effect of X-ray irradiation may be beneficial. Antibodies targeting EGFR have been shown to improve the efficacy of radiation therapy. Therefore, we analyzed cytotoxicity of ²¹³Bi-anti-EGFR immunoconjugates in combination with X-ray irradiation.

Methods

The monoclonal anti-EGFR antibody matuzumab was coupled to CHX-A鈥?DTPA forming stable complexes with ²¹³Bi. Cytotoxicity of X-ray radiation, of treatment with ²¹³Bi-anti-EGFR monoclonal antibodies (MAb) or of a combined treatment regimen was assayed using cell proliferation and colony formation assays in UD-SCC5 cells. Key proteins of cell-cycle arrest and cell death were examined by Western blot analysis. Cell cycle analysis was performed by flow cytometry. DNA double-strand breaks were detected via 纬H2AX and quantified using Definiens鈩?software.

Results

Irradiation with X-rays or treatment with ²¹³Bi-anti-EGFR-MAb resulted in median lethal dose (LD50) values of 12 Gy or 130 kBq/mL, respectively. Treatment with 37 kBq/mL of ²¹³Bi-anti-EGFR-MAb or 2 Gy of X-rays had only little effect on colony formation of UD-SCC5 cells. In contrast, a combined treatment regimen (37 kBq/mL plus 2 Gy) significantly decreased colony formation and enhanced the formation of DNA double-strand breaks. As revealed by flow cytometry, radiation treatments caused accumulation of cells in the G0/G1 phase. Both treatment with ²¹³Bi-anti-EGFR immunoconjugates and application of the combined treatment regimen triggered activation of genes of signaling pathways involved in cell-cycle arrest and induction of apoptosis like p21/Waf, GADD45, Puma and Bax, which were only marginally modulated by X-ray irradiation of cells.

Conclusions

²¹³Bi-anti-EGFR-MAb enhances cytotoxicity of X-ray irradiation in UD-SCC5 cells most probably due to effective induction of DNA double-strand breaks. Induction of genes involved in cell-cycle arrest and cell death is almost exclusively due to ²¹³Bi-anti-EGFR-MAb and seems to be independent of p53 function.