Pre-symptomatic Caspase-1 inhibitor delays cognitive decline in a mouse model of Alzheimer disease and aging.

Early therapeutic interventions are essential to prevent Alzheimer Disease (AD). The association of several inflammation-related genetic markers with AD and the early activation of pro-inflammatory pathways in AD suggest inflammation as a plausible therapeutic target. Inflammatory Caspase-1 has a significant impact on AD-like pathophysiology and Caspase-1 inhibitor, VX-765, reverses cognitive deficits in AD mouse models. Here, a one-month pre-symptomatic treatment of Swedish/Indiana mutant amyloid precursor protein (APPSw/Ind) J20 and wild-type mice with VX-765 delays both APPSw/Ind- and age-induced episodic and spatial memory deficits. VX-765 delays inflammation without considerably affecting soluble and aggregated amyloid beta peptide (Aß) levels. Episodic memory scores correlate negatively with microglial activation. These results suggest that Caspase-1-mediated inflammation occurs early in the disease and raise hope that VX-765, a previously Food and Drug Administration-approved drug for human CNS clinical trials, may be a useful drug to prevent the onset of cognitive deficits and brain inflammation in AD.

Phase 1 study of the MDM2 antagonist RO6839921 in patients with acute myeloid leukemia.

In acute myeloid leukemia (AML), TP53 mutations and dysregulation of wild-type p53 is common and supports an MDM2 antagonist as a therapy. RO6839921 is an inactive pegylated prodrug of the oral MDM2 antagonist idasanutlin (active principle [AP]) that allows for IV administration. This phase 1 monotherapy study evaluated the safety, pharmacokinetics, and pharmacodynamics of RO6839921 in patients with AML. Primary objectives identified dose-limiting toxicities (DLTs) and maximum tolerated dose (MTD). Secondary objectives assessed pharmacokinetic, pharmacodynamic, and antileukemic activity. A total of 26 patients received 120-300 mg AP of idasanutlin. The MTD was 200 mg, with DLTs at 250 (2/8 patients) and 300 mg (2/5). Treatment-related adverse events in >20% of patients were diarrhea, nausea, vomiting, decreased appetite, and fatigue. Six deaths (23.1%) occurred, all unrelated to treatment. Pharmacokinetics showed rapid and near-complete conversion of the prodrug to AP and dose-proportional exposure across doses. Variability ranged from 30%-47% (22%-54% for idasanutlin). TP53 was 21 (87.5%) wild-type and 3 mutant (12.5%). The composite response rate (complete remission [CR], CR with incomplete hematologic recovery/morphological leukemia-free state [CRi/MLFS], or CR without platelet recovery [CRp]) was 7.7%. Antileukemic activity (CR, CRi/MLFS, partial response, hematologic improvement/stable disease) was observed in 11 patients (disease control rate, 42%): 10/11 were TP53 wild-type; 1 had no sample. p53 activation was demonstrated by MIC-1 induction and was associated with AP exposure. There was not sufficient differentiation or improvement in the biologic or safety profile compared with oral idasanutlin to support continued development of RO6839921. NCT02098967.

Metabolic activation of TM5441 in vitro and in vivo: Formation of reactive metabolites and human enzymes involved.

TM5441, a furan-containing drug, is an inhibitor of plasminogen activator inhibitor-1 (PAI-1), which can induce intrinsic apoptosis of human cancer cell lines. The aim of this study was to identify the reactive metabolites of TM5441 and to reveal the bioactivation pathways that are associated with its hepatotoxicity. The reactive metabolites were trapped by using glutathione (GSH) or N-acetyl-lysine (NAL) in rat, dog, and human liver microsomal incubation system after exposure to TM5441. Two metabolic activation pathways were disclosed. The first bioactivation pathway was dominated by Cytochrome P450 enzymes (CYP450s); TM5441 was metabolized into cis-2-butene-1,4-dial derivative dependent on NADPH, which can be trapped in the liver microsomal incubations fortified with GSH or NAL as trapping agents. Five metabolites (M1, M2, M9, M12 and M13) associated with GSH and three metabolites (M4, M7 and M14) associated with NAL were identified by liquid chromatography-high resolution mass spectrometry. The second bioactivation pathway was catalyzed by UDP-glucuronosyltransferases (UGTs); TM5441 was conjugated with glucuronide to form acyl-glucuronide (M10), which further reacted with GSH, resulting in the identification of a TM5441-S-acyl-GSH adduct (M11) in liver microsomal incubations fortified with uridine-5'-diphosphoglucuronidc acid (UDPGA) and GSH. M9, M10, M11, M12 and M13 were also detected in bile samples of rats given TM5441. Compared with rat, dog would display closer bioactivation profiles to human. The CYP450 enzyme responsible for the bioactivation of TM5441 was mainly identified as CYP3A4, using human recombinant CYP450 enzymes and specific inhibitory studies. The UGT enzymes responsible for the bioactivation of TM5441 mainly involved UGT2B7, 1A1 and 1A4. These results facilitate the understanding of the bioactivation of TM5441 and potential toxicological implications.

Determination of benzonatate and its metabolite in human plasma by HPLC-MS/MS: A preliminary pharmacokinetic study in healthy Chinese volunteers after oral administration of benzonatate soft capsule.

Benzonatate has been used as a non-narcotic oral antitussive drug for many years. Its pharmacokinetics has never been reported due to the technical difficulties in detecting benzonatate by mass spectrometry. However, its concentration can be extrapolated based on the concentration of its metabolite, 4-(butylamino)benzoic acid (BBA). In this study, two sensitive high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) methods were developed and fully validated for the determination of the original 4-(butylamino)benzoic acid (method B) and total 4-(butylamino)benzoic acid (containing the original 4-(butylamino)benzoic acid and 4-(butylamino)benzoic acid converted from benzonatate after collection, method A). For both methods, one-step protein precipitation by methanol was performed to extract analytes from the plasma samples. Chromatographic separation was done on an InfinityLab Poroshell 120 Phenyl Hexyl column (2.1 mm × 50 mm, 2.7 µm, Agilent) with initial mobile phase consisting of 5 mM ammonium acetate containing 0.3% formic acid and acetonitrile (60:40, v/v) at a flow rate of 0.3 mL/min. Quantification was achieved by multiple reaction monitoring (MRM) in electron spray ionization (ESI) positive mode with the transitions of m/z 194.2 → 138.1 and 515.3 → 497.3 for 4-(butylamino)benzoic acid and telmisartan (the internal standard), respectively. The two methods exhibited good linearity over the concentration range of 10-10000 ng/mL. Both of the methods were successfully applied to the preliminary pharmacokinetic study in healthy Chinese volunteers after oral administration of benzonatate soft capsule at a single dose of 100 mg. The results showed that 4-(butylamino)benzoic acid and benzonatate were rapidly absorbed and reached a maximum concentration (Cmax) of 1708 ±â€¯457 ng/mL and 1063 ±â€¯460 ng/mL, respectively. The half-life (t1/2) were 1.32 ±â€¯0.29 h for 4-(butylamino)benzoic acid and 1.01 ±â€¯0.41 h for benzonatate. The area under the curve from 0 h to 10 h (AUC0-10) for 4-(butylamino)benzoic acid and benzonatate were 2103 ±â€¯918 ng/mL·h and 1097 ±â€¯559 ng/mL·h, respectively. And the data was valuable for further clinical study.

Phase 1 summary of plasma concentration-QTc analysis for idasanutlin, an MDM2 antagonist, in patients with advanced solid tumors and AML.

PURPOSE: Idasanutlin, a selective small-molecule MDM2 antagonist in phase 3 testing for refractory/relapsed AML, is a non-genotoxic oral p53 activator. The aim of this analysis is to examine the potential of idasanutlin to prolong the corrected QT (QTc) interval by evaluating the relationship between plasma idasanutlin concentration and QTc interval. METHOD: Intensive plasma concentration QTc interval data were collected at the same timepoints, from three idasanutlin (RO5503781) phase 1 studies in patients with solid tumors and AML. QTc data in absolute values and changes from baseline (Δ) were analyzed for a potential association with plasma idasanutlin concentrations with a linear mixed effect model. Categorical analysis was also performed. RESULTS: A total of 282 patients were exposed to idasanutlin and had at least one observation of QTc and idasanutlin plasma concentration. There was no apparent increase of QTcF or ΔQTcF in a wide idasanutlin plasma concentration range, even at concentrations exceeding the exposure matching the dose adopted in the ongoing phase 3 study (300-mg BID). Categorical analysis did not detect a potential signal of QT prolongation. CONCLUSION: The concentration-QTc analysis indicates that idasanutlin does not prolong the QT interval within the targeted concentration range currently in consideration for clinical development.

Population Pharmacokinetics of an Intranasally Administered Combination of Oxymetazoline and Tetracaine in Healthy Volunteers.

The primary objective of the current investigation was to establish the pharmacokinetic characteristics of oxymetazoline and tetracaine's primary metabolite, para-butylaminobenzoic acid (PBBA), after the intranasal administration of oxymetazoline/tetracaine. Thirty-six subjects contributing a total of 1791 plasma concentration results from 2 open-label trials were utilized. Model development was achieved using data from the second trial (N = 24) in which 0.3 mg oxymetazoline/18 mg tetracaine was administered. External model validation utilized data from the first trial (N = 12), which included doses of 0.3 mg oxymetazoline/18 mg tetracaine and 0.6 mg oxymetazoline/36 mg tetracaine. Oxymetazoline and PBBA dispositions were described by a 2-compartment model with first-order absorption. An allometric model for body weight was included on volumes and clearances to describe unexplained between-subject variability. The final oxymetazoline parameter estimates were ka 4.41 h-1 ; peripheral volume 418 L; clearance 66.4 L/h; central volume 6.97 L; and intercompartmental clearance 419 L/h for a 70-kg subject. The final PBBA parameter estimates were ka 8.51 h-1 ; peripheral volume 32.0 L; clearance 16.7 L/h; central volume 29.8 L; and intercompartmental clearance 2.43 L/h for a 70-kg subject. Between-subject variability ranged from 14% to 39% for oxymetazoline and from 10% to 94% for PBBA.

Identification of N-Hydroxy-para-aminobenzoic acid in a cyanotic child after benzocaine exposure.

CONTEXT: Methemoglobinemia (MetHb) after exposure to benzocaine (BZC) has been reported for more than 50 years, however the pathophysiologic mechanism has not been previously established. Direct administration of BZC to blood does not produce MetHb. After topical use, due to the lipophilicity and rapid acetylation in the tissue, little BZC reaches the liver for hepatic biotransformation. However, isolated human livers have been shown to produce MetHb forming N-hydroxyl metabolites from BZC. We report a case of BZC-induced MetHb with the first identification and quantification of the reactive metabolite responsible for the oxidative stress: N-Hydroxy-Para-amino benzoic acid (N-OH-PABA). CASE DETAILS: An 8 year old male was admitted to a hospital for an appendectomy. Several applications of BZC spray were used during multiple attempts at nasogastric tube placement. In various attempts to achieve local anesthesia, benzocaine spray was used in both nares and through the mouth aimed at the posterior oropharynx. The patient subsequently became cyanotic with an initial MetHb level of 32.9 %. Methylene blue was administered and the patient promptly responded with resolution of cyanosis. Blood taken within 20 min of the initial symptoms contained benzocaine (5.2ug/mL), bupivacaine (740ng/mL), lidocaine (530ng/mL), acetaminophen (12ug/mL), midazolam (60ng/mL), PABA and N-OH-PABA (35ng/mL). Serum was analyzed using Liquid Chromatography- Quadrupole Time-of-Flight Mass Spectrometry. Mass spectrometry was done using an electrospray ionization source run in negative and positive polarities. A reference standard for N-OH-PABA was synthesized for confirmation and quantification. DISCUSSION: The rare and idiopathic nature of methemoglobinemia after benzocaine use has made study of the pathophysiologic mechanism in humans difficult. Lack of understanding has brought calls for restriction of use of the widely used medication that may not be based on evidence. Our case presents several unique features: 1) benzocaine absorption after topical administration was documented with serum concentrations 2) confirmation of an in vivo formation of MetHb-forming n-hydroxyl-metabolite after benzocaine use and 3) the documentation of N-OH-PABA in humans within 20 min of MetHb post-benzocaine administration.