The annexins are a super-family of closely related calcium and membrane-binding proteins. They have a diverse range of cellular functions that include vesicle trafficking, cell division, apoptosis, calcium signalling and growth regulation. Many studies have shown the annexins to be among the genes whose expression are consistently differentially altered in neoplasia. Some annexins show increased expression in specific types of tumours, while others show loss of expression. Mechanistic studies relating the changes in annexin expression to tumour cell function, particularly tumour invasion and metastasis, angiogenesis and drug resistance, are now also emerging. Changes in the expression of individual annexins are associated with particular types of tumour and hence the annexins may also be useful biomarkers in the clinic.

2ZD, 2Department of Biochemistry and 3Department of Pharmacology, Marischal College, University of Aberdeen AB9 lAS 1 The localisation and distribution of cytochrome P-450 in human tissues has been studied by immunocytochemistry using a monoclonal antibody to a major form of human hepatic cytochrome P-450, P-45OhA7, which is closely related to cytochromes P-450 HLp and P-45ONF. 2 Strong immunoreactivity was identified in hepatocytes, columnar absorptive epithelial cells of the small intestine, polymorphonuclear leucocytes and their precursors in the bone marrow, and in mast cells.3 Weak immunoreactivity was present in the proximal tubules of the kidney, pancreatic acini, gall bladder epithelium, squamous epithelium and sebaceous glands of the skin, interstitial cells of the testis and luteal cells of the ovary. 4 Immunoreactivity could not be demonstrated in the adrenal gland, placenta, colonic epithelium and alveolar type II cells and Clara cells of the lung.

Summary Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme for degradation of 5-fluorouracil (5-FU). DPD activity is highly variable in liver and peripheral mononuclear cells (PMNCs) and it has not been well studied in human tumours. Characterization of DPD in colorectal cancer is of clinical interest through its role in the regulation of 5-FU, the main chemotherapeutic agent used in this disease. Therefore, DPD activity was analysed in colorectal tumour and adjacent normal tissue from 63 patients, including three liver metastasis. DPD activity was highly variable in all tissues studied (coefficient of variation 43-61 %) and was higher in normal tissue than in tumour. The tumournormal activity ratio ranged from 0.19 to 3.32 (median 0.76). PMNC DPD activity was available for 57 patients and was correlated with tumour activity (r5 = 0.29, P < 0.001). A higher correlation was observed between PMNCs and tumour samples that were both obtained in the morning (r, = 0.49), consistent with circadian variation in DPD activity. Normal tissue DPD activity was not correlated with either tumour (r5 = 0.11) or PMNC activity (rs = -0.06). This study provides the first analysis of DPD activity in colorectal cancer and illustrates the large degree of variation in tumour activity. The tumour-normal activity ratio results suggest that elevated tumour DPD can play a role in 5-FU resistance through increased inactivation in tumour cells, but is an uncommon event in colorectal tumours. The results support the use of PMNCs for monitoring tumour DPD activity, particularly when circadian variation is taken into account. As a large degree of the variation in tumour DPD activity is not explained by PMNC activity, more accurate alternatives are needed before DPD activity can be used for targeting 5-FU therapy.Keywords: dihydropyrimidine dehydrogenase; enzyme activity; colorectal cancer; 5-fluorouracil; pharmacogenetics 5-Fluorouracil (5-FU) is commonly used in the treatment of gastrointestinal, head and neck, and breast tumours. 5-FU is itself inactive and requires intracellular conversion to form cytotoxic nucleotides. Several cellular targets for fluoropyrimidines have been well characterized, including inhibition of thymidylate synthase (TS) by FdUMP and false-base incorporation into RNA or DNA. Most investigations into cellular resistance factors regulating 5-FU activity have focused on alterations in TS levels and reduced folate pools, the required cofactor for binding dUMP to TS (Wang et al, 1993;Johnston et al, 1995). However, most of an administered 5-FU dose undergoes metabolism to an inactive species through a threeenzyme process, which is initiated and rate limited by dihydropyrimidine dehydrogenase (DPD; EC 1.3.1.2). After a bolus injection of 5-FU, 80% is degraded via DPD 24 h after administration (Heggie et al, 1987). Studies of 19F-NMR spectroscopy in mice bearing colon tumours found that catabolites made up 51% of labelled drug in the tumour, compared with 26% for the anabolic products (Kamm et al, 1994). Ther...

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Summary The cytochromes P450 have a central role in the oxidative activation and detoxification of a wide range of xenobiotics, including many carcinogens and several anti-cancer drugs. Thus the cytochrome P450 enzyme system has important roles in both tumour development and influencing the response of tumours to chemotherapy. Stomach cancer is one of the commonest tumours of the alimentary tract and environmental factors, including dietary factors, have been implicated in the development of this tumour. This type of tumour has a poor prognosis and responds poorly to current therapies. In this study, the presence and cellular localization of several major forms of P450, CYPl A, CYP2E1 and CYP3A have been investigated in stomach cancer and compared with their expression in normal stomach. There was enhanced expression of CYP1 A and CYP3A in stomach cancer with CYP1 A present in 51% and CYP3A present in 28% of cases. In contrast, no P450 was identified in normal stomach. The presence of CYPlA and CYP3A in stomach cancer provides further evidence for the enhanced expression of specific forms of cytochrome P450 in tumours and may be important therapeutically for the development of anticancer drugs that are activated by these forms of P450.Keywords: cytochrome P450; neoplasm; stomach Stomach cancer is one of the commonest cancers of the alimentary tract and has a relatively poor prognosis with limited response to current modes of therapy (Thompson et al, 1993). Environmental factors, particularly dietary factors, are considered to be important in the aetiology and pathogenesis of this type of tumour. The current model for development of stomach cancer proposes that this type of tumour develops from normal stomach through different types of intestinal metaplasia (Correa, 1988).The cytochromes P450 (P450) are a multi-gene family (Nelson et al, 1996) of constitutive and inducible haem-containing enzymes with a critical role in the metabolism of a diverse range of xenobiotics, including many potential carcinogens (Shimada and Guengerich, 1991;Gonzalez and Gelboin, 1994; RobertsThomson et al, 1995) and various anti-cancer drugs (Kivisto et al, 1995a). Thus, the P450s are considered to have a central role in chemical carcinogenesis and are involved in tumour initiation and promotion as they can activate or deactivate most carcinogens (Gonzalez and Gelboin, 1994). Furthermore the P450s can influence the response of established tumours to anti-cancer drugs by metabolizing these drugs both in normal tissues and in tumour cells. In addition, P450s may have a role in cell regulation, in view of their involvement in the metabolism of physiological chemicals active in inter-and intracellular signalling, including steroid hormones, eicosanoids and fatty acids (Capdevila et al, 1992).The liver is the major normal tissue that expresses P450, while specific forms of P450 are expressed in several different normal extra-hepatic tissues, including small intestine, kidney and lung (Schwartzman et al, 1990;Kaminsky and Fasco, 1992; Shima...

Summary Reduced glutathione (GSH) has been demonstrated in benign and malignant human breast lesions using a newly.developed histofluorescence technique. GSH was present in every lesion and in each case was localised to the epithelium. A semi-quantitative assessment revealed a moderate amount of GSH in normal epithelium and fibroadenoma and a high level in apocrine metaplasia, epitheliosis and intraduct carcinoma. Invasive ductal carcinoma contained a variable amount of GSH. Correlation between fluorescence intensity and histological grade of ductal carcinomas was almost statistically significant but a relationship to oestrogen receptor status was not detected. The rapid assessment of GSH in breast cancer may aid in the selection of optimum chemotherapeutic regimens.Reduced glutathione (GSH) is a thiol containing tripeptide which is involved in a variety of cellular functions. GSH has a role in the detoxification of drugs and carcinogens, protection of cells from free radical damage and reactive oxygen compounds, participates in protein and DNA synthesis and in the regulation of enzyme activity (Meister & Anderson, 1983;Orrenius & Moldeus, 1984;Chasseaud, 1979). Altered levels of GSH are frequently observed in experimental animal neoplasia (Roomi et al., 1985, Fiala et al., 1976 but GSH levels in human neoplasia have been reported only for carcinomas of the stomach and colon (Siegers et al., 1983;Siegers et al., 1984). GSH in human breast tissue has not been investigated previously, although GSH-associated enzymes have been studied both biochemically (Di Ilio et al., 1985) and histochemically (Levine et al., 1983;Bard et al., 1986).Tumour GSH appears to be an important factor in determining the effectiveness of a variety of anti-cancer chemotherapeutic drugs, in general a low level of GSH being associated with increased chemotherapeutic efficiency (Arrick & Nathan, 1984;Romine and Kessel, 1986;Crook et al., 1986).We have studied the localisation and amount of GSH in normal and pathological human breast tissue using our recently developed specific histofluorescence method for GSH (Murray et al., 1986 (dichroic mirror) for epi-illumination (set at position 1, 50% transmission at 400 nm), a UGI excitation filter (band pass filter, peak transmission 365nm) and a K430 high pass emission filter (50% transmission at 430 nm).The distribution of fluorescence in each section was recorded and the intensity of GSH fluorescence assessed in semi-quantitatively on a 4-point scale (0=no fluorescence, + = slight fluorescence, + + =moderate fluorescence, + + + = bright fluorescence).Relationship between GSH concentration and histofluorescence Five known concentrations of authentic GSH (2, 4, 6, 8, 10mM) in 0.1 M formic acid were each mixed with 1% polyvinylpyrrolidone (PVP: 2 ml GSH solution plus 1 ml PVP), droplets applied to glass slides and then dried in air. The slides were then reacted with o-phthalaldehyde as described above, examined with fluorescence microscopy and the fluorescence intensity assessed semi-quantitati...

This study provides definitive evidence that eniluracil completely inactivates DPD activity in human solid tumors. The increased plasma uracil and decreased DPD activity are consistent with systemic inactivation of the enzyme. The mechanism of inactivation is at the catalytic level, because no changes in DPD protein or mRNA were observed. Treatment with eniluracil will eliminate DPD activity as a source of pharmacokinetic fluorouracil variability or resistance in human colorectal cancer.

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