Kristina M. Campanale

Pharmacodynamic comparison of LY3023703, a novel microsomal prostaglandin e synthase 1 inhibitor, with celecoxib

To assess the safety, tolerability, and pharmacology of LY3023703, a microsomal prostaglandin E synthase 1 (mPGES1) inhibitor, a multiple ascending dose study was conducted. Forty‐eight subjects received LY3023703, celecoxib (400 mg), or placebo once daily for 28 days. Compared with placebo, LY3023703 inhibited ex vivo lipopolysaccharide‐stimulated prostaglandin E2 (PGE2) synthesis 91% and 97% on days 1 and 28, respectively, after 30‐mg dosing, comparable to celecoxibs effect (82% inhibition compared to placebo). Unlike celecoxib, which also inhibited prostacyclin synthesis by 44%, LY3023703 demonstrated a maximal increase in prostacyclin synthesis of 115%. Transient elevations of serum aminotransferase were observed in one subject after 30‐mg LY3023703 dosing (10× upper limit of normal (ULN)), and one subject after 15‐mg dosing (about 1.5× ULN). Results from this study suggest that mPGES1 inhibits inducible PGE synthesis without suppressing prostacyclin generation and presents a novel target for inflammatory pain.

Discovery and Characterization of 2-Acylaminoimidazole Microsomal Prostaglandin E Synthase-1 Inhibitors.

As part of a program aimed at the discovery of antinociceptive therapy for inflammatory conditions, a screening hit was found to inhibit microsomal prostaglandin E synthase-1 (mPGES-1) with an IC50 of 17.4 μM. Structural information was used to improve enzyme potency by over 1000-fold. Addition of an appropriate substituent alleviated time-dependent cytochrome P450 3A4 (CYP3A4) inhibition. Further structure-activity relationship (SAR) studies led to 8, which had desirable potency (IC50 = 12 nM in an ex vivo human whole blood (HWB) assay) and absorption, distribution, metabolism, and excretion (ADME) properties. Studies on the formulation of 8 identified 8·H3PO4 as suitable for clinical development. Omission of a lipophilic portion of the compound led to 26, a readily orally bioavailable inhibitor with potency in HWB comparable to celecoxib. Furthermore, 26 was selective for mPGES-1 inhibition versus other mechanisms in the prostanoid pathway. These factors led to the selection of 26 as a second clinical candidate.

Disposition and Metabolism of LY2452473, a Selective Androgen Receptor Modulator, in Humans

The disposition and metabolism of isopropyl N-[(2S)-7-cyano-4-(2-pyridylmethyl)-2,3-dihydro-1H-cyclopenta[b]indol-2-yl]carbamate (LY2452473; a selective androgen receptor modulator) in humans was characterized after a single 15-mg (100 μCi) oral dose of [14C]LY2452473 to six healthy male subjects. LY2452473 was absorbed rapidly (time to reach maximum plasma concentration for both LY2452473 and total radioactivity was 2–3 h) and cleared slowly (plasma terminal t1/2 of 27 h for LY2452473 and 51 h for the total radioactivity). LY2452473 and metabolites S5 (acetylamine) and S12 (hydroxylation on the cyclopentene) were major circulating entities in plasma, accounting for approximately 42, 21, and 35% of the total radioactivity exposure, respectively, as calculated from relative area under the concentration versus time curves from zero to 48 h derived from the plasma radiochromatograms. The radioactive dose was almost completely recovered after 312 h with 47.9% of the dose eliminated in urine and 46.6% in feces. Minimal LY2452473 was detected in excreta, indicating that metabolic clearance was the main route of elimination. Multiple metabolic pathways were observed with no single metabolic pathway accounting for more than 30% of the dose in excreta. Metabolite S10 (a diol across the cyclopenta-indole linkage) was the largest excretory metabolite (approximately 14% of the dose). S10 displayed interesting chemical and chromatographic properties, undergoing conversion to the corresponding epoxide under acidic conditions and conversion back to the diol under neutral conditions. An in vitro phenotyping approach indicated that CYP3A4 was the largest contributor to LY2452473 depletion.

Disposition of LY333531, a selective protein kinase C β inhibitor, in the Fischer 344 rat and beagle dog

1. Studies were conducted in the Fischer 344 rat and beagle dog to determine the disposition of LY333531 and its equipotent active des-methyl metabolite, LY338522, both potent and selective inhibitors of the β-isozyme of protein kinase C. 2. Male Fischer 344 rats and female beagle dogs received a single 5-mgkg−1 oral dose of 14C-LY333531. Urine, faeces, bile and plasma were collected and analysed for 14C, LY333531 and LY338522. 3. LY333531 was eliminated primarily in the faeces (91% by 120 h in rat, 90% by 96h in dog). Bile contributed the majority of the radioactivity excreted in the faeces in rat (66% in the cannulated bile duct study) and a variable but significant proportion in dog. 4. Pharmacokinetics following a single 5 mg kg−1 oral dose of 14C-LY333531 to the male rat produced Cmax and AUC0-∞ for LY333531 of 14.7 ng ml−1 and 60.8ng h ml−1, respectively, with a half-life of 2.5 h. LY338522 and total radioactivity showed similar profiles. 5. In the female dog at the same dose, Cmax and AUC0-∞ of LY333531 were higher, producing 245 ± 94 ng ml−1 and 1419 ± 463ng h ml−1, respectively, with a half-life of 5.7 h. 6. The data indicate that the disposition of LY333531 is similar in rat and dog.

POTENT, ORALLY BIOAVAILABLE HIV-1 PROTEASE INHIBITORS CONTAINING NONCODED D-AMINO ACIDS

Abstract Novel noncoded D-amino acids have been combined with decahydroisoquinoline, octahydrothienopyridine, and urea hydroxyethylamine isosteres to provide potent HIV-1 protease inhibitors with excellent HIV-1 antiviral activity. LY314613 shows a promising combination of potency and oral bioavailability. Trends in the SAR and comparisons to other isostere derivatives will be discussed.

IN VIVO METABOLISM OF [14C]RUBOXISTAURIN IN DOGS, MICE, AND RATS FOLLOWING ORAL ADMINISTRATION AND THE STRUCTURE DETERMINATION OF ITS METABOLITES BY LIQUID CHROMATOGRAPHY/MASS SPECTROMETRY AND NMR SPECTROSCOPY

Ruboxistaurin (LY333531), a potent and isoform-selective protein kinase C β inhibitor, is currently undergoing clinical trials as a therapeutic agent for the treatment of diabetic microvascular complications. The present study describes the disposition and metabolism of [14C]ruboxistaurin following administration of an oral dose to dogs, mice, and rats. The study revealed that ruboxistaurin was highly metabolized in all species. Furthermore, the results from the bile duct-cannulated study revealed that ruboxistaurin was well absorbed in rats. The primary route of excretion of ruboxistaurin and its metabolites was through feces in all species. The major metabolite detected consistently in all matrices for all species was the N-desmethyl metabolite 1, with the exception of rat bile, in which hydroxy N-desmethyl metabolite 5 was detected as the major metabolite. Other significant metabolites detected in dog plasma were 2, 3, 5, and 6 and in mouse plasma 2, 5, and 19. The structures of the metabolites were proposed by tandem mass spectrometry with the exception of 1, 2, 3, 5, and 6, which were additionally confirmed either by direct comparison with authentic standards or by nuclear magnetic resonance spectroscopy. To assist identification by nuclear magnetic resonance spectroscopy, metabolites 3 and 5 were produced via biotransformation using recombinant human CYP2D6 and, likewise, metabolite 6 and compound 4 (regioisomer of 3 which did not correlate to metabolites found in vivo) were produced using a microbe, Mortierella zonata. The unambiguous identification of metabolites enabled the proposal of clear metabolic pathways of ruboxistaurin in dogs, mice, and rats.

Disposition of [14C] Ruboxistaurin in Humans

Ruboxistaurin is a potent and specific inhibitor of the β isoforms of protein kinase C (PKC) that is being developed for the treatment of diabetic microvascular complications. The disposition of [14C]ruboxistaurin was determined in six healthy male subjects who received a single oral dose of 64 mg of [14C]ruboxistaurin in solution. There were no clinically significant adverse events during the study. Whole blood, urine, and feces were collected at frequent intervals after dosing. Metabolites were profiled by high performance liquid chromatography with radiometric detection. The total mean recovery of the radioactive dose was approximately 87%, with the majority of the radioactivity (82.6 ± 1.1%) recovered in the feces. Urine was a minor pathway of elimination (4.1 ± 0.3%). The major route of ruboxistaurin metabolism was to the N-desmethyl ruboxistaurin metabolite (LY338522), which has been shown to be active and equipotent to ruboxistaurin in the inhibition of PKCβ. In addition, multiple hydroxylated metabolites were identified by liquid chromatography-mass spectrometry in all matrices. Pharmacokinetics were conducted for both ruboxistaurin and LY338522 (N-desmethyl ruboxistaurin, 1). These moieties together accounted for approximately 52% of the radiocarbon measured in the plasma. The excreted radioactivity was profiled using radiochromatography, and approximately 31% was structurally characterized as ruboxistaurin or N-desmethyl ruboxistaurin. These data demonstrate that ruboxistaurin is metabolized primarily to N-desmethyl ruboxistaurin (1) and multiple other oxidation products, and is excreted primarily in the feces.

Ly316340: A potent HIV-1 protease inhibitor containing a high affinity octahydrothienopyridine hydroxyethylamine isostere

Abstract Replacement of the decahydroisoquinoline group contained in Ro 31-8959 by a cis -octahydrothienopyridine moiety has provided a high affinity hydroxyethylamine isostere for use in HIV-1 protease inhibitors. Further gains in potency have been realized by incorporation of a sulfur atom into the P 1 benzyl group. Modification by a key P 2 ligand provided LY316440, a potent, orally absorbed inhibitor of HIV-1 protease.

Addition of 20-kDa PEG to insulin lispro alters absorption and decreases clearance in animals

ABSTRACTPurposeDetermine the pharmacokinetics of insulin peglispro (BIL) in 5/6-nephrectomized rats and study the absorption in lymph duct cannulated (LDC) sheep.MethodsBIL is insulin lispro modified with 20-kDa linear PEG at lysine B28 increasing the hydrodynamic size to 4-fold larger than insulin lispro. Pharmacokinetics of BIL and insulin lispro after IV administration were compared in 5/6-nephrectomized and sham rats. BIL was administered IV or SC into the interdigital space of the hind leg, and peripheral lymph and/or serum samples were collected from both LDC and non-LDC sheep to determine pharmacokinetics and absorption route of BIL.ResultsThe clearance of BIL was similar in 5/6-nephrectomized and sham rats, while the clearance of insulin lispro was 3.3-fold slower in 5/6-nephrectomized rats than in the sham rats. In non-LDC sheep, the terminal half-life after SC was about twice as long vs IV suggesting flip-flop pharmacokinetics. In LDC sheep, bioavailability decreased to <2%; most of the dose was absorbed via the lymphatic system, with 88% ± 19% of the dose collected in the lymph after SC administration.ConclusionThis work demonstrates that increasing the hydrodynamic size of insulin lispro through PEGylation can impact both absorption and clearance to prolong drug action.

Nasal Absorption of Enkephalins in Rats

In recent years, the possibility that the intranasal administration route might be useful for many compounds which are not absorbed orally has received a great deal of attention. For instance, the β-blocker propranolol (Hussain et al, 1979, 1980 a, b), the contraceptive agent progesterone (David et al, 1981; Hussain et al, 1981) and the anti-arrhythmic compound clofilium tosylate (Su et al, 1984) have been shown to be effectively absorbed via the intranasal route when compared to oral administration. These compounds undergo extensive degradation due to first-pass hepatic metabolism which can be minimized after nasal administration. For drugs which are poorly absorbed by the oral route such as sulbenicillin, cefazolin, and cephacetrile, it was demonstrated that the percent dose excreted in urine after nasal administration was nearly one-half of that after intramuscular administration (Hirai et al, 1981). The absorption of low molecular weight polypeptides, luteinising hormone-releasing hormone (LH-RH) and its analogues used as a contraceptive agent, was evaluated by the nasal route (Fink et al, 1974; Berquist et al, 1979; Gennser and Liedholm, 1974; London et al, 1973; Anik et al, 1984). Although the absorption efficiency by the nasal route was lower than the I.V. route for these polypeptides, the absorption was reproducible, and the advantage of non-parenteral route for such a compound was an important factor. Research has also been carried out on the nasal absorption of high molecular weight polypeptides such as insulin (Moses et al, 1983; Hirai et al, 1978, 1981 a,b), interferon (Greenberg et al, 1978; Harmon et al, 1976; 1977; Johnson et al, 1976) and growth hormone releasing factor (Evans et al, 1983).