Genetic Predictors of Azathioprine Toxicity and Clinical Response in Patients with Inflammatory Bowel Disease.

BACKGROUND: Thiopurines (Azathioprine (AZA) and 6-Mercaptopurine (6-MP) are considered a well-established therapy for patients with Inflammatory Bowel Disease (IBD) including ulcerative colitis (UC) and Crohn's Disease (CD). However, nearly 20% of patients discontinue thiopurines due to adverse events. Functional polymorphisms of several enzymes involved in the metabolism of thiopurines have been linked with toxicity. The clinical value of variant carriers such as TPMT, ITPA and GSTs in predicting toxicity and adverse events for IBD patients treated with thiopurines remains to be clarified. OBJECTIVES: To determine if variation in TPMT, ITPA and GST genotypes can predict adverse effects such as neutropenia, pancreatitis, liver enzyme elevation, as well as clinical response for patients with IBD treated with thiopurines. METHODS: Patients known to have IBD and treated with AZA or 6MP were enrolled. Adverse effects were calculated and their correlation with TPMT, ITPA and GST genotypes was evaluated. Further, the correlation between clinical response and TPMT, ITPA and GST genotypes were assessed. RESULTS: A total of 53 patients were enrolled. 16/53 patients (28.6%) responded to AZA therapy. 17 patients experienced adverse events with 10 having to discontinue treatment. Three patients (5.4%) developed severe myelosuppression (WBC< 2.0 or neutrophils <1.0). Loss of function TPMT genotype was associated with adverse events (OR 3.64, 95% CI 0.55 - 24.23, p=0.0313). ITPA and GST polymorphisms were not associated with toxicity. GSTM1 deletion was associated with poor clinical response to therapy (OR 3.75, 95% CI 0.940 - 14.97, p=0.1028), however, neither TPMT*3A nor ITPA polymorphisms were associated with clinical response. CONCLUSION: In addition to TPMT for adverse events, genotyping for GSTM1 appears to predict clinical response in IBD patients treated with thiopurines.

Prolonged neutropenia after irinotecan-based chemotherapy in a child with polymorphisms of UGT1A1 and SLCO1B1.

Genetic polymorphisms of uridine diphosphate glucuronosyl transferase 1A1 (UGT1A1), and SLCO1B1 coding organic anion-transporter polypeptide 1B1, are independent risk factors known to increase irinotecan toxicity in adults. Although combined occurrence of polymorphisms in these 2 genes is likely to influence susceptibility to irinotecan toxicity, data are scarce, especially in children. We report an 11-year-old female with severe and prolonged neutropenia after irinotecan-based chemotherapy. The patient's genotyping revealed polymorphisms in both UGT1A1 and SLCO1B1. To our knowledge, this is the first case report of combined genotyping of both UGT1A1 and SLCO1B1 in a child with severe irinotecan toxicity.

Induction of intestinal P-glycoprotein by St John's wort reduces the oral bioavailability of talinolol.

St John's wort (SJW) is known to induce cytochrome P450 (CYP) 3A4 and P-glycoprotein through pregnane X-receptor activation. Our study evaluated the effects of long-term SJW administration on oral and intravenous pharmacokinetics of the nonmetabolized in vivo probe of P-glycoprotein, talinolol, in relation to intestinal P-glycoprotein expression. In a controlled, randomized study (N=9), the pharmacokinetics of oral (50 mg) and intravenous talinolol (30 mg) was determined before and after 12 days SJW (900 mg daily, Jarsin 300). Duodenal biopsies were taken and MDR1 genotypes assessed. SJW reduced the oral talinolol bioavailability by 25% (P=0.049) compared with water control. A 93% increase in oral clearance (P=0.177) and a 31% reduction in area under the serum concentration time curve (AUC; P=0.030) were observed. Renal and nonrenal clearance (CLNR), elimination half-life, peak serum drug concentration (Cmax), and time to reach Cmax were not significantly altered. After intravenous talinolol, SJW affected only CLNR (35% increase compared with water, P=0.006). SJW increased MDR1 messenger ribonucleic acid (mRNA) as well as P-glycoprotein levels in the duodenal mucosa. Subjects with the combined MDR1 genotype comprising 1236C>T, 2677G>T/A, and 3435C>T polymorphisms had lower intestinal MDR1 mRNA levels and displayed an attenuated inductive response to SJW as assessed by talinolol disposition. Long-term SJW decreased talinolol AUC with a corresponding increase in intestinal MDR1 expression, suggesting that SJW has a major inductive effect on intestinal P-glycoprotein. Interestingly, the magnitude of induction appeared to be affected by MDR1 genotype.

Intestinal drug transporter expression and the impact of grapefruit juice in humans.

The goals of this study were to assess the extent of human intestinal drug transporter expression, determine the subcellular localization of the drug uptake transporter OATP1A2, and then to assess the effect of grapefruit juice consumption on OATP1A2 expression relative to cytochrome P450 3A4 and MDR1. Expression of drug uptake and efflux transporters was assessed using human duodenal biopsy samples. Fexofenadine uptake by different transporters was measured in a transporter-transfected cell line. We investigated the influence of grapefruit juice on pharmacokinetics of orally administered fexofenadine. The effect of grapefruit juice on the expression of intestinal transporters was determined using real-time polymerase chain reaction and Western blot analysis. In the duodenum of healthy volunteers, an array of CYP enzymes as well as uptake and efflux transporters was expressed. Importantly, uptake transporters thought to be liver-specific, such as OATP1B1 and 1B3, as well as OATP2B1 and 1A2 were expressed in the intestine. However, among OATP transporters, only OATP1A2 was capable of fexofenadine uptake when assessed in vitro. OATP1A2 colocalized with MDR1 to the brush border domain of enterocytes. Consumption of grapefruit juice concomitantly or 2 h before fexofenadine administration was associated with reduced oral fexofenadine plasma exposure, whereas intestinal expression of either OATP1A2 or MDR1 remained unaffected. In conclusion, an array of drug uptake and efflux transporters are expressed in the human intestine. OATP1A2 is likely the key intestinal uptake transporter for fexofenadine absorption whose inhibition results in the grapefruit juice effect. Although short-term grapefruit juice ingestion was associated with reduced fexofenadine availability, OATP1A2 or MDR1 expression was unaffected.

[Epidemiology of attempted suicide using drugs. An inquiry from the Dresden University Clinic]. / Epidemiologie medikamentöser Parasuizide. Eine Erhebung aus dem Universitätsklinikum Dresden.

BACKGROUND AND OBJECTIVE: Little representative data of the epidemiology of attempted suicide exists in Germany. In this study the frequency of parasuicidal drug intoxication, the distribution of age and gender, as well as the kind and origin of used drugs were evaluated. Furthermore the knowledge about used drugs and possible adverse effects of a previously given medication were analysed. PATIENTS AND METHODS: Over a period of 2 years (January 1998-December 1999) 155 patients (41 males, 114 females, average age 40.5 years) with drug intoxication by attempted suicide were recruited at the University Hospital of Dresden, Germany, for further retrospective analysis. RESULTS: 74 % of these patients were women. Sedatives and hypnotics were most frequently used for parasuicide (44 %), followed by analgesics (18 %) and antidepressants (12 %). Benzodiazepines and benzodiazepine-agonists were the most commonly used drugs (32 %). Moreover, 80 % of all drugs used had been prescribed by physicians. Approximately half of the patients were well-informed about drugs taken. In 43 (47 %) of 92 patients with long-term medication an adverse effect was considered as a possible cause of the attempted suicide. CONCLUSION: Our data underline the importance of attempted suicide in view of the frequency of their use, the need of hospitalization, the required intensive care and possible relapses. Because the majority of drugs used were prescribed by physicians, before giving any medication to their possible suicidal use should be considered.

Clinical relevance of genetic polymorphisms in the human CYP2C9 gene.

Cytochrome p450 (CYP) 2C9 hydroxylates about 16% of drugs in current clinical use. Of special interest are those with a narrow therapeutic index, such as S-warfarin, tolbutamide and phenytoin, where impairment in CYP2C9 metabolic activity might cause difficulties in dose adjustment as well as toxicity. Single-nucleotide polymorphisms (SNP) in the CYP2C9 gene have increasingly been recognized as determinants of the metabolic phenotype that underlies interindividual and ethnic differences. Apart from the wild-type protein CYP2C9*1 at least five allelic variants produce allozymes with reduced or deficient metabolic activity. Among white populations only CYP2C9*2 and CYP2C9*3 variants are of significance, with allelic frequencies of 0.08-0.14 and 0.04-0.16, respectively. In contrast, in Africans (African-Americans and Ethiopians) and Asians both variants are much less frequent (0.005-0.04), and CYP2C9*2 has not yet been detected in Asians. CYP2C9*4 has been exclusively identified in Japanese patients, and CYP2C9*5 and *6 were only found among African-Americans with a low allelic frequency of 0.017 and 0.006, respectively. Furthermore in Japanese a CYP2C9 promotor variant of four linked SNPs was correlated with reduced intrinsic clearance of phenytoin in vitro. Subjects who are carriers of one or more variant alleles may be at risk for adverse drug reactions/toxicities when prescribed drugs extensively metabolized by CYP2C9.

Identification of functionally variant MDR1 alleles among European Americans and African Americans.

MDR1 (P-glycoprotein) is an important factor in the disposition of many drugs, and the involved processes often exhibit considerable interindividual variability that may be genetically determined. Single-strand conformational polymorphism analysis and direct sequencing of exonic MDR1 deoxyribonucleic acid from 37 healthy European American and 23 healthy African American subjects identified 10 single nucleotide polymorphisms (SNPs), including 6 nonsynonymous variants, occurring in various allelic combinations. Population frequencies of the 15 identified alleles varied according to racial background. Two synonymous SNPs (C1236T in exon 12 and C3435T in exon 26) and a nonsynonymous SNP (G2677T, Ala893Ser) in exon 21 were found to be linked (MDR1*2 ) and occurred in 62% of European Americans and 13% of African Americans. In vitro expression of MDR1 encoding Ala893 (MDR1*1 ) or a site-directed Ser893 mutation (MDR1*2 ) indicated enhanced efflux of digoxin by cells expressing the MDR1-Ser893 variant. In vivo functional relevance of this SNP was assessed with the known P-glycoprotein drug substrate fexofenadine as a probe of the transporter's activity. In humans, MDR1*1 and MDR1*2 variants were associated with differences in fexofenadine levels, consistent with the in vitro data, with the area under the plasma level-time curve being almost 40% greater in the *1/*1 genotype compared with the *2/*2 and the *1/*2 heterozygotes having an intermediate value, suggesting enhanced in vivo P-glycoprotein activity among subjects with the MDR1*2 allele. Thus allelic variation in MDR1 is more common than previously recognized and involves multiple SNPs whose allelic frequencies vary between populations, and some of these SNPs are associated with altered P-glycoprotein function.

Identification and functional characterization of a new CYP2C9 variant (CYP2C9*5) expressed among African Americans.

CYP2C9 is a polymorphic gene for which there are four known allelic variants; CYP2C9*1, CYP2C9*2, CYP2C9*3, and CYP2C9*4. In the present study, DNA from 140 European Americans and 120 African Americans was examined by single-strand conformational polymorphism and restriction fragment length polymorphism analyses, resulting in the identification of a new CYP2C9 variant, CYP2C9*5. This variant is derived from a C1080G transversion in exon 7 of CYP2C9 that leads to an Asp360Glu substitution in the encoded protein. The CYP2C9*5 variant was found to be expressed only in African Americans, such that approximately 3% of this population carries the CYP2C9*5 allele. The variant was expressed in, and purified from, insect cells infected with a recombinant baculovirus. Comparative kinetic studies using the purified wild-type protein CYP2C9*1; the Ile359Leu variant, CYP2C9*3; and the Asp360Glu variant, CYP2C9*5 were carried out using (S)-warfarin, diclofenac, and lauric acid as substrates. The major effect of the Asp360Glu mutation was to increase the K(m) value relative to that of CYP2C9*1 for all three substrates: 12-fold higher for (S)-warfarin 7-hydroxylation, 5-fold higher for the 4'-hydroxylation of diclofenac, and 3-fold higher for the omega-1 hydroxylation of lauric acid. V(max) values differed less than K(m) values between the CYP2C9*1 and CYP2C9*5 proteins. In vitro intrinsic clearances for CYP2C9*5, calculated as the ratio of V(max)/K(m), ranged from 8 to 18% of CYP2C9*1 values. The corresponding ratio for CYP2C9*3 was 4 to 13%. Accordingly, the in vitro data suggest that carriers of the CYP2C9*5 allele would eliminate CYP2C9 substrates at slower rates relative to persons expressing the wild-type protein.

P-glycoprotein inhibitor erythromycin increases oral bioavailability of talinolol in humans.

OBJECTIVE: Increased bioavailability of the P-glycoprotein (Pgp) substrates digoxin and cyclosporin due to erythromycin has been observed in vivo. The aim of the present study was to investigate the effect of orally administered erythromycin on the oral bioavailability of the beta-blocker talinolol. Talinolol is a suitable model compound for Pgp drug-drug interaction studies due to its Pgp-related active intestinal secretion and lack of any significant metabolism. METHODS: In a randomized crossover study, the oral pharmacokinetics of talinolol (50 mg) after a concomitant single oral dose of erythromycin (2 g) or placebo were investigated in 9 healthy men. Concentrations of talinolol were measured in serum and urine by HPLC. RESULTS: The area under the curve of talinolol serum concentrations from 0 to 24 h (AUC(0-24)) and the maximum serum concentrations (Cmax) were significantly increased after administration of erythromycin compared to placebo. t(max) values were significantly reduced. The renal clearance (CLR) of talinolol was unchanged after co-administration of erythromycin and there was a small but statistically significant decrease in elimination half-life (t1/2). Serum pharmacokinetics correlate with the results derived from urine concentration measurement. One subject suffered from moderate diarrhea after erythromycin and was excluded from the analysis. CONCLUSION: We suggest that the increase in oral bioavailability of talinolol after concomitant erythromycin is caused by increased intestinal net absorption due to Pgp inhibition by erythromycin.

Regional drug information service.

BACKGROUND: Drug information centers (DICs) were established in Europe more than two decades ago. The majority of German DICs were created in the 90s. The regional University hospital-based DIC, which offers services to physicans, is now in operation for three and a half years . OBJECTIVE: To evaluate the types of enquiries received and the profile of the users of a drug information service. METHODS: The working procedure at a regional center in Dresden, Germany, is described. The topics for consultation (adverse reactions, pharmacokinetics, etc.) are presented, and the types of drugs involved are classified according to the Anatomical Therapeutic Chemical (ATC) classification. Users are grouped by medical specialty. Future plans for the DIC are discussed. RESULTS: A total of 516 enquiries were received. Questions concerning therapeutic use (34%), adverse drug reactions (28%), pregnancy/lactation (16%), and pharmacokinetics/dosage (15%) were asked most frequently. Cardiovascular drugs (20%), systemic antiinfectives (19%) as well as drugs targeting the central nervous system (15%) and alimentation/metabolism (9%) were the predominant foci of enquiries. The major users of the DIC were internists (19%), general practitioners (19%), pediatricians (18%), and gynecologists (11%). CONCLUSIONS: The types of questions and users of this service were generally similar to those recorded at many other European DICs. The service has begun producing educational bulletins on drug-related topics of clinical relevance.

Unexpected effect of verapamil on oral bioavailability of the beta-blocker talinolol in humans.

PURPOSE: To quantitate the effect of verapamil administered orally, a calcium channel blocker and potent inhibitor of P-glycoprotein on oral pharmacokinetics of the beta1-adrenergic receptor antagonist talinolol, a substrate of P-glycoprotein. SUBJECTS AND METHODS: In a randomized, crossover placebo-controlled study, oral pharmacokinetics of talinolol (50 mg) after concomitant administration of single doses of R-verapamil (120 mg) or placebo were investigated in 9 healthy volunteers. Concentrations of talinolol, verapamil, and its main metabolite norverapamil were measured in serum with HPLC. Concentrations of talinolol were also measured in urine by HPLC. Standard pharmacokinetic parameters were calculated with noncompartmental procedures. RESULTS: The area under the concentration-time curve for talinolol from 0 to 24 hours was significantly decreased after R-verapamil versus placebo (721+/-231 ng x h x mL(-1) versus 945+/-188 ng x h x mL(-1); P < .01). Maximum serum concentration of talinolol was reached significantly earlier after R-verapamil compared with placebo (P < .05). Coadministration of R-verapamil did not affect the renal clearance or half-life of talinolol. Serum pharmacokinetics are paralleled by the results derived from urine concentrations of talinolol. CONCLUSION: This is the first study to show a decreased oral bioavailability of a P-glycoprotein substrate (talinolol) in humans as a result of coadministration of verapamil. This effect is assumed to be caused by changes of the intestinal net absorption of talinolol because its renal clearance remains unaffected by administration of R-verapamil. This unexpected effect of R-verapamil is most likely dose dependent as a result of an interplay between intestinal P-glycoprotein and gut metabolism.

Outpatient costs of osteoporosis in a national health insurance population.

There are few valid data on the outpatient diagnosis and treatment of osteoporosis in Germany, despite the high prevalence of this disease and the high costs associated with its complications. We therefore conducted a retrospective cohort study to investigate the prevalence of documented osteoporosis and the use of health care resources in its outpatient treatment in a representative random sample of 7490 patients from the Dresden area who were insured under the national health insurance program for a 1-year period from the 3rd quarter of 1993 to the 2nd quarter of 1994. Documented cases of osteoporosis were identified by International Statistical Classification of Diseases, 10th Revision diagnostic codes M80 to M82, and the costs of diagnostic services for osteoporosis were calculated using a uniform fee schedule. Specific and nonspecific osteoporosis medications were classified using a published anatomic-therapeutic-chemical code, and their costs were calculated on the basis of pharmacy sales prices. Three age- and sex-matched controls without documented osteoporosis (n = 705) were assigned for each case patient in estimating the net use of resources. Data for the region, as well as age-standardized information for the overall German national health insurance system, were calculated. The 1-year prevalence of documented osteoporosis in the region was 3.14% (5.20% in women, 0.89% in men), and the age-standardized prevalence in the German national health insurance system was 2.25%. During the study period, 51.1% of the cases and 2.1% of the controls were examined by osteodensitometry. Patients received 106 defined daily doses of osteoporosis medications during the year; 37.0% of the prescribed daily doses were for sodium fluorophosphate/calcium combinations, 4.3% were for sodium fluoride, and 7.7% were for calcium alone. Sex hormones and calcitonin each accounted for 7.7% of the prescribed daily doses. Only a fraction of epidemiologically expected cases of osteoporosis have been identified and documented in the outpatient sector. Only approximately 50% of these receive osteoporosis-specific therapy, and compliance with therapy is low. To reduce osteoporosis-associated fracture rates, which are extremely cost intensive and greatly impair patients' quality of life, more consistent treatment is needed.