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Ex vivo delivery of suicide genes into melanoma cells using epidermal growth factor receptor-specific Fab immunogene
1Department of Molecular Biology, Keio University School of Medicine, Tokyo.
Abstract:
The Fab fragment of monoclonal antibody B4G7 against human epidermal growth factor (EGF) receptor was conjugated with cationic poly-L-lysine and the resulting conjugate was further complexed with reporter genes or therapeutic genes. This Fab/DNA complex was designated as "Fab immunogene." The Fab immunogene transfer in vitro was mediated through the EGF receptors in two melanoma cell lines. The frequency of cells expressing beta-galactosidase (beta-Gal) reporter gene was approximately 1%. The induction of suicide effects after Fab immunogene transfer of herpes simplex virus thymidine kinase (TK) or Escherichia coli cytosine deaminase (CD) gene was quite remarkable, and the growth of melanoma cells was inhibited for over 7 days in the presence of ganciclovir (GCV) or 5-fluorocytosine (5-FC). Similarly, when melanoma cells treated in vitro with the Fab immunogene carrying TK or CD were transplanted into the back of nude mouse, subsequent systemic administration of GCV or 5-FC effectively suppressed the growth of tumors, indicating the occurrence of in vivo suicide effects.
Insights
Researchers developed a "Fab immunogene" for targeted cancer therapy. This novel system effectively delivered therapeutic genes to melanoma cells, inhibiting tumor growth both in vitro and in vivo.
Area of Science:
- Biotechnology
- Molecular Biology
- Cancer Research
Background:
- Monoclonal antibody fragments (Fab) can target specific cell receptors.
- Gene therapy aims to deliver genetic material for therapeutic effects.
- Melanoma is a significant form of skin cancer with unmet therapeutic needs.
Purpose of the Study:
- To develop a novel gene delivery system targeting the epidermal growth factor (EGF) receptor.
- To evaluate the efficacy of this system in delivering reporter and therapeutic genes to melanoma cells.
- To assess the in vitro and in vivo anti-cancer effects of the gene delivery system.
Main Methods:
- Conjugation of B4G7 Fab fragment (targeting EGF receptor) with poly-L-lysine.
- Complexation of the conjugate with reporter (beta-galactosidase) or therapeutic genes (HSV-TK, E. coli-CD).
- In vitro gene transfer to melanoma cell lines and in vivo studies using nude mouse xenografts.
Main Results:
- Successful creation of the "Fab immunogene" complex for gene delivery.
- Demonstrated EGF receptor-mediated gene transfer into melanoma cells with ~1% reporter gene expression.
- Significant inhibition of melanoma cell growth in vitro (7+ days) and tumor suppression in vivo upon administration of specific drugs (ganciclovir or 5-fluorocytosine).
Conclusions:
- The Fab immunogene system is effective for targeted gene delivery to melanoma cells via EGF receptors.
- This approach mediates potent "suicide gene therapy" effects, leading to significant tumor growth inhibition.
- The Fab immunogene holds promise as a novel strategy for melanoma treatment.