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J Fischer

Publications and source records attributed to J Fischer.

At least 73 records · Page 4Linked to original sources
PubMed2001

Differential antiplatelet effects of various glycoprotein IIb-IIIa antagonists.

The blockade of platelet glycoprotein IIb-IIIa (GPIIb-IIIa) was recently introduced as a new antiplatelet strategy. At present, various GPIIb-IIIa inhibitors are available to treat patients with acute coronary syndrome or when undergoing percutaneous coronary interventions. The current study systematically evaluates the antiplatelet effects of GPIIb-IIIa inhibitors in clinical use. Using conformation-dependent monoclonal antibodies [ligand-induced binding sites (LIBS-1), PMI-1] and flow cytometry, we showed that the GPIIb-IIIa antagonists abciximab, integrelin, lamifiban, and tirofiban, but not EMD 122347 or YM 337, induced LIBS activity of platelet GPIIb-IIIa. The LIBS activity of GPIIb-IIIa antagonists correlates with a proaggregatory response of fixed platelets pretreated with GPIIb-IIIa antagonists (intrinsic activity). All tested GPIIb-IIIa antagonists completely inhibit concentration-dependent ADP (20 micromol/l)-induced aggregation. In contrast, substantial TRAP (25 micromol/l)-induced platelet aggregation still occurs even at high inhibitor concentrations of the tested GPIIb-IIIa antagonists. In addition, we show that GPIIb-IIIa antagonists are poor inhibitors of platelet release reaction (ATP and P-selectin secretion) especially when strong agonists such as TRAP are used to activate platelets. Inhibition of platelet procoagulant activity (thrombin generation) by GPIIb-IIIa antagonists is dependent on the type and concentration of antagonists and on the strength of stimulus (thrombin, tissue factor) used to induce platelet-dependent thrombin generation. The present data show that significant pharmacological differences exist between GPIIb-IIIa antagonists that may have consequences for antithrombotic strategies and for future drug development.

Animals↗
PubMed2001

Whole body positron emission tomography in the treatment of Hodgkin disease.

BACKGROUND: In Hodgkin disease (HD), accurate assessment of the extent of disease is essential because it provides the basis for different treatment strategies. In addition to conventional imaging methods (CIM), positron emission tomography with fluorine-18-fluorodeoxyglucose (FDG-PET) may permit reliable differentiation between lymphoma and nonmalignant tissue and thus improve determination of the stage of the disease. The aim of the current study was to assess the clinical value of FDG-PET for primary staging, treatment monitoring, and assessment in a suspected case of recurrent HD. METHODS: Eighty-one patients with HD underwent 106 FDG-PET studies using a dedicated whole body PET ring scanner. In 25 patients PET was part of the primary staging, 63 PET studies were undertaken for treatment monitoring after the completion of treatment, and in 18 patients PET was performed in cases of suspected recurrence of HD. PET scans were compared with CIM and verified histologically and/or by follow-up evaluation (mean follow-up duration, 20.4 months). RESULTS: With regard to primary staging, in a patient to patient analysis, both PET scans and CIM were positive (i.e., showed pathologic foci indicative of HD) in 24 of 25 cases. In a staging-relevant lesion to lesion analysis, accuracy in the determination of the stage of disease was 96% for PET versus 56% for CIM. PET led to a lower stage classification in 28% and a higher stage classification in 12% of cases, compared with the stage assumed with CIM. With regard to treatment monitoring, PET showed an accuracy of 91% compared with 62% for CIM. The negative predictive value of PET was 96%. With regard to suspected recurrence, PET findings were true-positive in 10 of 12 PET scans and true-negative in 5 of 6 PET scans, resulting in accuracy of 83%, which compares favorably with the accuracy rate of 56% for CIM. CONCLUSIONS: It may be concluded that FDG-PET is capable of determining the stage of HD with great accuracy and is capable of correctly detecting manifestations of HD in treatment monitoring and cases of suspected recurrence, in which CIM occasionally result in equivocal findings. The results of the current study suggest that FDG-PET should become a routine tool in the staging/restaging of HD.

Adult↗
PubMed2001

Detailed deletion mapping in sporadic breast cancer at chromosomal region 17p13 distal to the TP53 gene: association with clinicopathological parameters.

Chromosome 17p is among the most frequently deleted regions in a variety of human malignancies including breast cancer. This study has further refined the localization of a putative tumour suppressor gene (TSG) at 17p13 distal to the TP53 gene in breast carcinomas. It was found that 73% (37 of 51) of the breast tumours exhibited loss of heterozygosity (LOH) at one or more loci at 17p13. The allelic loss patterns of these tumours suggest the presence of at least seven commonly deleted regions on 17p13. The three most frequently deleted regions were mapped at chromosomal location 17p13.3-17p13.2 between the markers D17S831 and D17S1845 (56% LOH), at 17p13.1 between D17S1810 and D17S1832 (53% LOH), and at 17p13.1 between D17S938 and TP53 (55% LOH). A significant correlation was found between loss at 17p13 and tumour grade, size, proliferative activity, and oestrogen receptor (ER) status. Losses at 17p13 were seen more frequently in large and poorly differentiated tumours with high proliferative activity. These data support and extend previous reports on the presence of a putative TSG(s) at chromosomal region 17p13 distal to the TP53 gene and show that different subsets of LOH are associated with more aggressive tumour behaviour.

Adult↗
PubMed2001

Hypermethylation of the tumor necrosis factor receptor superfamily 6 (APT1, Fas, CD95/Apo-1) gene promoter at rel/nuclear factor kappaB sites in prostatic carcinoma.

DNA hypermethylation of CpG-rich promoter sequences is associated with tumor suppressor gene inactivation in many human cancers, notably in carcinoma of the prostate and the urinary bladder. Recently, the mouse homologue of the tumor necrosis factor receptor superfamily 6 (TNFRSF6) gene was reported to be inactivated by DNA methylation in various cell types. The Fas (CD95, Apo-1) protein encoded by the TNFRSF6 gene is an important mediator of apoptosis, which also is downregulated in different types of human carcinoma. We therefore investigated the methylation of the TNFRSF6 promoter in prostatic and bladder carcinomas and cell lines. In a restriction enzyme polymerase chain reaction assay, four of 32 prostatic carcinomas and three of 15 advanced bladder carcinomas showed evidence of hypermethylation at the rel/nuclear factor kappaB (NFkappaB) binding sites essential for promoter activity. The DU145 cell line derived from a metastasis of a prostate carcinoma also displayed hypermethylation in this assay, which was confirmed by bisulfite sequencing. Treatment of DU145 cells with the methylation inhibitor deoxyazacytidine slightly increased Fas protein expression, as detected by flow cytometry analysis. In vitro methylation of the TNFRSF6 promoter at the rel/NFkappaB sites completely abolished its activity. Thus, although the TNFRSF6 gene can be inactivated efficiently by DNA methylation, hypermethylation occurs neither frequently nor extensively in human carcinomas and appears to play a limited role in downregulation of Fas expression.

Aged↗
PubMed2001

The standardization of terminology for researchers in female pelvic floor disorders.

The lack of standardized terminology in pelvic floor disorders (pelvic organ prolapse, urinary incontinence, and fecal incontinence) is a major obstacle to performing and interpreting research. The National Institutes of Health convened the Terminology Workshop for Researchers in Female Pelvic Floor Disorders to: (1) agree on standard terms for defining conditions and outcomes; (2) make recommendations for minimum data collection for research; and (3) identify high priority issues for future research. Pelvic organ prolapse was defined by physical examination staging using the International Continence Society system. Stress urinary incontinence was defined by symptoms and testing; 'cure' was defined as no stress incontinence symptoms, negative testing, and no new problems due to intervention. Overactive bladder was defined as urinary frequency and urgency, with and without urge incontinence. Detrusor instability was defined by cystometry. For all urinary symptoms, defining 'improvement' after intervention was identified as a high priority. For fecal incontinence, more research is needed before recommendations can be made. A standard terminology for research on pelvic floor disorders is presented and areas of high priority for future research are identified.

Fecal Incontinence↗
PubMed2001

Evaluation of a simple method for minimizing iatrogenic blood loss from discard volumes in critically ill newborns and children.

OBJECTIVE: To validate a simple method avoiding discard volumes in pediatric patients with indwelling arterial and venous lines. DESIGN: Zero-discarding was achieved by passive extracorporeal arteriovenous backflow via standard single pressure transducer equipment. We tested backflow distances (10, 20 and 30 cm beyond the sampling port), corresponding to withdrawal volumes of 0.6 ml, 0.8 ml and 1.0 ml, respectively, in comparison to conventional sampling with discard of 0.6 ml. With the backflow technique, the "withdrawal volume" was flushed back to the patient after sampling. We enrolled 120 patients who were allocated to either of the following paired sampling procedures: 10 cm versus conventional, 20 cm versus conventional, 30 cm versus conventional and two paired conventional samples. The order of the sampling was randomly allocated. Bias and precision were determined using Bland-Altman diagrams and algorithms. RESULTS: No appreciable difference was found for blood gases, hemoglobin, potassium and calcium between the backflow technique and conventional sampling. Sodium results and blood glucose showed a bias towards higher values with the backflow technique (mean difference, sodium, 0.9 mmol/l; glucose, mean difference 0.5 mmol/l, standard deviation 0.44 mmol/l). CONCLUSIONS: The backflow technique provides reliable results for blood gases and electrolytes. However, in patients at risk of hypoglycemia, the backflow method should not be used to monitor blood glucose levels.

Blood Gas Analysis↗
PubMed2001

Outcome of women is impaired in patients undergoing emergency coronary artery bypass grafting for failed PTCA.

There is an ongoing debate whether female gender is associated with increased cardiovascular morbidity and mortality, especially after coronary interventions. The impact of gender on the outcome of patients undergoing emergency coronary artery bypass grafting (CABG) for failed PTCA was analyzed. Clinical and procedural data of all patients who underwent PTCA and subsequent emergency CABG at our institution from 1989 to 1998 were assessed. During these 10 years, 6681 PTCA procedures were performed, 1312 in women (19.6%). Subsequently, 110 patients underwent emergency CABG of whom 32 were females (29.1%). Postoperatively, 9 women and 5 men died (mortality 12.7%). Women presented with higher age (61.2 +/- 2.1 vs. 58.3 +/- 1.0 years, n.s.), smaller height (1.61 +/- 0.01 vs. 1.76 +/- 0.01 m, p < 0.0001), lower weight (67.7 +/- 2.4 vs. 82.1 +/- 1.2 kg, p < 0.0001), smaller body surface area (1.70 +/- 0.04 vs. 1.98 +/- 0.02 m2, p < 0.0001), and higher comorbidity as expressed by their Cleveland score (7.9 +/- 0.3 vs. 7.1 +/- 0.2, p = 0.013). The risk for failure of PTCA with subsequent emergency CABG was higher in women than in men (2.4% vs. 1.5%, p = 0.012, odds ratio 1.66) as well as for postoperative death (28.1% vs. 6.4%, p = 0.004, odds ratio 4.39). Women had longer in-hospital stays (19.7 +/- 4.2 vs. 12.9 +/- 1.3 days, p = 0.044). Logistic regression analyses found lower weight (p = 0.003), higher number of diseased coronary vessels (p = 0.024) and higher Cleveland score (p = 0.023) to be independent predictors of operative mortality. A Kaplan-Meier model (follow-up 5.3 +/- 2.5 years) showed an increased in-hospital mortality in women (p = 0.0034, log rang test), but a comparable long-term survival. Women had an increased risk for failure of PTCA and a markedly higher operative mortality after emergency CABG. In multivariate analyses, however, gender was not an independent predictor of postoperative death.

Angioplasty, Balloon, Coronary↗
PubMed2001

Hounsfield unit density in the determination of urinary stone composition.

OBJECTIVES: Noncontrast computed tomography (NCCT) has emerged as the diagnostic study of choice in the evaluation of acute flank pain. Recent in vitro studies have suggested that NCCT can be used to predict the composition of urinary stones on the basis of differences in radiodensity (measured in Hounsfield units, HUs). We sought to determine whether the analysis of in vivo urinary stones seen on NCCT could predict their composition. METHODS: Between March 1997 and August 1999, 100 pure stones from patients seen at the Wilford Hall Medical Center in San Antonio, Texas were submitted for analysis. All had been visualized by NCCT before stone passage or retrieval. A General Electric High-Speed Advantage CT scanner evaluated most of these patients by a "flank pain protocol" (ie, helical technique with breath-holding at 120 kV, 200 mA with 5 mm collimation). Each scan was interpreted by one of two staff radiologists who measured the HUs for each stone. A statistical comparison was made between the stone composition and radiodensity. To allow for subsequently observed increases in radiodensity with increasing stone size regardless of composition, the HU value was divided by each stone's largest transverse diameter in millimeters to give the HU density. A statistical comparison was then made between stone composition and HU density. RESULTS: No significant difference was noted between the HU values of calcium oxalate and calcium phosphate stones, and thus they were analyzed collectively as "calcium stones." When the HU values of calcium (n = 87), uric acid (n = 7), struvite (n = 4), and cystine (n = 2) stones were compared, the overlap of ranges precluded accurate identification, and the mean HU values were not significantly different from one another. There was less overlap noted when comparing the HU densities of the stones studied, and no noncalcium stone had an HU density greater than 76 HU/mm. Using one-way analysis of variance, significant differences were noted between the mean HU density of calcium (105 +/- 43) and uric acid (50 +/- 24) stones (P = 0.006). A trend toward significance was found between the mean HU density of the calcium and struvite stones (53 +/- 28, P = 0.073). No significant differences were found among the other stones. CONCLUSIONS: HU density compared with the HU value alone better characterized differences in radiodensities among urinary stones; calcium stones can be distinguished from uric acid stones on the basis of this value. However, neither the HU density nor the mean HU value was able to identify urinary stones in vivo.

Analysis of Variance↗
PubMed2001

Phenotypic and functional comparison of monocytes from cord blood and granulocyte colony-stimulating factor-mobilized apheresis products.

OBJECTIVE: It is well established that T cells are effector cells in graft-vs-host disease (GVHD), yet the contribution of graft monocytes is less well characterized. Therefore, monocytes in cord blood (CB) and granulocyte colony-stimulating factor-mobilized apheresis products (G-AP), two stem cell grafts associated with reduction of acute and chronic GVHD and relative reduction of acute GVHD, respectively, were compared phenotypically and functionally. MATERIALS AND METHODS: The frequencies, phenotypes, and pinocytosis activities of monocytes from CB and G-AP were determined by flow cytometry and their allostimulatory potential in a primary mixed leukocyte reaction. RESULTS: G-AP contained significantly more monocytes than CB (24.9% +/- 7.1% vs 8.8% +/- 1.5% CD14+ and 62.4 +/- 27.5 x 10(6) vs 0.9 +/- 0.2 x 10(6) CD14+ cells/mL). Monocytes from both sources revealed similar phenotypes. They expressed CD4, CD11a, CD11b, CD11c, CD18, CD32, CD33, CD45R0, CD48, CD50, CD54, CD58, CD64, CD86, CD102, CD116, CD123, and HLA-DR; showed no expression of CD1a and CD83; and weak expression of CD16, CD45RA, and CD80. The levels of CD80 and CD86 expression were comparable; however, in contrast to G-AP monocytes, CB monocytes lacked CD40. There was no difference in pinocytosis activity and allostimulatory capacity of CB and G-AP monocytes. CONCLUSIONS: Monocytes in CB and G-AP are phenotypically and functionally comparable. The only difference observed is the lack of CD40 on CB monocytes.

Antigen Presentation↗
PubMed2001

A prognostic index for 30-day mortality after stroke.

The objective of this study was to develop a simplified scoring system to predict 30-day mortality in patients with acute ischemic stroke. A retrospective cohort study was performed in a tertiary referral hospital in the Hunter Region of Australia. A prognostic index was created by assigning points to the variables in a Cox model. The index included impaired consciousness (5 points), dysphagia (3 points), urinary incontinence (4 points), admission body temperature higher than 36.5 degrees C (2 points), and hyperglycemia without a clinical history of diabetes (2 points). A score of 11 or more defined a high-risk group. The index achieved a sensitivity, specificity, and positive predictive value of 68%, 98% and 75%, respectively, in the derivation sample and 57%, 97% and 68%, respectively, in the validation sample. The results provide a simple risk stratification instrument for clinical research and practice. Further evaluation of the model in a prospective cohort is warranted

Aged↗
PubMed2001

Novel point mutations, deletions, and polymorphisms in the cathepsin C gene in nine families from Europe and North Africa with Papillon-Lefèvre syndrome.

Papillon-Lefèvre syndrome is an autosomal recessive disorder characterized by palmoplantar keratoderma, periodontitis, and premature loss of dentition. Mutations in the CTSC gene that encodes cathepsin C have been described in families affected with Papillon--Lefèvre syndrome. Cathepsin C is the least understood of the lysosomal cysteine proteases; it has been reported to participate in both intracellular and extracellular cleavage of proteins and activation of serine proteases in immune and inflammatory cells. We report here eight new mutations in Papillon-Lefèvre syndrome families: four deletions and four point mutations, including a missense mutation in the propeptide chain that could help elucidate structure-function relationships in this protein. We also found that the 458C > T mutation, first reported in two families by Hart et al (2000c), was a neutral polymorphism in our families, as suggested by Allende et al (Cathepsin C gene: first compound heterozygous patient with Papillon--Lefèvre syndrome and novel symptomless mutation. Hum Mutat 17:152-153, 2001).

Africa, Northern↗
PubMed2001

Use of the intubating laryngeal mask in children: an evaluation using video-endoscopic monitoring.

BACKGROUND AND OBJECTIVE: The intubating laryngeal mask is designed to act as a ventilatory device and as an aid to blind tracheal intubation in adults. The aim of this study was to evaluate the efficacy of the intubating laryngeal mask for ventilation of the lungs and tracheal intubation in children using video-endoscopic control. METHODS: The handling and efficacy of the size 3 intubating laryngeal mask for tracheal intubation in 80 children weighing > or = 25 kg were assessed under video-endoscopic control. Ease of intubating laryngeal mask insertion, adequacy of lung ventilation through the intubating laryngeal mask and airway sealing pressures were recorded. Tracheal intubation was performed blindly by the intubator, while the supervisor observed the procedure on the video display. If blind intubation failed at the first attempt, the monitor view was used to guide the tracheal tube into the trachea. The success rate and time required for successfully placing the tracheal tube were recorded. RESULTS: Insertion of the intubating laryngeal mask was easy in all children. Lung ventilation through the intubating laryngeal mask was uniformly excellent. Blind tracheal intubation at the first attempt was successful in 53 children (66%) within 18.8 +/- 4.1 s. Twenty-four of the 27 failed blind intubation attempts were successfully intubated with video-endoscopic guidance within 28.6 +/- 9.4 s. Two children required replacing the intubating laryngeal mask, one child had to be intubated conventionally. CONCLUSIONS: The size 3 intubating laryngeal mask provides an airway that is easy to establish in children > or = 25 kg with excellent ventilation conditions and allows blind tracheal intubation at the first attempt with a high success rate. Endoscopic monitoring improves its safety and intubation success rate.

Adolescent↗
PubMed2001

Start-up delays of infusion syringe pumps.

BACKGROUND: We performed a bench experiment to investigate the extent of start-up delays in fluid delivery for four different syringe pumps after initially placing the infusion syringe in the syringe pump. METHODS: Pump performance was determined at an infusion rate of 1 ml.h-1 with and without a fluid bolus delivered by the infusion pump prior to connecting the infusion line to the simulated patient. RESULTS: The time (mean +/- SD) from starting the pump up to first fluid delivery (t1) differed considerably between pumps (from 6.75 +/- 4.4 to 57.2 +/- 28.6 min) as did the time to steady state fluid delivery (t2) (from 19.6 +/- 9.3 to 76.3 +/- 29.0 min). Applying an initial bolus of 2 ml before connecting the line to the simulated patient practically eliminated the delay in fluid delivery (t1 ranging from 0.3 +/- 0.1 to 1.1 +/- 0.8 min). This manoeuvre also reduced the time to steady flow delivery (t2 from 6.0 +/- 3.1 to 11.1 +/- 4.3 min, P<0.001) and minimized the differences between syringe pumps. CONCLUSIONS: Syringe pump design affects start-up delay times because of free play of the syringe. These delays can be eliminated by a start-up bolus of 2 ml prior to connecting the infusion line to the patient.

Equipment Design↗
PubMed2001

The history of the Glasgow Coma Scale: implications for practice.

The Glasgow Coma Scale (GCS) has been the gold standard of neurologic assessment for trauma patients since its development by Jennett and Teasdale in the early 1970s. The GCS was found to be a simple tool to use. It became the method of choice for trauma care practitioners to document neurologic findings over time and predict functional outcome. Although the scale has been shown to be effective, many authors have cited weaknesses in the scale including the inability to predict outcome, variation in inter-rater reliability, and the inconsistent use by caregivers in the prehospital and hospital settings. This article outlines the components of the GCS and how practitioners can best use the scale, particularly in patients whose injuries and treatments make them difficult to assess.

Adult↗
PubMed2001

The C3435T mutation in the human MDR1 gene is associated with altered efflux of the P-glycoprotein substrate rhodamine 123 from CD56+ natural killer cells.

P-glycoprotein (PGP) is a membrane protein which determines drug disposition in humans (e.g. digoxin). It is also expressed in various leukocyte lineages with highest expression in CD56+ natural killer cells. Recently, a polymorphism in exon 26 (C3435T) of this gene was shown to correlate with intestinal PGP expression and function in humans. Carriers homozygous for this polymorphism (TT) showed more than two-fold lower PGP expression and higher digoxin plasma concentrations compared to the CC group. However, it is not known whether this mutation in the MDR1 gene is also associated with altered PGP function in peripheral blood cells. We therefore assessed efflux of the PGP-substrate rhodamine 123 from CD56+ natural killer cells. Leukocytes were isolated from whole blood of 10 CC, 10 CT and 11 TT healthy Caucasian individuals. Using flow cytometry, rhodamine fluorescence was determined in CD56+ cells. Moreover, MDRI mRNA was quantified in leukocytes by real-time polymerase chain reaction. Subjects with CC genotype revealed a significantly lower rhodamine fluorescence (i.e. higher PGP function) compared to individuals with TT genotype (51.1 +/- 11.4% versus 67.5 +/- 9.5%, p < 0.01). Heterozygous individuals had an intermediate rhodamine fluorescence (61.4 +/- 6.3%). MDR1 mRNA normalized for cyclophilin was lowest in the TT population (1.29 +/- 1.01), intermediate in heterozygous subjects (1.60 +/- 0.76) and highest in the CC group (1.91 +/- 0.94; not significant). In summary, subjects being homozygous for C in position 3435 of the MDR1 gene have a more pronounced efflux of rhodamine from CD56+ natural killer cells and a higher MDR1 mRNA expression in leukocytes than subjects with the TT genotype. Measurement of rhodamine efflux using flow-cytometry from peripheral blood cells allows assessment of genetically determined differences in P-glycoprotein function.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

Extensive genetic polymorphism in the human CYP2B6 gene with impact on expression and function in human liver.

The human cytochrome P450, CYP2B6, is involved in the metabolism of several therapeutically important drugs and environmental or abused toxicants. In this study, we present the first systematic investigation of genetic polymorphism in the CYP2B6 gene on chromosome 19. A specific direct sequencing strategy was developed based on CYP2B6 and CYP2B7 genomic sequence information and DNA from 35 subjects was completely analysed for mutations throughout all nine exons and their exon-intron boundaries. A total of nine novel point mutations were identified, of which five result in amino acid substitutions in exon 1 (C64T, Arg22Cys), exon 4 (G516T, Gln172His), exon 5 (C777A, Ser259Arg and A785G, Lys262Arg) and exon 9 (C1459T, Arg487Cys) and four are silent mutations (C78T, G216C, G714A and C732T). Polymerase chain reaction-restriction fragment length polymorphism tests were developed to detect each of the five nonsynonymous mutations in genomic DNA. By screening a population of 215 subjects the C64T, G516T, C777A, A785G and C1459T mutations were found at frequencies of 5.3%, 28.6%, 0.5%, 32.6% and 14.0%, respectively. Haplotype analysis revealed six different mutant alleles termed CYP2B6*2 (C64T), *3 (C777A), *4 (A785G), *5 (C1459T), *6 (G516T and A785G) and *7 (G516T, A785G and C1459T). By analysing a large number of human liver samples, significantly reduced CYP2B6 protein expression and S-mephenytoin N-demethylase activity were found in carriers of the C1459T (R487C) mutation (alleles *5 and *7). These data demonstrate that the extensive interindividual variability of CYP2B6 expression and function is not only due to regulatory phenomena, but also caused by a common genetic polymorphism.

Aryl Hydrocarbon Hydroxylases↗
PubMed2001

Comprehensive analysis of the genetic factors determining expression and function of hepatic CYP2D6.

Variable expression and function of the cytochrome P4502D6 (CYP2D6) leads to distinct phenotypes termed ultrarapid (UM), extensive (EM), intermediate (IM) and poor metabolizer (PM). Whereas the PM phenotype is known to be caused by two null-alleles leading to absence of functional CYP2D6 protein, the large variability among individuals with functional alleles remained largely unexplained. In this study, we systematically investigated 76 liver biopsies from individuals with known sparteine metabolic ratios (MRS) for the relationships between CYP2D6 genotype, microsomal protein expression, bufuralol 1'-hydroxylase activity and in-vivo phenotype. Average CYP2D6 protein levels ranged from undetectable in PMs (MRS > 20) to 2.6 +/- 2.7 pmol/mg microsomal protein in IMs (1.2 < MRS< 20), 7.6 +/- 4.7 in EMs (0.2 < MRS < 1.2) and 23.8 +/- 7.7 in UMs (MRS < 0.2), respectively. Analysis with respect to genotype demonstrated gradually increased expression and function for individuals with no, one, two or three functional gene copies per genome. The recently discovered -1584 C/G promoter polymorphism was identified as another major factor for expression and function with the mutant [-1584G] promoter type being consistently associated with significantly higher expression than [-1584C]. To investigate functional differences between the detected variant protein forms CYP2D6.1, 2D6.2, 2D6.9 and 2D6.10, we expressed them recombinantly in insect cells. The most significant difference was a decrease in the relative P450 holoprotein content of all allelic forms, including the common functional variant 2D6.2, in comparison to 2D6.1, whereas only modest Km changes were observed. Taken together, these data provide further insight into the complex mechanisms that govern the highly variable expression and function of CYP2D6.

Cytochrome P-450 CYP2D6↗