Preclinical development of AMG 139, a human antibody specifically targeting IL-23.

BACKGROUND AND PURPOSE: AMG 139 is a human anti-IL-23 antibody currently in a phase II trial for treating Crohn's disease. To support its clinical development in humans, in vitro assays and in vivo studies were conducted in cynomolgus monkeys to determine the pharmacology, preclinical characteristics and safety of this monoclonal antibody. EXPERIMENTAL APPROACH: The in vitro pharmacology, pharmacokinetics (PK), pharmacodynamics and toxicology of AMG 139, after single or weekly i.v. or s.c. administration for up to 26 weeks, were evaluated in cynomolgus monkeys. KEY RESULTS: AMG 139 bound with high affinity to both human and cynomolgus monkey IL-23 and specifically neutralized the biological activity of IL-23 without binding or blocking IL-12. After a single dose, linear PK with s.c. bioavailability of 81% and mean half-life of 8.4-13 days were observed. After weekly s.c. dosing for 3 or 6 months, AMG 139 exposure increased approximately dose-proportionally from 30 to 300 mg·kg(-1) and mean accumulation between the first and last dose ranged from 2- to 3.5-fold. Peripheral blood immunophenotyping, T-cell-dependent antigen responses and bone formation markers were not different between AMG 139 and vehicle treatment. No adverse clinical signs, effects on body weight, vital signs, ophthalmic parameters, clinical pathology, ECG, organ weights or histopathology were observed in the monkeys with the highest dose of AMG 139 tested (300 mg·kg(-1) s.c. or i.v.). CONCLUSIONS AND IMPLICATIONS: The in vitro pharmacology, PK, immunogenicity and safety characteristics of AMG 139 in cynomolgus monkeys support its continued clinical development for the treatment of various inflammatory diseases.

Safety toxicity study of plasmid-based IL-12 therapy in Cynomolgus monkeys.

We have investigated the potential toxicity of hlL-12 DNA plasmid formulated with 5% polyvinylpyrrolidone (PVP) administered twice weekly via subcutaneous injections to Cynomolgus monkeys for four weeks, and have evaluated recovery from any effects of the test article over a four-week treatment-free period. Doses of the formulated hIL-12 plasmid were selected based on anti-tumour efficacy studies previously conducted in mice. The duration of the study and the frequency of dosing were designed to support clinical trials. No clinical signs indicative of an adverse effect of administration of formulated hIL-12 plasmid were observed. There were no apparent effects of the formulated hIL-12 plasmid on body weights or on serum chemistry, haematology, coagulation or urinalysis parameters. No treatment-elated ocular abnormalities were evident. In addition, examination of the electrocardiograms from all monkeys at the pre-study, week-4, and week-8 time points did not reveal any treatment-related effects. No treatment-related gross lesions were noted at days 28 or 57. Slight histopathological changes associated with high doses of PVP vehicle were observed at both time points. These results suggested that the administration of formulated hIL-12 plasmid at a dose level up to 18 mg kg(-1) dose twice per week for four weeks to experimentally naïve Cynomolgus monkeys did not result in significant toxicity. These results support further testing of this gene therapy in clinical trials.

Evaluation of the renal effects of an antisense phosphorothioate oligodeoxynucleotide in monkeys.

Antisense phosphorothioate oligodeoxynucleotides are therapeutic agents that provide target specificity resulting from Watson-Crick base pairing. However, there are nonspecific effects that in some instances result in toxicity. These compounds accumulate in the kidney and induce renal proximal tubular degeneration at high doses. The relationship between accumulation of phosphorothioate oligodeoxynucleotides in the kidney, indicators of renal toxicity, and histomorphology were investigated in rhesus monkeys. Monkeys received vehicle or an escalating dose regimen of 3, 10, 40, and 80 mg/kg of ISIS 2105 and were then evaluated for changes in clinical pathology indices, urinalysis parameters, and renal histopathology. Urinalysis revealed an increase in protein levels and a slight increase in blood content following the third 40 mg/kg dose and continuing through the 80 mg/kg doses, whereas other urinary markers of renal toxicity were unchanged. Creatinine clearance was slightly decreased in monkeys during the 80 mg/kg dose cycle. Granulation in the cytoplasm of proximal tubular epithelial cells was evident by microscopic examination of kidney and was present at all doses examined and increased with dose. Immunohistochemical staining localized the oligodeoxynucleotide within these granules. Histopathologic changes consisting of minimal to moderate tubular degeneration were present only at the higher doses of 40 and 80 mg/kg and at high tissue concentrations, and these changes occurred concurrent with functional alterations, whereas lower doses (< or = 10 mg/kg) did not affect a pathologic or functional change.

Pharmacokinetics and urinary excretion of amikacin in low-clearance unilamellar liposomes after a single or repeated intravenous administration in the rhesus monkey.

Liposomal aminoglycosides have been shown to have activity against intracellular infections, such as those caused by Mycobacterium avium. Amikacin in small, low-clearance liposomes (MiKasome) also has curative and prophylactic efficacies against Pseudomonas aeruginosa and Klebsiella pneumoniae. To develop appropriate dosing regimens for low-clearance liposomal amikacin, we studied the pharmacokinetics of liposomal amikacin in plasma, the level of exposure of plasma to free amikacin, and urinary excretion of amikacin after the administration of single-dose (20 mg/kg of body weight) and repeated-dose (20 mg/kg eight times at 48-h intervals) regimens in rhesus monkeys. The clearance of liposomal amikacin (single-dose regimen, 0.023 +/- 0.003 ml min-1 kg-1; repeated-dose regimen, 0.014 +/- 0.001 ml min-1 kg-1) was over 100-fold lower than the creatinine clearance (an estimate of conventional amikacin clearance). Half-lives in plasma were longer than those reported for other amikacin formulations and declined during the elimination phase following administration of the last dose (from 81.7 +/- 27 to 30.5 +/- 5 h). Peak and trough (48 h) levels after repeated dosing reached 728 +/- 72 and 418 +/- 60 micrograms/ml, respectively. The levels in plasma remained > 180 micrograms/ml for 6 days after the administration of the last dose. The free amikacin concentration in plasma never exceeded 17.4 +/- 1 micrograms/ml and fell rapidly (half-life, 1.47 to 1.85 h) after the administration of each dose of liposomal amikacin. This and the low volume of distribution (45 ml/kg) indicate that the amikacin in plasma largely remained sequestered in long-circulating liposomes. Less than half the amikacin was recovered in the urine, suggesting that the level of renal exposure to filtered free amikacin was reduced, possibly as a result of intracellular uptake or the metabolism of liposomal amikacin. Thus, low-clearance liposomal amikacin could be administered at prolonged (2- to 7-day) intervals to achieve high levels of exposure to liposomal amikacin with minimal exposure to free amikacin.

Maintenance of morphology and function of canine proximal colon smooth muscle in organ culture.

We have determined that serum source plays a critical role in optimizing conditions for an organ culture model of canine proximal colon. Previous studies using equine serum in the medium have shown that some properties of canine colonic smooth muscle can be maintained in organ culture. However, many characteristics of the tissue were altered by the culture conditions. The aims of the present study were to determine whether serum isolated from canine blood would improve the preservation of physiological properties of canine proximal colon in organ culture. Strips of canine colonic smooth muscle were cultured in 10% canine serum medium, and electrical, mechanical, morphological, and molecular analyses were performed after 0, 3, and 6 days in culture. Unlike organ culture in equine serum, in which Na+-K+-adenosinetriphosphatase (Na+-K+-ATPase) expression declined, culture in canine serum maintained Na+-K+-ATPase expression, and resting membrane potential of smooth muscle cells along the submucosal surface of the circular muscle in cultured tissue remained unchanged during the culture period. Increased sensitivity in the contractile response to acetylcholine, previously observed with tissues cultured in equine serum, was not observed. However, the mechanical performance of the muscle (maximal contractile activity) declined over time in culture. Ultrastructural organization of cellular organelles and myofilaments remained intact in the majority of cells; however, some cells possessed regions devoid of contractile filaments. The results of these studies suggest that organ cultured strips of smooth muscle may provide a useful tool for evaluating electrical and mechanical events in conjunction with molecular analysis of functional components.