The ClC-1 chloride channel inhibitor NMD670 improves skeletal muscle function in rat models and patients with myasthenia gravis.

Myasthenia gravis (MG) is a neuromuscular disease that results in compromised transmission of electrical signals at the neuromuscular junction (NMJ) from motor neurons to skeletal muscle fibers. As a result, patients with MG have reduced skeletal muscle function and present with symptoms of severe muscle weakness and fatigue. ClC-1 is a skeletal muscle specific chloride (Cl-) ion channel that plays important roles in regulating neuromuscular transmission and muscle fiber excitability during intense exercise. Here, we show that partial inhibition of ClC-1 with an orally bioavailable small molecule (NMD670) can restore muscle function in rat models of MG and in patients with MG. In severely affected MG rats, ClC-1 inhibition enhanced neuromuscular transmission, restored muscle function, and improved mobility after both single and prolonged administrations of NMD670. On this basis, NMD670 was progressed through nonclinical safety pharmacology and toxicology studies, leading to approval for testing in clinical studies. After successfully completing phase 1 single ascending dose in healthy volunteers, NMD670 was tested in patients with MG in a randomized, placebo-controlled, single-dose, three-way crossover clinical trial. The clinical trial evaluated safety, pharmacokinetics, and pharmacodynamics of NMD670 in 12 patients with mild MG. NMD670 had a favorable safety profile and led to clinically relevant improvements in the quantitative myasthenia gravis (QMG) total score. This translational study spanning from single muscle fiber recordings to patients provides proof of mechanism for ClC-1 inhibition as a potential therapeutic approach in MG and supports further development of NMD670.

A deep-well plate enabled automated high-throughput cell line development platform.

Cell line development (CLD) plays a crucial role in the manufacturing process development of therapeutic biologics. Most biologics are produced in Chinese hamster ovary (CHO) cell. Because of the nature of random transgene integration in CHO genome and CHO's inherent plasticity, stable CHO transfectants usually have a vast diversity in productivity, growth, and product quality. Thus, we often must resort to screening a large number of cell pools and clones to increase the probability of identifying the ideal production cell line, which is a very laborious and resource-demanding process. Here we have developed a deep-well plate (DWP) enabled high throughput (DEHT) CLD platform using 24-well DWP (24DWP), liquid handler, and other automation components. This platform has capabilities covering the key steps of CLD including cell passaging, clone imaging and expansion, and fed-batch production. We are the first to demonstrate the suitability of 24DWP for CLD by confirming minimal well-to-well and plate-to-plate variability and the absence of well-to-well cross contamination. We also demonstrated that growth, production, and product quality of 24DWP cultures were comparable to those of conventional shake flask cultures. The DEHT platform enables scientists to screen five times more cultures than the conventional CLD platform, thus significantly decreases the resources needed to identify an ideal production cell line for biologics manufacturing.

Modeling immunogenecity data to establish screening bioassays cut point.

The response of immunogenecity anti-drug antibody (ADA) generally includes biological and analytical variability. The nature of biological and analytical variations may lead to a variety of symmetric and asymmetric ADA data. As a result, current statistical methods may yield unreliable results because these methods assume special types of symmetric or asymmetric ADA data. In this paper, we survey and compare parametric models that are useful for analyzing a variety of asymmetric data that have rarely been used to calculate assay cut points. These models include symmetric distributions as limiting case; therefore, they are useful in the analysis of a variety of symmetric data. We also investigate two nonparametric approaches that have received little attention in screening cut point calculations. A simulation study was conducted to compare the performance of the methods. We evaluate the methods using four published different types of data, and make recommendations concerning the use of the methods.

Statistical considerations for assessing precision of heterogeneous duplicate measurements: An application to pharmaceutical bioanalysis.

Duplicate analysis is a strategy commonly used to assess precision of bioanalytical methods. In some cases, duplicate analysis may rely on pooling data generated across organizations. Despite being generated under comparable conditions, organizations may produce duplicate measurements with different precision. Thus, these pooled data consist of a heterogeneous collection of duplicate measurements. Precision estimates are often expressed as relative difference indexes (RDI), such as relative percentage difference (RPD). Empirical evidence indicates that the frequency distribution of RDI values from heterogeneous data exhibits sharper peaks and heavier tails than normal distributions. Therefore, traditional normal-based models may yield faulty or unreliable estimates of precision from heterogeneous duplicate data. In this paper, we survey application of the mixture models that satisfactorily represent the distribution of RDI values from heterogeneous duplicate data. A simulation study was conducted to compare the performance of the different models in providing reliable estimates and inferences of percentile calculated from RDI values. These models are readily accessible to practitioners for study implementation through the use of modern statistical software. The utility of mixture models are explained in detail using a numerical example.

Development, validation, and implementation of a robust and quality control-friendly focused peptide mapping method for monitoring oxidation of co-formulated monoclonal antibodies.

Monoclonal antibody (mAb) coformulation containing two therapeutic proteins provides benefits of improved therapeutic efficacy and better patient compliance. Monitoring of the individual mAb stability in the coformulation is critical to ensure its quality and safety. Among post-translational modifications (PTMs), oxidation is often considered as one of the critical quality attributes (CQAs) as it potentially affects the structure and potency. Although hydrophobic interaction chromatography (HIC) and reversed phase liquid chromatography (RPLC) have been used to monitor overall protein oxidation, mass spectrometry of peptide digests resolved by LC methods can afford superior selectivity and sensitivity for specific PTMs. With the advent of the Quadrupole Dalton (QDa) mass spectrometer as an affordable add-on detector, implementation of targeted oxidation assays in development and quality control (QC) laboratories is now feasible. In this study, as the first effort to implement MS-based methods for antibody coformulation in QC laboratories, we developed and validated a high-throughput and robust focused peptide mapping method using QDa for simultaneous site-specific monitoring of oxidation of methionine and tryptophan residues in heavy-chain (HC) complementary determining regions (CDRs) of two co-formulated mAbs. The method was validated in terms of accuracy, precision, linearity, range, quantitation limit (QL), specificity, and solution stability per recommendations in ICH Q2. The method robustness was systematically assessed involving multiple sample preparation and instrument method parameters. The method met the validation criteria in GMP laboratories with excellent robustness and was implemented in both GMP and development environments.

Retrospective assessment of clonality of a legacy cell line by analytical subcloning of the master cell bank.

In recent years, assurance of clonality of the production cell line has been emphasized by health authorities during review of regulatory submissions. When insufficient assurance of clonality is provided, augmented control strategies may be required for a commercial production process. In this study, we conducted a retrospective assessment of clonality of a legacy cell line through analysis of subclones from the master cell bank (MCB). Twenty-four subclones were randomly selected based on a predetermined acceptance sampling plan. All these subclones share a conserved integration junction, thus providing a high level of assurance that the cell population in the MCB was derived from a single progenitor cell. However, Southern blot analysis indicates that at least four subpopulations possibly exist in the MCB. Additional characterization of these four subpopulations demonstrated that the resulting changes in product quality attributes of some subclones are not related to the genetic heterogeneity observed in Southern blot hybridization. Furthermore, process consistency, process comparability, and analytical comparability have been demonstrated in batches produced across varying manufacturing processes, scales, facilities, cell banks, and cell ages. Finally, process and product consistency together with a high level of assurance of clonal origin of the MCB helped clear the hurdle for regulatory approval without requirement of additional control strategies.

Development and comparison of three cell-based potency assays for anti-respiratory syncytial virus monoclonal antibody.

There is an increasing demand for monoclonal antibody (mAb) therapies to confer passive immunity against viral diseases. Respiratory syncytial virus (RSV) is the most common cause of bronchiolitis, lower respiratory tract infections, and hospitalization in infants. Currently, there is no RSV vaccine but a humanized mAb available for high risk infants. MK-1654 is a fully human mAb with YTE mutation in the fragment crystallizable (Fc) region to extend the half-life in circulation. It binds to a highly conserved epitope of RSV Fusion protein with high affinity and neutralizes RSV infection. A functional cell-based assay is a regulatory requirement for clinical development, commercial release, and stability testing of MK-1654. In this study, we have evaluated three RSV neutralization assays to test the potency of MK-1654, including an imaging-based virus reduction neutralization test (VRNT) and two reporter virus-based assays (RSV-GFP and RSV-NLucP). All three methods showed good dose response curves of MK-1654 with similar EC50 values. RSV-NLucP method was chosen for further development because it is simple and can be easily adapted to quality control testing laboratories. After optimization, the RSV-NLucP assay was pre-qualified with good linearity, relative accuracy, intermediate precision, and specificity, therefore suitable for a cell-based potency assay.

Manejo de la hemorragia digestiva alta en pacientes con infección por SARS-CoV-2 en un hospital de Lima, Perú / Management of upper gastrointestinal bleeding in patients with SARS-COV-2 infection in a hospital in Lima-Peru

Resumen Introducción: la pandemia por SARS-CoV-2 ha conllevado la reestructuración de las unidades de endoscopía digestiva en el mundo, lo cual ha limitado los procedimientos endoscópicos y priorizado indicaciones de emergencia como la hemorragia digestiva alta (HDA). No obstante, existe escasa evidencia respecto a su impacto en la evolución y resultados. Objetivo: evaluar el manejo de la HDA en el contexto de la pandemia del coronavirus por SARS-CoV-2. Materiales y métodos: estudio observacional, descriptivo, retrospectivo de marzo a agosto de 2020 en pacientes con diagnóstico de HDA e infección por SARS-CoV-2. Resultados: de 4320 pacientes con infección por SARS-CoV-2, 51 presentaron HDA al ingreso. La mediana de edad fue de 70 años. El 58,8 % era de sexo masculino. El 56,9 % tenía una puntuación de Glasgow-Blatchford (SGB) ≥12. El 21,6 % requirió soporte de oxígeno. Solo 34 pacientes (66,7 %) recibieron tratamiento médico; asimismo, 17 (33,3 %) recibieron tratamiento médico más endoscopia digestiva alta (EDA); de estos, a 6 (35,3 %) se les realizó endoscopia terapéutica. La enfermedad ulcerosa péptica fue el hallazgo más frecuente. Al comparar el tipo de tratamiento recibido, no hubo diferencias significativas entre el número de transfusiones de glóbulos rojos, resangrado, reingreso por HDA, estancia hospitalaria ni mortalidad secundaria a la HDA. La mortalidad global fue del 25,4 % (13 pacientes) y se debió, principalmente, al compromiso respiratorio por SARS-CoV-2. Conclusiones: se observa una reducción en el número de EDA de emergencia por HDA en la pandemia actual, así como un tiempo mayor al estándar para su realización. Más del 80 % de los pacientes que recibieron solo tratamiento médico evolucionaron favorablemente, y solo un tercio de los pacientes a quienes se les realizó una EDA requirió terapéutica endoscópica.

Abstract Introduction: The SARS-CoV-2 pandemic has led to the restructuring of digestive endoscopy units around the world, limiting endoscopic procedures and prioritizing emergency indications such as upper gastrointestinal hemorrhage (UGH). However, there is little evidence regarding its impact on evolution and outcomes. Objective: To evaluate the management of UGH in the context of the SARS-CoV-2 coronavirus pandemic. Materials and methods: Observational, descriptive, retrospective study carried out between March and August 2020 in patients with diagnosis of UGH and SARS-CoV-2 infection. Results: Of 4 320 patients with SARS-CoV-2 infection, 51 had UGH on admission. The median age of the population was 70 years and 58.8% were male. Glasgow-Blatchford Bleeding Score (GBS) of ≥12 was obtained in 56.9%. Oxygen support was required by 21.6%. 34 (66.7%) patients received medical treatment only, while 17 (33.3%) received medical treatment plus upper gastrointestinal endoscopy (UGE), of which 6 (35.3%) underwent therapeutic endoscopy. Peptic ulcer disease was the most frequent finding. When comparing the type of treatment received, there were no significant differences between the number of red blood cell transfusions, rebleeding, re-admission due to UGH, hospital stay, or mortality secondary to UGH. Overall mortality was 25.4% (13 patients), mainly due to respiratory failure due to SARS-CoV-2. Conclusions: A reduction in the number of emergency upper gastrointestinal endoscopies for UGH was observed during the current pandemic, as well as a longer than standard time for their performance. More than 80% of patients who received medical treatment alone evolved favorably and only one third of the patients who underwent UGE required endoscopic therapy.

The effectiveness of quality control samples in pharmaceutical bioanalysis.

The use of quality control (QC) samples in bioanalysis is well established and consistent with regulatory guidance. However, a systematic evaluation of whether QC samples serve the intended purpose of improving data quality has not been undertaken. The Translational and ADME Sciences Leadership Group (TALG) of the International Consortium for Innovation and Quality in Pharmaceutical Development (IQ) conducted an evaluation to assess whether closer agreement is observed when comparing pharmacokinetic data from two passed runs, than when comparing data from failed and passed (retest) runs. Analysis of data collected across organizations, molecular types and analytical platforms, revealed that bioanalytical methods are very reproducible; and that QC samples improve the overall quality of pharmacokinetic concentration data and justifies their continued use.

Models for Counts and Particle Size Distributions of Subvisible Particle Data.

Traditional statistical analyses of subvisible particle data are usually based on either descriptive statistics, normal-based methods, or standard Poisson models. These methods often do not adequately describe the counts or particle size distribution. They usually ignore relevant information represented in the data, such as count correlation. Therefore, any meaningful analyses of subvisible particle data require a reasonable representation of counts and particle size distribution and the correlation in the data. Such comprehensive approaches are not widely available or used when analyzing subvisible particle data. In this article, we propose the use of generalized linear mixed models to analyze the counts and the particle size distribution of subvisible particle data. These models make optimal use of the information in the data and allow flexible approaches for the analyses of a wide range of data structures. They are readily accessible to practitioners through the use of modern statistical software. These models are demonstrated with two numerical examples using two different data structures.

Recommendations for singlet-based approach in ligand binding assays: an IQ Consortium perspective.

Historically, ligand-binding assays for pharmacokinetic samples employed duplicate rather than singlet-based analysis. Herein, the Translational and absorption, distribution, metabolism and excretion (ADME) Sciences Leadership Group of the International Consortium for Innovation and Quality in Pharmaceutical Development (IQ) presents a study aiming to determine the value of duplicate versus singlet-based testing. Based on analysis of data collected from eight organizations for 20 drug candidates representing seven molecular types and four analytical platforms, statistical comparisons of validation and in-study quality controls and study unknown samples demonstrated good agreement across duplicate sets. Simulation models were also used to assess the impact of sample duplicate characteristics on bioequivalence outcomes. Results show that testing in singlet is acceptable for assays with %CV ≤15% between duplicates. Singlet-based approach is proposed as the default for ligand-binding assays while a duplicate-based approach is needed where imprecision and/or inaccuracy impede the validation of the assay.

Renal Replacement Therapy Preferences Survey: Is Allo-Hemodialysis an Acceptable Option for Patient Caregivers and Health Care Professionals?

End-stage kidney disease (ESKD) is a worldwide unsolved problem. Access to renal replacement therapies (RRT) is still a challenge in some developed countries and even more so in developing countries. Allo-hemodialysis (alloHD) is a recently proposed, still hypothetical, alternative RRT where the blood of a healthy subject ("buddy") flows countercurrent to the patient's blood through the dialyzer. Solutes and fluid are transferred to the buddy and then cleared by his/her healthy kidneys, making alloHD essentially a procedure where the buddy "donates" kidney function intermittently to the patient. Its drastically reduced complexity makes -alloHD particularly attractive for low-resource settings. The acceptance of alloHD by patients, caregivers, and health care professionals (HCP) is unknown. In this cross-sectional study, we surveyed the preferences and acceptance of alloHD in 3 groups: caregivers related to ESKD patients, nonrelated caregivers (nrCG), and HCP. Four areas were explored: RRT preferences, kidney organ donation for transplant acceptance, -alloHD acceptance as a potential RRT, and alloHD technique acceptance. Hemodialysis was the preferred form of RRT. Kidney donation acceptance was similar in all groups. Intermittent kidney function donation (i.e., alloHD) was mainly accepted by related and nrCG but less accepted by HCP (87, 90, and 60% respectively, p < 0.01). New RRT alternatives such as alloHD are expected to be better received and accepted once animal, and clinical studies have demonstrated their feasibility, safety, and benefits. New RRT strategies are required primarily in most vulnerable populations and should be explored.

Use of TEVI-R to describe passive vocabulary in children belonging to a Mapuche community. / Uso del TEVI-R para describir el vocabulario pasivo en niños de la etnia Mapuche.

PURPOSE: To describe the level of passive vocabulary in boys and girls belonging to the Mapuche ethnic group, using the Vocabulary Test in Images Revised version (TEVI-R). METHODS: A cross-sectional study was carried out. Twenty-six children, both gender, between 4 to 7 years old participated in the study. The performance of passive vocabulary was measured through the application of the TEVI-R, analyzing the variables age and gender, as well as performing an analysis of the items and the number of errors. RESULTS: The performance of these children is not influenced by gender or age. There are potential sources of error in the items of the instruments related to the cultural, geographical relevance and graphic quality of them. CONCLUSION: No association was observed by gender or differences by age in the study population. The possibility of developing new instruments or revising the available ones is discussed, given the characteristics of their native language, obtaining reliable results and respecting the elements that are part of their culture.

OBJETIVO: Describir el nivel de vocabulario pasivo en niños y niñas pertenecientes a la etnia mapuche, utilizando el Test de Vocabulario en Imágenes versión Revisada (TEVI-R). MÉTODO: Se llevó a cabo un estudio de corte transversal. Participaron 26 niños, de ambos géneros cuyas edades fluctuaron entre 4 y 7 años de edad. Se midió el desempeño de vocabulario pasivo mediante la aplicación del TEVI-R, analizando las variables edad y género, además de realizar un análisis de los ítems con mayor cantidad de errores. RESULTADOS: El rendimiento de estos niños no se ve influenciado ni por género, ni por edad. Existen potenciales fuentes de error en los ítems de los instrumentos relacionados a la pertinencia cultural, geográfica y a la calidad gráfica de los mismos. CONCLUSIÓN: No se observó asociación por género ni diferencias por edad en la población estudiada. Se discute la posibilidad de elaborar nuevos instrumentos o revisar los disponibles, dadas las características de su lengua materna, con la finalidad de recabar resultados fiables y respetar los elementos que forman parte de los marcadores propios de su cultura.

Validation of an ambient measurement system (AMS) for physical activities in a paediatric population.

Ambient measurement systems (AMSs) can enable continuous assessment of functional performance at home, increasing the availability of data for monitoring of neuromuscular disease. An AMS passively measures movement whenever someone is in range of the sensor, without the need for any wearable sensors. The current study evaluates the performance of an AMS for three metrics associated with functional assessments in Duchenne muscular dystrophy (DMD): ambulation speed, rise-to-stand speed and arm-raise speed. Healthy paediatric subjects performed a series of functional tasks and were graded by both a human rater and an AMS. Linear mixed-effect models were fit to calculate agreement between the two measurement methods. For all activities, the AMS and human rater supplied similar measurements of average speed, with correlation coefficients of 0.76-0.92 and systematic differences ranging in magnitude from 0 to 0.48 m per second. The largest systematic difference was for the 10-m run, which was likely due to human rater reaction time. Systematic differences in arm-raise measurements were due to incomplete execution of movements by test participants. These results are consistent with previous studies comparing automated and manual measurements of movement. This study demonstrates that an AMS device is able to measure ambulation speed, rise-to-stand speed and arm-raise speed in a paediatric population in a controlled setting without the need for complicated installation, calibration or worn sensors.

A comparative study of confidence intervals to assess biosimilarity from analytical data.

Assessment of analytical similarity of tier 1 quality attributes is based on a set of hypotheses that tests the mean difference of reference and test products against a margin adjusted for standard deviation of the reference product. Thus, proper assessment of the biosimilarity hypothesis requires statistical tests that account for the uncertainty associated with the estimations of the mean differences and the standard deviation of the reference product. Recently, a linear reformulation of the biosimilarity hypothesis has been proposed, which facilitates development and implementation of statistical tests. These statistical tests account for the uncertainty in the estimation process of all the unknown parameters. In this paper, we survey methods for constructing confidence intervals for testing the linearized reformulation of the biosimilarity hypothesis and also compare the performance of the methods. We discuss test procedures using confidence intervals to make possible comparison among recently developed methods as well as other previously developed methods that have not been applied for demonstrating analytical similarity. A computer simulation study was conducted to compare the performance of the methods based on the ability to maintain the test size and power, as well as computational complexity. We demonstrate the methods using two example applications. At the end, we make recommendations concerning the use of the methods.

Uso del TEVI-R para describir el vocabulario pasivo en niños de la etnia Mapuche / Use of TEVI-R to describe passive vocabulary in children belonging to a Mapuche community

RESUMEN Objetivo Describir el nivel de vocabulario pasivo en niños y niñas pertenecientes a la etnia mapuche, utilizando el Test de Vocabulario en Imágenes versión Revisada (TEVI-R). Método Se llevó a cabo un estudio de corte transversal. Participaron 26 niños, de ambos géneros cuyas edades fluctuaron entre 4 y 7 años de edad. Se midió el desempeño de vocabulario pasivo mediante la aplicación del TEVI-R, analizando las variables edad y género, además de realizar un análisis de los ítems con mayor cantidad de errores. Resultados El rendimiento de estos niños no se ve influenciado ni por género, ni por edad. Existen potenciales fuentes de error en los ítems de los instrumentos relacionados a la pertinencia cultural, geográfica y a la calidad gráfica de los mismos. Conclusión No se observó asociación por género ni diferencias por edad en la población estudiada. Se discute la posibilidad de elaborar nuevos instrumentos o revisar los disponibles, dadas las características de su lengua materna, con la finalidad de recabar resultados fiables y respetar los elementos que forman parte de los marcadores propios de su cultura.

ABSTRACT Purpose To describe the level of passive vocabulary in boys and girls belonging to the Mapuche ethnic group, using the Vocabulary Test in Images Revised version (TEVI-R). Methods A cross-sectional study was carried out. Twenty-six children, both gender, between 4 to 7 years old participated in the study. The performance of passive vocabulary was measured through the application of the TEVI-R, analyzing the variables age and gender, as well as performing an analysis of the items and the number of errors. Results The performance of these children is not influenced by gender or age. There are potential sources of error in the items of the instruments related to the cultural, geographical relevance and graphic quality of them. Conclusion No association was observed by gender or differences by age in the study population. The possibility of developing new instruments or revising the available ones is discussed, given the characteristics of their native language, obtaining reliable results and respecting the elements that are part of their culture.

Effect of the mGluR5-NAM Basimglurant on Behavior in Adolescents and Adults with Fragile X Syndrome in a Randomized, Double-Blind, Placebo-Controlled Trial: FragXis Phase 2 Results.

Preclinical data suggest that inhibition of the metabotropic glutamate receptor 5 (mGluR5) receptor might hold therapeutic benefits in Fragile X syndrome (FXS). Treatment of Fmr1 knockout mice with mGluR5-negative allosteric modulators (NAMs) has been reported to correct a broad range of phenotypes related to FXS. The early short-term clinical trials with mGluR5 NAMs, including basimglurant, assessing the effects in individuals with FXS, were supportive of further exploration in larger, well-controlled trials. We evaluated basimglurant, a potent and selective mGluR5 NAM, in a 12-week, double-blind, parallel-group study of 183 adults and adolescents (aged 14-50, mean 23.4 years) with FXS. Individuals with an FMR1 full mutation were randomized to placebo or one of two doses of basimglurant. The primary efficacy endpoint was the change from baseline in behavioral symptoms using the Anxiety Depression and Mood Scale (ADAMS) total score. All treatment arms showed marked behavioral improvements from baseline to week 12 with less improvement in the basimglurant 1.5 mg arm than placebo; however, basimglurant 0.5 mg was inferior to placebo in the ADAMs total score. Treatment with basimglurant was overall well-tolerated. A higher incidence of adverse events classified as psychiatric disorders were reported in patients treated with basimglurant, including three patients with hallucinations or psychosis. In this phase 2 clinical trial, basimglurant did not demonstrate improvement over placebo. Evaluation of the overall risk-benefit in younger patient populations is an important consideration for the design of potential further investigations of efficacy with this class of medications.

Discriminating Ability of Abbreviated Impactor Measurement Approach (AIM) to Detect Changes in Mass Median Aerodynamic Diameter (MMAD) of an Albuterol/Salbutamol pMDI Aerosol.

This article reports on results from a two-lab, multiple impactor experiment evaluating the abbreviated impactor measurement (AIM) concept, conducted by the Cascade Impaction Working Group of the International Pharmaceutical Aerosol Consortium on Regulation and Science (IPAC-RS). The goal of this experiment was to expand understanding of the performance of an AIM-type apparatus based on the Andersen eight-stage non-viable cascade impactor (ACI) for the assessment of inhalation aerosols and sprays, compared with the full-resolution version of that impactor described in the pharmacopeial compendia. The experiment was conducted at two centers with a representative commercially available pressurized metered dose inhaler (pMDI) containing albuterol (salbutamol) as active pharmaceutical ingredient (API). Metrics of interest were total mass (TM) emitted from the inhaler, impactor-sized mass (ISM), as well as the ratio of large particle mass (LPM) to small particle mass (SPM). ISM and the LPM/SPM ratio together comprise the efficient data analysis (EDA) metrics. The results of the comparison demonstrated that in this study, the AIM approach had adequate discrimination to detect changes in the mass median aerodynamic diameter (MMAD) of the ACI-sampled aerodynamic particle size distribution (APSD), and therefore could be employed for routine product quality control (QC). As with any test method considered for inclusion in a regulatory filing, the transition from an ACI (used in development) to an appropriate AIM/EDA methodology (used in QC) should be evaluated and supported by data on a product-by-product basis.

Efficacy and Safety of Basimglurant as Adjunctive Therapy for Major Depression: A Randomized Clinical Trial.

IMPORTANCE: Antagonism of the postsynaptic metabotropic glutamate subtype 5 receptor is a novel approach to modulate glutamatergic function and has proven efficacy in a number of preclinical behavioral models of depression. OBJECTIVE: To evaluate the safety and efficacy of basimglurant modified-release (MR) vs placebo as adjunctive therapy to ongoing antidepressant medication therapy in patients with MDD who had inadequate response within the current episode. DESIGN, SETTING, AND PARTICIPANTS: In this phase 2b, double blind, randomized clinical trial of 333 adult patients with a DSM-IV-TR diagnosis of MDD across 59 research clinics globally, patients were assigned to 1 of 2 doses of basimglurant MR (0.5 or 1.5 mg) or placebo once daily, adjunctive to ongoing antidepressant medication therapy (selective serotonin reuptake inhibitor or serotonin and norepinephrine reuptake inhibitor). Patients were enrolled from October 5, 2011, through July 26, 2013. INTERVENTIONS: Six-week treatment with 0.5 mg of basimglurant MR, 1.5-mg basimglurant MR, or placebo once daily, adjunctive to ongoing antidepressant medication therapy. MAIN OUTCOMES AND MEASURES: The primary end point was the mean change from baseline score on the Montgomery-Åsberg Depression Rating Scale (MADRS), as rated by the clinician at week 6. Other measures included patient-rated MADRS, Quick Inventory of Depressive Symptomatology-Self-Report, Clinical Global Impression-Improvement, Patient Global Impression-Improvement, and Clinical Global Impression-Severity Scales and adverse events. RESULTS: A total of 596 patients were screened, and 333 were randomized into the study (mean [SD] age, 47 [11.2] years; 216 female [65.1%]). The primary end point (mean change in clinician-rated MADRS score from baseline to end of treatment) was not met (effect size [ES] = 0.16, P = .42; intent-to-treat [ITT] mixed-effects model for repeated measures [MMRM] analysis for comparing 1.5-mg basimglurant MR and placebo). Across secondary and exploratory end points, 1.5-mg basimglurant MR revealed larger improvements vs placebo on the patient-rated MADRS (-16.2 vs -13.3, ES = 0.28, nominal P = .04), Quick Inventory of Depressive Symptomatology-Self-Report (-7.5 vs -5.8; ES = 0.37, nominal P = .009), Clinical Global Impression-Improvement mean score, and Patient Global Impression-Improvement mean score. Improvements were also seen in the patient-rated MADRS remission rate (36.0% vs 22.0%; nominal P = .03) and response rate (50.5% vs 40.4%; nominal P = .13), A 0.5-mg dose of basimglurant MR had no benefit over placebo in any of these measures. The most common adverse event was dizziness, which was mostly transient and of mild intensity. CONCLUSIONS AND RELEVANCE: No difference was observed on the study's primary outcome measure, the clinician-rated MADRS change from baseline to end of treatment, between adjunctive basimglurant MR vs placebo. Adjunctive 1.5-mg basimglurant MR daily revealed, however, an antidepressant effect across secondary end points, particularly in patient-rated measures. These findings combined with good tolerability warrant further investigation with this compound in depressive disorders. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01437657.

Statistical analysis of data from limiting dilution cloning to assess monoclonality in generating manufacturing cell lines.

Assurance of monoclonality of recombinant cell lines is a critical issue to gain regulatory approval in biological license application (BLA). Some of the requirements of regulatory agencies are the use of proper documentations and appropriate statistical analysis to demonstrate monoclonality. In some cases, one round may be sufficient to demonstrate monoclonality. In this article, we propose the use of confidence intervals for assessing monoclonality for limiting dilution cloning in the generation of recombinant manufacturing cell lines based on a single round. The use of confidence intervals instead of point estimates allow practitioners to account for the uncertainty present in the data when assessing whether an estimated level of monoclonality is consistent with regulatory requirements. In other cases, one round may not be sufficient and two consecutive rounds are required to assess monoclonality. When two consecutive subclonings are required, we improved the present methodology by reducing the infinite series proposed by Coller and Coller (Hybridoma 1983;2:91-96) to a simpler series. The proposed simpler series provides more accurate and reliable results. It also reduces the level of computation and can be easily implemented in any spreadsheet program like Microsoft Excel. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1061-1068, 2016.