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Binding of epidermal growth factor-dextran conjugates to cultured glioma cells
A Andersson1, A Holmberg, J Carlsson
1Department of Radiation Sciences, Uppsala University, Sweden.
Abstract:
Some gliomas, melanomas and squamous carcinomas have large numbers of EGF receptors which could, in these cases, be used for targeting with toxic agents. We investigated whether EGF could be conjugated to dextran, which is a suitable carrier for toxic agents, without losing its ability to bind to the receptor. Dextran of 20 kDa molecular weight was activated with I-cyano-4-dimethylamino pyridinium tetrafluoroborate (CDAP) yielding highly active pyridinium-isourea derivatives. EGF was coupled to the activated dextran through the amino terminus and glycine was added to block residual activity. The EGF-dextran conjugate was, after purification on Sephadex G25 and Sephacryl 200 columns, tested for its receptor binding properties on human malignant glioma, U343MGaC12:6, cells. The conjugate inhibited binding of 125I-EGF in a competitive assay, showing that the binding was receptor-specific. Dextran conjugated with glycine, without EGF, had no inhibitory effect. The conjugate was radio-labelled either on the EGF part with 125I or on the dextran part with 3H-glycine, and the internalization patterns were compared to the internalization of 125I-EGF. The radioactivity of the conjugates remained cell-associated for more than 20 hr, regardless of whether the radioactivity was on the EGF or on the dextran part, while the radioactivity of unconjugated EGF rapidly disappeared from the cells. Most of the cell-associated radioactivity was, at all analysed time intervals, located intracellularly. Thus, it seems promising to use dextran, conjugated with EGF, as a carrier of, for example, toxic radioactive nuclides.
Insights
This study shows that epidermal growth factor (EGF) can be conjugated to dextran, creating a carrier for targeted delivery of toxic agents to cancer cells expressing EGF receptors. The conjugate retains receptor binding and is internalized by cells, suggesting potential for cancer therapy.
Area of Science:
- Bioconjugation Chemistry
- Molecular Targeting
- Cancer Therapeutics
Background:
- Certain cancers like gliomas and melanomas overexpress Epidermal Growth Factor (EGF) receptors.
- This overexpression presents an opportunity for targeted drug delivery using EGF as a targeting moiety.
Purpose of the Study:
- To investigate the conjugation of EGF to dextran, a potential carrier for toxic agents.
- To evaluate the receptor binding and cellular internalization of the EGF-dextran conjugate.
Main Methods:
- Dextran activation using CDAP and subsequent conjugation with EGF.
- Purification of the conjugate using Sephadex G25 and Sephacryl 200 chromatography.
- Assessment of receptor binding via competitive assays and internalization studies using radiolabeled conjugates.
Main Results:
- The EGF-dextran conjugate demonstrated specific binding to EGF receptors on human malignant glioma cells.
- The conjugate showed prolonged cell association (>20 hr) and significant intracellular localization.
- Unconjugated EGF exhibited rapid clearance from cells, unlike the conjugate.
Conclusions:
- EGF-dextran conjugates are capable of specific receptor binding and cellular internalization.
- Dextran conjugated with EGF shows promise as a carrier system for targeted delivery of therapeutic agents, such as radioactive nuclides, to cancer cells.