title: Induction of apoptosis by the ADEPT agent ZD2767: comparison with the classical nitrogen mustard chlorambucil and a monofunctional ZD2767 analogue.
abstract: ZD2767P is a phenol mustard glutamate prodrug which is currently being developed for Antibody Directed Enzyme Prodrug Therapy (ADEPT). In ZD2767 ADEPT an active bi-functional alkylating drug, ZD2767D (4-[N,N-bis(2-iodoethyl)amino]phenol), is generated following cleavage of ZD2767P by the bacterial enzyme carboxypeptidase G2 (CPG2) which is targeted to the tumour by conjugation to the F(ab')(2)fragment of the anti-CEA antibody A5B7. The aim of the studies described here was to identify the mode of cell death induced by ZD2767P + CPG2 in comparison to the established nitrogen mustard chlorambucil. The contribution of bifunctional and monofunctional ZD2767 DNA lesions to cell death induction was investigated using a monofunctional ZD2767D analogue. Apoptosis in LoVo tumour cells was studied by three different methods (nuclear morphology, annexin V staining and TUNEL). Levels of apoptosis detected using the three assays were similar, and each drug treatment produced apoptosis at levels above those in control cells at concentrations which resulted in tumour cell growth inhibition. The bi-functional compounds, ZD2767P + CPG2 and chlorambucil, induced apoptosis in a concentration and time dependent manner, with equitoxic concentrations producing equivalent levels of apoptosis. In contrast, the mono-functional ZD2767D analogue at 100 microM resulted in the maximal level of apoptosis at 25 h with no further increase over the following 72 h. These studies have demonstrated that apoptosis is the mechanism of cell death induced by the ZD2767 ADEPT system, and that levels of apoptosis produced by ZD2767 are similar to those observed at equitoxic concentrations of the classical nitrogen mustard chlorambucil. The mono-functional ZD2767 analogue also induced apoptosis, but with a different time course and characteristics. In conjunction with previous data, these studies have shown that the potent activity of ZD2767 can be attributed to the ability of the compound to induce bifunctional DNA lesions and engage apoptosis.
authors.affiliation: Cancer Research Unit, University of Newcastle upon Tyne, Newcastle upon Tyne, UK.
authors.forename: N R
authors.initials: NR
authors.lastname: Monks
authors.forename: D C
authors.initials: DC
authors.lastname: Blakey
authors.forename: N J
authors.initials: NJ
authors.lastname: Curtin
authors.forename: S J
authors.initials: SJ
authors.lastname: East
authors.forename: A
authors.initials: A
authors.lastname: Heuze
authors.forename: D R
authors.initials: DR
authors.lastname: Newell
mesh_terms: D018906:Antineoplastic Agents, Alkylating / Q000494:pharmacology*; D017209:Apoptosis; D002699:Chlorambucil / Q000494:pharmacology*; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D009588:Nitrogen Mustard Compounds / Q000494:pharmacology*; D011355:Prodrugs / Q000494:pharmacology*; D013997:Time Factors; D014407:Tumor Cells, Cultured / Q000187:drug effects; D011623:gamma-Glutamyl Hydrolase / Q000494:pharmacology*
chemical_list: D018906:Antineoplastic Agents, Alkylating; D004273:DNA, Neoplasm; D009588:Nitrogen Mustard Compounds; D011355:Prodrugs; C097602:ZD 2767; D002699:Chlorambucil; D011623:gamma-Glutamyl Hydrolase