Tissue disposition and pharmacokinetics of recombinant human nerve growth factor after acute and chronic subcutaneous administration in monkeys.

In this study, we have characterized the metabolism, tissue disposition, excretion routes, and plasma pharmacokinetics of recombinant human nerve growth factor after single and multiple s.c. administration in male cynomolgus monkeys. Unlabeled nerve growth factor (NGF; 2 mg/kg) was administered three times a week for 4 weeks and a full pharmacokinetic profile was obtained for doses 1 and 12. For the tissue distribution studies, 0.8 microg/kg of trace (125)I-labeled recombinant human nerve growth factor was dosed. Histological analysis of emulsion-microautoradiography indicated that specific (125)I-NGF labeling was confined to sections of nerves most frequently localized adjacent to large vessels in sections of kidney, spleen, liver, and salivary gland. A small percentage of large neurons within the sympathetic ganglia were intensely labeled, as well as large neurons within the dorsal root ganglia. We found an increased disposition of (125)I-NGF in parts of the peripheral nervous system (including sympathetic ganglia) from 8 to 24 h postdose. In contrast, radioactivity in most non-neuronal tissues declined. This suggests specific uptake in these target tissues known to express specific receptors for NGF. We also identified changes in pharmacokinetic parameters after single versus chronic s. c. administration. These studies demonstrated that s.c. administration of NGF at 0.8 microg/kg doses in monkeys is capable of accessing and localizing in the target tissues.

Recombinant human Dnase I (rhDNase) in patients with lupus nephritis.

Systemic lupus erythematosus (SLE) is characterized by the production of pathogenic autoantibodies to nucleoprotein antigens, including double-stranded DNA (dsDNA). The deposition of IgG dsDNA immune complexes in glomeruli is thought to be crucial for disease pathogenesis and complement activation. rhDNase catalyzes the hydrolysis of extracellular DNA and has been shown to delay the development of dsDNA antibodies, reduce proteinuria, and delay mortality in a lupus-prone murine model. We conducted a 40d, phase Ib, randomized, double-masked, placebo-controlled trial to determine the safety and pharmacokinetics of rhDNase, and to measure any changes in markers of disease activity in 17 patients with lupus nephritis. Patients were assigned to receive either: (1) 25 microg/kg rhDNase (n = 8); (2) 125 microg/kg rhDNase (n=6); or (3) placebo (n = 3) initial single intravenous (IV) dose followed by 10 subcutaneous (SC) doses. Skin biopsies performed on nine patients pre- and post-treatment were studied for immune complex deposition by immunofluorescence. Serum cytokine levels (sIL2-R, IL-6, IL-10, and TNF-alpha) were analyzed by ELISA. Cytokine secretion and antibody production were measured by ELISPOT analysis and ELISA. Serum hydrolytic activity of rhDNase was achieved after IV administration at 25 and 125 microg/kg, but not after SC administration at either dose. A t 1/2 of 3-4h was estimated from serum concentration profiles following IV administration. Serum dsDNA antibodies were unchanged (mean values: 33 IU/mL vs 39 IU/mL [pre- and post-treatment] for the 25 microg/kg group, and 74 IU/mL vs 74 IU/mL for the 125 microg/kg group, and 14 IU/mL vs 20 IU/mL for the placebo group). Complement levels (C3 and C4) and circulating immune complexes did not change appreciably during the treatment period for any of the groups. Serum cytokine profiles by ELISA revealed no changes in sIL-2 receptor, IL-6, IL-10, or TNF-alpha. There was no change in the number of cells secreting either Th1 or Th2 specific cytokines, nor in the number of cells secreting dsDNA antibodies. Neutralizing antibodies to rhDNase were not detected in serum at any time during the study. Immune complex deposition was unchanged in pre- and post-treatment in skin biopsies in both dose groups. rhDnase was well tolerated without significant adverse events following administration, and treatment was not associated with the development of neutralizing antibodies to rhDNase. Serum rhDNase concentrations capable of hydrolytic activity of rhDNase were achieved for a few hours following IV, but not SC administration. Serum markers of disease activity were unchanged during the study period.

Predictability of the clinical potency of NSAIDs from the preclinical pharmacodynamics in rats.

OBJECTIVE AND DESIGN: Relevance of the preclinical pharmacodynamic, toxicity and pharmacokinetic parameters predicting the clinical potency of nonsteroidal antiinflammatory drugs (NSAIDs) was evaluated. MATERIAL: Data for oral potencies of 24 NSAIDs in rats were collected from the literature and from New Drug Applications with respect to the following parameters: antiinflammatory, analgesic, antipyretic, acute ulcerogenic activities, acute toxicity, in vitro inhibition of prostaglandin synthesis, acid dissociation constant (pKa), octanol-water partition coefficient and elimination half-life. TREATMENT: Data for most of the in vivo parameters in rats were collected following single dose administration with the exception of adjuvant arthritis. Single and daily clinical doses were considered. All of these NSAIDs have been approved for marketing although not all have been sold in the USA. METHODS: The preclinical data were compared to human dose (unit or daily doses) using single and multiple stepwise regression analyses. RESULTS: Analyses suggest that NSAIDs are effective in all models of preclinical tests for fever, pain and inflammation, however, carrageenin-induced rat paw edema model is clearly the best predictor of human dose. Rank order of preclinical models for predicting human dose is carrageenin > yeast induced fever > pressure induced pain = adjuvant arthritis in rats. The analysis suggested that the pain and adjuvant arthritis models in rats may also involve a prostaglandin independent mechanism. Of the two physicochemical factors tested, pKa contributed best to the carrageenin model towards predicting the clinical potency of NSAIDs. Mathematical relationships between human dose, carrageenin ED50 and pKa were established that may assist in the future clinical development of NSAIDs. CONCLUSIONS: Carrageenin-induced paw edema model in rats is the most robust predictor of the clinical potency of NSAIDs. Acid dissociation constant (pKa) appears to be a secondary contributor to the potency of NSAIDs. The relevance of the data analyses for developing cyclooxygenase-2 (COX-2) selective NSAIDs is discussed.

The interconversion of prednisone and prednisolone in human and rabbit organs in vitro is a function of tissue integrity.

The interconversion and elimination of prednisone (PO) and prednisolone (POH) were examined in human and rabbit liver, kidney, lung and skeletal muscle preparations to determine the effect of organ disruption on in vitro metabolism within multiple organs of two species. Results from microsomes, homogenates and minces were compared to determine the effect of various stages of tissue disruption on the reversible and irreversible metabolism of the glucocorticoids. Oxidation of POH to PO was enhanced by homogenization of all the organs examined; further disruption by subcellular fractionation to microsomes reduced oxidation relative to homogenates but yielded values greater than the original minces. The reverse reaction, reduction of PO to POH, was also enhanced by homogenization, but microsomal preparations were less active than the minces. The irreversible elimination of the glucocorticoids was progressively diminished by disruption of normal architecture in all organs. Results of in vitro studies of glucocorticoid metabolism have provided discrepant results, as a function of the laboratory, the species, the organ and the organ preparation examined. These results provide insights into the source of discrepant results regarding steroid metabolism, as it appears that results of experimentation with glucocorticoid conversion and/or elimination may be a function of the method used. Additionally, the data support the hypothesis that the enzyme system involved in the interconversion of the glucocorticoids may not be a simple single protein which acts bidirectionally.

Disposition of prednisone and prednisolone in the perfused rabbit liver: modeling hepatic metabolic processes.

The livers of 15 rabbits were perfused in situ with prednisone (PO) or prednisolone (POH) over a wide range of steady state concentrations, resulting in multiple experimental measurements per organ. Linearity of extraction, an apparent lack of oxidative conversion, and marked preference for the reduction of PO to POH was observed. Predictions of hepatic tissue concentrations were made using both the well-stirred and parallel-tube model approximations. Glucocorticoid disposition across the liver was described by a series of differential equations. Discrimination between the two models was accomplished by examining the effects of changes in flow rate upon the availability of the highly extracted drug PO. The well-stirred model very closely predicted the observed changes in availability of PO, whereas the parallel-tube model provided poor predictions. The intrinsic clearances of interconversion and elimination of PO and POH were subsequently calculated by population analysis using NONMEM. This method assumed the well-stirred model and resulted in intrinsic clearance estimates of 26 ml/min for the elimination of POH, 157 ml/min for reductive conversion of PO to POH, and 205 ml/min for the irreversible elimination of PO. A mechanism of intrahepatic disposition of these glucocorticoids was proposed using well-stirred model predictions of hepatic drug concentrations, the perfusion rate limitation to drug transport, and the assumption of no oxidative interconversion of POH to PO. In this case, the capacity for reduction of PO to POH approaches the elimination clearance of PO and the elimination of PO is about 13 times greater than the elimination clearance of POH.

Prednisolone and prednisone exhibit linear extraction in the perfused rabbit liver.

The rabbit and human both display nonlinear pharmacokinetics of prednisone (PO) and prednisolone (POH). The in vivo apparent clearances of these compounds increase with dose. To determine whether one source of the kinetic nonlinearity is due to hepatic metabolism, the isolated liver of the rabbit was perfused with widely varying concentrations of both PO and POH in the absence of corticosteroid binding globulin. Both compounds exhibited constant extraction ratios over a wide concentration range, even at concentrations exceeding 100 times those expected in portal venous blood during absorption of orally administered drug. Hepatic extraction of POH averaged 0.49 and that of PO was nearly complete at 0.96. The apparent hepatic blood clearance of POH was about 16 ml/min and that of PO approximated liver blood flow, at 30 ml/min. Furthermore, the liver was predominantly reductive toward these compounds: upon PO perfusion, POH was recovered as about half of the dose of PO administered. POH perfusion yielded no detectable PO in the exiting perfusate. The liver is believed to be the most significant organ involved in glucocorticoid biotransformation. Its capacity to eliminate PO and POH does not appear to be saturable, and greatly favors the pharmacologically active, reduced compound, POH. Hypotheses that attribute the nonlinear pharmacokinetics of PO and POH as partially due to saturable hepatic metabolism may be incorrect. It has been previously assumed that all metabolic organs produce both interconversion products. Since the liver apparently yields no PO (oxidized form), another organ must be responsible for the in vivo oxidation.