Anna Sjöström

HER-2--a possible target for therapy of metastatic urinary bladder carcinoma.

Human epidermal growth factor receptor 2, HER-2, is overexpressed in various tumours, e.g. breast- and bladder tumours. The aim of this study was to predict the potential use of HER-2 receptors as targets in systemic treatment of disseminated bladder tumours. HER-2 expression was assessed in bladder carcinoma metastases and the corresponding primary tumours, and subsequently compared with the EGFR expression. HER-2 and EGFR expression was analysed by immunohistochemistry in formalin-fixed, paraffin-embedded tissues from 21 patients with metastatic bladder carcinoma. HER-2 was overexpressed in 81% of the primary tumours and in 67% of the metastases. All HER-2-positive metastases were from HER-2-positive primary tumours. The results for EG FR were 71% of both primary and metastases-positive tumours. In 90% of the primary tumours and 86% of the metastases, at least one of the receptors was overexpressed. These results suggest that HER-2 targeted therapy can be considered as an alternative or a complement to other modalities in the treatment of metastatic urinary bladder carcinoma.

Expression of epidermal growth factor receptor in urinary bladder cancer metastases

Bladder cancers frequently exhibit an increased number of epidermal growth factor receptors (EGFR) in comparison to normal urothelium. The EGFR could potentially be a target for toxic conjugates. The aim of our study was to compare the expression of EGFR in metastases with concurrent or primary tumour in the urinary bladder using immunohistochemical techniques and a monoclonal antibody. Tumour material from 20 patients was investigated. The majority (13/20) of the metastases were homogeneously stained and showed a moderate to strong membranous staining for EGFR. The expression of EGFR in primary bladder tumours and metastases was similar. There was no indication that tumour tissue exposed to chemotherapy or radiation had a decreased number of EGFR. Targeting of the EGFR thus seems potentially applicable to metastatic disease. Int. J. Cancer 76:189–193, 1998.© 1998 Wiley‐Liss, Inc.

Cellular processing of 125I- and 111in-labeled epidermal growth factor (EGF) bound to cultured A431 tumor cells

Low molecular weight of epidermal growth factor (EGF) enables better intratumoral penetration in comparison with larger targeting proteins, but the cellular retention of EGF-associated radioactivity is poor for directly iodinated EGF. An attempt was made to improve intracellular retention by the use of metal-diethylenetriaminepentaacetic acid or nonphenolic linker (N-succinimidyl-para-iodobenzoate) as labeling agents. The use of nonphenolic linker did not improve retention of the radioactivity in A431 carcinoma cell line. The use of the radiometal label provided an appreciable prolongation of radioactivity residence inside the cell.

Direct astatination of a tumour-binding protein, human Epidermal Growth Factor, using nido-carborane as a prosthetic group

We have investigated a method for direct astatine labeling of proteins. Binding sites for astatine were created by coupling of a nido-carborane derivative to a protein, the human epidermal growth factor (hEGF), using two different conjugation methods - by glutaraldehyde cross-linking or by introduction of sulfohydryl groups by Trauts reagent with subsequent linking of ANC-1 with m-maleimidobenzoyl-N-hydroxysulfosuccinimide ester. The conjugates were astatinated using the Chloramine-T method in high yield. The best labeling was obtained by the glutaraldehyde conjugate with an average yield of 68±9%. In vitro stability tests indicated that the glutaraldehyde conjugated label was as stable as hEGF labeled with astatobenzoate.

Binding, internalization and degradation of EGF-dextran conjugates in two human bladder-cancer cell lines

Bladder carcinomas often have an increased number of epidermal growth factor (EGF) receptors. The EGF receptors can, in these cases, be targets for toxic conjugates. In this study, EGF‐dextran conjugates were used as targeting agents with therapeutic potential. The binding, internalization and degradation of 125I‐EGF‐dextran conjugates in J82 and RT4, 2 different bladder cancer cell lines, were investigated. The behaviour of 125I‐EGF was studied as a comparison. The 125I‐EGF‐dextran showed a continuous increase in binding up to 24 hr, while 125I‐EGF exhibited maximum binding after about 90 min. Both cell lines showed similar binding patterns. The binding of 125I‐EGF‐dextran and 125I‐EGF was, on both cell lines, receptor‐specific since the binding could be displaced with non‐radioactive EGF. Analysis of internalized and membrane‐bound 125I activity after administration of 125I‐EGF‐dextran showed that most of the activity was membrane‐bound. A large part of both the internalized and the membrane‐bound activity remained cell‐associated up to 24 hr. The internalized and membrane‐bound activity after administration of 125I‐EGF decreased rapidly and only a small fraction remained cell‐associated after 24 hr. 125I‐EGF‐dextran remained cell‐associated, with only a limited release of low‐ and high‐molecular‐weight radioactivity throughout the 24‐hr period, while 125I‐EGF was extensively degraded and released into the incubation medium as low‐molecular‐weight radioactivity during this time. Several qualities of the 125I‐EGF‐dextran conjugates might be favourable for targeted radiotherapy. Int. J. Cancer, 70:383–389, 1997.

Effects of dextranation on the uptake of peptides in micrometastases:studies on binding of EGF in tumor spheroids.

Four different types of radiolabelled dextranated EGF were added to spheroids consisting of the human glioma cells U-343MGaCl2:6. Binding was analysed both in the peripheral well-nourished regions and in the deeper regions containing mainly quiescent cells. The substances had different molecular weights, and they were characterized regarding hydrophilic properties and isoelectric points. Two of the analysed conjugates, 125I-EGF-dextran (CDAP) and 125I-EGF-allyldextran-BSH, gave very low 125I binding in the deeper regions even after 24 h of incubation while better binding in these regions was found for 125I-EGF-dextran-DTPA, 125I-EGF-allyldextran and for the reference substance 125I-EGF. The molecular weight seemed not to be of major importance for the binding properties and there were no clear relationships between binding and the hydrophilic properties or the isoelectric point values. The obtained differences could not be explained by differences in molecular weight or easily measured physicochemical parameters such as hydrophilic properties or isoelectric point values. Thus, other explanations must be found.

Direct astatination of a peptide, human epidermal growth factor, using nido-carborane as a prosthetic group

Direct astatination of a peptide, human epidermal growth factor, using nido-carborane as a prosthetic group

Cellular processing of 125I and 111In labeled epidermal growth factor (EGF) bound to cultured A431 tumor cells

Low molecular weight of epidermal growth factor (EGF) enables better intratumoral penetration in comparison with larger targeting proteins, but the cellular retention of EGF-associated radioactivity is poor for directly iodinated EGF. An attempt was made to improve intracellular retention by the use of metal-diethylenetriaminepentaacetic acid or nonphenolic linker (N-succinimidyl-para-iodobenzoate) as labeling agents. The use of nonphenolic linker did not improve retention of the radioactivity in A431 carcinoma cell line. The use of the radiometal label provided an appreciable prolongation of radioactivity residence inside the cell.

Expression of Epidermal Growth Factor Receptor in Urinary Bladder Cancer Metastases

Bladder cancers frequently exhibit an increased number of epidermal growth factor receptors (EGFR) in comparison to normal urothelium. The EGFR could potentially be a target for toxic conjugates. The aim of our study was to compare the expression of EGFR in metastases with concurrent or primary tumour in the urinary bladder using immunohistochemical techniques and a monoclonal antibody. Tumour material from 20 patients was investigated. The majority (13/20) of the metastases were homogeneously stained and showed a moderate to strong membranous staining for EGFR. The expression of EGFR in primary bladder tumours and metastases was similar. There was no indication that tumour tissue exposed to chemotherapy or radiation had a decreased number of EGFR. Targeting of the EGFR thus seems potentially applicable to metastatic disease.

Boron Neutron Capture Therapy Against Tumor Cells with Overexpression of the EGF-Receptor

The binary nature of boron neutron capture therapy, BNCT, is an advantage because the tumor-seeking substance can be activated at any chosen time and because the neutron field can be delivered to selected areas so that exposure of critical healthy organs, which might contain significant amounts of boron, can be avoided. The tumor selective action should work in spite of the fact that tumor cells often have an infiltrative growth pattern being mixed with populations of normal cells. The targeting principle should be based on well-characterized properties of the tumor cells such as appearence of tumor-associated antigens or overexpression of receptors. The targeting agent could be antibodies, antibody-fragments or receptor ligands. Presently, mainly monoclonal antibodies are considered as targeting substances but it has been claimed that current approaches are limited by low uptake in the tumors studied. Thus, it seems necessary to also consider other principles such as growth factor mediated targeting (1–4).