Pharmacological and biochemical determinants of the antitumour activity of the indoloquinone EO9.
Cummings J. Spanswick VJ. Gardiner J. Ritchie A. Smyth JF.
Imperial Cancer Research Fund, Medical Oncology Unit, Western General Hospital, Edinburgh, UK.
EO9 is a novel bioreductive drug which has recently undergone extensive clinical evaluation. Its mechanism of action remains to be clearly defined. Antitumour activity of EO9 has been determined in 2 human colon cancer xenografts (HT-29 and BE) and 2 murine colon adenocarcinomas (MAC 16 and 26) after intratumoural injection of 250 microg of drug. Levels of the major bioreductive enzymes (DT-diaphorase, cytochrome P-450 reductase and cytochrome b5 reductase) were measured in tumours using cytochrome c reduction and menadione as the intermediate electron acceptor. There was no correlation between chemosensitivity (T/C: HT-29, 15%; BE, 27%; MAC 16, 33% and MAC 26, 60%) and enzyme activity (r2 = 0.47 for DT-diaphorase, r2 = 0.1 for cytochrome P-450 reductase and r2 = 0.52 for cytochrome b5 reductase). Drug metabolism was followed in vitro using tumour homogenates incubated under aerobic and anaerobic conditions. Four metabolites were identified by HPLC and characterised bv UV-visible spectroscopy. With the exception of the hydrolysis product EO5A, all other metabolites appeared to be drug adducts. No correlation was observed between the kinetics of metabolite formation and antitumour activity. A good correlation (r2 = 0.86) was found with the rate of disappearance of parent drug and antitumour activity. These data show that the overall capacity of a tumour to metabolise EO9 is the most important determinant of antitumour activity rather than the expression of the major bioreductive enzymes and that the parent drug rather than a metabolite leads to the active form of the drug.
Cellular interactions of 5-fluorouracil and the camptothecin analogue CPT-11 (irinotecan) in a human colorectal carcinoma cell line.
Guichard S. Hennebelle I. Bugat R. Canal P.
Groupe de Pharmacologie Clinique et Experimentale, Institut Claudius Regaud, Toulouse, France.
CPT-11 (irinotecan) is a DNA topoisomerase I inhibitor active against metastatic colorectal carcinoma. We investigated, in a human colon carcinoma cell line, HT-29, the effects of CPT-11 and 5-fluorouracil (5FU) combinations. A strong synergism between CPT-11 and 5FU was observed after sequential exposure and only additivity or antagonism after simultaneous exposure. When cells were first exposed to 5FU, the product of cellular CPT-11 concentrations versus time (CxT) was 6895 +/- 1020 pmol x hr/10(6) cells, while it was 3875 +/- 121 pmol x hr/10(6) cells with CPT-11 alone (p < 0.01). The same phenomenon was observed with SN-38: 148.2 +/- 49.5 versus 83.4 +/- 23.6 pmol x hr/10(6) cells (p < 0.05). Consequently, the formation of protein-DNA complexes was 1.4 times greater with 5FU pretreatment than with CPT-11 alone (p = 0.03). Moreover, the incorporation of 5FU derivatives into DNA was multiplied by a factor of 1.5 24 hr after CPT-11 exposure. When cells were first incubated with CPT-11, the decrease in thymidylate synthase (TS) activity was identical to that obtained after 5FU exposure (1.09 to 0.023 pmol/min/mg protein), but this decrease persisted for 24 hr (0.014 pmol/min/mg protein) (p = 0.035). At the same time, a 1.8-fold increase in the incorporation of 5FU derivatives into DNA and a 2-fold increase in DNA-protein complex formation were evidenced. With the two sequential associations, we observed a persistent S-phase arrest, as compared with CPT-11 alone. These results suggest that CPT-11 and 5FU combinations are of clinical interest, and mechanisms of interaction between the two drugs seem to be multifactorial.