Safety, pharmacokinetics, and immunogenicity of single-dose rFXIII administration to healthy volunteers.

BACKGROUND: Factor XIII (FXIII) is a transglutaminase that cross-links fibrin and other proteins to improve clot strength and resistance to fibrinolysis. Both congenital and acquired FXIII deficiency may result in a bleeding diathesis, and plasma-derived FXIII has been used to treat many of these clinical conditions. OBJECTIVES: A clinical study was designed and performed to evaluate the safety, pharmacokinetics, and immunogenicity of recombinant FXIII (rFXIII) administration to healthy adult volunteers. PATIENTS AND METHOD: Fifty healthy adult volunteers were enrolled in this randomized, double-blinded, placebo-controlled study. A single dose of rFXIII, ranging from 2 U kg(-1) to 50 U kg(-1), or placebo was administered. Safety was evaluated by capturing adverse events, clinical safety laboratory studies, and clinical score for deep venous thrombosis. Blood samples were taken for pharmacokinetic and immunogenicity analysis throughout the 28-day follow-up period. RESULTS: Recombinant FXIII was well tolerated, with no serious adverse events or dose-related toxicities. Following a single i.v. injection of 50 U kg(-1) rFXIII, the estimated terminal half-life was 270-320 h, the volume of distribution ranged from 40 to 75 mL kg(-1), and FXIII activity increased 1.77% per 1 U kg(-1) rFXIII administered. Increase in circulating A2B2 and decrease in free FXIII-B subunit indicate in vivo formation of FXIII heterotetramer. An immunogenic response to rFXIII or yeast, the production host, was not observed. CONCLUSIONS: Recombinant FXIII was well tolerated at doses of up to 50 U kg(-1) in healthy adult volunteers. The safety, pharmacological and immunological profile of rFXIII suggests it should be studied in patients with congenital FXIII deficiency as well as evaluated as a systemic hemostat in patients with acquired FXIII deficiency or hemorrhage.

A study of therapy-related acute leukaemia after mitoxantrone therapy for multiple sclerosis.

To evaluate the incidence of therapy-related acute leukaemia (t-AL) after single-agent mitoxantrone (MITO) treatment, we reviewed medical records of patients in three studies of single-agent MITO therapy for multiple sclerosis (MS) and existing literature on MITO therapy in MS, leukaemia, and solid tumors. Of 1378 MITO recipients in the three MS studies (mean cumulative dose of 60 mg/m2 and mean follow-up of 36 months), one patient had t-AL, an observed incidence proportion of 0.07% [95% confidence interval (CI) = 0.00-0.40%]. There were no cases of t-AL in published reports of nine additional studies of single-agent MITO therapy for MS. There was one published case report of acute promyelocytic leukoemia detected five years after initiating MITO therapy for MS. The observed incidence proportion of t-AL is very low in patients who received MITO as single-agent therapy for MS. Although these observations provide preliminary reassurance, extended follow-up of these patients and those who receive higher cumulative doses of MITO is required to define the long-term risk of t-AL after MITO therapy for MS.

Cardiac adverse effects associated with mitoxantrone (Novantrone) therapy in patients with MS.

BACKGROUND: Mitoxantrone (MITO) is associated with dose-related cardiotoxicity when administered concomitantly with other cytotoxic agents with or without radiotherapy for leukemia and solid tumors. OBJECTIVE: To review observed cardiotoxicity of single-agent MITO therapy for MS. METHODS: Records of 1,378 patients from three clinical trials of MITO treatment for MS were reviewed for signs and symptoms of cardiac dysfunction and left ventricular ejection fraction (LVEF) results. Duration of follow-up was a median of 29 months (4,084 patient-years). RESULTS: No patients experienced congestive heart failure (CHF) before treatment. Cumulative MITO doses ranged from 2 to 183 mg/m(2) (mean 60.5 mg/m(2), median 62.5 mg/m(2)), and 141 patients received >100 mg/m(2). Two of 1,378 patients experienced CHF after initiating MITO therapy. Of 1,378 patients, 779 completed baseline and scheduled follow-up LVEF testing. Baseline LVEF was >50% in all 779 patients. Seventeen of 779 patients had asymptomatic LVEF of <50% (incidence proportion = 2.18%, 95% CI = 1.28 to 3.47%). Although the incidence of asymptomatic LVEF of <50% was not significantly related to monthly versus 3-monthly therapy, duration of therapy, age, or gender, asymptomatic LVEF of <50% trended higher with a cumulative dose of >/=100 mg/m(2) (5.0%) than with <100 mg/m(2) (1.8%) (p = 0.06). CONCLUSIONS: The observed incidence of CHF in patients with MS who received a mean cumulative dose of 60.5 mg/m(2) MITO was <0.20%. Continued monitoring of patients with MS who are receiving MITO is needed to determine whether the incidence of CHF increases with higher cumulative MITO doses and prolonged follow-up.

Phase I study of recombinant human CD40 ligand in cancer patients.

PURPOSE: To determine the toxicity, maximum-tolerated dose (MTD), and pharmacokinetics of recombinant human CD40 ligand (rhuCD40L) (Avrend; Immunex Corp, Seattle, WA), suggested in preclinical studies to mediate cytotoxicity against CD40-expressing tumors and immune stimulation. PATIENTS AND METHODS: Patients with advanced solid tumors or intermediate- or high-grade non-Hodgkin's lymphoma (NHL) received rhuCD40L subcutaneously daily for 5 days in a phase I dose-escalation study. Subsequent courses were given until disease progression. RESULTS: Thirty-two patients received rhuCD40L at three dose levels. A total of 65 courses were administered. The MTD was 0.1 mg/kg/d based on dose-related but transient elevations of serum liver transaminases. Grade 3 or 4 transaminase elevations occurred in 14%, 28%, and 57% of patients treated at 0.05, 0.10, and 0.15 mg/kg/d, respectively. Other toxicities were mild to moderate. At the MTD, the half-life of rhuCD40L was calculated at 24.8 +/- 22.8 hours. Two patients (6%) had a partial response on study (one patient with laryngeal carcinoma and one with NHL). For the patient with laryngeal cancer, a partial response was sustained for 12 months before the patient was taken off therapy and observed on no additional therapy. Three months later, the patient was found to have a complete response and remains biopsy-proven free of disease at 24 months. Twelve patients (38%) had stable disease after one course, which was sustained in four patients through four courses. CONCLUSION: The MTD of rhuCD40L when administered subcutaneously daily for 5 days was defined by transient serum elevations in hepatic transaminases. Encouraging antitumor activity, including a long-term complete remission, was observed. Phase II studies are warranted.

Evaluation of solid phase radiolabels of dog food for gastric emptying.

99mTc-Pertechnetate, 99mTc-mebrofenin, 99mTc-disofenin, 99mTc-sulfur colloid, and 99mTc-Dowex resin beads were evaluated for in vitro stability as a label for both dry extruded, and canned dog food for gastric emptying scintigraphy. A sample of each radiolabeled diet was added to water, gastric juice, intestinal juice, or gastric juice followed by intestinal juice for in vitro digestion. After a 3-hour digestion period, tubes were centrifuged and percentage solid phase retention (%SPR) was calculated. The experiment was repeated three times over a 14-day period to minimize day-to-day variation. For dry dog food, 99mTc-mebrofenin and 99mTc-disofenin had similar %SPR in water (96 and 93%, respectively) and gastric juice (>95% each) and were significantly higher than other labels. For canned dog food, mebrofenin had a 91% SPR for the water or gastric juice digestions, and 99mTc-Dowex had a %SPR of >99%. 99mTc-mebrofenin and 99mTc-Dowex were also tested in vivo, where 99mTc-Dowex had poor stability, and 99mTc-Mebrofenin had excellent stability. 99mTc-Mebrofenin is a suitable label for dog food for gastric emptying scintigraphy.

Actin polymerization enhances Pasteurella haemolytica leukotoxicity.

Pasteurella haemolytica leukotoxin is cytotoxic to bovine leukocytes, causing increased cell membrane permeability, osmotic swelling, release of cytosolic proteins and cell lysis. These studies were designed to test if leukotoxin causes release of the cytoskeletal protein, actin, from bovine leukemia cells and if purified actin-influenced bacterial growth or leukotoxin production. Culture supernatants caused a 7-fold decrease in viability of bovine leukemia cells and increased cell permeability that was accompanied by release of beta-actin into the cell culture supernatant. Exposing P. haemolytica to purified actin solutions induced the conversion of monomeric G-actin to polymerized F-actin. This conversion was partially inhibited by bovine P. haemolytica immune, but not pre-immune, serum. Loss of streptomycin resistance following treatment of the organism with acridine orange ablated the polymerizing activity. Incubation of P. haemolytica in the presence of purified F-actin did not affect growth but resulted in culture supernatant that had 3.0-3.9-fold greater leukotoxicity compared to medium alone or medium containing G-actin, heat-denatured actin or albumin. The effect of actin on leukotoxicity was concentration-dependent and directly associated with increases in secreted leukotoxin. The interaction between P. haemolytica and actin is potentially detrimental to the host by inducing polymerization of actin into insoluble filaments and by enhancing leukotoxicity.

Comparison of gastric emptying times in healthy cats simultaneously evaluated with radiopaque markers and nuclear scintigraphy.

Gastric emptying in 18 healthy cats was assessed simultaneously using scintigraphy and barium-impregnated polyethylene spheres (BIPS). Canned Prescription Diet Feline c/d (Hill's Pet Nutrition, Inc., Topeka, KS) labeled with 99mTc-disofenin (Hepatolite, DuPont Merck Pharmaceutical Co., Billerica, Mass.) was fed on four separate days. Scintigraphic images were obtained at time 0 and then every 30 minutes to 6 hours. On the fourth scan day, 30 small (1.5 mm) and 10 large (5 mm) BIPS (Chemstock Animal Health Ltd., Christchurch, New Zealand) were mixed with the labeled meal, and in addition to scintigraphy, radiographs were made at 60-minute intervals for 6 hours. Gastric emptying was 11 to 15% slower on the day of simultaneous radiography as compared with the 3 days when only scintigraphy was performed (p < or = .05). Percentage retention of 1.5 mm BIPS in the stomach was significantly greater than the percentage retained gastric activity at hours 1, 2, 3, 4, 5, and 6 (p < or = .05). BIPS were clustered in the pyloric region of the stomach by 3 hours in all cats. In 10/18 animals, all BIPS were retained in pyloric region of the stomach at 6 hours, despite observable decreased size of the gastric silhouette and < or =15% retained gastric activity. In conclusion, gastric emptying of 1.5-mm BIPS does not parallel gastric emptying of 99mTc-disofenin labeled canned Prescription Diet Feline c/d. Stress associated with radiography may delay gastric emptying. Diet type should be considered when evaluating clinical radiographic studies where BIPS have been used.

Scintigraphic evaluation of dogs with acute synovitis after treatment with glucosamine hydrochloride and chondroitin sulfate.

OBJECTIVE: To evaluate the effects of orally administered glucosamine hydrochloride (GlAm)-chondroitin sulfate (CS) and GlAm-CS-S-adenosyl-L-methionine (SAMe) on chemically induced synovitis in the radiocarpal joint of dogs. ANIMALS: 32 adult mixed-breed dogs. PROCEDURE: For 21 days, all dogs received a sham capsule (3 groups) or GlAm-CS (prior treatment group) in a double-blinded study. Unilateral carpal synovitis was induced by injecting the right radiocarpal joint with chymopapain and the left radiocarpal joint (control joint) with saline (0.9% NaCl) solution. Joints were injected on alternate days for 3 injections. After induction of synovitis, 2 groups receiving sham treatment were given GlAm-CS or GlAm-CS-SAMe. Another group continued to receive sham capsules (control group). Joint inflammation was quantified, using nuclear scintigraphy, before injection of joints and days 13, 20, 27, 34, 41, and 48 after injection. Lameness evaluations were performed daily. RESULTS: Dogs given GlAm-CS before induction of synovitis had significantly less scintigraphic activity in the soft-tissue phase 48 days after joint injection, significantly less uptake in the bone phase 41 and 48 days after joint injection, and significantly lower lameness scores on days 12 to 19, 23, and 24 after injection, compared with other groups. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis of results of this study suggest that prior treatment with GlAm-CS for 21 days had a protective effect against chemically induced synovitis and associated bone remodeling. Prior treatment with GlAm-CS also reduced lameness in dogs with induced synovitis.

Effect of percutaneous endoscopic gastrostomy on gastric emptying in clinically normal cats.

OBJECTIVE: To assess the effect of percutaneous endoscopic gastrostomy (PEG) tube placement on gastric emptying in clinically normal cats. ANIMALS: 8 healthy adult 3- to 5-year-old cats. PROCEDURE: Cats were accommodated to the diet for 2 weeks prior to scintigraphy. Caloric needs were divided into 3 feedings/d. Food was withheld for 24 hours after tube placement, then was fed as a third of the caloric needs on day 1, two-thirds on day 2, and full caloric requirements thereafter. Gastric emptying was measured via nuclear scintigraphy. Labeled meals contained 111 MBq (3 mCi) of 99mTc-labeled disofenin. Sixty-second ventral scintigraphic images were acquired immediately, every 20 minutes for the first hour, then every 30 minutes for 4 hours after feeding. Each cat was evaluated 3 times prior to PEG tube placement. Cats were anesthetized, and 16-F mushroom-tipped Pezzar gastrostomy tubes were placed, using a video endoscope. Scintigraphy was repeated on days 1, 4, 7, 11, 14, and 21 after PEG tube placement. RESULTS: Gastric emptying was faster with a PEG tube in place. Percentage of retained gastric activity was significantly lower after PEG for 150, 180, 210, and 240 minutes versus time before PEG tube placement. CONCLUSION: Placement of a PEG tube does not delay gastric emptying in clinically normal cats. CLINICAL RELEVANCE: Gastric retention of food, vomiting, and aspiration pneumonia after PEG tube placement may not be related to delayed gastric emptying.

Comparison of serologic and protective responses induced by two Pasteurella vaccines.

Vaccine development for the prevention of pneumonic pasteurellosis remains a critical issue for the feedlot industry. Most currently available Pasteurella vaccines are formulated to stimulate immunity by either providing an adequate antigenic mass in the administered dose, or by relying on subsequent production of antigens by in vivo growth of live organisms. The ability of these different types of vaccines to stimulate rapid and high titres to key antigens is a key factor that will influence subsequent resistance to disease. The serologic and protective responses to a streptomycin-dependent, modified-live vaccine and a killed (bacterin-toxoid) vaccine against experimental pneumonic pasteurellosis were compared. Calves were vaccinated with a single injection of either a test vaccine or phosphate-buffered saline, challenged 14 d later by transthoracic injection with Pasteurella haemolytica, and euthanized 3 d post-challenge to evaluate the severity of pneumonia. On days 0, 7, and 14, serologic responses to various P. haemolytica antigens, including cell-associated and soluble antigens, were determined by enzyme-linked immunosorbent assays, and anti-leukotoxin antibody levels were determined by leukotoxin neutralization. The bacterin-toxoid elicited significantly greater serologic responses compared to controls for all antigens. The modified-live vaccine elicited a significantly greater response compared to controls for a whole-cell antigen preparation. Lesion scores were significantly smaller (greater protection) in calves that received the bacterin-toxoid, but not the modified-live vaccine, compared to controls.

Scintigraphic assessment of gastric emptying of canned and dry diets in healthy cats.

OBJECTIVE: To characterize factors that affect solid-phase gastric emptying in healthy cats by use of nuclear scintigraphy and to assess differences in emptying patterns of dry and canned diets. ANIMALS: 20 healthy cats. PROCEDURE: 2 groups of 10 cats each were fed dry or canned diet for at least 2 weeks before scintigraphy was done. Diets were labeled with 99mTc-disofenin. After ingestion of labeled meals, scintigraphic images were obtained at 0, 15, 30, 45, and 60 minutes, then every 30 minutes to 6 hours. Gastric emptying scans were obtained 3 times for each cat for each diet, in a complete crossover design. The T90, T50, and T20 (times when 90, 50, and 20% of initial meal activity remained in the stomach, respectively) were derived from gastric emptying curves fit to nonlinear models. A mixed models approach was used for data analysis. RESULTS: Gastric emptying was well described by a nonlinear model. Meal size, water intake, and diet type significantly (P < 0.05) effected gastric emptying. The T90, T50, and T20 increased with meal size, regardless of diet type or water intake. Gastric emptying of a dry diet meal took significantly (P < 0.05) longer than that of an isocaloric meal of canned diet, except when meal size was small. Differences in gastric emptying of dry and canned diets varied with the phase (T90 vs T50 vs T20) of emptying. CONCLUSION: Water intake, meal size, and diet type significantly influence gastric emptying in healthy cats, and these factors must be considered in analysis of gastric emptying data.

Population profile of stable flies (Diptera: Muscidae) caught on Alsynite traps in various feedlot habitats.

Cylindrical traps made from Alsynite fiberglass were placed in 4 habitats in a confined cattle feedlot environment from 2 May to 30 October 1996 to evaluate abundance, sex ratio, physiological age structure, and blood-fed status of trapped adult stable flies, Stomoxys calcitrans (L.). Significantly more stable flies were caught on the trap located between host cattle and trees. The abundance of stable flies decreased geometrically with increasing distance from the host cattle in the open area. The spatial distribution of adult flies in cattle feedlot habitats became less heterogeneous as the population density increased. Male stable flies clearly dominated the total collection (66.7:33.3, male:female). A higher proportion (38.9%) of female stable flies was found on the trap close to a major immature developing area (manure pile). Most of the trapped flies (64.8%) showed a positive blood-fed status, but only 331 (8.8%) of 3,767 flies had recently fed. A majority (72.6%) of the attracted females was at the early nulliparous stage. Trap positioning did not have any significant influence on specific physiological age groups of the captured flies.

Effect of stanozolol on body composition, nitrogen balance, and food consumption in castrated dogs with chronic renal failure.

OBJECTIVE: To determine the effect of stanozolol on body composition, nitrogen balance, and food consumption in castrated dogs with chronic renal failure. DESIGN: Blinded crossover trial. ANIMALS: 22 castrated Beagles with experimentally induced chronic renal failure. PROCEDURE: Dogs were divided into 2 groups of 11 dogs each. During each of two 6-week treatment periods, dogs in 1 group received stanozolol, and those in the other group received a control agent. Nitrogen balance, body composition, and food consumption were determined. RESULTS: During administration of stanozolol, the amount of food consumed per dog, lean body mass, and nitrogen balance increased. Stanozolol did not have a significant effect on body fat, bone mineral content, or food consumption per kilogram of body weight. CLINICAL IMPLICATIONS: For dogs with mild-to-moderate, nonuremic, experimentally induced, chronic renal failure, stanozolol had positive effects on nitrogen balance and lean body mass.

Evaluation of solid-phase labels for gastric emptying studies in cats.

UNLABELLED: Development of appropriate radiolabeled diets for solid-phase gastric emptying studies in experimental animals is important for testing the effects of disease, drugs, surgical procedures and stress. This study evaluates the in vitro and in vivo stability of various radiolabels in commercially available dry, extruded and canned cat foods. METHODS: Dry, extruded cat food was labeled with 99mTc-pertechnetate, 99mTc-sulfur colloid or 99mTc-disofenin. Canned cat food was labeled with 99mTc-Dowex resin beads, 99mTc-pertechnetate, 99mTc-sulfur colloid or 99mTc-disofenin. A sample of each labeled diet and 99mTc-sulfur colloid-labeled egg was digested in water, gastric juice, intestinal juice or gastric juice followed by intestinal juice. The samples were centrifuged and the activity in the samples counted before and after removal of the supernatant. Based on in vitro results, three labeled diets were fed to 10-12 cats for in vivo testing. RESULTS: 99mTc-Dowex beads had the best labeling efficiency in vitro, but were not stable in vivo, resulting in unacceptable levels of circulating 99mTc. Technetium-99m-disofenin labeling resulted in in vitro percent solid-phase retention of 92.5% and 89.5% in water and gastric juice, respectively, for dry food and 86% and 94.9% in water and gastric juice, respectively, for canned food. CONCLUSION: Technetium-99m-disofenin is a suitable label for solidphase gastric emptying studies using commercially available cat foods.

Vitamin E is not degraded by ruminal microorganisms: assessment with ruminal contents from a steer fed a high-concentrate diet.

Using an in vitro incubation system containing undiluted ruminal contents from a steer fed a high-concentrate, corn-based diet, we examined microbial degradation of DL-alpha-tocopherol acetate (TA). Gas production, pH, and fermentation acid profiles were done in an initial experiment to ensure conditions for reproducible, viable cultures over 24 h. The pH decreased from 5.7 to 4.9, gas production averaged 3.4 mL/mL of ruminal contents, and > 300 mM fermentation acids were produced. We then monitored the fate of TA added to bottles containing ruminal contents. Three methods of TA extraction were tried, of which two were used in experiments. The two methods used were 1) hot ethanol in a Soxhlet apparatus and 2) chloroform/methanol. Each of these was used to extract added TA from a set of three in vitro experiments. Concentrations of TA were determined at 0 h and after 4, 8, and 24 h at 39 degrees C. In the three hot ethanol extracted experiments, TA recoveries were 85% at 0 h. With time of incubation, TA levels either 1) remained constant, 2) decreased then returned to the initial value, or 3) decreased by approximately 50%. These inconsistent results indicated that this extraction method was unacceptable. In the latter three experiments we used a chloroform/methanol extraction method. Recoveries of added TA averaged 96% overall. Thus, the level of TA remained constant during the 24-h period, suggesting that microbial destruction of TA does not occur. Rather, the previously reported losses of vitamin E may be attributable to incomplete extraction of tocopherol from high-concentrate ruminal contents.