A Systematic Approach to Revision Total Knee Arthroplasty.

With the recent increase in primary total knee arthroplasties and the associated rise in failures of the index operation, there has been growing demand for orthopaedic surgeons to perform revision procedures. The orthopaedic surgeon performing revision total knee arthroplasty should be knowledgeable about the various etiologies of primary total knee arthroplasty failure, the steps for proper patient evaluation, and important factors in the preoperative planning process. A systematic methodology for obtaining surgical exposure, strategies for reconstruction, fundamentals of soft-tissue closure, and postoperative care also should be reviewed.

Developing Prediction Models Using Near-Infrared Spectroscopy to Quantify Cannabinoid Content in Cannabis Sativa.

Cannabis is commercially cultivated for both therapeutic and recreational purposes in a growing number of jurisdictions. The main cannabinoids of interest are cannabidiol (CBD) and delta-9 tetrahydrocannabidiol (THC), which have applications in different therapeutic treatments. The rapid, nondestructive determination of cannabinoid levels has been achieved using near-infrared (NIR) spectroscopy coupled to high-quality compound reference data provided by liquid chromatography. However, most of the literature describes prediction models for the decarboxylated cannabinoids, e.g., THC and CBD, rather than naturally occurring analogues, tetrahydrocannabidiolic acid (THCA) and cannabidiolic acid (CBDA). The accurate prediction of these acidic cannabinoids has important implications for quality control for cultivators, manufacturers and regulatory bodies. Using high-quality liquid chromatography-mass spectroscopy (LCMS) data and NIR spectra data, we developed statistical models including principal component analysis (PCA) for data quality control, partial least squares regression (PLS-R) models to predict cannabinoid concentrations for 14 different cannabinoids and partial least squares discriminant analysis (PLS-DA) models to characterise cannabis samples into high-CBDA, high-THCA and even-ratio classes. This analysis employed two spectrometers, a scientific grade benchtop instrument (Bruker MPA II-Multi-Purpose FT-NIR Analyzer) and a handheld instrument (VIAVI MicroNIR Onsite-W). While the models from the benchtop instrument were generally more robust (99.4-100% accuracy prediction), the handheld device also performed well (83.1-100% accuracy prediction) with the added benefits of portability and speed. In addition, two cannabis inflorescence preparation methods were evaluated: finely ground and coarsely ground. The models generated from coarsely ground cannabis provided comparable predictions to that of the finely ground but represent significant timesaving in terms of sample preparation. This study demonstrates that a portable NIR handheld device paired with LCMS quantitative data can provide accurate cannabinoid predictions and potentially be of use for the rapid, high-throughput, nondestructive screening of cannabis material.

High-Throughput Quantitation of Cannabinoids by Liquid Chromatography Triple-Quadrupole Mass Spectrometry.

The high-throughput quantitation of cannabinoids is important for the cannabis industry. As medicinal products increase, and research into compounds that have pharmacological benefits increase, and the need to quantitate more than just the main cannabinoids becomes more important. This study aims to provide a rapid, high-throughput method for cannabinoid quantitation using a liquid chromatography triple-quadrupole mass spectrometer (LC-QQQ-MS) with an ultraviolet diode array detector (UV-DAD) for 16 cannabinoids: CBDVA, CBDV, CBDA, CBGA, CBG, CBD, THCV, THCVA, CBN, CBNA, THC, Δ8-THC, CBL, CBC, THCA-A and CBCA. Linearity, limit of detection (LOD), limit of quantitation (LOQ), accuracy, precision, recovery and matrix effect were all evaluated. The validated method was used to determine the cannabinoid concentration of four different Cannabis sativa strains and a low THC strain, all of which have different cannabinoid profiles. All cannabinoids eluted within five minutes with a total analysis time of eight minutes, including column re-equilibration. This was twice as fast as published LC-QQQ-MS methods mentioned in the literature, whilst also covering a wide range of cannabinoid compounds.

Absorption, Metabolism, and Excretion, In Vitro Pharmacology, and Clinical Pharmacokinetics of Ozanimod, a Novel Sphingosine 1-Phosphate Receptor Modulator.

Ozanimod is approved for the treatment of relapsing forms of multiple sclerosis. Absorption, metabolism, and excretion of ozanimod were investigated after a single oral dose of 1.0 mg [14C]ozanimod hydrochloride to six healthy subjects. In vitro experiments were conducted to understand the metabolic pathways and enzymes involved in the metabolism of ozanimod and its active metabolites. The total mean recovery of the administered radioactivity was ∼63%, with ∼26% and ∼37% recovered from urine and feces, respectively. Based on exposure, the major circulating components were active metabolite CC112273 and inactive metabolite RP101124, which together accounted for 50% of the circulating total radioactivity exposure, whereas ozanimod accounted for 6.7% of the total radioactive exposure. Ozanimod was extensively metabolized, with 14 metabolites identified, including two major active metabolites (CC112273 and CC1084037) and one major inactive metabolite (RP101124) in circulation. Ozanimod is metabolized by three primary pathways, including aldehyde dehydrogenase and alcohol dehydrogenase, cytochrome P450 isoforms 3A4 and 1A1, and reductive metabolism by gut microflora. The primary metabolite RP101075 is further metabolized to form major active metabolite CC112273 by monoamine oxidase B, which further undergoes reduction by carbonyl reductases to form CC1084037 or CYP2C8-mediated oxidation to form RP101509. CC1084037 is oxidized rapidly to form CC112273 by aldo-keto reductase 1C1/1C2 and/or 3ß- and 11ß-hydroxysteroid dehydrogenase, and this reversible oxidoreduction between two active metabolites favors CC112273. The ozanimod example illustrates the need for conducting timely radiolabeled human absorption, distribution, metabolism, and excretion studies for characterization of disproportionate metabolites and assessment of exposure coverage during drug development. SIGNIFICANCE STATEMENT: Absorption, metabolism, and excretion of ozanimod were characterized in humans, and the enzymes involved in complex metabolism were elucidated. Disproportionate metabolites were identified, and the activity of these metabolites was determined.

Timing and Risk Factors for Venous Thromboembolism After Rotator Cuff Repair in the 30-Day Perioperative Period.

PURPOSE: To analyze the American College of Surgeons National Surgical Quality Improvement Program database to evaluate the incidence of deep venous thrombosis and pulmonary embolism in patients undergoing rotator cuff repair surgery. In addition, we aim to identify risk factors associated with the development of thromboembolic events following rotator cuff repair. METHODS: A retrospective review of the American College of Surgeons National Surgical Quality Improvement Program database was performed. Current Procedural Terminology codes were used to identify patients who underwent rotator cuff repair between 2005 and 2017. The presence of deep venous thrombosis or pulmonary embolism during the 30-day perioperative period were the primary outcomes assessed. Logistic regression analysis was performed to identify risk factors for postoperative venous thromboembolic events (VTEs). RESULTS: In total, 39,825 rotator cuff repairs (RCRs) were performed and 117 (0.3%) VTE events occurred. VTE was identified at a mean of 11.5 ± 7.4 days. A total of 31,615 RCRs were performed arthroscopically. There was no significant difference of VTE between groups comparing arthroscopic RCR VTE 0.3% (94) with open RCR 0.3% (23) (P = .81). RCR in patients with an American Society of Anesthesiologists classification of III or IV was associated with >1.5-fold increase risk of VTE (odds ratio [OR] 1.68, 95% confidence interval [CI] 1.14-2.45). Increased risks of VTE included surgery >80 minutes (OR 2.10, 95% CI 1.42-3.15), performed under general anesthesia (OR 4.38, 95% CI 1.18-36.6), and in the outpatient setting (OR 6.09, 95% CI 1.06-243.7), male sex (OR 1.53, 95% CI 1.01-2.33), bleeding disorders (OR 2.87, 95% CI 1.17-7.05), or dyspnea (OR 1.51, 95% CI 1.02-2.23). The biggest risk for VTE was unplanned reoperation OR 16.6 (95% CI 5.13-53.5). CONCLUSIONS: Venous thromboembolism is a rare complication following rotator cuff repair 0.3%. Understanding the risk factors: duration of surgery >80 minutes, male sex, body mass index >30 kg/m2, ASA III or IV, RCR as an inpatient under general anesthesia, bleeding disorder, or dyspnea may be useful in guiding treatment to prevent VTE. The largest risk for VTE is a patient with unplanned reoperation. RCR surgery performed in an outpatient setting resulted in a significantly lower incidence of VTE. LEVEL OF EVIDENCE: III Retrospective Comparative Study.

Therapeutic protein-drug interaction assessment for daclizumab high-yield process in patients with multiple sclerosis using a cocktail approach.

AIMS: To characterize the potential effect of daclizumab high-yield process (DAC HYP), a monoclonal antibody that blocks the high-affinity interleukin-2 receptors for treatment of multiple sclerosis, on activity of cytochrome P450 (CYP) enzymes. METHODS: Twenty patients with multiple sclerosis received an oral cocktail of probe substrates of CYP1A2 (caffeine 200 mg), CYP2C9 (warfarin 10 mg/vitamin K 10 mg), CYP2C19 (omeprazole 40 mg), CYP2D6 (dextromethorphan 30 mg) and CYP3A (midazolam 5 mg) on two sequential occasions: 7 days before and 7 days after subcutaneous administration of DAC HYP 150 mg every 4 weeks for three doses. Serial pharmacokinetic blood samples up to 96 h post dose and 12-h urine samples were collected on both occasions. Area under the curve (AUC) for caffeine, S-warfarin, omeprazole and midazolam, and urine dextromethorphan to dextrorphan ratio were calculated. Statistical analyses were conducted on log-transformed parameters using a linear mixed-effects model. RESULTS: The 90% confidence intervals (CIs) for the geometric mean ratio (probe substrate with DAC HYP/probe substrate alone) for caffeine AUC from 0-12 h (0.93-1.15), S-warfarin AUC from 0 to infinity (AUC[0-inf]) (0.95-1.06), omeprazole AUC(0-inf) (0.88-1.13) and midazolam AUC(0-inf) (0.89-1.15) were within the no-effect boundary of 0.80-1.25. The geometric mean ratio for urine dextromethorphan to dextrorphan ratio was 1.01, with the 90% CI (0.76-1.34) extending slightly outside the no-effect boundary, likely due to high variability with urine collections and CYP2D6 activity. CONCLUSIONS: DAC HYP treatment in patients with multiple sclerosis had no effect on CYP 1A2, 2C9, 2C19, 2D6 and 3A activity.

Population PK-PD analyses of CD25 occupancy, CD56bright NK cell expansion, and regulatory T cell reduction by daclizumab HYP in subjects with multiple sclerosis.

AIM: Daclizumab high yield process (HYP) is a humanized IgG1 monoclonal antibody that binds to the α-subunit of the interleukin-2 receptor and is being developed for treatment of multiple sclerosis (MS). This manuscript characterized the pharmacokinetic-pharmacodynamic (PK-PD) relationships of daclizumab HYP in subjects with MS. METHODS: Approximately 1400 subjects and 7000 PD measurements for each of three biomarkers [CD25 occupancy, CD56bright natural killer (NK) cell count, regulatory T cell (Treg) count] from four clinical trials were analyzed using non-linear mixed effects modelling. Evaluated regimens included 150 or 300 mg subcutaneous (s.c.) every 4 weeks. RESULTS: CD25 occupancy was characterized using a sigmoidal maximum response (Emax ) model. Upon daclizumab HYP treatment, CD25 saturation was rapid with complete saturation occurring after approximately 7 h and maintained when daclizumab HYP serum concentration was ≥5 mg l-1 . After the last 150 mg s.c. dose, unoccupied CD25 returned to baseline levels in approximately 24 weeks, with daclizumab HYP serum concentration approximately ≤1 mgl-1 1L. CD56bright NK cell expansion was characterized using an indirect response model. Following daclizumab HYP 150 mg s.c. every 4 weeks, expansion plateaus approximately at week 36, at which the average maximum expansion ratio is 5.2. After the last dose, CD56bright NK cells gradually declined to baseline levels within 24 weeks. Treg reduction was characterized by a sigmoidal Emax model. Average maximum reduction of 60% occurred approximately 4 days post 150 mg s.c. dose. After the last dose, Tregs were projected to return to baseline levels in approximately 20 weeks. CONCLUSIONS: Robust PK-PD models of CD25 occupancy, CD56bright NK cell expansion and Treg reduction by daclizumab HYP were developed to characterize its key pharmacodynamic effects in the target patient population.

Efficacy and safety of danoprevir-ritonavir plus peginterferon alfa-2a-ribavirin in hepatitis C virus genotype 1 prior null responders.

Danoprevir (DNV) is a hepatitis C virus (HCV) protease inhibitor that achieves high sustained virologic response (SVR) rates in combination with peginterferon alfa-2a-ribavirin in treatment-naive HCV genotype 1 (G1)-infected patients. This study explored the efficacy and safety of ritonavir-boosted DNV (DNVr) plus peginterferon alfa-2a-ribavirin in G1-infected prior peginterferon-ribavirin null responders. Null responders (<2-log10 reduction in HCV RNA level at week 12) were given an open-label combination of 100 mg of ritonavir and 100 mg of DNV (100/100 mg DNVr) every 12 h (q12h) plus peginterferon alfa-2a-ribavirin for 12 weeks. All patients achieving an early virologic response (EVR; ≥2-log10 decrease in HCV RNA by week 12) continued treatment with peginterferon alfa-2a-ribavirin; those without an EVR discontinued all study drugs. Twenty-four prior null responders were enrolled; 16 patients (67%) were infected with HCV G1b, and 8 (33%) were infected with G1a. Ninety-six percent of patients had an IL28B non-CC genotype. A sustained virologic response at 24 weeks posttreatment (SVR24) was achieved in 67% of patients, with a higher rate in G1b-infected (88%) than G1a-infected (25%) patients. Resistance-related breakthrough occurred in 4/8 G1a and 1/16 G1b patients through the DNV resistance-associated variant (RAV) NS3 R155K. NS3 R155K was also detected in 2/2 G1a patients who relapsed. Treatment was well tolerated. Two patients withdrew prematurely from study medications due to adverse events. Two serious adverse events were reported; both occurred after completion of DNVr therapy and were considered unrelated to treatment. No grade 3 or 4 alanine aminotransferase (ALT) elevations were observed. DNVr plus peginterferon alfa-2a-ribavirin demonstrated high SVR24 rates in HCV G1b-infected prior null responders and was well tolerated. (This study has been registered at ClinicalTrials.gov under registration no. NCT01185860.).

Two-way interaction study between ritonavir boosted danoprevir, a potent HCV protease inhibitor, and ketoconazole in healthy subjects.

BACKGROUND: Danoprevir is a potent, highly selective, macrocyclic, orally bioavailable inhibitor of the hepatitis C virus protease, and a substrate of cytochrome P450 (CYP) 3A. It is co-administered with low-dose ritonavir, a potent CYP3A inhibitor, to enhance danoprevir pharmacokinetics. Ketoconazole is a substrate for and potent selective inhibitor of CYP3A. METHODS: In this open-label, 3-period study, the 2-way interaction potential between ritonavir-boosted danoprevir (danoprevir/r) and ketoconazole was investigated in 18 healthy subjects. Subjects initially received ketoconazole 200 mg q24h for 4 days (Period 1) followed by a 7-day washout period. Danoprevir/r 100/100 mg q12h was then given for 10 days (Period 2) followed by a further 4 days of danoprevir/r 100/100 mg q12h plus ketoconazole 200 mg q24h (Period 3). Serial blood samples were collected for the determination of danoprevir, ritonavir and/or ketoconazole plasma concentrations, and calculation of pharmacokinetic parameters. Safety and tolerability were monitored throughout the study. RESULTS: Co-administration of ketoconazole with danoprevir/r modestly increased the danoprevir AUCτ by 1.44-fold, with no effect on danoprevir Cmax. Co-administration of danoprevir/r with ketoconazole substantially increased ketoconazole AUCτ and Cmax by 3.71-fold and 1.73-fold, respectively. Danoprevir/r was well tolerated when administered alone or with ketoconazole. CONCLUSIONS: These results indicate that the effect of potent CYP3A inhibitors, such as ketoconazole, on danoprevir/r pharmacokinetics is not likely to be clinically relevant.

Concentration-QTc modeling of sitravatinib in patients with advanced solid malignancies.

Sitravatinib (MGCD516) is an orally available, small molecule, tyrosine kinase inhibitor that has been evaluated in patients with advanced solid tumors. Concentration-corrected QT interval (QTc; C-QTc) modeling was undertaken, using 767 matched concentration-ECG observations from 187 patients across two clinical studies in patients with advanced solid malignancies, across a dose range of 10-200 mg, via a linear mixed-effects (LME) model. The effect on heart rate (HR)-corrected QT interval via Fridericia's correction method (QTcF) at the steady-state maximum concentration (Cmax,ss) for the sitravatinib proposed therapeutic dosing regimen (100 mg malate once daily [q.d.]) without and with relevant intrinsic and extrinsic factors were predicted. No significant changes in HR from baseline were observed. Hysteresis between sitravatinib plasma concentration and change in QTcF from baseline (ΔQTcF) was not observed. There was no significant relationship between sitravatinib plasma concentration and ΔQTcF. The final C-QTc model predicted a mean (90% confidence interval [CI]) ΔQTcF of 3.92 (1.95-5.89) ms and 2.94 (0.23-6.10) ms at the proposed therapeutic dosing regimen in patients with normal organ function (best case scenario) and patients with hepatic impairment (worst-case scenario), respectively. The upper bounds of the 90% CIs were below the regulatory threshold of concern of 10 ms. The results of the described C-QTc analysis, along with corroborating results from nonclinical safety pharmacology studies, indicate that sitravatinib has a low risk of QTc interval prolongation at the proposed therapeutic dose of 100 mg malate q.d.

A randomised study of the effect of danoprevir/ritonavir or ritonavir on substrates of cytochrome P450 (CYP) 3A and 2C9 in chronic hepatitis C patients using a drug cocktail.

PURPOSE: The aim of this study was to evaluate the effects of danoprevir in combination with low-dose ritonavir (danoprevir/r) and placebo plus low-dose ritonavir on the pharmacokinetics of probe drugs for cytochrome P450 (CYP) 3A and CYP2C9, in patients with chronic hepatitis C. METHODS: A total of 54 patients infected with hepatitis C virus genotype 1 received an oral drug cocktail (2 mg midazolam, 10 mg warfarin and 10 mg vitamin K) before and after 14 days of dosing with either danoprevir/r or placebo plus low-dose ritonavir (placebo/r). Serial pharmacokinetic samples were collected up to 24 (midazolam) and 72 (S-warfarin) h post-dose. Plasma concentrations of midazolam, α-hydroxymidazolam and S-warfarin were measured using validated assays. Pharmacokinetic parameters were estimated using non-compartmental analysis, and geometric mean ratios (GMRs) and 90 % confidence intervals (CIs) for the differences between baseline and post-dosing values were calculated. RESULTS: Danoprevir/r and placebo/r significantly increased midazolam area under the time-concentration curve (AUC0-∞) and reduced the midazolam metabolic ratio while S-warfarin AUC0-∞ was modestly decreased. When danoprevir data were pooled across doses, the midazolam GMR (90 % CI) AUC0-∞ was 9.41 (8.11, 10.9) and 11.14 (9.42, 13.2) following danoprevir/r and placebo/r dosing, respectively, and the S-warfarin GMR (90 % CI) AUC0-∞ was 0.72 (0.68, 0.76) and 0.76 (0.69, 0.85), respectively. The effects of danoprevir/r and placebo/r appeared to be qualitatively similar. CONCLUSIONS: Substantial inhibition of CYP3A- and modest induction of CYP2C9- activity were observed with danoprevir/r and low-dose ritonavir.

Assessing Technology Platforms for Global Health Engagement to Support Integration of Efforts Across Geographic Combatant Commands.

Global health engagement (GHE) is an integral part of the cooperation efforts of the U.S. Department of Defense (DoD) and the geographic combatant commands (GCCs) with partner nations and provides support in training and preparing their military and civilian health systems. These activities encompass a wide spectrum of engagements-military-to-military, military-to-civilian, and multilateral-and support joint missions of humanitarian aid and disaster response, deterrence, access and presence, counterterrorism, and homeland defense. Global health engagements and activities require extensive planning, funding, and resource allocation within the GCCs and component commands. For a continuously growing breadth of GHE and the need to support joint exercises with partner military and civilian medical professionals for partner capacity-building, GHE also requires a robust information technology infrastructure. In this study, the authors assess the technology and process requirements to support the life cycle of GHE activities and assessments-from planning to evaluation-and the information- and knowledge-sharing needs of the GHE community. To do so, they conducted a literature review related to GHE activities, funding sources, and stakeholders; the evolution of technology solutions to support GHE; systems in use by GHE practitioners; and technology solutions in the market, focusing particularly on cloud infrastructure and services and cloud service providers. They held discussions with GHE subject-matter experts to document and analyze GHE technology platform requirements. And they assessed the available and planned platforms according to their features, enhancements, support and maintenance, data integration, interoperability, and future road maps.

Systematic Exposure in Revision Total Hip Arthroplasty: The Posterior Approach.

As indications for total hip arthroplasty (THA) continue to expand, and patients continue to live longer with more active lifestyles, the incidence of revision THA is expected to rise. General orthopaedic surgeons are now beginning to consider doing revision THA surgery because of the increased revision burden being experienced nationwide. While classical approaches to the hip can be used for simple revisions, extensile exposure techniques in conjunction with selective soft-tissue releases are often required for adequate visualization for more complex revision cases. This review provides a systematic approach to surgical exposure for revision THA using the posterior approach. The surgeon should follow a stepwise progression to obtain safe, adequate, and reproducible visualization of both the acetabulum and the proximal femur.

The Cannabis Plant as a Complex System: Interrelationships between Cannabinoid Compositions, Morphological, Physiological and Phenological Traits.

Maintaining specific and reproducible cannabinoid compositions (type and quantity) is essential for the production of cannabis-based remedies that are therapeutically effective. The current study investigates factors that determine the plant's cannabinoid profile and examines interrelationships between plant features (growth rate, phenology and biomass), inflorescence morphology (size, shape and distribution) and cannabinoid content. An examination of differences in cannabinoid profile within genotypes revealed that across the cultivation facility, cannabinoids' qualitative traits (ratios between cannabinoid quantities) remain fairly stable, while quantitative traits (the absolute amount of Δ9-tetrahydrocannabinol (THC), cannabidiol (CBD), cannabichromene (CBC), cannabigerol (CBG), Δ9-tetrahydrocannabivarin (THCV) and cannabidivarin (CBDV)) can significantly vary. The calculated broad-sense heritability values imply that cannabinoid composition will have a strong response to selection in comparison to the morphological and phenological traits of the plant and its inflorescences. Moreover, it is proposed that selection in favour of a vigorous growth rate, high-stature plants and wide inflorescences is expected to increase overall cannabinoid production. Finally, a range of physiological and phenological features was utilised for generating a successful model for the prediction of cannabinoid production. The holistic approach presented in the current study provides a better understanding of the interaction between the key features of the cannabis plant and facilitates the production of advanced plant-based medicinal substances.

The Workplace and Psychosocial Experiences of Australian Senior Doctors during the COVID-19 Pandemic: A Qualitative Study.

The coronavirus disease 2019 (COVID-19) pandemic has had significant mental health impacts among healthcare workers (HCWs), related to resource scarcity, risky work environments, and poor supports. Understanding the unique challenges experienced by senior doctors and identifying strategies for support will assist doctors facing such crises into the future. A cross-sectional, national, online survey was conducted during the second wave of the Australian COVID-19 pandemic. Inductive content analysis was used to examine data reporting workplace and psychosocial impacts of the pandemic. Of 9518 responses, 1083 senior doctors responded to one or more free-text questions. Of the senior doctors, 752 were women and 973 resided in Victoria. Four themes were identified: (1) work-life challenges; (2) poor workplace safety, support, and culture; (3) poor political leadership, planning and support; and (4) media and community responses. Key issues impacting mental health included supporting staff wellbeing, moral injury related to poorer quality patient care, feeling unheard and undervalued within the workplace, and pandemic ill-preparedness. Senior doctors desired better crisis preparedness, HCW representation, greater leadership, and accessible, authentic psychological wellbeing support services from workplace organisations and government. The pandemic has had significant impacts on senior doctors. The sustainability of the healthcare system requires interventions designed to protect workforce wellbeing.

Antiviral activity, safety, and pharmacokinetics of danoprevir/ritonavir plus PEG-IFN α-2a/RBV in hepatitis C patients.

BACKGROUND & AIMS: Danoprevir (RG7227; ITMN-191) is a potent inhibitor of the HCV NS3/4A serine protease. The aims of this double-blind, placebo-controlled, multiple-ascending dose phase Ib study were to evaluate safety, tolerability, antiviral activity, resistance, and pharmacokinetics of once- and twice-daily danoprevir in the presence of low-dose ritonavir (danoprevir/r) and in combination with peginterferon alfa-2a (40KD)/ribavirin in treatment-naive HCV genotype 1 patients. METHODS: Thirty eligible patients were enrolled into three cohorts and treated with danoprevir/r or placebo/r all in combination with peginterferon alfa-2a (40KD)/ribavirin for 15 days. Cohort 1 received danoprevir/r at 100/100mg twice daily; Cohort 2 200/100mg once daily; and Cohort 3 200/100mg twice daily. RESULTS: The median reductions in HCV RNA from baseline after 14 days of treatment (day 15) were -5.1, -4.8, and -4.6 log(10)IU/ml in Cohorts 1, 2, and 3, respectively, and -2.7 log(10) in placebo/r and peginterferon alfa-2a (40KD)/ribavirin recipients. Viral breakthrough was not observed in any patient. On day 15, HCV RNA was undetectable (<15IU/ml) in 6/9 (67%), 4/8 (50%), and 8/8 (100%) patients in Cohorts 1, 2, and 3, respectively. When co-administered with low dose ritonavir, danoprevir concentrations reached a steady state between 6 to 10 days of dosing. Danoprevir exposures increased more than dose proportionally between 100/100mg and 200/100mg. Danoprevir/r plus peginterferon alfa-2a (40KD)/ribavirin was well-tolerated with no safety-related discontinuations. CONCLUSIONS: Danoprevir/r plus peginterferon alfa-2a (40KD)/ribavirin provides profound and robust reductions in serum HCV RNA, at substantially lower systemic exposures compared to those observed with higher doses of danoprevir in the absence of ritonavir. These results support further studies of danoprevir/r.

Structural-based analysis of dihydrofolate reductase evolution.

The evolution of dihydrofolate reductase (DHFR) was studied through a comprehensive structural-based analysis. An amino acid sequence alignment was generated from a superposition of experimentally determined X-ray crystal structures of wild-type (wt) DHFR from the Protein Data Bank (PDB). Using this structure-based alignment of DHFR, a metric was generated for the degree of conservation at each alignment site - not only in terms of amino acid residue, but also secondary structure, and residue class. A phylogenetic tree was generated using the alignment that compared favorably with the canonical phylogeny. This structure-based alignment was used to confirm that the degree of conservation of active-site residues in terms of both sequence as well as structure was significantly greater than non-active site residues. These results can be used in helping to understand the likely future evolution of DHFR in response to novel therapies.

Physiological modeling and assessments of regional drug bioavailability of danoprevir to determine whether a controlled release formulation is feasible.

Danoprevir, a potent, selective inhibitor of HCV NS3/4A protease, has a short half-life in humans. Therefore, the feasibility of a controlled release (CR) formulation to allow less frequent dosing was investigated using experimental approaches and physiological modeling to examine whether danoprevir is absorbed in the colon. Danoprevir absorption was studied in portal-vein-cannulated monkeys and in monkeys surgically modified to make intraduodenal, intrajejunal, intracolonic and oral administration possible. In portal-vein-cannulated monkeys, absorption was apparent up to 24 h after administration. The observed relative bioavailability from intracolonic delivery in the monkey was approximately 30% relative to oral administration, consistent with the model prediction of 40%. Human relative bioavailability for a tablet delivered to the colon compared with an immediate release (IR) formulation was predicted to be 4-28%. Preclinical data and modeling suggested that CR development would be challenging for this Biopharmaceutics Classification System Class IV compound. Therefore, a confirmative study in healthy volunteers was conducted to investigate the relative bioavailability of danoprevir in various regions of the gastrointestinal tract. In a randomized, open-label, crossover study, subjects received 100 mg danoprevir IR soft gel capsule, 100 mg danoprevir solution delivered to the distal small bowel and colon via an Enterion™ capsule (a remotely activated capsule for regional drug delivery) and 100 mg danoprevir powder to the colon via an Enterion™ capsule. The relative bioavailability of danoprevir (compared with IR) delivered to the colon was 6.5% for a solution and 0.6% for a powder formulation, indicating that a CR formulation is not feasible.

Concentration-QTc Modeling of Ozanimod's Major Active Metabolites in Adult Healthy Subjects.

Ozanimod, approved by regulatory agencies in multiple countries for the treatment of adults with relapsing multiple sclerosis, is a sphingosine 1-phosphate (S1P) receptor modulator, which binds with high affinity selectively to S1P receptor subtypes 1 and 5. The relationships between plasma concentrations of ozanimod and its major active metabolites, CC112273 and CC1084037, and the QTc interval (C-QTc) from a phase I multiple-dose study in healthy subjects were analyzed using nonlinear mixed effects modeling. QTc was modeled linearly as the sum of a sex-related fixed effect, baseline, and concentration-related random effects that incorporated interindividual and residual variability. Common linear, power, and maximum effect (Emax ) functions were assessed for characterizing the relationship of QTc with concentrations. Model goodness-of-fit and performance were evaluated by standard diagnostic tools, including a visual predictive check. The placebo-corrected change from baseline in QTc (ΔΔQTc) was estimated based on the developed C-QTc model using a nonparametric bootstrapping approach. QTc was better derived using a study-specific population formula (QTcP). Among the investigated functions, an Emax function most adequately described the relationship of QTcP with concentrations. Separate models for individual analytes characterized the C-QTcP relationship better than combined analytes models. Attributing QT prolongation independently to CC1084037 or CC112273, the upper bound of the 95% confidence interval of the predicted ΔΔQTcP was ~ 4 msec at the plateau of the Emax curves. Therefore, ΔΔQTcP is predicted to remain below 10 msec at the supratherapeutic concentrations of the major active metabolites.