Sharon A. Chen

Interspecies scaling of clearance and volume of distribution data for five therapeutic proteins

The clearance and volume of distribution of five human proteins (recombinant CD4, CD4 immuno-globulin G, growth hormone, tissue-plasminogen activator, and relaxin) in humans and laboratory animals were analyzed as a function of body weight using allometric scaling techniques. These proteins cover a 16-fold range of molecular weight (6 to 98 kD), are produced by recombinant or synthetic methods, and may be cleared by different mechanisms. The analyses revealed that the clearance and volume data for each protein were satisfactorily described by an allometric equation (Y = a Wb). The allometric exponent (b) for clearance (ml/min) ranged from 0.65 to 0.84, the allometric exponent for the initial volume of distribution (ml) ranged from 0.83 to 1.05, and the allometric exponent for the volume of distribution at steady state (ml) ranged from 0.84 to 1.02. Exponent values from 0.6 to 0.8 for clearance and 0.8 to 1.0 for volumes are frequently cited for small molecules and are expected based on empirical interspecies relationships. When the preclinical data were analyzed separately, the pre-clinical allometric relationships were usually predictive of the human results. These findings indicate that the clearance and volume of distribution of select biomacromolecules follow well-defined, size-related physiologic relationships, and preclinical pharmacokinetic studies provide reasonable estimates of human disposition. Employing this methodology during the early phases of drug development may provide a more rational basis for dose selection in the clinical environment.

Pharmacokinetics and Tissue Distribution of Recombinant Human Transforming Growth Factor Beta1 After Topical and Intravenous Administration in Male Rats

Recombinant human transforming growth factor beta (rhTGF-β1) enhances the healing process after topical application to various animal wound models. A detailed pharmacokinetic and tissue distribution study was performed to support the clinical development of rhTGF-β1 for wound healing indications. Rats received radioiodinated or unlabeled rhTGF-β1 as an intravenous (iv) bolus or as a topical formulation applied to a full thickness wound. Plasma concentrations of TGF-β1 were estimated from TCA-precipitable radioactivity or were measured by ELISA. Following iv administration, the initial half-life was rapid (<11 min), regardless of whether radi-olabeled or unlabeled rhTGF-β1 was used. The terminal half-life was long (163 min) when the test material was radioiodinated and administered as a trace dose and relatively short (≤61 min) when given at high doses and assayed by ELISA. Analysis of plasma radioactivity by SDS-PAGE revealed a time-dependent clearance of the 25-kDa parent molecule without a significant appearance of lower molecular weight radiolabeled metabolites. The majority of the radioactivity was associated with highly perfused organs, known iodide elimination pathways, and the thyroid at 1 and 8 hr after iv injection. After topical administration of a high dose (0.8 mg/kg), no immunoreactive TGF-β1 was detectable in plasma samples taken over a 48-hr period. However, trace amounts (≤0.05 ng/mL) of acid-precipitable radioactivity were detected in plasma after topical application of [125I]rhTGF-β1 (1 µg/kg, 126 µCi/kg). A significant portion (35%) of the [125I]rhTGF-β1 persisted intact (25 kDa) at the wound site 24 hr after application. In conclusion, rhTGF-β1 was rapidly cleared after iv bolus administration and distributed primarily to the liver, lungs, kidney, and spleen. Little systemic exposure was observed after applying a single topical dose of rhTGF-β1 to a wound, and the intact molecule persisted at the wound site.

Phase I study of continuous-infusion soluble CD4 as a single agent and in combination with oral dideoxyinosine therapy in children with symptomatic human immunodeficiency virus infection

To determine the safety and pharmacokinetics of recombinant soluble CD4 (sCD4) administered by continuous intravenous infusion to children with symptomatic human immunodeficiency virus type 1 infection, we conducted a phase I study at the National Cancer Institute. Three dose levels of sCD4 were evaluated: 100, 300, and 1000 micrograms/kg per day. After an initial 12 weeks of treatment with sCD4 alone, dideoxyinosine at a dose of 90 mg/m2 every 8 hours was added and subjects were observed for an additional 12 weeks. Combination therapy was continued in patients in whom it was well tolerated. In addition to toxicity and pharmacokinetic monitoring, surrogate markers of antiviral activity were evaluated. Eleven children were enrolled in the study. During the 12 weeks of treatment with sCD4 alone, and during subsequent sCD4 plus dideoxyinosine combination therapy, no significant toxic reaction attributable to sCD4 or dideoxyinosine was encountered. Low-level anti-CD4 antibodies developed in two patients. Steady-state sCD4 levels increased proportionately at higher doses. The CD4 cell counts and serum p24 antigen levels did not provide evidence of antiviral activity. We conclude that sCD4 was well tolerated at doses up to 1000 micrograms/kg per day when administered by continuous intravenous infusion; however, evidence of in vivo antiviral activity was not observed in this study.

The Pharmacokinetics of Recombinant Human Relaxin in Nonpregnant Women After Intravenous, Intravaginal, and Intracervical Administration

The pharmacokinetics of recombinant human relaxin (rhRlx) after intravenous (iv) bolus administration and the absorption of rhRlx after intracervical or intravaginal administration were determined in nonpregnant women. The study was conducted in two parts. In part I, 25 women received 0.01 mg/kg rhRlx iv. After a minimum 7-day washout period, these women were dosed intracervically (n = 10) or intravaginally (n = 15) with 0.75 or 1.5 mg rhRlx, respectively, in 3% methylcellulose gel. Part II was a double-blind, randomized, three-way crossover study in 26 women. At 1-month intervals, each woman received one of three intravaginal treatments consisting of 0 (placebo), 1, or 6 mg rhRlx in 3% methylcellulose gel. The serum concentrations of relaxin following iv administration were described as the sum of three exponentials. The mean (±SD) initial, intermediate, and terminal half-lives were 0.09 ± 0.04, 0.72 ± 0.11, and 4.6 ± 1.2 hr, respectively. Most of the area under the curve was associated with the intermediate half-life. The weight-normalized clearance was 170 ± 50 mL/hr/kg. The observed peak concentration was 98 ± 29 ng/mL, and the weight-normalized initial volume of distribution was 78 ± 40 mL/kg, which is approximately equivalent to the serum volume. If central compartment elimination was assumed, the volume of distribution at steady state (Vss/W) was 280 ± 100 mL/kg, which is approximately equivalent to extracellular fluid volume. Vss/W could be as large as 1300 ± 400 mL/kg without this assumption. After intravaginal administration of the placebo gel, endogenous relaxin concentrations were evident (i.e., ≥20 pg/mL) in 9 of the 26 women (maximum concentrations, 23–234 pg/mL). A similar proportion of women (approximately 35–40%) exhibited measurable serum concentrations of relaxin following intravaginal rhRlx treatment; this proportion increased to 90% following intracervical rhRlx treatment. For both routes of administration, the maximum serum concentrations of relaxin were usually within the range of values observed for endogenous relaxin, suggesting that the absorption of rhRlx was minimal.

The Disposition of a Human Relaxin (hRlx-2) in Pregnant and Nonpregnant Rats

The pharmacokinetics and tissue distribution of a human relaxin were investigated after intravenous (iv) bolus administration to pregnant or nonpregnant rats. Human gene-2 relaxin (hRlx-2) serum concentrations after iv bolus administration were described as the sum of three exponentials. The pharmacokinetics were comparable in pregnant and nonpregnant rats. The serum clearance (CL) was 7.4–10.2 ml/min/kg at doses of 46–93 µg/kg and was linear in this range. The half-lives were 1.1–2.0, 15.1–16.4, and 53.7–67.9 min, respectively. The volume of the central compartment (Vc) was 48–79 ml/kg and the volume of distribution at steady state (Vss) was 271–336 ml/kg. Increasing the dose to 463 µg/kg increased the dose-corrected area under the serum concentration–time curve and significantly decreased CL and Vss. The distribution of radioactivity in the tissues of pregnant rats was followed after iv bolus dosing with hRlx-2 internally labeled with 35S-cysteine. Comparison of the extent of organ uptake of radiolabel after 35S-hRlx-2 or 35S-cysteine administration suggested that the kidneys were the principal site of uptake; the liver was of secondary importance. In perfusion experiments utilizing livers isolated from pregnant or nonpregnant rats, 36–52% of the dose of hRlx-2 was cleared from the perfusate in 2 hr. These studies showed that the pharmacokinetics of hRlx-2 in rats appeared to be unaffected by pregnancy and suggested that the kidneys and liver both play a role in the elimination of hRlx-2.

The pharmacokinetics and absorption of recombinant human relaxin in nonpregnant rabbits and rhesus monkeys after intravenous and intravaginal administration.

Recombinant human relaxin (rhRlx) is being developed as a potential cervical ripening agent to be applied intravaginally or intracervically prior to parturition. The pharmacokinetics and absorption of rhRlx were determined in nonpregnant female rabbits and rhesus monkeys after intravenous bolus (iv) and intravaginal administration of 0.1 mg/kg; additionally, rabbits were dosed with 0.5 mg/kg intravaginally. In rabbits (n = 6), mean (±SD) peak concentrations following iv bolus administration were 1554 ± 296 ng/mL. The weight-normalized clearance (CL/W) was 5.9 ± 0.4 mL/min/kg, initial volume of distribution (V1/W) was 57 ± 9 mL/kg, and volume of distribution at steady state (VSS/W), assuming central compartment elimination, was 240 ± 20 mL/kg. Vss/W could be as large as 2000 ± 400 mL/kg without this assumption. The estimated amounts of rhRlx absorbed in rabbits following intravaginal administration of 0.1 and 0.5 mg/kg (n = 5/dose) were 3.1 ± 1.4 and 0.7 ± 0.3%, respectively; peak concentrations were 600 ± 297 and 1066 ± 584 pg/mL, respectively. In rhesus monkeys (n = 5) after iv administration, peak concentrations were 971 ± 277 ng/mL; CL/W was 4.1 ± 0.6 mL/ min/kg, V1/W was 78 ± 25 mL/kg, and Vss/W, assuming central compartment elimination, was 690 ± 220 mL/kg. The upper limit for Vss/W was 1600 ± 200 mL/kg when no assumptions were made regarding site (compartment) of elimination. After intravaginal administration (n = 6), two monkeys had undetectable rhRlx concentrations throughout the 48-hr sampling interval; one monkey had only one sample containing measurable rhRlx (51 pg/mL) at 24 hr; and three monkeys absorbed <2% of the 0.1 mg/kg dose. Peak concentrations in these three animals ranged from 64 to 1475 pg/mL. The absorption of rhRlx was low and variable in both species, and similar results have been observed in women.

PHARMACOLOGICAL COMPARISON OF TWO HYBRID RECOMBINANT DNA-DERIVED HUMAN LEUKOCYTE INTERFERONS

ABSTRACT. Two hybrid human leukocyte interferons differing in only three amino acid residues show marked differences in various pharmacological properties. One hybrid, IFN-αAD (Bgl) shows greater antiviral activity in vitro and in vivo and suppresses P-450 metabolism of drugs including hexobarbital and acetaminophen. The pharmacokinetics of this hybrid and IFN-αAD (Pvu) are also distinct.

Bioequivalence of Two Recombinant Human Growth Hormones in Healthy Male Volunteers after Subcutaneous Administration.

The bioequivalence of recombinant human growth hormone (rhGH) (somatropin) and its N-methionyl variant (Met-hGH) [Protropin® (somatrem for injection)] was determined in 42 healthy male volunteers (n = 21 per treatment) who were randomized to receive either protein by subcutaneous administration of 0.1 mg kg−1. Serum samples were collected over 24 h after the injection, and the concentrations of human growth hormone (hGH) were determined by an immunoradiometric assay. Bioequivalence of the two proteins was assessed by determining whether the 90% confidence limits for the ratio of geometric means using logarithmically transformed AUC and Cmax parameters (log0AUCo0–24, log0AUCo0-x, and log10Cmax) were within the 80–125% range. The bioequivalence of the two treatments was also tested by calculating a bioequivalence index (ζ2) that measured the difference between the two mean concentration-time profiles. The 90% confidence intervals for the ratios of the geometric means for AUC were within the prescribed 80–125% range for bioequivalence. The upper limit of the 90% confidence interval for the ratio of the geometric means for Cmax fell slightly outside the 125% criterion even though the geometric mean itself, 106.6%, was very close to the ideal of 100%. There was a larger standard error associated with Cmax than with the AUCs, and this marginally larger confidence limit for Cmax resulted more from the variance among the subjects than to the difference in the means. In fact, the bioequivalence index, ζ2, was 0.075, indicating that the mean curves after rhGH and Met-hGH are essentially superimposable.