Uncovering residues that regulate cyclin D1 proteasomal degradation.

Cyclin D1 regulates G1 cell-cycle progression and is aberrantly expressed in carcinogenesis. Proteasomal degradation of cyclin D1 was highlighted as a cancer chemopreventive mechanism. To understand this mechanism better, residues responsible for degradation and ubiquitination of cyclin D1 were investigated. Eighteen lysines in cyclin D1 had single, double or multiple mutations engineered before transfection into BEAS-2B human bronchial epithelial (HBE) cells to evaluate stabilities after all-trans-retinoic acid (RA) or cycloheximide treatments. Specific mutations stabilized cyclin D1, including substitutions of lysines surrounding the cyclin box domain that inhibited RA-mediated degradation and extended the cyclin D1 half-life. Mutation of all cyclin D1 lysines blocked polyubiquitination. N-terminus (but not C-terminus) modification stabilized cyclin D1. Ubiquitination-resistant mutants preferentially localized cyclin D1 to the nucleus, directly implicating subcellular localization in regulating cyclin D1 degradation. Taken together, these findings uncover specific residues conferring ubiquitination of cyclin D1. These provide a mechanistic basis for proteasomal degradation of cyclin D1.

Induction of thymidylate synthase as a 5-fluorouracil resistance mechanism.

Thymidylate synthase (TS) is a key enzyme in the de novo synthesis of 2'-deoxythymidine-5'-monophosphate (dTMP) from 2'-deoxyuridine-5'-monophosphate (dUMP), for which 5,10-methylene-tetrahydrofolate (CH(2)-THF) is the methyl donor. TS is an important target for chemotherapy; it is inhibited by folate and nucleotide analogs, such as by 5-fluoro-dUMP (FdUMP), the active metabolite of 5-fluorouracil (5FU). FdUMP forms a relatively stable ternary complex with TS and CH(2)THF, which is further stabilized by leucovorin (LV). 5FU treatment can induce TS expression, which might bypass dTMP depletion. An improved efficacy of 5FU might be achieved by increasing and prolonging TS inhibition, a prevention of dissociation of the ternary complex, and prevention of TS induction. In a panel of 17 colon cancer cells, including several variants with acquired resistance to 5FU, sensitivity was related to TS levels, but exclusion of the resistant variants abolished this relation. For antifolates, polyglutamylation was more important than the intrinsic TS level. Cells with low p53 levels were more sensitive to 5FU and the antifolate raltitrexed (RTX) than cells with high, mutated p53. Free TS protein down-regulates its own translation, but its transcription is regulated by E2F, a cell cycle checkpoint regulator. Together, this results in low TS levels in stationary phase cells. Although cells with a low TS might theoretically be more sensitive to 5FU, the low proliferation rate prevents induction of DNA damage and 5FU toxicity. TS levels were not related to polymorphisms of the TS promoter. Treatment with 5FU or RTX rapidly induced TS levels two- to five-fold. In animal models, 5FU treatment resulted in TS inhibition followed by a two- to three-fold TS induction. Both LV and a high dose of 5FU not only enhanced TS inhibition, but also prevented TS induction and increased the antitumor effect. In patients, TS levels as determined by enzyme activity assays, immunohistochemistry and mRNA expression, were related to a response to 5FU. 5FU treatment initially decreased TS levels, but this was followed by an induction, as seen with an increased ratio of TS protein over TS-mRNA. The clear retrospective relation between TS levels and response now forms the basis for a prospective study, in which TS levels are measured before treatment in order to determine the treatment protocol.

Transcriptional activation of the nuclear receptor corepressor RIP140 by retinoic acid: a potential negative-feedback regulatory mechanism.

Through the use of microarray analysis it was discovered that the nuclear receptor coregulator, receptor interacting protein 140 (RIP140), was induced early during all-trans retinoic acid (RA)-induced differentiation of human embryonal carcinoma cells. A rapid, fourfold induction of RIP140 mRNA was detected within 3 h of RA treatment in human embryonal carcinoma and MCF-7 human breast cancer cells. RIP140 protein levels were induced within 6 h of RA treatment. The RA induction of RIP140 mRNA did not require de novo protein synthesis, consistent with RIP140 being a direct transcriptional target of retinoid receptors. Promoter/enhancer elements directly upstream of the RIP140 coding region supported RA-induced transcription of a luciferase gene. In addition the ability of overexpressed RIP140 to repress ligand activated retinoid receptors was confirmed. The finding that RIP140 is a direct transcriptional target of RA is one of the first examples of acute transcriptional regulation of a nuclear receptor coactivator or corepressor. These data are consistent with a model by which RA induction of RIP140 supplies a negative feedback signal toward ligand-activated retinoid receptors.

The pharmacovigilance of olanzapine: results of a post-marketing surveillance study on 8858 patients in England.

Olanzapine is an 'atypical' antipsychotic indicated for the treatment of schizophrenia. We analysed adverse events (AEs) reported in primary practice in England. Dispensed prescriptions issued between December 1996 and May 1998 provided exposure data. Questionnaires sent to general practitioners provided outcomes. Frequently reported AEs were: drowsiness/sedation (n = 19), extrapyramidal disorder (n = 13) and unspecified side-effects (n = 33). Events with highest incidence density in first month and reason for stopping were: drowsiness/sedation [n = 153, incidence density (ID)1 18.9], weight gain (n = 117, ID1 8.9) and malaise/lassitude (n = 65, ID1 5.2). Extrapyramidal disorders were more common in elderly population (> 70 years, ID1 3.6, risk 26.0 per 1,000 patients) compared to < 70 years (ID1 1.1, risk 8.4 per 1,000 patients). Serious suspected adverse reactions were neuroleptic malignant syndrome (n = 1) and angioneurotic ooedema (n = 2). There were eight reports of diabetes mellitus assessed as possibly due to olanzapine. Diabetes mellitus was an unlabelled AE and possible signal generated by prescription-event monitoring.

Cyclin proteolysis as a retinoid cancer prevention mechanism.

The retinoids, natural and synthetic derivatives of vitamin A, are active in cancer therapy and prevention. Their biological effects are mediated through ligand-dependent interactions with retinoid receptors that associate with specific co-regulators. A better understanding of retinoid chemopreventive mechanisms is needed. Our prior work revealed that all-trans-retinoic acid (RA) prevented tobacco-specific carcinogenic transformation of cultured human bronchial epithelial cells. RA signaled G1 arrest that permitted repair of genomic DNA damage caused by these carcinogens. RA triggered G1 arrest at least partly through proteasome-dependent degradation of cyclin D1. Proteasomal inhibitors blocked RA-mediated cyclin D1 degradation. To confirm that a specific proteolysis pathway was induced by RA-treatment, a degradation assay was established using in vitro translated cyclin D1 and cellular extracts from RA-treated or untreated human bronchial epithelial cells. Incubation of RA-treated but not the control cellular extracts with in vitro translated cyclin D1 led to cyclin degradation. This degradation depended on the PEST domain of cyclin D1, implicating ubiquitination in this retinoid degradation. Retinoid receptor selective agonists demonstrated that retinoic acid receptor (RAR)beta and retinoid X receptor (RXR) but not RARalpha- or RARgamma-dependent pathways signaled this cyclin degradation. Findings were extended to the NT2/D1 human embryonal carcinoma differentiation model where a similar pathway was activated by RA-treatment. To determine whether G1 cyclins were involved directly in bronchial preneoplasia, immunohistochemical expression profiles for cyclins D1 and E were examined. Aberrant expression of these cyclins was frequent in bronchial preneoplasia. Taken together, these findings indicate that ubiquitin-dependent proteolysis of G1 cyclins is a retinoid chemoprevention mechanism. Whether the retinoids represent the optimal agents to activate this pathway is the subject of ongoing work. These findings provide a rationale for combining the retinoids in chemoprevention trials with other agents that do not activate this proteolysis pathway. What is now known about the retinoids as cancer prevention agents will be reviewed. Emphasis is placed on retinoid effects on cell cycle progression at G1.

The rates of common adverse events reported during treatment with proton pump inhibitors used in general practice in England: cohort studies.

AIMS: To estimate the rates of common adverse events in patients treated with the proton pump inhibitors omeprazole, lansoprazole and pantoprazole in general practice in England. METHODS: In prescription-event monitoring cohort studies, data on dispensed prescriptions prescribed by general practitioners in England soon after each drug was launched were linked to subsequent clinical events recorded by the prescriber. 16 205 patients prescribed omeprazole between June 1989 and June 1990, 17 329 patients prescribed lansoprazole between May and November 1994, and 11 541 patients prescribed pantoprazole between December 1996 and June 1997 were studied. RESULTS: The commonest adverse events in the omeprazole, lansoprazole and pantoprazole cohorts were diarrhoea (incidence: 0. 18, 0.39 and 0.23 per 1000 days of exposure, respectively); nausea/vomiting (incidence: 0.16, 0.22 and 0.18 per 1000 days of exposure, respectively); abdominal pain (incidence: 0.17, 0.21 and 0. 17 per 1000 days of exposure, respectively); and headache (incidence rates: 0.10, 0.17 and 0.15 per 1000 days of exposure, respectively). The remaining adverse events occurred at rates of less than 0.11 per 1000 days of exposure. There were little absolute differences in the rates of most events between the three proton pump inhibitors. However, diarrhoea was more commonly associated with lansoprazole compared with omeprazole (rate difference: 0.21 per 1000 days of exposure; 95% CI 0.17, 0.25; rate ratio: 2.11; 1.78, 2.51), and there was a clear age-response relationship. CONCLUSIONS: Adverse events occurred relatively infrequently in all three cohorts. There were only small absolute differences in event rates between the three drugs, although these data suggest the hypothesis that lansoprazole is associated with more frequent occurrence of diarrhoea, particularly in the elderly.

The pharmacovigilance of tamsulosin: event data on 12484 patients.

OBJECTIVE: To determine drug effectiveness and adverse effects in a noninterventional observational cohort study of over 10 000 patients treated with tamsulosin in general medical practice. METHODS: Using prescription-event monitoring, data were collected of all prescriptions for tamsulosin issued nationally during June 1996 to January 1998. For each patient entered into the cohort a computerized longitudinal record of exposure was constructed. The outcome data, patient information and an opinion about the effectiveness of the drug were provided by the prescriber, using a standard questionnaire sent 6 months after the initial prescription for tamsulosin. The incidence of each of almost 2000 events listed in the Drug Safety Research Unit computerized dictionary was calculated and scrutinized by medical assessors for possible adverse reactions, and any difference determined between the incidence of each event in the first month and subsequent months of exposure. All deaths were followed up to detect possibly drug-related causes. RESULTS: Event data were obtained on 12484 patients, from the 52.9% of questionnaires returned and that contained valid event data. Tamsulosin was reported to have been effective in 7428 (78.3%) of the 9487 patients in whom the general practitioners expressed an opinion about effectiveness. Suspected adverse drug reactions were reported in only 171 (1.4%) of the cohort. Dizziness, headache, malaise and hypotension were common to the reported adverse reactions, reasons for stopping the drug and events of greatest incidence density. None of the 282 deaths that occurred in this elderly cohort were attributed to the drug. CONCLUSION: This study suggests that tamsulosin has a highly acceptable benefit-to-risk ratio. No untoward features not already mentioned in the prescribing guidance were identified.

Losartan: a study of pharmacovigilance data on 14,522 patients.

The objective of this study was to undertake a post- marketing safety surveillance study of losartan, the first specific angiotensin-II receptor antagonist to be marketed in England. It is a non-interventional observational cohort study using the technique of prescription-event monitoring. Subjects who took part in the study were from 14 522 patients treated by general practitioners in England. We recorded adverse drug reactions, adverse events, reasons for stopping the drug, pregnancies and deaths. The results were that general practitioners considered the drug to have been effective in 85.9% of the patients evaluated for effectiveness. Dizziness, malaise, lassitude, nausea, cough and headache were among the most frequently reported adverse drug reactions, reasons for stopping the drug and events of highest incidence density. None of the deaths that occurred were attributed to losartan and no serious unexpected adverse reactions or interactions were identified. In conclusion losartan has an acceptable safety profile and the study confirms the safety information given in the current Summary of Product characteristics for losartan.

Retinoic acid promotes ubiquitination and proteolysis of cyclin D1 during induced tumor cell differentiation.

Mechanisms by which differentiation programs engage the cell cycle are poorly understood. This study demonstrates that retinoids promote ubiquitination and degradation of cyclin D1 during retinoid-induced differentiation of human embryonal carcinoma cells. In response to all-trans-retinoic acid (RA) treatment, the human embryonal carcinoma cell line NT2/D1 exhibits a progressive decline in cyclin D1 expression beginning when the cells are committed to differentiate, but before onset of terminal neuronal differentiation. The decrease in cyclin D1 protein is tightly associated with the accumulation of hypophosphorylated forms of the retinoblastoma protein and G(1) arrest. In contrast, retinoic acid receptor gamma-deficient NT2/D1-R1 cells do not growth-arrest or accumulate in G(1) and have persistent cyclin D1 overexpression despite RA treatment. Notably, stable transfection of retinoic acid receptor gamma restores RA-mediated growth suppression and differentiation to NT2/D1-R1 cells and restores the decline of cyclin D1. The proteasome inhibitor LLnL blocks this RA-mediated decline in cyclin D1. RA treatment markedly accelerates ubiquitination of wild-type cyclin D1, but not a cyclin D1 (T286A) mutant. Transient expression of cyclin D1 (T286A) in NT2/D1 cells blocks RA-mediated transcriptional decline of a differentiation-sensitive reporter plasmid and represses induction of immunophenotypic neuronal markers. Taken together, these findings strongly implicate RA-mediated degradation of cyclin D1 as a means of coupling induced differentiation and cell cycle control of human embryonal carcinoma cells.

Cohort study on calcium channel blockers, other cardiovascular agents, and the prevalence of depression.

AIMS: Some reports have suggested that calcium channel blockers may be associated with an increased incidence of depression or suicide. There is a paucity of evidence from large scale studies. The aim of this study was to assess rates of depression with calcium channel antagonists using data from prescription event monitoring studies. METHODS: Observational studies on large cohorts of patients using lisinopril, enalapril (ACE inhibitors), nicardipine (type 2 calcium channel blocker) and diltiazem (type 3 calcium channel blocker) were conducted, using prescription-event monitoring. Rates of depression in the different drugs and rate ratios (95% CI) were computed. RESULTS: The crude overall rates of depression during treatment were 1.89, 1.92 and 1.62 per 1000 patient months for the ACE inhibitors, diltiazem and nicardipine, respectively. Using the ACE inhibitors as the reference group, the rate ratios for depression were 1.07 (0. 82-1.40) and 0.86 (0.69-1.08) for diltiazem and nicardipine, respectively. CONCLUSIONS: This study does not support the hypothesis that calcium channel blockers are associated with depression, when considering patients treated in general practice in the UK.

Nicorandil and diabetes: a nested case-control study to examine a signal generated by prescription-event monitoring.

OBJECTIVE: To investigate a signal generated by an observational cohort study that treatment with the potassium channel opener drug, nicorandil, is associated with an increased incidence of diabetes mellitus (DM). METHODS: Nested case-control study within cohorts used for prescription-event monitoring. RESULTS: The cohort study of nicorandil showed that there was a significant difference between the event rate for DM in the first month after starting nicorandil, compared to months 2-6 (difference in rates 1.93, 99% CI 0.7-3.1, per 1000 patient months of treatment). However, the adjusted odds ratio from the case-control study was 1.42 (95% CI 0.66-3.07) for incidence of newly diagnosed DM in nicorandil versus four comparator drugs. CONCLUSIONS: The nested case-control study was used as a hypothesis-testing instrument for following up a signal of a possible drug reaction to nicorandil. It showed no evidence that nicorandil is associated with increased incidence of newly diagnosed DM.

Newer antidepressants: a comparison of tolerability in general practice.

BACKGROUND: An increasing number of antidepressants have been released on the United Kingdom market in recent years, and these are being prescribed more frequently in general practice. Clinical trials suggest that such agents have similar efficacy and the choice of drug is probably based on tolerability, toxicity in overdose, and cost. AIM: To compare the tolerability and safety profile of six, newly marketed antidepressants used in general practice. METHOD: Studies have been conducted for six antidepressants: fluoxetine, sertraline, paroxetine, moclobemide, venlafaxine, and nefazodone, using the technique of prescription-event monitoring. Patients were identified using incident dispensed prescription data. Questionnaires were sent to patients' general practitioners six months after the date of first prescription. Questionnaires asked for date of birth, sex, indication for prescribing each drug, and all events entered in the patients' records after the date of first prescription. RESULTS: Each cohort exceeded 10,000 patients. Nausea/vomiting was the most frequently reported event for all drugs. The difference in incidence rates for drowsiness/sedation, male sexual dysfunction, and hypertension is shown. Mortality data are also reported. CONCLUSION: Frequently reported events were similar for all six drugs but there were clinically and statistically significant differences for less frequently reported events. The adjusted mortality rate was identical between the six drugs. This study provides valuable comparative data for six, widely used antidepressants in general practice.

Nefazodone in community use: the results of prescription-event monitoring.

PURPOSE: This study was a pharmacovigilance exercise which aimed to determine the post-marketing event profile of nefazodone, a newly marketed antidepressant, in community use. METHODS: Information was collected on patients included in a non-interventional observational cohort study conducted by means of Prescription-Event Monitoring in England. Incidence densities were calculated for all reported events. RESULTS: Information was obtained for 11 834 patients. Nausea and dizziness were the most frequent adverse events that led to stopping nefazodone and the most frequently reported events during the first month of treatment. Unsteadiness and falls were reported more frequently in the elderly. Hepatic events, involuntary movements, thrombocytopenia, hallucinations and withdrawal reactions were reported rarely but were possibly associated with nefazodone. Eight overdoses involving nefazodone alone were reported with no serious clinical sequelae. Two premature births, one low birth weight term baby and two babies with renal abnormalities were outcomes in 38 pregnancies exposed in the first trimester to nefazodone. One death in a woman aged 71 years followed an illness with serotonergic features. CONCLUSIONS: Event data are presented for patients dispensed nefazodone in the community. The findings are discussed.

Safety profile of nicorandil--prescription-event monitoring (PEM) study.

PURPOSE: A PEM study of nicorandil (Ikorel) was undertaken to assess the drug's overall safety in everyday clinical practice. METHODS: The prescription data used covered the period December 1994 to October 1996. The event data, which were based on a minimum observation period of 6 months, came from questionnaires returned by the prescribing general practitioners. Incidence densities (IDs) were calculated for all events occurring during month 1, months 2-6, and the overall treatment period. Selected events were followed up. RESULTS: The study was based on a cohort of 13,260 patients and 86,760 patient-months of nicorandil treatment. Major indications for use were angina (8744) and ischaemic heart disease (846). Adverse reactions (258) were reported in 175 (1.3%) of patients--the most frequent being headache (58; 0.4%) and unspecified side effects (36; 0.3%). The most common reasons for stopping treatment (excluding those confounded by indication) were headache (477; 3.5%), dizziness (88; 0.65%) and 'not effective' (491; 3.6%). The number of patients still being prescribed nicorandil after 6 months was 74.3%. In those cases where an opinion on effectiveness was given, nicorandil was reported to be effective in 80% (8713) of patients. The event of headache/migraine had the highest ID in the first month of treatment (49.4 per 1000 patient-months) and was not confounded by indication. Follow-up of selected events was reassuring overall; rare side effects included angioneurotic oedema and photosensitivity (3 cases each). CONCLUSION: This PEM study provides information on the 'real-world' use of nicorandil and shows generally that the drug is safe when used in the recommended dosage.

Age and sex distribution of suspected adverse drug reactions to newly marketed drugs in general practice in England: analysis of 48 cohort studies.

AIMS: Little is known about the frequency with which suspected adverse drug reactions are seen by general practitioners after the prescription of newly marketed drugs. We investigated age and sex specific incidence rates of suspected adverse drug reactions recorded by general practitioners in England after the prescription of selected newly marketed drugs. METHODS: Information was collected from 48 national cohort studies of newly marketed drugs studied by prescription-event monitoring. Questionnaires were sent to prescribers asking for details of events and suspected adverse drug reactions recorded in the patients' notes occurring after the drugs were prescribed. RESULTS: During the 48 cohort studies, a total of 513608 patients were investigated, of which 221781 (43.2%) were males and 285862 (55.7%) were females. The overall incidence of suspected adverse drug reactions in males was 12.9 per 10000 patient-months of exposure (95% confidence limits 12.3 to 13.5), and in females was 20.6 per 10000 patient-months of exposure (95% confidence limits 19.9 to 21.3). The overall age-standardised relative risk of an adverse drug reaction in females compared with males was 1.6 (1.5-1.7). This sex difference was significant in all age groups above 19 years of age, and was relatively consistent across all age groups (chi2 test for heterogeneity = 9.2, P=0.3). The highest rate of recording in males was in the 50-59 year age group, and in females was in the 30-39 year age group. CONCLUSIONS: In general practice in England, suspected adverse drug reactions to newly marketed drugs are recorded more often in adults aged between 30 and 59 years of age and are 60% more common in women than in men. The sex difference occurs in all age groups over 19 years of age.

Risk of non-fatal cardiac failure and ischaemic heart disease with long acting beta 2 agonists.

BACKGROUND: The long term safety of beta agonists, particularly in patients with heart disease, has not been fully established. METHODS: This study accessed the results of three cohort studies involving: 12,294 patients receiving at least one prescription for nedocromil between November 1986 and September 1988; 15,407 patients prescribed salmeterol between December 1990 and May 1991; and 8098 patients prescribed bambuterol between February 1993 and December 1995. Details of all dispensed prescriptions for these drugs prescribed by general practitioners in England soon after their launch were provided in confidence by the Prescription Pricing Authority. Questionnaires were sent to the prescriber asking for details of events occurring after the first prescription (prescription event monitoring). Rates and relative risks of non-fatal cardiac failure and ischaemic heart disease were calculated, comparing bambuterol and salmeterol with the reference drug nedocromil. RESULTS: The age and sex adjusted relative risk of non-fatal cardiac failure associated with bambuterol was 3.41 (95% confidence limits (CL) 1.99 to 5.86) when compared with nedocromil. When salmeterol was compared with nedocromil the adjusted relative risk of non-fatal cardiac failure was 1.10 (95% CL 0.63 to 1.91). The adjusted relative risk of non-fatal ischaemic heart disease was 1.23 (95% CL 0.73 to 2.08) and 1.07 (95% CL 0.69 to 1.66) for bambuterol and salmeterol, compared with nedocromil, respectively. However, in the first month of exposure the adjusted relative risk of non-fatal ischaemic heart disease was 3.95 (95% CL 1.38 to 11.31) when bambuterol was compared with nedocromil. CONCLUSIONS: Caution should be exercised when prescribing long acting oral beta agonists to patients at risk of cardiac failure. More definitive evidence would come from prospective randomised trials.

United Kingdom experience with alendronate and oesophageal reactions.

Alendronate is indicated for the treatment of osteoporosis in post-menopausal women. Although the drug has been associated with reports of severe oesophagitis, there have been no studies establishing the incidence of such reactions. Information was collected on 1523 patients included in a study conducted by means of prescription-event monitoring. Dyspepsia, nausea/vomiting, and abdominal pain were the most frequently reported events in the first month of treatment. After follow-up, 20 patients (1.3%) experienced oesophageal events that were considered to be possibly related to alendronate.

The role of prescribing and referral bias in studies of the association between third generation oral contraceptives and increased risk of thromboembolism.

Recently published papers have shown a slightly increased risk for venous thromboembolism (VTE) among users of third generation oral contraceptive pills (OC) as compared with users of second generation OC. The extra risk is small (about twofold) and could be explained possibly by biases introduced into the original case-control studies. This paper examines the possible role of prescribing bias (i.e. that doctors would prescribe third generation OC preferentially to women whom they knew to be at risk of VTE) and referral bias (i.e. that doctors would preferentially refer women on third generation OC to hospital for investigations of symptoms suggestive of VTE). A cross-sectional survey was carried out of 106 physicians, to record their attitude to the prescription of OC in certain risk situations. Each doctor then extracted data on OC prescription from approximately 12 patients under his care. A total of 1192 patients were included. Data collected related to known risk factors to which the patient was prone, and the type of OC that the woman was taking. The results from the doctors' attitudinal survey showed that third generation oral contraceptives seemed to be very popular among doctors in England and that they would prefer to prescribe third generation to second generation oral contraceptives in virtually all risk situations, and also in situations where there was no particular risk of VTE. The results from the patients' survey, however, did not correspond with the attitudinal survey, and there was no evidence of a systematic bias running through this sample of patients, associating third generation OC prescription with particular risk factors. The results from the doctors' attitudinal survey on referral did not show any evidence of potential bias towards referring patients on third generation OC to hospitals. Comparison of this survey with an equivalent survey done in Germany suggests that there may be different factors governing doctors' behaviour in Germany and England, with reference to choice of OC prescription.