A novel in vitro allometric scaling methodology for aldehyde oxidase substrates to enable selection of appropriate species for traditional allometry

1.?Failure to predict human pharmacokinetics of aldehyde oxidase (AO) substrates using traditional allometry has been attributed to species differences in AO metabolism.

2.?To identify appropriate species for predicting human in vivo clearance by single-species scaling (SSS) or multispecies allometry (MA), we scaled in vitro intrinsic clearance (CL_int) of five AO substrates obtained from hepatic S9 of mouse, rat, guinea pig, monkey and minipig to human in vitro CL_int.

3.?When predicting human in vitro CL_int, average absolute fold-error was ≤2.0 by SSS with monkey, minipig and guinea pig (rat/mouse >3.0) and was <3.0 by most MA species combinations (including rat/mouse combinations).

4.?Interspecies variables, including fraction metabolized by AO (F_m,AO) and hepatic extraction ratios (E) were estimated in vitro. SSS prediction fold-errors correlated with the animal:human ratio of E (r²?=?0.6488), but not F_m,AO (r²?=?0.0051).

5.?Using plasma clearance (CL_p) from the literature, SSS with monkey was superior to rat or mouse at predicting human CL_p of BIBX1382 and zoniporide, consistent with in vitro SSS assessments.

6.?Evaluation of in vitro allometry, F_m,AO and E may prove useful to guide selection of suitable species for traditional allometry and prediction of human pharmacokinetics of AO substrates.     

细胞色素P450的工具药选择及种属差异的研究进展

药物对代谢酶的影响以及代谢产生的药物相互作用与药物安全性密切相关。细胞色素P450(CYP)是参与内、外源性化合物I相代谢反应的重要超家族酶系。特异性探针、诱导剂、抑制剂以及实验动物模型广泛应用于CYP介导的代谢途径以及药物相互作用研究。已知CYP底物存在明显重叠性,且表达调控机制种属差异显著,故研究中选择适当的实验动物以及特异性的探针、诱导剂和抑制剂,成为影响数据外推的关键问题。本文简要综述了CYP1A2,CYP2C9,CYP2C19,CYP2D6,CYP3A4/5的常用体内外探针、诱导剂、抑制剂以及动物种属表达调控的差异。     

Species differences in bile acids II. Bile acid metabolism

One of the mechanisms of drug‐induced liver injury (DILI) involves alterations in bile acid (BA) homeostasis and elimination, which encompass several metabolic pathways including hydroxylation, amidation, sulfation, glucuronidation and glutathione conjugation. Species differences in BA metabolism may play a major role in the failure of currently used in vitro and in vivo models to predict reliably the DILI during the early stages of drug discovery and development. We developed an in vitro cofactor‐fortified liver S9 fraction model to compare the metabolic profiles of the four major BAs (cholic acid, chenodeoxycholic acid, lithocholic acid and ursodeoxycholic acid) between humans and several animal species. High‐ and low‐resolution liquid chromatography–tandem mass spectrometry and nuclear magnetic resonance imaging were used for the qualitative and quantitative analysis of BAs and their metabolites. Major species differences were found in the metabolism of BAs. Sulfation into 3‐O‐sulfates was a major pathway in human and chimpanzee (4.8%–52%) and it was a minor pathway in all other species (0.02%–14%). Amidation was primarily with glycine (62%–95%) in minipig and rabbit and it was primarily with taurine (43%–81%) in human, chimpanzee, dog, hamster, rat and mice. Hydroxylation was highest (13%–80%) in rat and mice followed by hamster, while it was lowest (1.6%–22%) in human, chimpanzee and minipig. C6‐β hydroxylation was predominant (65%–95%) in rat and mice, while it was at C6‐α position in minipig (36%–97%). Glucuronidation was highest in dog (10%–56%), while it was a minor pathway in all other species (<12%). The relative contribution of the various pathways involved in BA metabolism in vitro were in agreement with the observed plasma and urinary BA profiles in vivo and were able to predict and quantify the species differences in BA metabolism. In general, overall, BA metabolism in chimpanzee is most similar to human, while BA metabolism in rats and mice is most dissimilar from human.     

Case report of extensive metabolism by aldehyde oxidase in humans: Pharmacokinetics and metabolite profile of FK3453 in rats,dogs, and humans

1. We describe the preclinical and clinical pharmacokinetic profiles of FK3453 [6-(2-amino-4-phenylpyrimidin-5-yl)-2-isopropylpyridazin-3(2H)-one] and the mechanism responsible for poor oral exposure of FK3453 in humans.

2. FK3453 showed favourable profiles in preclinical pharmacokinetic studies, including satisfactory absolute bioavailability and total body clearance in animals (30.5%–41.4%, 54.7%–68.2%, and 71.3%–93.4% and 10.8–17.6, 1.9–17.1, and 5.0?mL/min/kg in male rats, female rats, and dogs, respectively), and good metabolic stability in liver microsomes (42.3, 14.5, and 1.1?mL/min/kg in male rats, dogs, and humans, respectively).

3. However, despite these promising preclinical findings, plasma concentrations of FK3453 in humans were extremely low, with the oxidative metabolite of the aminopyrimidine moiety (M4) identified as a major metabolite. Given that aldehyde oxidase (AO) and xanthine oxidase (XO) were presumed to be the enzymes responsible for M4 formation, we investigated the mechanism of M4 formation using human liver subcellular fractions.

4. M4 was detected in the incubation mixture with S9 and cytosol but not with microsomes, and M4 formation was inhibited by AO inhibitors (menadione, isovanillin) but not by cytochrome P-450 inhibitor (1-aminobenzotiazole) or XO inhibitor (allopurinol). These results suggest M4 formation is catalyzed by AO, and therefore, its poor exposure in humans was attributed to extensive AO metabolism.

     

9-硝基20（S）喜树碱在大鼠体内的药物动力学

目的:研究9-硝基喜树碱在大鼠体内的药物动力学及排泄。方法:利用高效液相色谱法对静脉注射或灌胃给药后的大鼠血浆及排泄物样品进行分析。绘制血浆药物浓度-时间曲线,并进行非室模型分析及房室模型拟合。利用线性回归评价药物浓度-时间曲线下面积(AUC)与剂量之间、血浆药物峰浓度(C_(max))与剂量之间的线性关系;不同剂量下的药物半衰期及清除率通过方差分析进行比较。计算原形药物自大鼠体内的排泄量。结果:大鼠分别以1.5、3、6mg/kg静脉给药后,AUC_(o-t)分别为633、1606和3011h·μg·L~(-1);t_(1/2)分别为0.5、0.5和0.7h;大鼠分别以3、6、12mg/kg灌胃给药后,C_(max)分别为203、417和1150μg/L,T_(max)均在0.3h左右,AUC_(o-t)分别为269、439和881h·μg·L~(-1);t1/2分别为1.7、0.9和0.9h。9-硝基喜树碱在大鼠体内的绝对生物利用度为14.6％,这与灌胃及静脉注射两种给药途径下原形药物(胆汁和尿中)累积排泄量之比值相一致。结论:9-硝基喜树碱在大鼠体内动力学过程符合二室模型。静脉给药后,药物在大鼠体内的动力学不依赖于剂量,肾排泄为原形药物的主要排泄途径;灌胃给药后,药物绝对生物利用度低,原形药物大部分经粪排泄。     

Acute effect of glipizide on glucose tolerance in obesity and diabetes mellitus (NIDDM)

Summary The polymorphic cytochrome P-450 DB1 (P-450 IID6) is responsible for the O-demethylation of codeine to morphine by human liver microsomes. The influence of P-450 DB1 variable activity on the bioactivation of codeine in vivo to morphine and on its analgesic effect was investigated in phenotyped healthy volunteers — 7 extensive [EM] and 1 poor [PM] metabolizer of debrisoquine. After pretreatment with oral placebo or quinidine sulphate 50 mg, codeine phosphate 100 mg or placebo were administered orally according to a double-blind randomized crossover design.In EM subjects the plasma morphine C_max was 17.9 nmol/l, whereas virtually no morphine was detectable after quinidine pretreatment (1.5 nmol/l), and in the PM subject (0.60 nmol/l). In EM codeine significantly increased subjective (VAS) and objective (R-III reflex) pain thresholds in response to selective transcutaneous nerve stimulation, whereas no significant analgesia was detected after placebo, or after codeine with quinidine pretreatment, or in the PM. In PM of genetic origin, or due to environmental alteration of the phenotypic expression (i.e. drug interaction), codeine is not activated into morphine and is an inefficient analgesic.Presented in part at the 2nd International Congress on Cancer Pain, New York, N.Y., July 1988, and at the Nineteenth Annual Meeting of the American Society For Clinical Pharmacology and Therapeutics, Nashville, Tennessee, March 1989     

Systematic structure modifications of imidazo[1,2-a]pyrimidine to reduce metabolism mediated by aldehyde oxidase (AO)

N-{trans-3-[(5-Cyano-6-methylpyridin-2-yl)oxy]-2,2,4,4-tetramethylcyclobutyl}imidazo[1,2-a]pyrimidine-3-carboxamide (1) was recently identified as a full antagonist of the androgen receptor, demonstrating excellent in vivo tumor growth inhibition in castration-resistant prostate cancer (CRPC). However, the imidazo[1,2-a]pyrimidine moiety is rapidly metabolized by aldehyde oxidase (AO). The present paper describes a number of medicinal chemistry strategies taken to avoid the AO-mediated oxidation of this particular system. Guided by an AO protein structure-based model, our investigation revealed the most probable site of AO oxidation and the observation that altering the heterocycle or blocking the reactive site are two of the more effective strategies for reducing AO metabolism. These strategies may be useful for other drug discovery programs.     

Species-dependent hepatic metabolism of immunosuppressive agent tacrolimus (FK-506)

1. The current study aims to investigate species-related differences in the in-vitro hepatic metabolism of tacroliums using liver microsomes obtained from rat, hamster, guinea pig, rabbit, pig, dog, baboon and humans.

2. Tacrolimus metabolism was characterized using high-performance liquid chromatography- ultraviolet light (HPLC-UV) and two soft ionization mass spectrometric techniques; matrix-assisted lasers desorption/ionization (MALDI) and time-of-flight-secondary ion mass spectrometry (TOF-SIMS).

3. The extent of tacrolimus metabolism, when normalized to the cytochrome P-450 content, was in the order: rat < hamster < rabbit < pig < guinea pig < dog < human < baboon. Tacrolimus metabolism exhibited significant qualitative and quantitative differences between the animal species tested.

4. Desmethyl- (MI–MIII), didesmethyl- (MIV–MVI), monohydroxy- (MVII), dihydroxy- (MVIII), epoxide- (MIX), dihydrodiol- (MX), monodesmethyl and monohydroxy- (MXI–MXIII), and didesmethyl and monohydroxy- (MXIV–MXVI) tacroliums metabolites were identified in the species tested. MI–MX were identified in all the species tested; MXI–MXVI were identified in all species except rat, rabbit and guinea pig; and MXIV–MXVI were identified only in baboon.

5. The current investigation was unable to detect any phase II metabolites due to the limitations of the test system used.

6. The analytical methods were not able to differentiate optical and positional isomers of metabolites due to the nature of the analytical tools used, therefore groups of metabolites were identified based on their molecular weights and available information.

7. From the current in-vitro metabolism studies, the pattern of tacroliums metabolism in baboons closely resembled that in humans and thus it is ideal for studying tacroliums metabolism-related work of clinical relevance.

     

Terfenadine metabolism of human cytochrome P450 2J2 containing genetic variations (G312R,P351L and P115L)

The human cytochrome P450 2J2 is involved in several metabolic reactions, including the oxidation of important therapeutics and epoxidation of endogenous arachidonic acid. At least ten genetic variations of P450 2J2 have been identified, but their effects on enzymatic activity have not been clearly characterized. Here, we evaluated the functional effects of three genetic variations of P450 2J2 (G312R, P351L, and P115L). Recombinant enzymes of wild-type and three variant P450 2J2 were heterologously expressed in Escherichia coli and purified. P450 expression levels in the wild-type and two variants (P351L and P115L) were 142–231 nmol per liter culture, while the G312R variant showed no holoenzyme peak in the CO-binding spectra. Substrate binding titrations to terfenadine showed that the wild-type and two variants displayed K_d values of 0.90–2.2 μM, indicating tight substrate binding affinities. Steady-state kinetic analysis for t-butyl methyl hydroxylation of terfenadine indicated that two variant enzymes had similar k_cat and K_m values to wild-type P450 2J2. The locations of mutations in three-dimensional structural models indicated that the G312R is located in the I-helix region near the formal active site in P450 2J2 and its amino acid change affected the structural stability of the P450 heme environment.     

Regio-Selectivity of Purified Forms of Rabbit Liver Microsomal Cytochrome P-450 in the Metabolism of Benzo(a)pyrene,n-Hexane and 7-Ethoxyresorufin

Abstract: The specificity of electrophoretically homogeneous preparations of rabbit liver microsomal cytochrome P-450lm _2–4 towards oxygenation of n-hexane, 7-ethoxyresorufin and benzo(a)pyrene was examined using a reconstituted system consisting of cytochrome P-450, NADPH-cytochrome P-450 reductase and dilauroylphosphatidylcholine. Epoxide hydrase was included when benzo(a)pyrene was used as substrate. Cytochrome P-450lm ₂ was most active in n-hexane and benzo(a)pyrene oxygenation especially with regard to the formation of 2-hexanol, B(a)P-4,5-dihydrodiol and B(a)P-phenol metabolites. 7-Ethoxyresorufin was, however, a very poor substrate for cytochrome P-450lm ₂. Cytochrome P-450lm ₃ had less activity towards the investigated substrates while cytochrome P-450lm ₄ preferentially formed 2- and 3-hexanol, resorufin and B(a)P-9,10-dihydrodiol. Cytochrome P-450lm ₄ isolated after pretreatment with 3-methylcholanthrene or pheno-barbital showed roughly the same characteristics except in the formation of 1-hexanol where cytochrome P-450lm ₄ isolated after phenobarbital treatment was the most effective. The formation of B(a)P-4,5- and ?9,10-dihydrodiols was greatly increased by incorporation of epoxide hydrase. Our results indicate a certain specificity of the different forms of cytochrome P-450 in the liver microsomes although some overlap in activities was observed.     

Accumulation of Mutagenic Xenobiotics in Fresh Water (Lake Baikal) and Marine (Hornoya Island) Ecosystems

Extracts from tissues of a wide range of aquatic organisms (plants, plankton, decapods, molluscs, fish, Baikal seals, and fish-eating birds and their eggs) from Lake Baikal and from the Selenga River estuary and tissue extracts of birds breeding on Hornoya Island (northern Norway) were assayed for mutagenicity using the Ames Salmonella/microsome test. The activities of cytochrome P-450 and the enzymes of phase II of detoxification were also studied in the liver of fish and birds. Evidence was found for the accumulation of mutagens in the food chain. The relationship of bioaccumulation to the levels of enzyme activities possessing both detoxification and activation functions is discussed in the cases of fish and birds. The accumulation of mutagens was found to depend on the activity and the level of induction of the enzymes providing the detoxification and metabolic activation in the livers of fish and birds.     

Species differences in the CYP3A-catalyzed metabolism of TPN729, a novel PDE5 inhibitor

TPN729 is a novel phosphodiesterase 5(PDE5)inhibitor used to treat erectile dysfunction in men.Our previous study shows that the plasma exposure of metabolite M3(N-dealkylation of TPN729)in humans is much higher than that of TPN729.In this study,we compared its metabolism and pharmacokinetics in different species and explored the contribution of its main metabolite M3 to pharmacological effect.We conducted a combinatory approach of ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry-based metabolite identification,and examined pharmacokinetic profiles in monkeys,dogs,and rats following TPN729 administration.A remarkable species difference was observed in the relative abundance of major metabolite M3:i.e.,the plasma exposure of M3 was 7.6-fold higher than that of TPN729 in humans,and 3.5-,1.2-,1.1-fold in monkeys,dogs,and rats,respectively.We incubated liver S9 and liver microsomes with TPN729 and CYP3A inhibitors,and demonstrated that CYP3A was responsible for TPN729 metabolism and M3 formation in humans.The inhibitory activity of M3 on PDE5 was 0.78-fold that of TPN729(The IC50 values of TPN729 and M3 for PDE5A were 6.17±0.48 and 7.94±0.07 nM,respectively.).The plasma protein binding rates of TPN729 and M3 in humans were 92.7%and 98.7%,respectively.It was astonishing that the catalyzing capability of CYP3A4 in M3 formation exhibited seven-fold disparity between different species.M3 was an active metabolite,and its pharmacological contribution was equal to that of TPN729 in humans.These findings provide new insights into the limitation and selection of animal model for predicting the clinical pharmacokinetics of drug candidates metabolized by CYP3A4.     

Species differences in enantioselective 2-oxidations of RS-8359, a selective and reversible MAO-A inhibitor, and cinchona alkaloids by aldehyde oxidase

The 2-oxidation activity on the pyrimidine ring of RS-8359, a MAO-A inhibitor, is the major metabolic pathway catalysed by aldehyde oxidase. This study investigated the species differences in the 2-oxidation activity by using liver cytosolic fractions from rats, mice, guinea-pigs, rabbits, dogs, monkeys and humans. The Vmax/Km value for the (S)-enantiomer of RS-8359 was extremely high in monkeys and humans, moderate in guinea-pigs, and low in rats and mice. Dogs were deficient in 2-oxidation activity. The (R)-enantiomer was only oxidized at a very low rate in guinea-pigs, monkeys and humans, and not oxidized in rats, mice and rabbits. Thus, marked species differences and enantioselectivity were obvious for the 2-oxidation of the (S)-enantiomer of RS-8359. The in vitro results were in good accordance with previously reported in vivo excretion data of the 2-keto metabolite and the non-detectable plasma concentrations of the (S)-enantiomer in monkeys and humans after administration of racemic RS-8359. Enantioselectivity was also observed for the oxidation of cinchona alkaloids catalysed by aldehyde oxidase. Among the four cinchona alkaloids studied, the oxidation activity of cinchonidine, which has no substituents at the 6-hydroxy group but bears (8S,9R)-configurations, was highest. As opposed to the (S)-enantiomer, an extremely high catalytic activity of cinchonidine was confirmed in rabbits, but not in monkeys or humans. Rabbit liver aldehyde oxidase was suggested to have characteristic properties around the active site.     

Use of Freund's Complete Adjuvant (FCA) in inflammatory pain models: Consequences on the metabolism and pharmacokinetics of the non-peptidic delta receptor agonist SNC80 in the rat

This study was initiated to characterize the metabolism and pharmacokinetics of SNC80 in rats and to evaluate the impact of Freund's complete adjuvant (FCA)-induced inflammation on its body disposition. In vitro, the disappearance and intrinsic clearance (CL_int) of SNC80 were measured following incubations in recombinant rat CYPs and in phenotyped liver microsomes from naive and 24-h FCA-treated rats. The unbound fraction (f_u) was assessed by ultrafiltration. Based on the Clint values, in vivo blood clearance of 3.35 and 2.48 L/h/kg were predicted in naive and FCA-treated rats. In vivo, SNC80 was administered to naive and 24-h FCA-treated rats at 10?µmol/kg i.v. and 50?µmol/kg p.o. The naive animals showed high plasma clearance (3.1 L/h/kg), low renal clearance (<0.02 L/h/kg) and poor bioavailability (<4%). Following i.v. administration, plasma clearance was lower (22%) in FCA-treated vs. untreated rats. Despite the decreases in f_u (～30%) and CL_int (～40%) in vitro, in vivo the apparent bioavailability and oral clearance were not significantly different between FCA-treated and naive rats. Hepatic and possibly intestinal losses contribute to the low bioavailability of SNC80. Non-hepatic mechanisms may compensate for the decrease in plasma clearance found in FCA-treated rats, preventing an increase in the oral bioavailability of SNC80.     

Pulmonary Delivery of Powders and Solutions Containing Recombinant Human Granulocyte Colony-Stimulating Factor (rhG-CSF) to the Rabbit

Two powder formulations (MMAD <4 µm) containing rhG-CSF were insufflated (IF) via an endotracheal tube at doses of 5, 75 or 500 µg/kg to New Zealand white rabbits. Doses of 5 and 500 µg/kg of solutions were administered by intratracheal instillation (IT), subcutaneous (SC) injection in the thigh and intravenous injection (IV) via the marginal ear vein. Blood samples were removed at regular intervals from an indwelling jugular catheter. Blood was analyzed directly for total white blood cell counts (WBC). Plasma was assayed for rhG-CSF by a specific ELISA. The distribution of radioactive dose in lung tissue was found after administering Tc99m HSA in solution or when incorporated into powders. The pharmacokinetics and pharmacodynamics were determined for all routes of administration. High dose IV concentration vs. time profiles declined biexponentially (t_1/2 = 0.6 ± 0.2 hrs, t_1/2 = 4.6 ± 0.2 hrs, n = 8). Clearance was dose dependent (11.6 ± 2.6 [500 µg/kg, n = 8] vs. 21.8 ± 3.3 ml/hr/kg [5 µg/kg, n = 5]). A normal systemic response was obtained after IF, indicating that rhG-CSF retains activity in the solid state. Dissolution and absorption of rhG-CSF from the powders were not rate limiting. The plasma concentration vs. time profiles peaked at similar times to those after IT (T_max 1 -2 hrs) but were earlier than obtained after SC (T_max 6-10 hrs). Powders were less efficiently dosed to the lung lobes after insufflation compared with instillates (14.7 ± 10.5 vs. 60.1 ± 10.6%), resulting in bioavailabilities ranging from 5 to 33%. Bioavailability after SC was 11.0 ± 7.0% and 95.3 ± 7.9% (n = 6) for the low and high doses, respectively.     

Inter-species variation in the metabolism and inhibition of N-[(2'-dimethylamino)ethyl]acridine-4-carboxamide (DACA) by aldehyde oxidase

N-[(2'-Dimethylamino)ethyl]acridine-4-carboxamide (DACA) is a new anticancer agent currently undergoing clinical trials. The metabolism of DACA to acridone metabolites by aldehyde oxidase (AO) (EC 1.2.3.1) appears to play a major role in its elimination in human patients and rodents. The aim of this study was to compare the ability of human, guinea pig, and rat AO preparations to metabolise DACA, and to determine if either animal model was appropriate for predicting AO-mediated DACA-drug interactions in humans. Both human and rodent liver samples were homogenised in buffer before sequential centrifugation to produce the cytosol fraction. Human supernatant underwent an additional ammonium sulphate precipitation procedure, which produced a 2-fold increase in enzyme activity per milligram of protein. After incubations with DACA (range, 0-200 microM), DACA-9(10H)-acridone formation was determined by HPLC analysis. Michaelis-Menten parameters, Km and Vmax, were determined from the best fit curves by nonlinear regression. Three of the four human liver preparations had similar DACA intrinsic clearance values (Vmax/Km) ranging from 0.27 to 0.35 mL/min/mg protein, whereas both the rat and guinea pig had approximately 7- and 160-fold greater intrinsic clearances, due to lower Km values in rats (4.5 +/- 0.7 microM) and guinea pigs (0.15 +/- 0.1 microM) compared with humans (28.3 +/- 8.3 microM, N = 4). Amsacrine, menadione, and 7-hydroxy-DACA were potent inhibitors of DACA metabolism in all three species, but 10-fold differences in IC50 values were apparent between species. In addition, SKF-525A was a potent inhibitor of the metabolism of DACA in rat cytosol but caused minimal inhibition in the guinea pig or human preparations. These results suggest that neither rat nor guinea pig AO preparations are suitable for predicting AO-mediated DACA-drug interactions in humans.     

Species differences in metabolism of EPZ015666, an oxetane-containing protein arginine methyltransferase-5 (PRMT5) inhibitor

Abstract

1.?Metabolite profiling and identification studies were conducted to understand the cross-species differences in the metabolic clearance of EPZ015666, a first-in-class protein arginine methyltransferase-5 (PRMT5) inhibitor, with anti-proliferative effects in preclinical models of Mantle Cell Lymphoma. EPZ015666 exhibited low clearance in human, mouse and rat liver microsomes, in part by introduction of a 3-substituted oxetane ring on the molecule. In contrast, a higher clearance was observed in dog liver microsomes (DLM) that translated to a higher in vivo clearance in dog compared with rodent.

2.?Structure elucidation via high resolution, accurate mass LC-MSⁿ revealed that the prominent metabolites of EPZ015666 were present in hepatocytes from all species, with the highest turnover rate in dogs. M1 and M2 resulted from oxidative oxetane ring scission, whereas M3 resulted from loss of the oxetane ring via an N-dealkylation reaction.

3.?The formation of M1 and M2 in DLM was significantly abrogated in the presence of the specific CYP2D inhibitor, quinidine, and to a lesser extent by the CYP3A inhibitor, ketoconazole, corroborating data from human recombinant isozymes.

4.?Our data indicate a marked species difference in the metabolism of the PRMT5 inhibitor EPZ015666, with oxetane ring scission the predominant metabolic pathway in dog mediated largely by CYP2D.     

Toxicokinetic Study of Recombinant Human Heparin-Binding Epidermal Growth Factor-Like Growth Factor (rhHB-EGF) in Female Sprague Dawley Rats

Purpose To determine the toxicity and pharmacokinetics of recombinant heparin-binding epidermal growth factor-like growth factor in female Sprague Dawley rats following intra-bladder and intravenous administration. Materials and Methods rhHB-EGF was administered once daily for 6 or 27 days at doses of 3, 10, or 30 μg/kg. ¹²⁵I-rhHB-EGF was administered on day 7 or 28 for pharmacokinetic analysis. Toxicity was assessed by general appearance and behavior, gross necropsy, blood chemistry and microscopic evaluation. Results Plasma AUCss of [¹²⁵I] rhHB-EGF equivalents following IB administration for 7 days were 4.28 ± 2.29, 7.75 ± 2.70, and 7.11 ± 1.42 ng ml⁻¹ h⁻¹ at doses of 3, 10, and 30 μg/kg, respectively. Following IV administration, the AUCss on day 7 increased from 27.0 ± 2.66 to 124 ± 5.09 and 385.11 ± 7.57 ng ml⁻¹ h⁻¹ with increasing the dose from 3 to 10 and 30 μg/kg. Similar AUCss data was obtained after 28 day administration. No toxicity was evident upon gross examination. Histologic examination revealed subacute inflammation and lymphocytic infiltration of the urinary bladder in animals from all groups dosed by the IB route. Conclusions Plasma and bladder concentrations of recombinant human [¹²⁵I] rhHB-EGF equivalents were significantly lower following the IB route than following IV administration. Histologic tissue examination indicated no toxicity attributable to rhHB-EGF.     

Species differences in the nephrotoxic response to S-(1,2-dichlorovinyl)glutathione.

The present study was carried out to investigate the species differences in the nephrotoxic response to S-(1,2-dichlorovinyl)glutathione (DCVG) using rats, hamsters and guinea-pigs. DCVG was given intraperitoneally in physiological saline to groups of 5 animals at doses 0, 165 and 330 μmol/kg. Urine was collected for 24 h and the animals were then sacrificed. Significantly increased levels of urinary glucose, , γ-glutamyl transpeptidase, proteins and blood urea nitrogen were observed in rats at both dose levels of DCVG. An increase, but not of similar magnitude, of these biochemical parameters was noted in hamsters only at the higher dose of DCVG. Guinea-pigs showed significant increases in these biochemical parameters at the lower dose, but not at the higher dose. Light-microscopic studies showed increasing proximal tubular necrosis (PTN) in rats with increasing dose of DCVG, but PTN involving straight tubules only was observed at the higher dose in hamsters. PTN was again observed in guinea-pigs at the lower dose, but not at the higher dose of DCVG.     

Species differences in the metabolism of di(2-ethylhexyl) phthalate (DEHP) in several organs of mice, rats, and marmosets

To clarify species differences in the metabolism of di(2-ethylhexyl) phthalate (DEHP) we measured the activity of four DEHP-metabolizing enzymes (lipase, UDP-glucuronyltransferase (UGT), alcohol dehydrogenase (ADH), and aldehyde dehydrogenase (ALDH)) in several organs (the liver, lungs, kidneys, and small intestine) of mice (CD-1), rats (Sprague–Dawley), and marmosets (Callithrix jacchus). Lipase activity, measured by the rate of formation of mono(2-ethylhexyl) phthalate (MEHP) from DEHP, differed by 27- to 357-fold among species; the activity was highest in the small intestines of mice and lowest in the lungs of marmosets. This might be because of the significant differences between Vmax/Km values of lipase for DEHP among the species. UGT activity for MEHP in the liver microsomes was highest in mice, followed by rats and marmosets. These differences, however, were only marginal compared with those for lipase activity. ADH and ALDH activity also differed among species; the activity of the former in the livers of marmosets was 1.6–3.9 times greater than in those of rats or mice; the activity of the latter was higher in rats and marmosets (2–14 times) than in mice. These results were quite different from those for lipase or UGT activity. Because MEHP is considered to be the more potent ligand to peroxisome proliferator-activated receptor involved in different toxic processes, a possibly major difference in MEHP-formation capacity could be also considered on extrapolation from rodents to humans.