ErbB-2 Blockade and Prenyltransferase Inhibition Alter Epidermal Growth Factor and Epidermal Growth Factor Receptor Trafficking and Enhance In-111-DTPA-hEGF Auger Electron Radiation Therapy

TY - JOUR

T1 - ErbB-2 Blockade and Prenyltransferase Inhibition Alter Epidermal Growth Factor and Epidermal Growth Factor Receptor Trafficking and Enhance In-111-DTPA-hEGF Auger Electron Radiation Therapy

AU - Cornelissen, Bart

AU - Darbar, Sonali

AU - Hernandez, Rebecca

AU - Kersemans, Veerle

AU - Tullis, Iain

AU - Barber, Paul R.

AU - Smart, Sean

AU - Vojnovic, Boris

AU - Reilly, Raymond

AU - Vallis, Katherine A.

PY - 2011/5

Y1 - 2011/5

N2 - The intracellular distribution of Auger electron-emitting radiopharmaceuticals is a determinant of cytotoxicity. However, the mechanisms by which these agents are routed through the cell are ill understood. The aim of this study was to investigate how trafficking of In-111-labeled human epidermal growth factor (In-111-DTPA-hEGF) relates to that of the EGF receptor (EGFR) and whether coadministration of agents that modulate EGFR signaling alters the efficacy of In-111-DTPA-hEGF. Methods: The spatiotemporal interaction between AlexaFluor488-EGF (AF488-EGF) and Cy3-conjugated anti-EGFR antibody (Cy3-anti-EGFR) was studied in the breast cancer cell line MDA-MB-468 using fluorescence resonance energy transfer and 2-photon fluorescence lifetime imaging. In-111 internalization and nuclear fractionation assays were performed to investigate the effect of the ErbB-2-blocking antibody trastuzumab and a prenyltransferase inhibitor, L-778,123, on the subcellular localization of In-111-DTPA-hEGF in MDA-MB-468 (1.3 x 10(6) EGFR per cell; ErbB-2 negative) and 231-H2N (0.2 x 10(6) EGFR per cell; 0.4 x 10(5) ErbB-2 per cell) cell lines. The cytotoxicity of In-111-DTPA-hEGF (0-64 nM) plus trastuzumab (0-50 mu g/mL) or L-778,123 (0-22.5 mu M) was measured using clonogenic assays in a panel of breast cancer cell lines that express different levels of EGFR and ErB-2. Clonogenic survival data were used to calculate combination indices. Tumor growth inhibition was measured in vivo in 231-H2N xenograft-bearing mice treated with In-111-DTPA-hEGF plus trastuzumab or L-788,123. Results: Using fluorescence resonance energy transfer, we showed that EGF interacts with EGFR in the cytoplasm and nucleus after internalization of the ligand-receptor complex in MDA-MB-468 cells. Nuclear localization of In-111-DTPA-hEGF is enhanced by trastuzumab and L-788,123. Trastuzumab and L-788,123 sensitized 231-H2N cells to In-111-DTPA-hEGF. Nuclear localization and cytotoxicity of In-111-DTPA-hEGF were significantly increased in 231-H2N xenografts by cotreatment with L-788,123 (P <0.0001). Conclusion: The therapeutic efficacy of In-111-DTPA-hEGF is increased through the coadministration of selected molecularly targeted drugs that modulate EGFR signaling and trafficking.

AB - The intracellular distribution of Auger electron-emitting radiopharmaceuticals is a determinant of cytotoxicity. However, the mechanisms by which these agents are routed through the cell are ill understood. The aim of this study was to investigate how trafficking of In-111-labeled human epidermal growth factor (In-111-DTPA-hEGF) relates to that of the EGF receptor (EGFR) and whether coadministration of agents that modulate EGFR signaling alters the efficacy of In-111-DTPA-hEGF. Methods: The spatiotemporal interaction between AlexaFluor488-EGF (AF488-EGF) and Cy3-conjugated anti-EGFR antibody (Cy3-anti-EGFR) was studied in the breast cancer cell line MDA-MB-468 using fluorescence resonance energy transfer and 2-photon fluorescence lifetime imaging. In-111 internalization and nuclear fractionation assays were performed to investigate the effect of the ErbB-2-blocking antibody trastuzumab and a prenyltransferase inhibitor, L-778,123, on the subcellular localization of In-111-DTPA-hEGF in MDA-MB-468 (1.3 x 10(6) EGFR per cell; ErbB-2 negative) and 231-H2N (0.2 x 10(6) EGFR per cell; 0.4 x 10(5) ErbB-2 per cell) cell lines. The cytotoxicity of In-111-DTPA-hEGF (0-64 nM) plus trastuzumab (0-50 mu g/mL) or L-778,123 (0-22.5 mu M) was measured using clonogenic assays in a panel of breast cancer cell lines that express different levels of EGFR and ErB-2. Clonogenic survival data were used to calculate combination indices. Tumor growth inhibition was measured in vivo in 231-H2N xenograft-bearing mice treated with In-111-DTPA-hEGF plus trastuzumab or L-788,123. Results: Using fluorescence resonance energy transfer, we showed that EGF interacts with EGFR in the cytoplasm and nucleus after internalization of the ligand-receptor complex in MDA-MB-468 cells. Nuclear localization of In-111-DTPA-hEGF is enhanced by trastuzumab and L-788,123. Trastuzumab and L-788,123 sensitized 231-H2N cells to In-111-DTPA-hEGF. Nuclear localization and cytotoxicity of In-111-DTPA-hEGF were significantly increased in 231-H2N xenografts by cotreatment with L-788,123 (P <0.0001). Conclusion: The therapeutic efficacy of In-111-DTPA-hEGF is increased through the coadministration of selected molecularly targeted drugs that modulate EGFR signaling and trafficking.

KW - EGF

KW - EGFR

KW - FRET

KW - radioimmunotherapy

KW - trastuzumab

KW - prenyltransferase inhibitor

KW - In-111

KW - Auger electrons

KW - BREAST-CANCER CELLS

KW - IN-VIVO

KW - RAS

KW - TRASTUZUMAB

KW - ENDOCYTOSIS

KW - XENOGRAFTS

KW - KINASE

KW - LOCALIZATION

KW - METHOTREXATE

U2 - 10.2967/jnumed.110.084392

DO - 10.2967/jnumed.110.084392

M3 - Article

SN - 0161-5505

VL - 52

SP - 776

EP - 783

JO - Journal of Nuclear Medicine

JF - Journal of Nuclear Medicine

IS - 5

M1 - N/A

ER -