Roger Becker

mdm2 mRNA expression is associated with survival in ovarian cancer

Expression of mdm‐2 mRNA was measured in 90 ovarian‐cancer tissue specimens using the S1 nuclease assay, to investigate a possible association between MDM2 expression and prognosis. mdm‐2 mRNA expression was an independent prognostic factor for patients with primary ovarian cancer, FIGO (International Federation of Gynecology and Obstetrics) stages III and IV (n = 57), who all received chemotherapy with carboplatin or cisplatin and cyclophosphamide. Median survival time for patients (FIGO stages III and IV) with no detectable expression of mdm‐2 mRNA (n = 14) was 171 days, as compared with 839 days for patients (n = 43) with detectable mdm‐2 mRNA (p = 0.0194; log‐rank test). However, no association between mdm‐2 mRNA expression and survival was observed for patients with FIGO stages I and II who did not receive chemotherapy. mdm‐2 expression was not associated with FIGO stage, residual disease, histologic grade and type. Our results suggest that mdm‐2, which is known to disrupt p53 function, sensitizes ovarian‐cancer cells to cisplatin/cyclophosphamide, possibly by inhibition of p53‐mediated G1 cell‐cycle arrest and p53‐stimulated nucleotide‐excision repair. Int. J. Cancer 74:438–442, 1997.

cAMP‐dependent phosphorylation of CYP2B1 as a functional switch for cyclophosphamide activation and its hormonal control in vitro and in vivo

An important feature of cytochrome P450 (CYP) 2B1 is its high ability to convert the prodrug cyclophosphamide (CPA) to therapeutically cytotoxic metabolites, resulting in interstrand DNA‐cross‐linking and cell death. We have examined whether and how the phosphorylation of CYP2B1 influences CPA metabolic activation in vitro and in vivo. We found first that only part of the total CYP2B1 pool undergoes phosphorylation. This part is fully inactivated. Second, phosphorylation of CYP2B1 in intact hepatocytes reduced by up to 75% toxification of CPA to mutagenic metabolites (totally dependent on the same preferentially CYP2B‐catalyzed 4‐hydroxylation of CPA as is the generation of highly cytotoxic species). Third, the phosphoacceptor‐serine 128 of CYP2B1 in the consensus sequence for interaction with the protein kinase A represents an on/off switch for the activation of CPA depending on the phosphorylation conditions in the cell. Fourth, evidence is presented that the above‐described events also occur in vivo. In conclusion, a successful therapy with CPA, helped by forced expression of CYP2B1 in tumor cells (as recently proposed) will, in addition, be profoundly modified by its phosphorylation status.

Investigating the Role of the Microsomal Epoxide Hydrolase Membrane Topology and Its Implication for Drug Metabolism Pathways

The microsomal epoxide hydrolase (mEH) catalyzes the hydrolysis of reactive epoxides which are formed by the action of cytochromes P450 from xenobiotics. In addition the mEH has been found to mediate the transport of bile acids. For the mEH it has been shown that it is cotranslationally inserted into the endoplasmic reticulum. Here we demonstrate that the amino-terminal twenty amino acid residues of this protein serve as its single membrane anchor signal sequence and that the function of this sequence can be also supplied by a cytochrome P450 (CYP2B1) anchor signal sequence.

Stable Expression of Heterologous Microsomal Epoxide Hydrolase in BHK21 Cells: Influence on the Mutagenicity of Benzo[a]pyrene 4,5-Oxide

Most environmental mutagens and carcinogens require metabolic activation to electro- philic intermediates capable of reacting with cellular target structures, such as DNA. These electrophilic intermediates are in addition subject to metabolic detoxification. This metabolism is mainly controlled by enzymes whose expression is very variable. Among other things, various enzymes are inducible by environmental chemicals. Understanding the toxicology of chemicals (for example, species differences, idiosyncrasias, organotropisms) therefore requires knowledge of critical host factors. One approach towards this goal involves the use of purified enzymes in metabolism and toxicological studies (Glatt et al., 1983; Glatt and Oesch, 1986). This approach, however, is laborious, the purification may alter enzymatic properties, and it may be difficult to rule out the influence of impurities in the enzyme preparation, especially when large families of structurally closely related enzymes exist, as is common in xenobiotic metabolism.