Alpha 1-acid Glycoprotein Concentration Research Articles

Critical Assessment of the Diagnostic Value of Feline α1-Acid Glycoprotein for Feline Infectious Peritonitis Using the Likelihood Ratios Approach

Alpha-1-acid glycoprotein (AGP) increases in the blood of cats with feline infectious peritonitis (FIP), a lethal disease caused by feline coronavirus (FCoV). However, the diagnostic potential of AGP might be limited because AGP also increases in pathophysiological conditions other than FIP. In this retrospective study, the diagnostic potential of serum AGP concentration was evaluated on the basis of the pretest probability of disease, according to the Bayesian approach. Serum AGP levels from cats with FIP (group 1; n = 58) and without FIP (group 2; n = 104) were evaluated. Non-FIP cats were further subgrouped as follows: 2a) inflammation (n = 26), 2b) asymptomatic FCoV infection (n = 49), 2c) injection-site sarcoma (n = 19), 2d) postvaccination (n = 7), and 2e) specific pathogen free (n = 3). Standard descriptive analyses by group and empirical receiver-operating characteristic (ROC) curve estimation were performed. Ordinary logistic regression analysis was performed to derive an estimate of the continuous likelihood ratio to produce the posttest probability of disease for any combination of pretest probability and serum AGP value. The comparison of serum AGP levels in the different groups and the analysis of the ROC curve confirmed that serum AGP is a powerful discriminating marker for FIP. The Bayesian approach demonstrated that when the pretest probability of FIP is high, based on history and clinical signs (groups 1 or 2a), moderate serum AGP levels (1.5-2 mg/ml) can discriminate cats with FIP from others, while only high serum AGP levels (>3 mg/ml) can support a diagnosis of FIP in cats with a low pretest probability of disease (groups 2b to 2e).

Broiler chicken health, welfare and fluctuating asymmetry in organic versus conventional production systems

The aim of this study was to test three predictions: (1) that a combination of animal-based parameters indicates better health and welfare in broilers from organic versus conventional farms, (2) that broilers from organic farms show less fluctuating asymmetry (FA) than broilers from conventional farms, and (3) that, at the level of the individual bird, the relationship between FA and welfare is negative and strongest in conventional broilers. On 140 slaughter-age birds randomly selected from seven organic and seven conventional flocks in Belgium we measured standardised FA and five conventional animal-based welfare indicators (tonic immobility duration, latency-to-lie, and condition of the foot pad, hock, and breast). The caeca from the birds from four organic and four conventional flocks were removed for assessing the presence of two bacterial pathogens ( Salmonella and Campylobacter) and the concentration of the health-promoting lactic acid bacteria. Finally, the blood serum concentration of the acute phase protein alpha-1-acid glycoprotein (AGP), a non-specific indicator of immunological stress, was determined. Salmonella was found in the caeca content of two (2.5%) birds only (both from the same conventional farm), whereas 44 (55.7%) birds were infected with Campylobacter. The prevalence of Campylobacter, the concentration of lactic acid bacteria, the duration of tonic immobility, and the condition of the breast and foot pad did not differ between both production systems. Apart from a higher concentration of AGP, organic birds had better scores for hock condition and a longer latency-to-lie indicating better leg health. In addition, organic birds scored better on the aggregated welfare index (i.e. the average of the five standardised welfare indicators). As both production systems differed in many aspects (e.g. slower-growing genotypes, slaughter age, feed, stocking density, group size) it is impossible to assign differences in welfare/health indicators to a single factor. Whatever the causes may be our findings suggest that, despite the potentially elevated risk of immunological challenge, broiler chicken welfare is generally superior in organic farms as compared with conventional farms in Belgium. Regarding the validity of FA as welfare indicator, the prediction of lower FA in the population with highest aggregated welfare score was confirmed but, at the level of the individual, no associations between FA and the aggregated welfare index were found irrespective of whether data from organic and conventional broilers were analysed separately or combined.

Safety of 96-Hour Incision-Site Continuous Infusion of Ropivacaine for Postoperative Analgesia After Bowel Cancer Resection

The aim of this study was to examine the safety of ropivacaine given to patients as a continuous infusion [0.2% (2 mg/mL), 5 mL/h for 96 hours] into a right lateral transverse incision using a portable elastomeric infusion pump after colon cancer resection. Blood samples were collected throughout the infusion and up to 12 hours after infusion and were analyzed by high-performance liquid chromatography (HPLC) for total and unbound ropivacaine concentrations in plasma. Alpha1 acid glycoprotein (AAG) concentrations were measured at 0 and 48 hours to assess possible changes in ropivacaine protein binding after surgery. Postoperative pain control was assessed using 12 hour visual analog scale (VAS) scores. Patient-controlled analgesia (PCA) using fentanyl was freely available in parallel for breakthrough pain, with usage and consumption compared with a historical cohort. The mean +/- SD Cmax total plasma ropivacaine concentration (n = 5) from 12 hours to the end of the infusion was 4.5 +/- 2.6 mg/L, comparable with the previously published threshold for CNS toxicity in the most sensitive patient studied (3.4 mg/L). However, the corresponding maximum unbound ropivacaine concentration (ie, the pharmacologically active moiety) of 0.07 +/- 0.01 mg/L ranged from four- to sevenfold below the reported toxicity threshold (0.34 mg/L). The apparently greater safety margin seen with unbound ropivacaine may have resulted from a significant increase (mean 63%, P < 0.05) in AAG concentrations measured at 48 hours after surgery. This reduction resulted from the known AAG reaction after surgical intervention, resulting in a reducing unbound ropivacaine fraction throughout the 96 hour infusion in all patients. Mean subjective 12 hour pain scale scores at rest and on movement showed large variability between patients. No signs or symptoms of ropivacaine toxicity were observed or reported on questioning. In this limited sample, extending the infusion period from the presently approved 48 hours to 96 hours seems to be a safe alternative and/or adjunct to standard opiate analgesia after colorectal surgery using a right lateral transverse incision, hence reducing the incidence of opiate adverse effects and enhancing recovery. Unbound ropivacaine concentrations suggest there is scope for testing elevated doses to enhance efficacy further.

Effects of Adequate or Marginal Excess of Dietary Methionine Hydroxy Analogue Free Acid on Growth Performance, Edible Meat Yields and Inflammatory Response in Female Broiler Chickens

Two experiments were conducted to determine the effects of adequate or marginal excess of dietary DL-2-hydroxy-4-(methylthio) butanoic acid (MHA) on productive performance and inflammatory response induced by E. coli lipopolysaccharide (LPS). In first experiment, day-old-female chicks (Ross strain) were provided for 48 days with four experimental diets; (1) AB diet: a diet with antibiotics (AB) and DL-methionine (DLM) to meet the total sulfur amino acid (TSAA) requirements of broiler chickens, (2) DLM diet: the first diet without AB, (3) MHA diet: a diet supplemented with MHA in place of DLM of the diet 2 to meet the TSAA requirement, (4) excess MHA diet: a diet with 1.6 folds higher than the content of MHA in the MHA diet. All diets were formulated with ingredients of plant origin. There was no difference in growth performance among the dietary treatments. Edible meat yields in chicks fed the excess MHA diet were better than that in chicks fed the DLM or MHA diet. Chicks fed the excess MHA or MHA diet showed lower plasma ceruloplasmin and alpha-1 acid glycoprotein concentration following LPS injection than those fed the DLM diet at 23 days of age. The concentrations in chicks fed the excess MHA diet were comparable to those chicks fed the AB diet. In second experiment, 7-days -old female chicks were fed either of the DLM or MHA diet for 14 days. Chicks fed the MHA showed higher excreta dry matter content than chicks fed the DLM diet. Heterophil to lymphocyte ratio in blood after LPS injection was lower in chicks fed the MHA diet than that in chicks fed the DLM diet. These results suggest that marginal excess supplementation of MHA, in place of DLM, to diet improved meat production in female broiler chicks and dietary MHA has potential to alleviate certain stress responses.

Morphine in Children with Cancer

The aim of this study was to contrast protein binding of morphine and morphine-6 glucuronide in cord blood and children with adults and examine impact of chemotherapy and other factors. Morphine binding was measured in spiked samples from 18 adults and 18 neonates (cord blood), and compared with six children with cancer receiving morphine. The influence of the following was examined: Human serum albumin (HSA), alpha-1 acid glycoprotein (AAG), non-esterified fatty acids (NEFA); palmitic acid and oleic acid, pH, vincristine, methotrexate, 6-mercaptopurine and M6G. binding correlated with concentrations of albumin and alphal acid glycoprotein. In vitro, binding was not altered by vincristine, 6-mercaptopurine, methotrexate or M6G. Compared with HSA alone, AAG increased binding, palmitic acid reduced it and oleic acid had no effect. Binding was unaffected by pH in samples from patients. Morphine binding was influenced by concentrations of albumin, AAG and morphine itself, but not by age.

Acute phase protein response in an experimental model of ovine caseous lymphadenitis

BackgroundCaseous lymphadenitis (CLA) is a disease of small ruminants caused by Corynebacterium pseudotuberculosis. The pathogenesis of CLA is a slow process, and produces a chronic rather than an acute disease state. Acute phase proteins (APP) such as haptoglobin (Hp) serum amyloid A (SAA) and α1 acid glycoprotein (AGP) are produced by the liver and released into the circulation in response to pro-inflammatory cytokines. The concentration of Hp in serum increases in experimental CLA but it is not known if SAA and AGP respond in parallel or have differing response profiles.ResultsThe concentration in serum of Hp, SAA and AGP in 6 sheep challenged with 2 × 105 cells of C. pseudotuberculosis showed significant increases (P < 0.05) compared to 3 unchallenged control sheep. By day 7 post infection. (p.i.) the Hp and SAA concentrations reached mean (± SEM) values of 1.65 ± 0.21 g/L and 18.1 ± 5.2 mg/L respectively. Thereafter, their concentrations fell with no significant difference to those of the control sheep by day 18 p.i.. In contrast, the serum AGP concentration in infected sheep continued to rise to a peak of 0.38 ± 0.05 g/L on day 13 p.i., after which a slow decline occurred, although the mean concentration remained significantly higher (P < 0.05) than the control group up to 29 days p.i.. Specific IgG to phospholidase D of C. pseudotuberculosis became detectable at 11 days p.i. and continued to rise throughout the experiment.ConclusionThe serum concentrations of Hp, SAA and AGP were raised in sheep in an experimental model of CLA. An extended response was found for AGP which occurred at a point when the infection was likely to have been transforming from an acute to a chronic phase. The results suggest that AGP could have a role as a marker for chronic conditions in sheep.

Microheterogeneity of acute phase proteins in patients with ulcerative colitis.

To estimate the serum alpha1-antichymotrypsin (ACT), alpha1-acid glycoprotein (AGP) and transferrin (Tf) concentrations and to evaluate the microheterogeneity of these acute phase proteins in patients with ulcerative colitis. Twenty-seven patients with ulcerative colitis (UC) and 17 healthy control subjects were studied. The patients were categorised as severe (n = 9), moderate (n = 10) and mild groups (n = 8) using Truelove and Witts' classification of ulcerative colitis. Microheterogeneity of ACT, AGP and Tf was analysed by crossed immunoaffinity electrophoresis (CIAE) with concanavalin A. In all serum samples standard electrophoresis of serum proteins was performed, iron (Fe) concentration, total iron binding capacity (TIBC) and C-reactive protein (CRP) were also measured. Our patients suffering from ulcerative colitis had significantly higher serum ACT and AGP concentrations and lower serum transferrin concentration in comparison to healthy subjects. Changes in concentrations of acute phase proteins were dependent on the activity of the inflammatory process. The glycosylation patterns of transferrin were related to the inflammation status. We also observed the correlation between ACT and AGP concentrations, patterns of transferrin glycosylation and changes in standard protein electrophoresis or blood cell count. The glycosylation patterns of transferrin obtained from patients suffering from ulcerative colitis are highly branched and sialylated compared with those obtained from healthy subjects. In contrast, the glycosylation patterns of transferrin do not differ according to the activity index of ulcerative colitis. The microheterogeneity patterns of AGP and ACT are similar in ulcerative colitis patients and healthy subjects.

Population pharmacokinetics of imatinib and the role of α1‐acid glycoprotein

The aims of this observational study were to assess the variability in imatinib pharmacokinetics and to explore the relationship between its disposition and various biological covariates, especially plasma alpha1-acid glycoprotein concentrations. A population pharmacokinetic analysis was performed using NONMEM based on 321 plasma samples from 59 patients with either chronic myeloid leukaemia or gastrointestinal stromal tumours. The influence of covariates on oral clearance and volume of distribution was examined. Furthermore, the in vivo intracellular pharmacokinetics of imatinib was explored in five patients. A one-compartment model with first-order absorption appropriately described the data, giving a mean (+/-SEM) oral clearance of 14.3 l h-1 (+/-1.0) and a volume of distribution of 347 l (+/-62). Oral clearance was influenced by body weight, age, sex and disease diagnosis. A large proportion of the interindividual variability (36% of clearance and 63% of volume of distribution) remained unexplained by these demographic covariates. Plasma alpha1-acid glycoprotein concentrations had a marked influence on total imatinib concentrations. Moreover, we observed an intra/extracellular ratio of 8, suggesting substantial uptake of the drug into the target cells. Because of the high pharmacokinetic variability of imatinib and the reported relationships between its plasma concentration and efficacy and toxicity, the usefulness of therapeutic drug monitoring as an aid to optimizing therapy should be further investigated. Ideally, such an approach should take account of either circulating alpha1-acid glycoprotein concentrations or free imatinib concentrations.

Controlled Self-Assembly of Collagen Fibrils by an Automated Dialysis System

In vitro self-assembled collagen fibrils form a variety of different structures during dialysis. The self-assembly is dependent on several parameters, such as concentrations of collagen and alpha1-acid glycoprotein, temperature, dialysis time, and the acid concentration. For a detailed understanding of the assembly pathway and structural features like banding pattern or mechanical properties it is necessary to study single collagen fibrils. In this work we present a fully automated system to control the permeation of molecules through a membrane like a dialysis tubing. This allows us to ramp arbitrary diffusion rate profiles during the self-assembly process of macromolecules, such as collagen. The system combines a molecular sieving method with a computer assisted control system for measuring process variables. With the regulation of the diffusion rate it is possible to control and manipulate the collagen self-assembly process during the whole process time. Its performance is demonstrated by the preparation of various collagen type I fibrils and native collagen type II fibrils. The combination with the atomic force microscope (AFM) allows a high resolution characterization of the self-assembled fibrils. In principle, the represented system can be also applied for the production of other biomolecules, where a dialysis enhanced self-assembly process is used.

Urinary alpha‐1‐acid glycoprotein as a biomaker of ARI‐induced inflammation

We enrolled 17 patients, 19–30 y-old, 10 males and 7 females with acute respiratory infection (ARI) and followed them for two weeks. Patients provided blood and urine samples at 0, 7 and 14 d. Serum C-reactive protein (CRP), alpha-1-acid glycoprotein (AGP), and alpha-1 antichymotrypsin (ACT) were significantly higher at 0 than at 14 d, but their dynamics differed from each other. CRP declined abruptly from 0 to 14 d whereas AGP was gradual, and ACT lagged compared to the other proteins. Serum CRP >10 mg/L or AGP > 1.0 g/L were used to diagnose inflammation. Serum creatinine concentrations and glomerular filtration rates were within normal limits for this age group, and did not differ significantly between acute infection and convalescence. Mean urinary AGP concentration was three times higher during the acute phase of infection than at 7 and 14 d after (P< 0.05). Sensitivity and specificity analyses of inflammation based on urinary AGP vs. inflammation based on the combination of serum CRP or AGP at 0 and 14 d showed 77-75% sensitivity, respectively, and 100-90% specificity, respectively. Collection of urine samples required no invasive or painful procedures, and a small urine volume, 15 μl, was necessary to measure AGP by radial immunodiffusion. These results showed that urinary AGP is a simple non-invasive method of diagnosing inflammation due to ARI. Support: USAID cooperative agreement.

Phase I/II study of weekly docetaxel dose escalation in combination with fixed weekly cisplatin and concurrent thoracic radiotherapy in locally advanced non-small cell lung cancer

We conducted a phase I study to determine the maximum-tolerated dose (MTD) and dose-limiting toxicities (DLT) of weekly docetaxel and cisplatin (DOC/CDDP) with concurrent thoracic radiotherapy (TRT) in patients with unresectable stage III non-small-cell lung cancer (NSCLC). The DOC/CDDP administration schedules consisted of a split schedule (SS) with administration in 3 out of every 4 weeks, and a continuous schedule (CS) with administration every week. TRT was given to a total dose of 60 Gy at 2 Gy per fraction over 6 weeks. Twenty-one patients entered the study. The patient characteristics were: PS 0/1/2, 6/13/2; Sq/Ad, 16/5; stage IIIA/IIIB, 4/17. The principal DLT was grade3 esophagitis. The MTD of DOC on the SS and CS in combination with CDDP (25 mg/m2/week) was 25 and 20 mg/m(2)/week, respectively. We determined the RD and schedule of DOC/CDDP on the SS to be 20/25 mg/m(2)/week. The serum alpha-1-acid glycoprotein (AAG) concentration values were found to be negatively correlated with the grade of esophagitis. The median survival time was 23.1 months. The chemoradiation regimen tested in this study has promising activity and manageable toxicity. The continuous schedule could not be recommended due to excessive toxicity. The main DLT was esophagitis, and it significantly correlated with the plasma AAG concentration.

High-performance liquid chromatography determination of α1-acid glycoprotein in small volumes of plasma from neonates

In order to investigate how the alpha1-acid glycoprotein (AGP) concentrations of neonates change in response to surgical stress, a simple high-performance liquid chromatography (HPLC)-assay for the measurement of alpha1-acid glycoprotein levels was developed. A fraction containing alpha1-acid glycoprotein was isolated from the bulk of plasma protein by addition of 0.6M perchloric acid and was then analysed directly on a short PLRP-S 4000A reversed phase column column. The method was validated by analysis of pooled plasma from healthy adults both in comparison with a calibration curve and by standard additions. The procedure was able to isolate alpha1-acid glycoprotein rapidly (<30 min) and required only 50 microl of plasma. The mean extraction recovery was 79.1% (CV 6.4%). The within-run precision for the analysis of three replicates of quality control sample ranged from +/-1.2 to +/-3.8% and the between-run precision was +/-6.1%. The method was linear (r(2)=0.988) over a concentration range from 6 to 100.0 mg/100 ml. The AGP levels in neonatal samples ranged from 25 to 93 mg/100 ml.

An immunoturbidimetric assay for rapid quantitative measurement of feline alpha‐1‐acid glycoprotein in serum and peritoneal fluid

Alpha-1-acid glycoprotein (AGP) is an acute phase protein that increases in concentration in infectious and inflammatory conditions. The serum and peritoneal fluid concentrations of AGP may be useful in the diagnosis of feline infectious peritonitis (FIP), a lethal disease of cats. Currently AGP can be measured by radioimmunodiffusion (RID) assays, which are time consuming and difficult. The objectives of this study were to develop a rapid immunoturbidimetric assay for measurement of AGP in feline serum and peritoneal fluid and to compare the results with those obtained by RID. AGP was purified by perchloric acid precipitation and ion-exchange chromatography from a pool of peritoneal fluid obtained from cats with FIP, as determined by a panel of laboratory tests, including serum AGP concentration, albumin: globulin ratio, and total protein concentration, anti-coronavirus antibody titers, and effusion analysis. The purified AGP in a complete Freund's adjuvant and Tween 20 mixture was injected into a sheep and blood was collected at monthly intervals. Anti-AGP antiserum, as confirmed by ELISA and Western blot techniques, and a pool of peritoneal fluid from cats with FIP were used to prepare standards. Clinical samples of feline peritoneal fluid (n=55) and serum (n=59) were assayed for AGP and results from the immunoturbidimetric and RID methods were compared. Significant correlation (P < .001) was obtained between methods for both peritoneal fluid (R2=.9259) and serum (R2=.9448) samples. Coefficients of variation for the immunoturbidimetric method were <5%. This rapid immunoturbidimetric assay for measurement of feline AGP in serum and peritoneal fluid may be of value in the diagnosis of FIP and possibly other inflammatory diseases in cats.

Prevention of Bone Loss by Phloridzin, an Apple Polyphenol, in Ovariectomized Rats under Inflammation Conditions

Aging and sex hormones related changes lead to inflammatory and oxidant conditions, which are involved in the pathogenesis of osteoporosis. Recent studies have suggested that polyphenols may exert a protective effect in such conditions. We assessed the effect of phloridzin (Phlo), a flavonoid exclusively found in apple, on bone metabolism in ovariectomized (OVX) or sham-operated (SH) rats with and without inflammation. Six-month-old Wistar rats were allocated to two equal groups that received either a control diet or a diet supplemented with 0.25% Phlo for 80 days. Three weeks before necropsy, inflammation was induced by subcutaneous injection of talc in 10 animals of each group. At necropsy, ovariectomy decreased both total (T-BMD) and metaphyseal (M-BMD) femoral bone mineral density (P < 0.01). Inflammation conditions, checked by an increase in the spleen weight and alpha1-acid glycoprotein concentration in OVX rats, exacerbated the decrease in T-BMD (g/cm2) (as well as M-BMD) observed in castrated animals (P < 0.05). Daily Phlo intake prevented ovariectomy-induced bone loss in conditions of inflammation as shown by T-BMD and M-BMD (P < 0.05). At the diaphyseal site, BMD was improved by Phlo in OVX rats with or without inflammation (P < 0.05). These results could be explained by changes in bone remodeling as the increased urinary deoxypyridinoline excretion in OVX and OVXinf animals was prevented by the polyphenol-rich diet (P < 0.001), while plasma osteocalcin concentration was similar in all experimental groups. In conclusion, Phlo consumption may provide protection against ovariectomy-induced osteopenia under inflammation conditions by improving inflammation markers and bone resorption.

Nitric Oxide Is Not a Mediator of Inflammation-Induced Resistance to Atracurium

Resistance to atracurium as a result of increased drug binding to alpha1-acid glycoprotein is associated with increased inducible nitric oxide synthase activity and increased nitric oxide levels in plasma. We investigated if the inhibition of inducible nitric oxide synthase and suppression of nitric oxide can reverse the resistance to atracurium. As a model of alpha1-acid glycoprotein and nitric oxide increase, 84 male Sprague-Dawley rats received an IV injection of either 60 mg/kg Corynebacterium parvum (CP) or saline (control). The 2 groups (CP/Control) were further divided into subgroups, receiving the selective inducible nitric oxide synthase inhibitor, N-Iminolysine, via drinking water at different concentrations. On day 4 post-CP injection, the pharmacodynamics of atracurium were determined. Plasma concentrations of nitric oxide, atracurium, and alpha1-acid glycoprotein were measured and acetylcholine receptor numbers were quantified. In the CP groups, N-Iminolysine suppressed nitric oxide levels in a dose-dependent manner. Resistance to atracurium persisted. alpha1-acid glycoprotein serum levels remained increased in all CP groups with no differences in acetylcholine receptor expression. Our results suggest that the mechanism leading to increased expression of alpha1-acid glycoprotein and consecutive increased protein binding of atracurium is not mediated by inducible nitric oxide synthase induction and nitric oxide expression.

Identification of the bovine α1-acid glycoprotein in colostrum and milk

Alpha1-acid glycoprotein (AGP) is an immunomodulatory protein expressed by hepatocytes in response to the systemic reaction that follows tissue damage caused by inflammation, infection or trauma. This paper presents the detection of bovine AGP (boAGP) in mammary secretions (colostrum and milk) and mammary gland tissue. Bovine AGP was detected by Western blotting in all the samples analysed, and could be quantified in colostrum at 162 (+/- 63.7) microg/mL and 114.5 (+/- 67.8) microg/mL during the first 12 h and 24 h respectively. In mature milk, the boAGP concentration clearly decreased and was no longer detectable using the Radial Immunodiffusion (RID) technique. The concentration of mature milk boAGP was therefore semi-quantified using an anion-exchange chromatographic procedure that allowed the concentration of the protein to be determined. The presence of AGP in bovine milk was confirmed by the internal sequence analysis performed following purification to homogeneity of the protein from milk. The concentration of AGP in bovine milk with low SCC (< 250,000) was very similar to that from bovine milk with high SCC (> 250,000). In order to investigate the origin of AGP in bovine milk, a search for mRNA was carried out in somatic cells and mammary gland tissue: mRNA expression of the boAGP gene was detected in mammary gland tissue, but not in somatic cells. Finally, the cDNA sequence of the boAGP was determined, and is hereby presented.

Phase I Trial of the Cyclin-Dependent Kinase Inhibitor and Protein Kinase C Inhibitor 7-Hydroxystaurosporine in Combination With Fluorouracil in Patients With Advanced Solid Tumors

Preclinical studies indicate that the cyclin-dependent kinase and protein kinase C inhibitor 7-hydroxystaurosporine (UCN-01) potentiates the cytotoxic effects of fluorouracil (FU). We designed a phase I clinical trial of FU in combination with UCN-01. FU was administered as a weekly 24-hour infusion. Doses were escalated in successive cohorts according to a modified Fibonacci design. UCN-01 was administered once every 4 weeks, immediately after disconnection from FU, at a dose of 135 mg/m(2) over 72 hours in cycle 1 and 67.5 mg/m(2) over 36 hours in subsequent cycles. FU and UCN-01 pharmacokinetics were obtained on all patients, and thymidylate synthetase (TS) activity was measured in peripheral-blood mononuclear cells by reverse-transcriptase polymerase chain reaction. We escalated the weekly FU dose to 2,600 mg/m(2) in combination with once a month infusions of UCN-01. Dose-limiting toxicity included arrhythmia and syncope. Other toxicities included hyperglycemia, headache, and nausea and vomiting. The mean maximal plasma concentration of UCN-01 was 33.5 micromol/L. There was significant interpatient variability, which correlated with plasma concentrations of alpha-1 acid glycoprotein. FU was rapidly cleared and the dose had no effect on the area under the curve of UCN-01. Changes in TS expression were detectable in peripheral-blood mononuclear cells after administration of UCN-01 but did not correlate with toxicity or activity. We observed no objective response, although seven patients had stable disease, six of whom had received prior fluoropyrimidines. The combination of weekly infusions of FU and monthly UCN-01 can be administered safely and warrants further study in phase II trials. The recommended phase II dose of FU in combination with monthly UCN-01 is 2,600 mg/m(2).

Effect of Protein Binding of Pilsicainide on the Pharmacokinetics

To evaluate the effect of protein binding of pilsicainide on its clearance and the contribution of protein binding to optimized pilsicainide therapy, clinical laboratory and pharmacokinetic data were studied in 160 Japanese inpatients (Study 1) and 18 Japanese inpatients (Study 2). To determine the relation between protein concentration and the protein binding ratio of pilsicainide in vitro, the effect of human alpha1-acid glycoprotein (AAG) and human albumin on the binding ratio was studied. The mean ratio of serum pilsicainide concentration to dose per body weight (C/D) increased with increases in the C-reactive protein (CRP) concentration in Study 1. The AAG level increased with increases in the CRP concentration and the binding ratio increases in the AAG concentration in the Study 2. The binding ratios increased with increased AAG and albumin concentrations; the AAG concentration relative to the ratio was particularly large in vitro study. These results suggest C/D is increased in patients with high CRP levels because of binding of pilsicainide to protein, resulting decreased clearance.

Small gallstones, preserved gallbladder motility, and fast crystallization are associated with pancreatitis†‡

Acute pancreatitis is a severe complication of gallstones with considerable mortality. We sought to explore the potential risk factors for biliary pancreatitis. We compared postprandial gallbladder motility (via ultrasonography) and, after subsequent cholecystectomy, numbers, sizes, and types of gallstones; gallbladder bile composition; and cholesterol crystallization in 21 gallstone patients with previous pancreatitis and 30 patients with uncomplicated symptomatic gallstones. Gallbladder motility was stronger in pancreatitis patients than in patients with uncomplicated symptomatic gallstones (minimum postprandial gallbladder volumes: 5.8 +/- 1.0 vs. 8.1 +/- 0.7 mL; P = .005). Pancreatitis patients had more often sludge (41% vs. 13%; P = .03) and smaller and more gallstones than patients with symptomatic gallstones (smallest stone diameters: 2 +/- 1 vs. 8 +/- 2 mm; P = .001). Also, crystallization occurred much faster in the bile of pancreatitis patients (1.0 +/- 0.0 vs. 2.5 +/- 0.4 days; P < .001), possibly because of higher mucin concentrations (3.3 +/- 1.9 vs. 0.8 +/- 0.2 mg/mL; P = .04). No significant differences were found in types of gallstones, relative biliary lipid contents, cholesterol saturation indexes, bile salt species composition, phospholipid classes, total protein or immunoglobulin (G, M, and A), haptoglobin, and alpha-1 acid glycoprotein concentrations. In conclusion, patients with small gallbladder stones and/or preserved gallbladder motility are at increased risk of pancreatitis. The potential benefit of prophylactic cholecystectomy in this patient category has yet to be explored.

Cholesterol saturation rather than phospholipid/bile salt ratio or protein content affects crystallization sequences in human gallbladder bile.

In model biles, cholesterol crystallization (an important factor in gallstone formation) mainly depends on phospholipid/bile salt ratios with characteristic sequences of plate-like (monohydrate) vs. non-plate-like (presumed anhydrous: arcs, needles, tubules, spirals) cholesterol crystals. We now investigate whether the same phenomenon occurs in human bile. Appearances of plate-like and non-plate-like cholesterol crystals were determined in filtered bile of 80 cholesterol gallstone patients, and related to biliary lipid and pro-nucleating protein composition. Non-plate-like crystals appeared before plate-like crystals in 9 biles, on the same day in 24 biles, and after plate-like crystals in 31 biles. In 16 biles only plate-like crystals were observed. Crystal sequences did not depend on biliary lipid or protein composition. Cholesterol saturation indexes were higher in biles with than without non-plate-like crystals (150 +/- 6 vs. 125 +/- 12, P = 0.02). In contrast, phospholipid/(bile salt + phospholipid) ratios, bile salt species, phospholipid classes, concentrations of mucin, IgG, IgM, IgA, haptoglobin and alpha-1 acid glycoprotein did not differ. Cholesterol crystallization sequences in human bile depend on cholesterol saturation index rather than on phospholipid/bile salt ratio.