Using apoptosis for targeted cancer therapy by a new gonadotropin releasing hormone-DNA fragmentation factor 40 chimeric protein.
OBJECTIVE
GnRH-based chimeric proteins have been shown to specifically target and kill adenocarcinomas both in vitro and in vivo. The purpose of this study is to construct a new GnRH-based chimeric protein for the treatment of adenocarcinomas in humans.


RESULTS
In this study, we constructed and characterized a new chimeric protein, GnRH-DFF40, composed of a new human killing moiety: the apoptotic DNase-DFF40 (DNA fragmentation factor), known also as caspase-activated DNase (CAD). GnRH-DFF40 exhibits DNase activity in vitro. We found that this chimeric protein can target and kill adenocarcinoma cells. Such death occurs via apoptotic pathways, resulting in an increase in the sub-G(1) population, DNA fragmentation, terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL)-positive cells, and morphology typical of apoptotic cells. These apoptotic events involve the mitochondria because we found cytochrome c depletion and caspase-9 and caspase-3 activation. Preliminary in vivo results showed that treatment of colon adenocarcinoma xenografts in nude mice with the new chimeric protein caused a reduction in tumor weight.


CONCLUSIONS
Because GnRH-DFF40 is a whole human-based chimeric protein when applied to humans, the nonspecific toxicity and immunogenicity seen with bacterial/plant-based chimeric proteins should be avoided. Thus, GnRH-DFF40 is a promising candidate for the treatment of adenocarcinomas in humans.
Ben-Yehudah
Ahmi
A
Department of Cellular Biochemistry and Human Genetics, Hebrew University-Hadassah Medical School, Jerusalem 91120, Israel.
Aqeilan
Rami
R
Robashkevich
Dana
D
Lorberboum-Galski
Haya
H
D000230:Adenocarcinoma / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D001483:Base Sequence; D015153:Blotting, Western; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002455:Cell Division; D002469:Cell Separation; D002470:Cell Survival; D003574:Cytochrome c Group / Q000378:metabolism; D053938:DNA Fragmentation; D003851:Deoxyribonucleases / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling; D051379:Mice; D008819:Mice, Nude; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000378:metabolism* / Q000628:therapy*; D010957:Plasmids / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured
D003574:Cytochrome c Group; D007987:Gonadotropin-Releasing Hormone; D003851:Deoxyribonucleases; C109663:caspase-activated deoxyribonuclease; C505407:CASP3 protein, human; C506433:CASP9 protein, human; C505409:Casp3 protein, mouse; C506434:Casp9 protein, mouse; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases