Uridine diphosphate glucuronosyltransferases (UGTs) catalyze the glucuronidation of a wide range of xenobiotics and endogenous substrates. However, there is a lack of information concerning the ...response of human UGTs to inducers, and this observation prompted the current investigation. The glucuronidation of estradiol (3- and 17-positions), naphthol, propofol, and morphine (3- and 6-positions) was assessed against a battery of recombinant human UGTs to determine selective glucuronidation reactions for induction studies. The potential induction of the glucuronidation of estradiol at the 3-position, naphthol, propofol, and morphine at the 3-position was subsequently investigated in cultured primary human hepatocytes against a range of prototypic inducers including dexamethasone, 3-methylcholanthrene (3-MC), phenobarbital, rifampicin, and omeprazole. Treatment with 3-MC induced estradiol-3-glucuronidation (up to 2.5-fold) in four of five donors investigated. Statistically significant increases in naphthol glucuronidation (up to 1.7-fold) were observed following treatment with carbamazepine. UGT1A9-mediated propofol glucuronidation was induced by phenobarbital (up to 2.2-fold) and rifampicin (up to 1.7-fold). However, treatment with alpha-naphthoflavone and tangeretin resulted in a decrease in propofol glucuronidation (30% of control values). Statistically significant induction of morphine-3-glucuronidation was observed in at least three donors following treatment with phenobarbital, rifampicin, and carbamazepine. Each UGT isoform investigated displayed a distinct induction profile. Although statistically significant increases in glucuronidation were observed for each reaction studied, the level of induction was less than that observed for CYP1A2 or CYP3A4 and exhibited a large interdonor variability. The clinical relevance of the induction responses obtained in this study is unclear.
Application of hypothetical strategies in acute pain Zhong, Jinglin; Petullo, David
Pharmaceutical statistics : the journal of the pharmaceutical industry,
May/June 2024, 2024 May-Jun, 2024-05-00, 20240501, Letnik:
23, Številka:
3
Journal Article
Recenzirano
Since the publication of ICH E9 (R1), “Addendum to statistical principles for clinical trials: on choosing appropriate estimands and defining sensitivity analyses in clinical trials,” there has been ...a lot of debate about the hypothetical strategy for handling intercurrent events. Arguments against the hypothetical strategy are twofold: (1) the clinical question has limited clinical/regulatory interest; (2) the estimation may need strong statistical assumptions. In this article, we provide an example of a hypothetical strategy handling use of rescue medications in the acute pain setting. We argue that the treatment effect of a drug that is attributable to the treatment alone is the clinical question of interest and is important to regulators. The hypothetical strategy is important when developing non‐opioid treatment as it estimates the treatment effect due to treatment during the pre‐specified evaluation period whereas the treatment policy strategy does not. Two widely acceptable and non‐controversial clinical inputs are required to construct a reasonable estimator. More importantly, this estimator does not rely on additional strong statistical assumptions and is considered reasonable for regulatory decision making. In this article, we point out examples where estimators for a hypothetical strategy can be constructed without any strong additional statistical assumptions besides acceptable clinical inputs. We also showcase a new way to obtain estimation based on disease specific clinical knowledge instead of strong statistical assumptions. In the example presented, we clearly demonstrate the advantages of the hypothetical strategy compared to alternative strategies including the treatment policy strategy and a composite variable strategy.
In the studies reported here, the ability of atomoxetine hydrochloride (Strattera) to inhibit or induce the metabolic capabilities of selected human isoforms of cytochrome P450 was evaluated. ...Initially, the potential of atomoxetine and its two metabolites, N-desmethylatomoxetine and 4-hydroxyatomoxetine, to inhibit the metabolism of probe substrates for CYP1A2, CYP2C9, CYP2D6, and CYP3A was evaluated in human hepatic microsomes. Although little inhibition of CYP1A2 and CYP2C9 activity was observed, inhibition was predicted for CYP3A (56% predicted inhibition) and CYP2D6 (60% predicted inhibition) at concentrations representative of high therapeutic doses of atomoxetine. The ability of atomoxetine to induce the catalytic activities of CYP1A2 and CYP3A in human hepatocytes was also evaluated; however, atomoxetine did not induce either isoenzyme. Based on the potential of interaction from the in vitro experiments, drug interaction studies in healthy subjects were conducted using probe substrates for CYP2D6 (desipramine) in CYP2D6 extensive metabolizer subjects and CYP3A (midazolam) in CYP2D6 poor metabolizer subjects. Single-dose pharmacokinetic parameters of desipramine (single dose of 50 mg) were not altered when coadministered with atomoxetine (40 or 60 mg b.i.d. for 13 days). Only modest changes (approximately 16%) were observed in the plasma pharmacokinetics of midazolam (single dose of 5 mg) when coadministered with atomoxetine (60 mg b.i.d. for 12 days). Although at high therapeutic doses of atomoxetine inhibition of CYP2D6 and CYP3A was predicted, definitive in vivo studies clearly indicate that atomoxetine administration with substrates of CYP2D6 and CYP3A does not result in clinically significant drug interactions.
On 16 December 2020, FDA approved Benlysta® (belimumab) for both the intravenous (IV) and subcutaneous (SC) administration routes for the treatment of adult patients with active lupus nephritis (LN) ...who are receiving standard therapy. This approval represents the first FDA approved treatment of patients with active LN.
The approved IV dosing regimen (10 mg/kg dose Q2W for three doses, then 10 mg/kg Q4W thereafter) was based on a randomized double-blind placebo controlled clinical trial in adult patients with LN. For the approval of the SC dosing regimen (400 mg dose QW for four doses, then 200 mg QW thereafter), efficacy was supported solely by pharmacokinetics (PK) modeling and simulation which estimated a matched steady state average concentration and higher trough concentrations for the SC administration route, for bridging to the efficacy of IV belimumab in adults with LN. The safety and immunogenicity profile of the SC administration route has been assessed in the SLE studies.
In a population PK analysis, higher proteinuria was associated with greater belimumab clearance and lower belimumab exposure. In an exposure response analysis, the efficacy of belimumab as evaluated by renal response was mainly driven by patients with lower proteinuria at baseline regardless of other baseline characteristics (e.g. baseline renal function, renal biopsy classification), induction therapies, or belimumab exposure levels (within 10 mg/kg dosing regimen), etc. However, post hoc analyses showed that belimumab had activity in LN patients with higher proteinuria at baseline. There is no adequate information to suggest that a higher dose would provide additional benefit for patients with lower exposure (e.g. higher proteinuria).
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, OILJ, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
The gold standard for estimating the ability of a new chemical entity to induce the human cytochromes P450 (P450s) is to determine the increase in catalytic activity of these enzymes after treatment ...of primary cultures of human hepatocytes with the potential inducer. The limited availability of fresh human hepatocytes makes these studies impossible to perform on demand. A substrate cassette strategy in which probe substrates for three different P450s were simultaneously added to hepatocyte preparations was studied to determine whether this would be a viable method to increase the capacity of induction studies. The biotransformations of phenacetin (CYP1A2), diclofenac (CYP2C9), and midazolam (CYP3A) were compared when administered to four different hepatocyte preparations (+/-inducer) individually or simultaneously as a cassette. The determinations of fold-induction in response to known inducers were not significantly affected, although slight differences were occasionally observed between the various P450 activities, whether determined individually or in the cassette. In total, for three P450 activities in the four hepatocyte preparations with and without inducer, no trend demonstrating a drug-drug interaction among any of the three probes was detected. The lack of interactions between the probe substrates demonstrates that this cassette strategy may be used in primary human hepatocyte induction studies without concern that the interactions between the substrates may be affecting the results, as has been seen in other cassette dosing experiments. Therefore, this substrate cassette is an excellent method for increasing the capacity and throughput of human hepatocyte induction studies by combining three experiments into one.
Introduction: In order to examine CYP3A-mediated metabolism in vitro, a unique analytical assay was developed to detect the formation of 6β-hydroxytestosterone (6β-OHT). This assay has been ...determined to be useful for the study of both inhibition- and induction-related drug–drug interactions in vitro and involves simple incubation and sample handling procedures.
Methods: A primary and three backup sets of analytical conditions were developed to detect interference between a test compound and either 6β-OHT or the internal standard.
Results: The primary set of conditions was validated with a three-batch validation, and the remaining sets of conditions were validated with one-batch validations, all in human liver microsomes. The primary assay was also validated with a single batch for CYP3A induction studies in primary human hepatocytes. Enzyme kinetic parameters of 6β-OHT formation (
K
m,
V
max) were determined to be reproducible in human liver microsomes.
Discussion: Utility of the assay in inhibition studies and induction studies, respectively, was confirmed with the test compounds ketoconazole and rifampicin. In addition, superiority to existing methods was demonstrated in three areas: ease of sample preparation, short run times, and low detection limits.
Oxytetracycline (OTC) is an important antimicrobial used in aquaculture. However, residues of OTC have been isolated from nontarget aquatic organisms, sediments, and water located near aquaculture ...facilities. Identifying OTC in plant material is particularly difficult due to interference from pigments and polyphenol substances but is important especially for algae since they are a primary food source for fish in early life stages. In this study, we describe the effect of OTC (0.1, 1, 10, 25, 50, 100 μg · mL⁻¹) on cell growth, and the localization of OTC (0, 1, 25, 100 μg · mL⁻¹) in vacuoles of Chlamydomonas reinhardtii P. A. Dang. (wildtype, ATCC 18798). We also present a method for semiquantifying OTC in living cells using fluorescent microscopy and Adobe Photoshop. We exposed algal cells to OTC and sampled after 2 or 7 d exposure. On day 7, OTC significantly inhibited algal growth at 1, 10, 25, 50, and 100 μg · mL⁻¹. When viewed with fluorescent microscopy, cells exposed to the 25 and 100 μg · mL⁻¹ contained yellow fluorescent areas, less-than or equal to1 μm in diameter that were easily discernable against the red fluorescence of the intracellular chl. The fluorescent areas corresponded to small spherical vacuoles (i.e., polyphosphate bodies that contain calcium and magnesium complexed with polyphosphate) seen in the cells by LM. Since OTC has a high affinity for divalent cations, we suggest that OTC is localized in these vacuoles.
Oxytetracycline (OTC) is an important antimicrobial used in aquaculture. However, residues of OTC have been isolated from nontarget aquatic organisms, sediments, and water located near aquaculture ...facilities. Identifying OTC in plant material is particularly difficult due to interference from pigments and polyphenol substances but is important especially for algae since they are a primary food source for fish in early life stages. In this study, we describe the effect of OTC (0.1, 1, 10, 25, 50, 100 μg · mL−1) on cell growth, and the localization of OTC (0, 1, 25, 100 μg · mL−1) in vacuoles of Chlamydomonas reinhardtii P. A. Dang. (wildtype, ATCC 18798). We also present a method for semiquantifying OTC in living cells using fluorescent microscopy and Adobe Photoshop. We exposed algal cells to OTC and sampled after 2 or 7 d exposure. On day 7, OTC significantly inhibited algal growth at 1, 10, 25, 50, and 100 μg · mL−1. When viewed with fluorescent microscopy, cells exposed to the 25 and 100 μg · mL−1 contained yellow fluorescent areas, ≤1 μm in diameter that were easily discernable against the red fluorescence of the intracellular chl. The fluorescent areas corresponded to small spherical vacuoles (i.e., polyphosphate bodies that contain calcium and magnesium complexed with polyphosphate) seen in the cells by LM. Since OTC has a high affinity for divalent cations, we suggest that OTC is localized in these vacuoles.
Fibroblast growth factor-13 (FGF-13), novel member of FGF family has recently been molecularly cloned as a result of high throughput sequencing of a ovarian cancer cell, hippocampal, and kidney cDNA ...libraries. The human gene encodes for a protein with a molecular weight of 22 kDa that is most homologous to FGF-8 (70% similarity). In the current study, we tested the effects of intravenously administered FGF-13 in a model of permanent focal cerebral ischemia in Sprague–Dawley rats. FGF-13 or the vehicle was administered systematically via the tail vein 30 min prior, and 30 min and 24 h after the occlusion of the left middle cerebral artery (MCAo). Animals were weighed and evaluated behaviorally prior to and at 24 and 48 h after MCAo. The volume of cerebral infarct and swelling were determined using an image analysis system (BioQuant) and cresyl violet stained sequential sections from the forebrain region. Histopathology was evaluated to compare the therapeutic effects. We found a 63% reduction in infarct volume in FGF-13- vs. vehicle-treated animals (infarct volume was 21.9±3.8% in vehicle- and 8.1±1.6% in FGF-13-treated rats,
p=0.0016) and a moderate inhibition of brain swelling by FGF-13. The reduction in infarct volume and brain swelling were associated with improvement of clinical deficits in FGF-13 treated animals (
p<0.001). Histopathological examination determined that nervous tissue was better preserved in FGF-13 treated rats than those of controls. These data show that pretreatment with intravenous FGF-13 reduces infarct size and ameliorates neurological deficits following permanent focal cerebral ischemia in rats.
