Ada H. C. Kung

A Phase I Safety And Pharmacokinetic Study Of A Recombinant Amino Terminal Fragment Of Bactericidal/ Permeability-increasing Protein In Healthy Male Volunteers

A phase I pharmacokinetic and safety clinical trial of rBPI23, a recombinant amino terminal fragment of bactericidal/permeability-increasing protein, was conducted in healthy male volunteers. rBPI23 was administered as a 5 or 30 min infusion at doses of .1 to 1 mg/kg. The pharmacokinetics of rBPI23 in human subjects were described by a bi-exponential disposition function with evidence of concentration-dependent kinetics. The α half-life increased significantly with increasing dose, from 4–5 min at .1 mg/kg to 7–8 min at 1 mg/kg. The β half-life varied between 18 and 29 min regardless of dose and the clearance varied from 5 to 10 mL/min/kg. Very little, if any, of the administered rBPI23 was excreted intact in the urine. Electrocardiograms, ionized calcium concentration, prothrombin and partial prothrombin times, hematologic parameters, and blood chemistries remained normal. Furthermore, no antibody response to rBPI23 was observed in any of the subjects.

Protective effect of a recombinant fragment of bactericidal/permeability increasing protein against carbohydrate dyshomeostasis and tumor necrosis factor-α elevation in rat endotoxemia

Endotoxin (lipopolysaccharide, LPS), a component of the gram-negative bacterial cell wall, induces carbohydrate dyshomeostasis and the release of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) when administered to experimental animals. Bactericidal/permeability increasing protein (BPI), a cationic protein found in human neutrophil granules, binds with high affinity to LPS and is capable of neutralizing its biological activity. The present study was designed to determine if a recombinant N-terminal fragment of BPI, rBPI23, attenuates LPS-induced alterations in serum glucose, lactate, and TNF-alpha in rats. In anesthetized animals challenged with a 30 min infusion of Escherichia coli O111:B4 LPS (0.25 mg/kg), there was an early transient increase in serum levels of glucose followed by a drop to 60% of those found in saline control rats. A prolonged elevation in serum levels of lactate and a transient, but marked, elevation of TNF-alpha were also observed following LPS infusion. These LPS-induced changes were inhibited significantly by simultaneous infusion of rBPI23. Different dose-response profiles of rBPI23 on LPS-induced alterations in glucose, lactate and TNF-alpha were observed. When rBPI23 was infused 30 min after the initiation of LPS infusion, it significantly inhibited the alterations in glucose and lactate, but not TNF-alpha. The rise in TNF-alpha was reduced significantly with a 15 min delayed infusion of rBPI23. A control protein failed to alter any responses to LPS. The results indicate that rBPI23 can provide significant protection against the metabolic disturbances and TNF-alpha release associated with endotoxemia. In addition, the results suggest that LPS-induced metabolic alterations in glucose and lactate are at least partially independent of TNF-alpha release.

Protective effects of a recombinant N-terminal fragment of bactericidal/permeability increasing protein on endotoxic shock in conscious rabbits.

Endotoxin (lipopolysaccharide, LPS) can induce shock, multiple organ failure, and death. A recombinant N-terminal fragment of bactericidal/permeability increasing protein, rBPI23, binds with high affinity to gram-negative bacterial LPS and neutralizes its biological activity. We sought to determine the effect of rBPI23 on LPS-induced respiratory dysfunction and cardiovascular depression in conscious rabbits. Rabbits were injected with Escherichia coli O113 LPS (6 micrograms/kg) and treated with rBPI23 (2 mg/kg), vehicle, or control protein after recovery from surgery performed to implant catheters for hemodynamic assessments and intravenous injections. LPS challenge caused respiratory dysfunction including tachypnea, significant decreases in arterial O2 tension (PO2), arterial oxygen content, and an increase in alveolar-arterial O2 gradient (A-aDO2). LPS administration also resulted in profound and prolonged decreases in mean arterial blood pressure and cardiac index. Treatment with rBPI23 prevented LPS-induced respiratory dysfunction and significantly ameliorated the cardiovascular depression. 5 of 16 LPS-challenged animals died of respiratory failure and acidosis, whereas none died in the rBPI23 treated group (p = .11). The results demonstrate that rBPI23 protects animals against LPS-induced cardiopulmonary depression in endotoxic shock.

ALTERATION OF THE PHARMACOKINETICS OF SMALL PROTEINS BY IODINATION

The pharmacokinetics of several proteins were investigated using two different assays. A 23 kDa recombinant protein fragment of bactericidal/permeability-increasing protein (rBPI23) was radiolabeled with 125I using Iodo-beads and administered rats. Plasma samples were collected and assayed for 125I-rBPI23 by radioactivity. In a separate experiment, rBPI23 was administered to rats and plasma samples were assayed for rBPI23 by ELISA. The clearance determined from plasma concentrations of 125I-rBPI23 measured by radioactivity was about 2.5-fold lower than that of rBPI23 determined by ELISA. In addition, the steady state volumes of distribution and mean residence times of 125I-rBPI23 measured by radioactivity were four-fold and 10-fold greater, respectively, compared to those measured by the ELISA method. By studying several proteins with a range of molecular weights, we found that the pharmacokinetics of proteins below about 60 kDa were different when assayed by radioactivity or ELISA, but those of proteins with molecular weights of at least 80 kDA revealed only minor differences. To determine which assay method yielded the correct plasma pharmacokinetic profile, rBPI23 was metabolically labeled with 35S-methionine and administered to rats, and plasma samples were assayed by radioactivity. The concentration-time profile assessed by this method was very close to that determined by ELISA. Exposing rBPI23 to chloramine-T (the oxidant used in the iodination process) and measuring its plasma concentration by ELISA revealed pharmacokinetics similar to those of the iodinated protein measured by radioactivity. In contrast, radiolabeling rBPI23 using iodinated Bolton-Hunter reagent (which avoids exposing the protein to oxidant), and measuring 125I-rBPI23 by radioactivity, yielded pharmacokinetics that were similar, although not identical, to the pharmacokinetics of rBPI23 measured by ELISA. Thus, our data suggest that directly iodinating low-molecular-weight proteins by oxidation procedures alters their clearance from the blood, preventing reliable determination of pharmacokinetic parameters.

The Role of Liver and Kidney on the Pharmacokinetics of a Recombinant Amino Terminal Fragment of Bactericidal/Permeability-Increasing Protein in Rats

AbstractPurpose. The pharmacokinetics of rBPI23, a recombinant amino terminal fragment of bactericidal/permeability-increasing protein that binds to and neutralizes endotoxin, was investigated. Methods. rBPI23 was administered to rats at doses 0.01−10 mg/kg and plasma rBPI23 levels were measured by ELISA. rBPI23 was also administered to bilaterally nephrectomized rats. In addition, rBPI23 was administered intra-hepatically via the pyloric vein to determine the first-pass effect by the liver. rBPI23 concentrations were also simultaneously measured in the right atrium and aorta to determine the removal of rBPI23 by the lungs. Results. The concentration-time profile of rBPI23 was described by a 3-compartmental model with parallel first order and Michaelis-Menten (saturable) elimination. The clearance of rBPI23 was not altered by bilateral nephrectomy. Clearance of intra-hepatically administered rBPI23 was 4.5 fold lower than intra-femorally administered rBPI23. The concentration difference of rBPI23 between aortic and right atrial blood was no greater than 11%. Clearance of rBPI23 in rats could be reduced up to 10 fold by co-administration of heparin. Uptake by liver of intra-hepatically administered rBPI23 was prevented by co-administration of heparin. Conclusions. rBPI23 is not significantly cleared by the kidneys, and no more than 11% of the rBPI23 was removed by the lungs with each pass. The liver could remove 78% of the rBPI23 from the hepatic circulation. Studies with heparin suggest rBPI23 is cleared by binding to heparan sulfate sites in the liver.

Preclinical Development of Xomen™-E5

The use of monoclonal antibodies (Mabs) as human therapeutic agents, either in their native form or conjugated to radioisotopes or toxins, is still in its infancy. Only one such product, the murine anti-CD3 Mab, Orthoclone OKT3®, is approved for use in the United States, although several other products are in clinical trials or have completed clinical testing and are awaiting approval. Thus, little information has been published on the pharmacokinetic and toxicologic evaluation of monoclonal antibody products as part of their preclinical evaluation prior to approval for human use. Such studies are well-established for traditional drugs and provide essential information for the proper design of clinical trials. For example, pharmacokinetic characteristics influence frequency-of-dosing decisions, and toxicologic information alerts clinical investigators to areas of potential toxicity.