Maximum Clinical Dose Research Articles

Gadolinium-based coronary CT angiography on a clinical photon-counting-detector system: a dynamic circulating phantom study

BackgroundCoronary computed tomography angiography (CCTA) offers non-invasive diagnostics of the coronary arteries. Vessel evaluation requires the administration of intravenous contrast. The purpose of this study was to evaluate the utility of gadolinium-based contrast agent (GBCA) as an alternative to iodinated contrast for CCTA on a first-generation clinical dual-source photon-counting-detector (PCD)-CT system.MethodsA dynamic circulating phantom containing a three-dimensional-printed model of the thoracic aorta and the coronary arteries were used to evaluate injection protocols using gadopentetate dimeglumine at 50%, 100%, 150%, and 200% of the maximum approved clinical dose (0.3 mmol/kg). Virtual monoenergetic image (VMI) reconstructions ranging from 40 keV to 100 keV with 5 keV increments were generated on a PCD-CT. Contrast-to-noise ratio (CNR) was calculated from attenuations measured in the aorta and coronary arteries and noise measured in the background tissue. Attenuation of at least 350 HU was deemed as diagnostic.ResultsThe highest coronary attenuation (441 ± 23 HU, mean ± standard deviation) and CNR (29.5 ± 1.5) was achieved at 40 keV and at the highest GBCA dose (200%). There was a systematic decline of attenuation and CNR with higher keV reconstructions and lower GBCA doses. Only reconstructions at 40 and 45 keV at 200% and 40 keV at 150% GBCA dose demonstrated sufficient attenuation above 350 HU.ConclusionCurrent PCD-CT protocols and settings are unsuitable for the use of GBCA for CCTA at clinically approved doses. Future advances to the PCD-CT system including a 4-threshold mode, as well as multi-material decomposition may add new opportunities for k-edge imaging of GBCA.Relevance statementPatients allergic to iodine-based contrast media and the future of multicontrast CT examinations would benefit greatly from alternative contrast media, but the utility of GBCA for coronary photon-counting-dector-CT angiography remains limited without further optimization of protocols and scanner settings.Key PointsGBCA-enhanced coronary PCD-CT angiography is not feasible at clinically approved doses.GBCAs have potential applications for the visualization of larger vessels, such as the aorta, on PCD-CT angiography.Higher GBCA doses and lower keV reconstructions achieved higher attenuation values and CNR.Graphical

A randomized placebo controlled trial demonstrates the effect of dl-methylephedrine on brain functions is weaker than that of pseudoephedrine

Intellectual drug doping in athletics by using stimulants that affect central nervous system functions has been diversified. Stimulants are regulated by the World Anti-Doping Agency according to their levels of urinary concentration. Positron emission tomography could evaluate how stimulants affect central nervous system functions. We aimed to evaluate the effect of stimulants on brain function by examining the difference in brain dopamine transporter occupancy by PET after administration of dl-methylephedrine or pseudoephedrine at the clinical maximum daily dose. Four PET scans without and with drug administration (placebo, dl-methylephedrine 150 mg and pseudoephedrine 240 mg) were performed. The concentrations of dl-methylephedrine and pseudoephedrine in plasma and urine were measured. DAT occupancies in the striatum with placebo, dl-methylephedrine and pseudoephedrine were calculated by PET images. The urinary concentration of dl-methylephedrine (12.7 µg/mL) exceeded the prohibited concentration (10 µg/mL), but the DAT occupancy with dl-methylephedrine (6.1%) did not differ (p = 0.92) from that with placebo (6.2%). By contrast, although the urinary concentration of pseudoephedrine (144.8 µg/mL) was below the prohibited concentration (150 μg/mL), DAT occupancy with pseudoephedrine was 18.4%, which was higher than that with placebo (p = 0.009). At the maximum clinical dose, dl-methylephedrine was shown to have weaker effects on brain function than pseudoephedrine.

Process validation and preclinical development of a new PET cerebral blood flow tracer [11C]MMP for initial clinical trials

Background2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) is commonly used for diagnosis of dementia because brain glucose metabolism reflects neuronal activity. However, as [18F]FDG is an analogue of glucose, accumulation of tracer in the brain is affected by plasma glucose levels. In contrast, cerebral blood flow (CBF) tracers are theoretically unaffected by plasma glucose levels and are therefore expected to be useful alternatives for the diagnosis of dementia in patients with diabetes. The techniques currently used for CBF imaging using single photon emission computed tomography (SPECT) and [15O]H2O positron emission tomography (PET), but these are limited by their insufficient resolution and sensitivity for regional brain imaging, especially in patients with brain atrophy. N-isopropyl-4-[11C]methylamphetamine ([11C]MMP) is a possible CBF tracer with high resolution and sensitivity that exhibits comparable performance to that of [15O]H2O in conscious monkey brains. We performed process validation of the radiosynthesis and preclinical development of [11C]MMP prior to clinical translation.ResultsThe decay-corrected yields of [11C]MMP at the end of synthesis were 41.4 ± 6.5%, with 99.7 ± 0.3% radiochemical purity, and 192.3 ± 22.5 MBq/nmol molar activity. All process validation batches complied with the product specifications. The acute toxicity of MMP was evaluated at a dose of 3.55 mg/kg body weight, which is 10,000 times the potential maximum clinical dose of [11C]MMP. The acute toxicity of [11C]MMP injection at 150 or 200 times, to administer a postulated dose of 740 MBq of [11C]MMP, was also evaluated after the decay-out of 11C. No acute toxicity of MMP and [11C]MMP injection was found. No mutagenic activity was observed for MMP. The effective dose calculated according to the Medical Internal Radiation Dose (MIRD) method was 5.4 µSv/MBq, and the maximum absorbed dose to the bladder wall was 57.6 µGy/MBq. MMP, a derivative of phenylalkylamine, showed binding to the sigma receptor, but had approximately 1/100 of the affinity of existing sigma receptor imaging agents. The affinity for other brain neuroreceptors was low.Conclusions[11C]MMP shows acceptable pharmacological safety at the dose required for adequate PET imaging. The potential risk associated with [11C]MMP PET imaging is well within the acceptable dose limit.

A “toxic window” study on the hippocampal development of mice offspring exposed to azithromycin at different doses, courses, and time during pregnancy

BackgroundAzithromycin, one of the new-generation macrolides, is an effective medicine for the treatment of mycoplasma infection during pregnancy. Epidemiological studies have reported adverse pregnancy outcomes with prenatal azithromycin exposure (PAzE). However, the effect of PAzE on fetal hippocampal development is unclear. This study aimed to explore the effects and potential mechanism of PAzE-induced fetal hippocampal development at different doses, courses, and time. MethodPregnant mice were administered azithromycin by gavage at different doses (50, 100 or 200 mg/kg.d), different courses (gestational day (GD)15–17 for three consecutive days, or GD17 once a day) and different time (GD10-12, GD15-17). ResultsCompared with the control group, morphological development damage of the fetal hippocampus was observed in the PAzE group, with a dysbalance in neuronal proliferation and apoptosis, decreased expression of the neuronal-specific marker Snap25, NeuN, PSD95 and Map2, increased expression of the glial-specific marker Iba1, GFAP, and S-100β, and decreased expression of P2ry12. The PAzE-induced hippocampal developmental deficiency varied based on different doses, courses, and time, and the developmental toxicity was most significant in the late pregnancy, high dose, multi-course group (AZHT). The significant reduction of SOX2 and Wnt, which were related to regulation of neural progenitor cells (NPCs) proliferation in PAzE fetus compared with the control group indicated that the SOX2/Wnt signaling may be involved in PAzE-induced hippocampal developmental toxicity. ConclusionIn this study, PAzE was associated with hippocampal developmental toxicity in a variety of nerve cells. Hippocampal developmental toxicity due to azithromycin was most significant in the late pregnancy, high-dose (equivalent to maximum clinical dose) and multi-course group (AZHT). The findings provide an experimental and theoretical foundation for guiding the sensible use of medications during pregnancy and effectively assessing the risk of fetal hippocampal developmental toxicity.

Effect of budesonide on pulmonary activity of multidrug resistance-associated protein 1 assessed with PET imaging in rats

Multidrug resistance-associated protein 1 (MRP1/ABCC1) is a highly abundant efflux transporter in the lungs, which protects cells from toxins and oxidative stress and has been implicated in the pathophysiology of chronic obstructive pulmonary disease and cystic fibrosis. There is evidence from in vitro studies that the inhaled glucocorticoid budesonide can inhibit MRP1 activity. We used positron emission tomography (PET) imaging with 6-bromo-7-[11C]methylpurine ([11C]BMP), which is transformed in vivo into a radiolabeled MRP1 substrate, to assess whether intratracheally (i.t.) aerosolized budesonide affects pulmonary MRP1 activity in rats. Three groups of rats (n = 5–6 each) underwent dynamic PET scans of the lungs after i.t. aerosolization of either [11C]BMP alone, or [11C]BMP mixed with either budesonide (0.04 mg, corresponding to the maximum soluble dose) or the model MRP1 inhibitor MK571 (2 mg). From PET-measured radioactivity concentration-time curves, the rate constant describing radioactivity elimination from the right lung (kE,lung) and the area under the curve (AUClung) were calculated from 0 to 5 min after start of the PET scan as measures of pulmonary MRP1 activity. Co-administration of MK571 resulted in a pronounced decrease in kE,lung (25-fold, p < 0.0001) and an increase in AUClung (5.3-fold, p < 0.0001) when compared with vehicle-treated animals. In contrast, in budesonide-treated animals kE,lung and AUClung were not significantly different from the vehicle group. Our results show that i.t. aerosolized budesonide at an approximately 5 times higher dose than the maximum clinical dose leads to no change in pulmonary MRP1 activity, suggesting a lack of an effect of inhaled budesonide treatment on the MRP1-mediated cellular detoxifying capacity of the lungs. However, the strong effect observed for MK571 raises the possibility for the occurrence of transporter-mediated drug-drug interactions at the pulmonary epithelium with inhaled medicines.

Validation of Risk-Stratification Method for the Chronic Atrioventricular Block Cynomolgus Monkey Model and Its Mechanistic Interpretation Using 6 Drugs With Pharmacologically Distinct Profile.

Validation of risk-stratification method for the chronic atrioventricular block cynomolgus monkey model and its mechanistic interpretation was performed using 6 pharmacologically distinct drugs. The following drugs were orally administered in conscious state, astemizole: 1, 5, and 10 mg/kg (n = 6); haloperidol: 1, 10, and 30 mg/kg (n = 5); amiodarone: 30 mg/kg (n = 4); famotidine: 10 mg/kg (n = 4); levofloxacin: 100 mg/kg (n = 4); and tolterodine: 0.2, 1, and 4.5 mg/kg (n = 4). Astemizole of 5 and 10 mg/kg significantly prolonged ΔΔQTcF, whereas no significant change was observed by the others. Torsade de pointes (TdP) was induced by astemizole of 5 and 10 mg/kg in 3/6 and 6/6, and by haloperidol of 10 and 30 mg/kg in 1/5 and 1/5, respectively, which was not observed in the others. Torsadogenic risk of the drugs was quantified using the criteria for the monkey model specified in our previous study. Namely, high-risk drugs induced TdP at ≤ 3 times of their maximum clinical daily dose. Intermediate-risk drugs did not induce TdP at this dose range, but induced it at higher doses. Low/no-risk drugs never induced TdP at any dose tested. The magnitude of risk was intermediate for astemizole and haloperidol, and low/no risk for the others. The prespecified, risk-stratification method for the monkey model may solve the issue existing between nonclinical models and patients with labile repolarization, which can reinforce the regulatory decision-making and labeling at time of marketing application of nondouble-negative drug candidate (hERG assay positive and/or in vivo QT study positive).

Automated synthesis, preclinical toxicity, and radiation dosimetry of [18F]MC225 for clinical use: a tracer for measuring P-glycoprotein function at the blood-brain barrier

Introduction[18F]MC225 is a selective substrate for P-glycoprotein (P-gp) that has good metabolic stability and shows higher baseline uptake compared with other P-gp substrates such as (R)-[11C]Verapamil. Prior to clinical translation, it is necessary to perform process validation of the radiosynthesis, assessment of preclinical toxicity, and radiation dosimetry.MethodsThe production of [18F]MC225 was automated on a CFN-MPS200 multipurpose synthesizer. The acute toxicity of MC225 was evaluated at a dose of 2.5 mg/kg bodyweight, which is more than 10,000-fold the postulated maximum clinical dose of [18F]MC225. The acute toxicity of [18F]MC225 injection at a 200-fold dose, to administer a postulated dose of 185 MBq of [18F]MC225, was also evaluated after the decay-out of 18F. The mutagenicity of MC225 was studied by a reverse mutation test using Salmonella typhimurium and Escherichia coli (Ames test). In vivo biodistribution and dosimetry studies of [18F]MC225 were carried out in normal mice. Human dosimetry was estimated using OLINDA software.ResultsThe mean decay-corrected yields of [18F]MC225 at end of synthesis were 13%, with > 99% radiochemical purity, > 1000 GBq/μmol molar activity, and ≤ 1.5 μg/185 MBq of total chemical contents. All process validation batches complied with the product specifications and the process was confirmed to be appropriate for the production of [18F]MC225. No acute toxicity of MC225 or [18F]MC225 injection was found. No mutagenic activity was observed for MC225. The biodistribution study demonstrated both hepatobiliary and renal excretion of radioactivity. The most critical organ was the pancreas, with (63.8 μGy/MBq) or without urination (63.9 μGy/MBq) at 360 min after injection. The estimated effective dose (μSv/MBq) with and without urination at 360 min after injection was calculated as 15.7 and 16.9, respectively.Conclusion[18F]MC225 shows acceptable pharmacological safety at the dose required for adequate PET imaging. The potential risk associated with [18F]MC225 PET imaging is well within acceptable dose limits.

A Simple and Rapid LC-MS/MS Method for Therapeutic Drug Monitoring of Lenalidomide

Immunomodulatory drugs lenalidomide (LENA) and pomalidomide (POMA) are synthetic compounds derived by modifying the chemical structure of thalidomide to improve its potency and reduce its side effects. LENA is used as a treatment for myeloma and blood disorders called myelodysplastic syndromes. The maximum clinical dose of LENA for some haematological cancers is generally ≤25 mg. Therapeutic drug monitoring (TDM) is important for the individualization of drug dosage. A highly sensitive and high performance liquid chromatography tandem mass spectrometry (LC–MS/MS) assay was developed and validated for the quantification of LENA in human plasma. LENA was extracted from human plasma by liquid-liquid extraction by ethyl acetate and analysed using a reversed phase isocratic elution on a Poroshell 120 EC-C18, (4.6 - 50 mm, 2.7μm) column. 0.1% formic acid: methanol (10:90% v/v), was used as mobile phase and detection was performed by triple quadrupole mass spectrometry LC-MS/MS using jet stream electrospray ionization in negative mode. POMA was used as the internal standard (IS). Analyte and IS were detected by tandem mass spectrometry using MRM of precursor–product ion transitions with 0.100 s dwell time, at m/z 258.0 > 213.0 for LENA and m/z 272.0 > 161.0 for POMA. The calibration curves were consistently accurate and precise over the concentration range of 20 ng/mL to 1000 ng/mL in plasma for LENA. This novel LC–MS/MS method competes with all the regulatory requirements and shows satisfactory accuracy and precision. It is sufficiently sensitive for the performance of pharmacokinetic, bioequivalence and TDM studies in humans.

A six-weekly dosing schedule for pembrolizumab in patients with cancer based on evaluation using modelling and simulation

BackgroundPembrolizumab is approved for multiple cancer types at 200 mg and 2 mg/kg dose every 3 weeks (Q3W). We used a model-based approach to compare the exposure of pembrolizumab 400 mg dose every 6 weeks (Q6W) with the Q3W regimens. MethodsThe Q6W dose was selected by matching exposure with the 200 mg and 2 mg/kg Q3W doses. Concentration-time profiles were simulated using the established population pharmacokinetic model of pembrolizumab based on 2993 subjects from five clinical trials across tumour types. Efficacy was bridged by evaluating projections of average concentration over the dosing interval (Cavg) and trough concentration (Cmin) at steady state (ss). Safety was bridged by ensuring that concentrations were below those at 10 mg/kg dose every 2 weeks (Q2W), the maximum clinical dose. ResultsThe 400 mg Q6W dose had similar predicted exposure (Cavg,ss, geometric mean ∼1% higher) as the 200 mg Q3W dose. Fewer than 1% of subjects had transiently lower Cmin,ss than that observed for 200 mg and 2 mg/kg Q3W. Despite these reductions, similar target saturation is expected. The predicted peak concentrations (Cmax,ss) for 400 mg Q6W were substantially (∼65%) lower than the 10 mg/kg Q2W dose. ConclusionsExposures expected for pembrolizumab 400 mg Q6W were similar to the 200 mg and 2 mg/kg Q3W and below the 10 mg/kg Q2W regimens. Established exposure-response relationships for pembrolizumab over a 5-fold dose range (2 mg/kg Q3W to 10 mg Q2W) support that clinical efficacy and safety of 400 mg Q6W would be similar to the 200 mg and 2 mg/kg Q3W doses across tumour types. Clinical trial registration numbersNCT01295827, NCT01704287, NCT01866319, NCT01905657, NCT02142738.

Concentration-QTc (C-QTc) Analysis of the MDM2 Inhibitor KRT-232 (formerly AMG 232) in Subjects with Advanced Solid Tumors, Multiple Myeloma, or Acute Myeloid Leukemia

Introduction: KRT-232 is a potent and selective, targeted small molecule inhibitor of human mouse double minute 2 (MDM2) homolog interactions with tumor protein 53 (p53). MDM2 prevents p53 activation and reduces p53-mediated transcription and cell cycle control. KRT-232 is under development by Kartos Therapeutics for treatment of myelofibrosis, polycythemia vera, acute myeloid leukemia (AML) and Merkel cell carcinoma (see NCT03662126, NCT03669965, NCT03787602). The KRT-232 no effect-level for in vitro inhibition of hERG function (10 μM) was approximately 147- and 73-fold greater than KRT-232 unbound Cmax concentrations for steady state doses of 240 mg and 480 mg, respectively, based on population pharmacokinetic (PK)-derived parameters for subjects with AML (Ma et al. submitted, ASH 2019). The primary objective of this analysis was to evaluate the relationship between KRT-232 plasma concentration and changes in heart rate-corrected QT interval duration (QTc) in oncology patients treated in Amgen studies 20120106 (Gluck et al. Invest New Drugs; in press, NCT01723020) and 20120234 (Erba et al. Blood Adv 2019; NCT02016729). Methods Study 20120106 was a 2-part Phase 1 dose-exploration and dose-expansion monotherapy study in advanced solid tumors or multiple myeloma. KRT-232 doses of 15 mg (n=3), 30 mg (n=3), 60 mg (n=4), 120 mg (n=7), 240 mg (n=76), 300 mg (n=4), 360 mg (n=4) and 480 mg (n=6) were administered daily (QD) for 7 days in 21-day cycles. Subjects received up to 31 cycles of treatment. Study 20120234 was a Phase 1b study evaluating KRT-232 alone and in combination with trametinib in relapsed/refractory AML. Subjects received the following KRT-232 doses: 60 mg (n=14; n=10 co-administered with 2 mg trametinib daily [excluded from C-QTc analysis]); n=4 as single agent), 90 mg (n=4), 180 mg (n=5), 240 mg (n=3), and 360 mg (n=10). Doses were administered QD for 7 days in 14-day cycles. Subjects received up to 46 cycles of treatment. In both studies, time-matched PK and ECG measurements were collected intensively during Cycle 1 and less frequently at other visits. Triplicate 12-lead ECG data (N=3) were read by a central laboratory. A linear mixed effects model using R (v 3.5.2) was used to analyze the relationship between KRT-232 plasma concentrations and the QT interval corrected using Fridericia's method (QTcF). Effects of baseline QTcF, study, sex and tumor type on C-QTc were investigated. The upper bound of 2-sided 90% CIs for the mean QTcF change from baseline (ΔQTcF) predicted at Cmax was compared to the 10 ms threshold of regulatory concern (FDA Guidance: E14(R3) 2017; Garnett et al. Pharmacokinet Pharmacodyn 2018). Results ECG and PK data for this analysis were available from 130 subjects. The final model was a linear mixed-effect model with parameters for intercept, KRT-232 concentration-ΔQTcF slope, and baseline QTcF effect on the intercept. Diagnostic plots indicated an adequate model fit. The final C-QTc model was used to predict mean ΔQTcF and associated 2-sided 90% CI mean steady-state KRT-232 Cmax at doses up to the maximum clinical dose of 480 mg QD, in subjects with AML or solid tumors. The mean and upper bound of the 90% CI of ΔQTcF were predicted not to exceed 10 ms at doses of up to 480 mg QD in subjects with AML, multiple myeloma or solid tumors. Mean (90% CI) predicted ΔQTcF values at 480 mg QD were 2.040 (0.486, 3.595) ms for subjects with solid tumors and 4.521 (2.348, 6.693) ms for subjects with AML (Figure A). The KRT-232 concentrations at which the upper bounds of 90% CI of mean ΔQTcF are predicted to reach 10 ms and 20 ms are 4298 ng/mL and 7821 ng/mL, respectively. These concentrations are 2.2- and 4-fold higher, respectively, than the predicted mean steady-state Cmax for 480-mg KRT-232 in subjects with solid tumors, and 1.4- and 2.5-fold higher, respectively, than the corresponding mean steady-state Cmax in subjects with AML. Conclusion Since the mean and upper bound of the 90% CI of mean ΔQTcF were predicted not to exceed 10 ms at KRT-232 doses of up to 480 mg QD in solid tumor or AML patients, KRT-232 should not result in clinically meaningful QT prolongation at the doses currently under investigation in Kartos clinical trials. Disclosures Taylor: Certara Strategic Consulting: Consultancy, Employment. Allard:Certara Strategic Consulting: Consultancy, Employment. Kresja:Kartos Therapeutics: Employment, Equity Ownership. Lee:Kartos Therapeutics: Employment, Equity Ownership. Slatter:Kartos Therapeutics: Employment, Equity Ownership. OffLabel Disclosure: KRT-232 (formerly AMG 232) is a small molecule MDM2 inhibitor

Nonclinical Safety of SHP 826 (BAX 826), a Next Generation Extended Half-Life Recombinant Factor VIII Product

Hemophilia A is a rare genetic bleeding disorder caused by missing or defective factor VIII (FVIII), a crucial factor in blood coagulation. Baxalta Innovations / Shire is currently developing SHP 826 (BAX 826), as the next generation EHL rFVIII replacement for the treatment of hemophilia A. SHP 826 is the first EHL rFVIII to utilize novel technology in which polysialic acid (PSA) is conjugated to Baxalta's licensed rFVIII product (Advate) to extend the circulation half-life of FVIII.The potential toxicity of SHP 826 was evaluated in Sprague Dawley rats and cynomolgus monkeys. Acute toxicity was assessed in a dose-escalation study in monkeys that were intravenously administered 350 and 1800 IU rFVIII/kg SHP 826. Repeat dose toxicity was assessed in rats at doses of 80, 350, and 800 IU rFVIII/kg, in monkeys at doses of 800 IU rFVIII/kg every 5 days for 4 weeks, and in monkeys at doses of 80, 350, or 600 IU rFVIII/kg every five days for 31 days. Cardiovascular and respiratory safety was assessed in one of the repeat dose toxicity studies in monkeys. The thrombogenic potential of SHP 826 was evaluated after a single intravenous administration in rabbits (900 rFVIII/kg BW) using a stasis model developed by Wessler et al. (1959). The licensed rFVIII product was used as a comparator in the acute toxicity study and in the Wessler test.Parameters evaluated included local tolerance at the injection site, body weight, clinical pathology, blood gas analysis, analysis of neutralizing and binding antibodies in plasma, organ weight, necropsy observation, and histopathological evaluation. In addition, seminology (epididymal and testicular sperm) was assessed in rats, and urinalysis, ophthalmic and telemetric cardiovascular examination and respiratory rate measurement was performed in monkeys.SHP 826 showed a favorable safety/toxicity profile in both species used, with no abnormalities directly caused by test item administration noted in vivo or during clinical or histopathological examination. The NOAEL was set to the highest dose tested in each study (i.e. 800 IU rFVIII/kg BW in rats and 600 IU rFVIII/ kg BW in monkeys). Toxicokinetic evaluation indicated dose proportional PK. Exposure to FVIII decreased after repeated treatments due to appearance of anti-FVIII antibodies in both species. Development of anti-FVIII antibodies is an expected immune response after repeated application of heterologous proteins, and is not predictive of any potential for triggering development of anti-FVIII antibodies in humans.Safety pharmacology studies with SHP 826 revealed no evidence of thrombogenic potential in rabbits. SHP 826 administered intravenously was well tolerated in conscious telemetered cynomolgous monkeys (part of the repeat dose toxicity study) and did not cause any adverse clinical, cardiovascular, or respiratory effects.The results of these safety studies indicate sufficient safety margins to support the anticipated maximum clinical dose, and therefore entry into clinical development. DisclosuresEhrlich:Shire: Employment. Leidenmuehler:Shire: Employment. Dietrich:Shire: Employment. Hoebarth:Shire: Employment. Ruthsatz:Shire: Employment. Turecek:Shire: Employment. Wolfsegger:Shire: Employment, Equity Ownership. Weber:Shire: Employment. Gritsch:Shire: Employment. Hoellriegl:Shire: Employment. Turecek:Shire: Employment.

Is There a Causal Relation between Maternal Acetaminophen Administration and ADHD?

ObjectiveRecent epidemiological studies reported an association between maternal intake of acetaminophen (APAP) and attention deficit hyperactivity disorder (ADHD) in their children. However, none of these studies demonstrated causality. Our objective was to determine whether exposure to APAP during pregnancy result in hyperkinetic dysfunctions in offspring, using a murine model.Material and MethodsPregnant CD1 mice (N = 8/group) were allocated to receive by gavage either APAP (150 mg/kg/day, equivalent to the FDA-approved maximum human clinical dose), or 0.5% carboxymethylcellulose (control group), starting on embryonic day 7 until delivery. Maternal serum APAP and alanine transaminase (ALT) concentrations were determined by ELISA and kinetic colorimetric assays, respectively. Open field locomotor activity (LMA) in the 30-day old mouse offspring was quantified using Photobeam Activity System. Mouse offspring were then sacrificed, whole brains processed for magnetic resonance imaging (MRI; 11.7 Tesla magnet) and for neuronal quantification using Nissl stain. The association between APAP exposure and LMA in mouse offspring was analyzed using a mixed effects Poisson regression model that accounted for mouse offspring weight, gender, random selection, and testing time and day. We corrected for multiple comparisons and considered P<0.008 as statistically significant.ResultsMaternal serum APAP concentration peaked 30 minutes after gavage, reaching the expected mean of 117 μg/ml. Serum ALT concentrations were not different between groups. There were no significant differences in vertical (rearing), horizontal, or total locomotor activity between the two rodent offspring groups at the P level fixed to adjust for multiple testing. In addition, no differences were found in volumes of 29 brain areas of interest on MRI or in neuronal quantifications between the two groups.ConclusionThis study refutes that hypothesis that prenatal exposure to APAP causes hyperkinetic dysfunction in mouse offspring. Due to lack of accurate assessment of ADHD in murine models, our results should be taken with caution when compared to the reported clinical data.

Hydroxyurea Adherence By Personal Best HbF and the 2-Site “HABIT” Intervention Trial for Pediatric Sickle Cell Disease

IntroductionHydroxyurea (HU) induction of fetal hemoglobin (HbF) is a major therapeutic effect for sickle cell disease (SCD) HbSS or HbSB0thalassemia. Non-adherence to a HU daily regimen is a barrier to its full effect. As no uniform level of HbF induction exists, we hypothesized that a child’s historical “personal best” (PB) HbF level at maximum clinical dose can be used as a marker for HU adherence. The NIH-funded “HABIT” study is a two-site randomized trial to improve patient-centered HU adherence in adolescents age 10-18 years. Eligibility is determined by fall-off from PB HbF over the previous year. Adherence measures include tracking HbF, pharmacy refill data and self-report. MethodsIn a cross-sectional analysis of the entire 2-site clinic-based sample of 95 youth with SCD on HU, ages 10-18 years, and from the subset enrolled in HABIT to date, we assessed demographics, HU use and HbF at pre-treatment, PB and a recent time point. Here, HU non-adherence was defined as ≥20% deviation from PB HbF. Data were analyzed using descriptive statistics and Pearson correlation; groups were stratified by site and participation in HABIT, and were compared using chi-square and student’s t-tests. ResultsOnly ethnicity and mean HU dosing significantly differed by site (more Latinos/other and lower HU dosing at Columbia vs. Einstein). Only 39 (41%) youth remained within 20% of their PB HbF, with no significant differences in deviation from PB HbF by: site, gender; ethnicity; age (< or ≥14 years); higher induced PB (HbF of ≥20%); or time to PB (≤ or >1 year). For the 39 adherent to HU, mean deviation from PB HbF was 7.3±8.4% (median 7.2); for the 56 non-adherent, mean deviation was 40.5±16.4% (median 36.8; p<0.001). Greater deviation from PB was associated with: 1) Younger age at HU initiation (r = -0.24, p = 0.03); and 2) Longer duration of HU use (r = 0.28, p=0.009). For the 13 HABIT subjects, significant differences from the other 82 patients were limited to the greater proportion of Latinos/other and slightly older ages. Analyses will be performed to examine relationships between deviation from PB HbF, alterations in MCV, acute clinical events and use of urgent medical services (ER use/admissions). ConclusionOur definition of HU non-adherence as ≥20% deviation from Personal Best HbF identified two non-overlapping patient groups ages 10-18 years, with the majority found to be non-adherent. These findings also suggest that HU initiation at younger ages may increase risk of under-adherence during adolescence. PB HbF appears to be a useful marker for HU adherence, and may aid in clinical and patient-centered assessment and intervention. These results underscore the need to assess HU adherence in adolescent patients and for intervention trials such as HABIT.Abstract 1383. Table:Characteristics of the 2-site sample on hydroxyurea and subjects enrolled in the HABIT studyTotal(N =95)Columbia (N=39)Einstein (N=56)P valueHABIT (N=13)P1 valueN%N%N%N%Female gender4345.31846.22544.60.88538.50.60Ethnicity Hispanic/other2 African Amer 25 5926.3 62.1 22 17 56.4 43.6 14 42 25.0 75.0 0.0029 469.2 30.8 0.03Age group 10-13 years 14-18 years44 5146.3 53.721 1853.9 46.123 3341.1 58.90.223 1023.1 76.90.07MeanSDMeanSDMeanSDMeanSDCurrent age (years)14.32.614.12.714.42.50.6315.72.30.03Age at HU initiation (years)9.23.78.84.39.53.70.449.14.80.91Age at Personal Best (years)11.93.311.23.712.33.00.1413.23.70.12HU duration (years)4.63.24.93.54.32.90.366.04.20.10Pre-HU HbF HU7.74.87.24.68.14.90.416.62.00.42Time to Personal Best (years)2.52.32.52.72.42.10.964.03.30.09Personal Best HbF18.76.618.56.618.76.70.8718.05.80.72HU dose at Personal Best (mg/kg/day)24.13.822.64.125.13.30.00225.03.80.32Recent HbF13.66.313.26.413.86.30.6212.15.90.36HbF increase from pre-HU to PB10.86.011.64.710.26.80.2911.07.20.92Recent HU dose (mg/kg/day)23.94.021.94.525.42.7<0.00123.53.60.74Deviation from PB HbF28.821.728.821.726.220.50.5734.121.50.21N%N%N%N%Proportion Who Deviate from PB0.670.16Adherent (≤20% deviation)3941.11538.52442.93 23.1Non-adherent (>20% deviation)5658.92461.53257.11076.91Comparing HABIT subjects to the remaining 82 patients in the sample.2Designation of “other” includes multi-racial HABITHydroxyurea Adherence for Personal Best in Sickle Cell Treatment Funding: 5R21NR013745 (PIs NSG and AMS) DisclosuresOff Label Use: Hydroxyurea is not FDA approved for use in children..

Combined actions of ivabradine and ranolazine reduce ventricular rate during atrial fibrillation.

Ventricular rate during atrial fibrillation (AF) can be reduced by slowing atrioventricular (AV) node conduction and/or by decreasing dominant frequency of AF. We investigated whether combined administration of ivabradine and ranolazine reduces ventricular rate during AF. Ivabradine (maximum clinical dose, 0.25 mg/kg, and 0.10 mg/kg, i.v.) and ranolazine (2.4 mg/kg, i.v., bolus followed by 0.135 mg/kg/min) were studied in an anesthetized pig (N = 16) model of AF. Combined administration of 0.25 mg/kg ivabradine with ranolazine reduced ventricular rate during AF by 51.9 ± 9.7 beats/min (23%, P = 0.017) and dominant frequency of AF by 2.8 ± 0.5 Hz (32%, P = 0.005). It increased PR (P = 0.0002, P = 0.0007) and A-H intervals (P = 0.047, P = 0.002) during pacing at 130 and 180 beats/min, respectively, to a greater degree than additive effects of single agents. Combined administration of 0.1 mg/kg ivabradine with ranolazine exceeded additive effects of single agents on A-H intervals and dominant frequency of AF. Moreover, ranolazine potentiated low-dose ivabradine's reduction in ventricular rate, as combined administration more than doubled effects of the higher ivabradine dose alone and was similar to the combination with the higher dose. Neither drug nor their combination affected contractility (left ventricular [LV] dP/dt), QT or His-ventricular (H-V) intervals, or mean arterial pressure during sinus rhythm or AF. Combined administration of ivabradine and ranolazine at clinically safe levels decreases ventricular rate during AF by reducing AV node conduction and AF dominant frequency without QT prolongation or depression in contractility. Targeting these actions offers intrinsic advantages over conventional nodal agents, which can reduce contractility.

Carcinogenicity in rats of the SGLT2 inhibitor canagliflozin

The carcinogenicity potential of canagliflozin, an inhibitor of SGLT2, was evaluated in a 2-year rat study (10, 30, and 100mg/kg). Rats showed an increase in pheochromocytomas, renal tubular tumors, and testicular Leydig cell tumors. Systemic exposure multiples at the highest dose relative to the maximum clinical dose were 12- to 21-fold. Pheochromocytomas and renal tubular tumors were noted in both sexes at 100mg/kg. Leydig cell tumors were observed in males in all dose groups and were associated with increased luteinizing hormone levels. Hyperplasia was increased in the adrenal medulla at 100mg/kg, but only a limited increase in simple tubular hyperplasia was observed in the kidney of males at 100mg/kg. Hyperostosis occurred and was accompanied by substantial effects on calcium metabolism, including increased urinary calcium excretion and decreased levels of calcium regulating hormones (1,25-dihydroxyvitamin D and parathyroid hormone). A separate study with radiolabeled calcium confirmed that increased urinary calcium excretion was mediated via increased calcium absorption from the gastrointestinal tract. It was hypothesized that, at high doses, canagliflozin might have inhibited glucose absorption in the intestine via SGLT1 inhibition that resulted in glucose malabsorption, which increased calcium absorption by stimulating colonic glucose fermentation and reducing intestinal pH. Pheochromocytomas and adrenal medullary hyperplasia were attributed to altered calcium homeostasis, which have a known relationship in the rat. In conclusion, Leydig cell tumors were associated with increased luteinizing hormone levels and pheochromocytomas were most likely related to glucose malabsorption and altered calcium homeostasis. Renal tubular tumors may also have been linked to glucose malabsorption.

Nonclinical safety of the sodium-glucose cotransporter 2 inhibitor empagliflozin.

Empagliflozin, a selective inhibitor of the renal tubular sodium-glucose cotransporter 2, was developed for treatment of type 2 diabetes mellitus. Nonclinical safety of empagliflozin was studied in a battery of tests to support global market authorization. Safety pharmacology studies indicated no effect of empagliflozin on measures of respiratory or central nervous system function in rats or cardiovascular safety in telemeterized dogs. In CD-1 mouse, Wistar Han rat, or beagle dogs up to 13, 26, or 52 weeks of treatment, respectively, empagliflozin exhibited a toxicity profile consistent with secondary supratherapeutic pharmacology related to glucose loss and included decreased body weight and body fat, increased food consumption, diarrhea, dehydration, decreased serum glucose and increases in other serum parameters reflective of increased protein catabolism, gluconeogenesis, and electrolyte imbalances, and urinary changes such as polyuria and glucosuria. Microscopic changes were consistently observed in kidney and included tubular nephropathy and interstitial nephritis (dog), renal mineralization (rat) and tubular epithelial cell karyomegaly, single cell necrosis, cystic hyperplasia, and hypertrophy (mouse). Empagliflozin was not genotoxic. Empagliflozin was not carcinogenic in female mice or female rats. Renal adenoma and carcinoma were induced in male mice only at exposures 45 times the maximum clinical dose. These tumors were associated with a spectrum of nonneoplastic changes suggestive of a nongenotoxic, cytotoxic, and cellular proliferation-driven mechanism. In male rats, testicular interstitial cell tumors and hemangiomas of the mesenteric lymph node were observed; both tumors are common in rats and are unlikely to be relevant to humans. These studies demonstrate the nonclinical safety of empagliflozin.

Impact of Microdose Clinical Trials in the Preclinical Stage

A microdose clinical trial may be useful as a safe early-phase exploratory study using doses as low as 100 μg or less for determination of the disposition of a candidate compound in humans in a short period of time. This may increase confidence in candidate compounds, especially those for which it is difficult to predict disposition based on the results of in vitro or preclinical studies. In this study, we examined microdose trials performed in the preclinical stage for two first-in-class compounds with a new mechanism of action. These compounds showed species difference in first pass metabolism in the digestive tract and liver, causing uncertainty in prediction of disposition in humans. For this reason, first-in-human microdose clinical trials were performed. The results showed that the two compounds had effective blood concentrations after oral administration at a dose of 100 mg qd. Administration of an extremely small dose of one (14)C-labeled compound permitted identification of major metabolites. No toxic metabolites were detected. The preclinical toxic dose was determined based on prediction of blood exposure at the estimated maximum clinical dose. For the other candidate compound, the findings of the microdose trial indicated a high bioavailability after oral administration and low hepatic clearance after intravenous administration. These results suggested only a small risk of a change in disposition in patients with hepatic disorder. The data obtained for the two compounds suggest that microdose clinical trials can be useful for improving the process of candidate selection in the preclinical stage.

Safety of recombinant human platelet-derived growth factor-BB in Augment® Bone Graft

This article discusses nonclinical and clinical data regarding the safety of recombinant human platelet-derived growth factor-BB as a component of the Augment® Bone Graft (Augment). Augment is a bone graft substitute intended to be used as an alternative to autologous bone graft in the fusion of hindfoot and ankle joints. Nonclinical studies included assessment of the pharmacokinetic profile of intravenously administered recombinant human platelet-derived growth factor-BB in rat and dog, effects of intravenous administration of recombinant human platelet-derived growth factor-BB in a reproductive and development toxicity study in rats, and chronic toxicity and carcinogenicity of Augment in a 12-month implantation model. These studies showed that systemic exposure was brief and clearance was rapid. No signs of toxicity, carcinogenicity, or tumor promotion were observed even with doses far exceeding the maximum clinical dose. Results of clinical trials (605 participants) and commercial use of recombinant human platelet-derived growth factor-BB containing products indicate that these products are not associated with increased incidence of adverse events or cancer. The safety data presented provide evidence that recombinant human platelet-derived growth factor-BB is a safe therapeutic when used in combination products as a single administration during surgical procedures for bone repair and fusion. There is no evidence associating use of recombinant human platelet-derived growth factor-BB in Augment with chronic toxicity, carcinogenicity, or tumor promotion.

The impact of dose of the angiotensin-receptor blocker valsartan on the post-myocardial infarction ventricular remodeling: study protocol for a randomized controlled trial.

BackgroundAngiotensin-converting enzyme inhibitors and the angiotensin-receptor blocker valsartan ameliorate ventricular remodeling after myocardial infarction (MI). Based on previous clinical trials, a maximum clinical dose is recommended in practical guidelines. Yet, has not been clearly demonstrated whether the recommended dose is more efficacious compared to the lower dose that is commonly used in clinical practice.Method/DesignValsartan in post-MI remodeling (VALID) is a randomized, open-label, single-blinded multicenter study designed to compare the efficacy of different clinical dose of valsartan on the post-MI ventricular remodeling. This study also aims to assess neurohormone change and clinical parameters of patients during the post-infarct period. A total of 1116 patients with left ventricular dysfunction following the first episode of acute ST-elevation MI are to be enrolled and randomized to a maximal tolerable dose (up to 320 mg/day) or usual dose (80 mg/day) of valsartan for 12 months in 2:1 ratio. Echocardiographic analysis for quantifying post-MI ventricular remodeling is to be conducted in central core laboratory. Clinical assessment and laboratory test are performed at fixed times.DiscussionVALID is a multicenter collaborative study to evaluate the impact of dose of valsartan on the post-MI ventricular remodeling. The results of the study provide information about optimal dosing of the drug in the management of patients after MI. The results will be available by 2012.Trial registrationNCT01340326

Evaluation of the potential for pharmacokinetic and pharmacodynamic interactions between dutogliptin, a novel DPP4 inhibitor, and metformin, in type 2 diabetic patients

Objective:Dutogliptin is a novel, orally available, potent, and selective DPP4 inhibitor that improves glycemic control in type 2 diabetic patients. The objective of this study was to evaluate the potential pharmacokinetic and pharmacodynamic interactions, as well as the tolerability, of dutogliptin and metformin alone and in combination in type 2 diabetic patients.Methods:This was a single-center, randomized, open-label, 3-way, crossover study in type 2 diabetic patients. All patients received three treatment regimens, each of 5 days duration in order to reach steady state: 400 mg once daily of dutogliptin (the anticipated clinical dose); 1000 mg metformin twice daily (maximum effective clinical dose); and concomitant administration of 400 mg dutogliptin once daily and 1000 mg metformin twice daily.Results:Co-administration of dutogliptin and metformin did not alter the pharmacokinetics of either agent. The geometric mean ratio, GMR (dutogliptin + metformin /dutogliptin) of the area under the plasma concentration-time curve (AUC0–24h) at steady state was 0.91 (90% CI: 0.79–1.06; p = 0.29); the GMR of the maximum plasma concentrations (Cmax) was 0.95 (90% CI: 0.76–1.19; p = 0.70); the time to maximum plasma concentrations (Tmax) was essentially the same for dutogliptin with or without metformin. The GMR (dutogliptin + metformin/metformin) of AUC0–12h at steady state was 0.99 (90% CI: 0.84–1.17; p = 0.93); the GMR of Cmax was 0.91 (90% CI: 0.79–1.04; p = 0.18); Tmax was comparable for metformin with or without dutogliptin. Metformin added to dutogliptin had no effect on plasma DPP4 inhibition. All three treatment regimens were well tolerated.Conclusions:In this small, multiple dose study, the steady state pharmacokinetics of either dutogliptin or metformin were not altered by co-administration of the two agents. Dutogliptin and metformin were well tolerated either alone or in combination and co-administered metformin did not alter the ex vivo DPP4 inhibition by dutogliptin. There is no need to consider pharmacokinetic and pharmacodynamic interactions when determining the dosage of either agent for co-administration. A phase 3 clinical trial is underway to provide more definitive data on the safety and efficacy of dutogliptin administered on a background of metformin treatment.