Abstract | UNLABELLED: METHODS: Western blot analysis was used to determine the optimum concentration of gefitinib to abolish EGFR activation. Internalization and nuclear translocation of fluorescein isothiocyanate-labeled hEGF were evaluated by confocal microscopy in MDA-MB-468 cells (1.3 x 10(6) EGFRs/cell) in the presence or absence of 1 microM gefitinib. The proportion of radioactivity partitioning into the cytoplasm and nucleus of MDA-MB-468 cells after incubation with (111)In-DTPA-hEGF for 24 h at 37 degrees C in the presence or absence of 1 microM gefitinib was measured by cell fractionation. DNA double-strand breaks caused by (111)In were quantified using the gamma-H2AX assay, and radiation-absorbed doses were estimated. Clonogenic survival assays were used to measure the cytotoxicity of (111)In-DTPA-hEGF alone or in combination with gefitinib. RESULTS:
Gefitinib (1 microM) completely abolished EGFR phosphorylation in MDA-MB-468 cells. Internalization and nuclear translocation of fluorescein isothiocyanate-labeled EGF were not diminished in gefitinib-treated cells compared with controls. The proportion of internalized (111)In that localized in the nucleus was statistically significantly greater when (111)In-DTPA-hEGF was combined with gefitinib compared with (111)In-DTPA-hEGF alone (mean +/- SD: 26.0% +/- 5.5% vs. 14.6% +/- 4.0%, respectively; P < 0.05). Induction of gamma-H2AX foci was greater in MDA-MB-468 cells that were treated with (111)In-DTPA-hEGF (250 ng/mL, 1.5 MBq/mL) plus gefitinib (1 microM ) compared with those treated with (111)In-DTPA-hEGF alone (mean +/- SD: 35 +/- 4 vs. 24 +/- 5 foci per nucleus, respectively). In clonogenic assays, a significant reduction in the surviving fraction was observed when (111)In-DTPA-hEGF (5 ng/mL, 6 MBq/microg) was combined with gefitinib (1 microM ) compared with (111)In-DTPA-hEGF alone (42.9% +/- 5.7% vs. 22.9% +/- 3.6%, respectively; P < 0.01). CONCLUSION:
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Authors | Kristy E Bailey, Danny L Costantini, Zhongli Cai, Deborah A Scollard, Zhuo Chen, Raymond M Reilly, Katherine A Vallis |
Journal | Journal of nuclear medicine : official publication, Society of Nuclear Medicine
(J Nucl Med)
Vol. 48
Issue 9
Pg. 1562-70
(Sep 2007)
ISSN: 0161-5505 [Print] United States |
PMID | 17704253
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
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Chemical References |
- 111In-DTPA-human epidermal growth factor
- Antineoplastic Agents
- Fluorescent Dyes
- Indium Radioisotopes
- Quinazolines
- Radiopharmaceuticals
- Epidermal Growth Factor
- Pentetic Acid
- ErbB Receptors
- Fluorescein-5-isothiocyanate
- Gefitinib
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Topics |
- Antineoplastic Agents
(pharmacokinetics, pharmacology)
- Cell Compartmentation
- Cell Line, Tumor
- Cell Nucleus
(metabolism)
- Electrons
- Epidermal Growth Factor
(metabolism, pharmacokinetics, pharmacology)
- ErbB Receptors
(antagonists & inhibitors, metabolism)
- Fluorescein-5-isothiocyanate
- Fluorescent Dyes
- Gefitinib
- Humans
- Indium Radioisotopes
- Pentetic Acid
(analogs & derivatives, pharmacokinetics, pharmacology)
- Phosphorylation
- Protein Transport
- Quinazolines
(pharmacology)
- Radiopharmaceuticals
(pharmacokinetics, pharmacology)
- Signal Transduction
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