Physiologically Based Biopharmaceutics Modeling (PBBM): Best Practices for Drug Product Quality, Regulatory and Industry Perspectives: 2023 Workshop Summary Report.

Physiologically based biopharmaceutics modeling (PBBM) is used to elevate drug product quality by providing a more accurate and holistic understanding of how drugs interact with the human body. These models are based on the integration of physiological, pharmacological, and pharmaceutical data to simulate and predict drug behavior in vivo. Effective utilization of PBBM requires a consistent approach to model development, verification, validation, and application. Currently, only one country has a draft guidance document for PBBM, whereas other major regulatory authorities have had limited experience with the review of PBBM. To address this gap, industry submitted confidential PBBM case studies to be reviewed by the regulatory agencies; software companies committed to training. PBBM cases were independently and collaboratively discussed by regulators, and academic colleagues participated in some of the discussions. Successful bioequivalence "safe space" industry case examples are also presented. Overall, six regulatory agencies were involved in the case study exercises, including ANVISA, FDA, Health Canada, MHRA, PMDA, and EMA (experts from Belgium, Germany, Norway, Portugal, Spain, and Sweden), and we believe this is the first time such a collaboration has taken place. The outcomes were presented at this workshop, together with a participant survey on the utility and experience with PBBM submissions, to discuss the best scientific practices for developing, validating, and applying PBBMs. The PBBM case studies enabled industry to receive constructive feedback from global regulators and highlighted clear direction for future PBBM submissions for regulatory consideration.

PBBM Considerations for Base Models, Model Validation, and Application Steps: Workshop Summary Report.

The proceedings from the 30th August 2023 (Day 2) of the workshop "Physiologically Based Biopharmaceutics Models (PBBM) Best Practices for Drug Product Quality: Regulatory and Industry Perspectives" are provided herein. Day 2 covered PBBM case studies from six regulatory authorities which provided considerations for model verification, validation, and application based on the context of use (COU) of the model. PBBM case studies to define critical material attribute (CMA) specification settings, such as active pharmaceutical ingredient (API) particle size distributions (PSDs) were shared. PBBM case studies to define critical quality attributes (CQAs) such as the dissolution specification setting or to define the bioequivalence safe space were also discussed. Examples of PBBM using the credibility assessment framework, COU and model risk assessment, as well as scientific learnings from PBBM case studies are provided. Breakout session discussions highlighted current trends and barriers to application of PBBMs including: (a) PBBM credibility assessment framework and level of validation, (b) use of disposition parameters in PBBM and points to consider when iv data are not available, (c) conducting virtual bioequivalence trials and dealing with variability, (d) model acceptance criteria, and (e) application of PBBMs for establishing safe space and failure edges.

Safety assessment of drug impurities for patient safety: A comprehensive review.

Drug impurities are undesirable but unavoidable chemicals which can occur throughout the drug life cycle. The safety implications of drug impurities can be significant given that they can impact safety, quality, and efficacy of drug products and that certain drug impurities are mutagenic, carcinogenic, or teratogenic. The characteristics of drug impurities could be specific to drug modalities (e.g., small molecules vs. biologics). The commonly encountered drug impurities include elemental impurity, residual solvent, organic impurity, host cell protein and DNA, residual viral vector, extractable and leachable, and particle. They can cause various adverse effects such as immunogenicity, infection, genotoxicity, and carcinogenicity upon significant exposure. Therefore, the effective control of these drug impurities is central for patient safety. Regulations and guidelines are available for drug developers to manage them. Their qualification is obtained based on authoritative qualification thresholds or safety assessment following the classic toxicological risk assessment. The current review focuses on the safety assessment science and methodology used for diverse types of drug impurities. Due to the different nature of diverse drug impurities, their safety assessment represents a significant challenge for drug developers.

Analysis of non-mutagenic substances in the context of drug impurity assessment - Few are potent toxicants.

ICH Q3A/B guidelines provide qualification thresholds for impurities or degradation products in new drug substances and products. However, the guidelines note that certain impurities/degradation products may warrant further safety evaluation for being unusually potent or toxic. The purpose of this study was to confirm that especially toxic non-mutagenic compounds are rare and to identify classes of compounds that could warrant lower qualification thresholds. A total of 2815 compounds were evaluated, of which 2213 were assessed as non-mutagenic. For the purpose of this analysis, compounds were considered potent when the point of departure was ≤0.2 mg/kg/day based on the qualification threshold (1 mg/day or 0.02 mg/kg/day for a 50 kg human) in a new drug substance, with an additional 10-fold margin. Only 54 of the entire set (2.4%) would be considered potent based on this conservative potency analysis, confirming that the existing ICH Q3A/B qualification thresholds are appropriate for the majority of impurities. If the Q3A/B threshold, without the additional 10-fold margin is used, 14 compounds (0.6%) are considered "highly potent". Very few non-mutagenic structural classes were identified, including organothiophosphates and derivatives, polychlorinated benzenes and polychlorinated polycyclic aliphatics, that correlate with potential high potency, consistent with prior publications.

To Dilute or Not to Dilute: Nominal Titer Dosing for Genetic Medicines.

Recombinant adeno-associated virus (AAV) has become a popular platform for many gene therapy applications. The strength of AAV-based products is a critical quality attribute that affects the efficacy of the drug and is measured as the concentration of vector genomes, or physical titer. Because the dosing of patients is based on the titer measurement, it is critical for manufacturers to ensure that the measured titer of the drug product is close to the actual concentration of the batch. Historically, dosing calculations have been performed using the measured titer, which is reported on the drug product label. However, due to recent regulatory guidance, sponsors are now expected to label the drug product with nominal or "target" titer. This new expectation for gene therapy products can pose a challenge in the presence of process and analytical variability. In particular, the manufacturer must decide if a dilution of the drug substance is warranted at the drug product stage to bring the strength in line with the nominal value. In this paper, we present two straightforward statistical methods to aid the manufacturer in the dilution decision. These approaches use the understanding of process and analytical variability to compute probabilities of achieving the desired drug product titer. We also provide an approach for determining an optimal assay replication strategy for achieving the desired probability of meeting drug product release specifications.

Accelerated Predictive Stability Study of a Pediatric Drug Product for a Supplemental New Drug Application.

In this paper, we report two Accelerated Stability Assessment Program (ASAP) studies for a pediatric drug product. Whereas the first study using a generic design failed to establish a predictive model, the second one was successful after troubleshooting the first study and customizing the study conditions. This work highlighted important lessons learned from designing an ASAP study for formulations containing excipients that could undergo phase change at high humidity levels. The stability predictions by the second ASAP model were consistent with available long-term stability data of the drug product under various storage conditions in two different packaging configurations. The ASAP model was part of the justifications accepted by the health authority to submit a stability package with reduced long-term stability data from the primary stability batches for a Supplemental New Drug Application (sNDA).

Strengths and weaknesses of current and future prospects of artificial intelligence-mounted technologies applied in the development of pharmaceutical products and services.

Starting from drug discovery, through research and development, to clinical trials and FDA approval, artificial intelligence (AI) plays a vital role in planning, developing, assessing modelling, and optimization of product attributes. In recent decades, machine-learning algorithms integrated into artificial neural networks, neuro-fuzzy logic and decision trees have been applied to tremendous domains related to drug formulation development. Optimized formulations were transformed from lab to market based on optimized properties derived from AI Technologies. Research and development in pharmaceutical industry rely upon computer-driven equipment and machine learning technology to extract data, perform simulations, modelling, and optimization to get optimum solutions. Merging AI technologies in various steps of pharmaceutical manufacture is a major challenge due to lack of in-house technologies. In silico studies based on artificial intelligence are widely applied as effective tools to screen the market needs of medications and pharmaceutical services through inspecting scientific literature and prioritizing medicines for specific illnesses or a particular patient. Specialized personnel who excel in scientific and data science with analytical knowledge are essential for transformation to smart manufacturing and offering services. However, privacy, cybersecurity, AI-dependent unemployment, and ownership rights of AI technologies require proper regulations to gain the benefits and minimize the drawbacks.

Advanced mass spectrometry workflows for accurate quantification of trace-level host cell proteins in drug products: Benefits of FAIMS separation and gas-phase fractionation DIA.

Therapeutic monoclonal antibodies (mAb) production relies on multiple purification steps before release as a drug product (DP). A few host cell proteins (HCPs) may co-purify with the mAb. Their monitoring is crucial due to the considerable risk they represent for mAb stability, integrity, and efficacy and their potential immunogenicity. Enzyme-linked immunosorbent assays (ELISA) commonly used for global HCP monitoring present limitations in terms of identification and quantification of individual HCPs. Therefore, liquid chromatography tandem mass spectrometry (LC-MS/MS) has emerged as a promising alternative. Challenging DP samples show an extreme dynamic range requiring high performing methods to detect and reliably quantify trace-level HCPs. Here, we investigated the benefits of adding high-field asymmetric ion mobility spectrometry (FAIMS) separation and gas phase fractionation (GPF) prior to data independent acquisition (DIA). FAIMS LC-MS/MS analysis allowed the identification of 221 HCPs among which 158 were reliably quantified for a global amount of 880 ng/mg of NIST mAb Reference Material. Our methods have also been successfully applied to two FDA/EMA approved DPs and allowed digging deeper into the HCP landscape with the identification and quantification of a few tens of HCPs with sensitivity down to the sub-ng/mg of mAb level.

Sterile filtration of a multi-serotype glycoconjugate vaccine drug product.

Glycoconjugate vaccines consisting of multiple serotypes of the bacterial capsular polysaccharide can provide strong protection against infection by significant pathogens. Previous studies of the sterile filtration behavior of these glycoconjugates have been limited to experiments with individual serotypes even though the formulated vaccines contain several different serotypes to provide broad immunization. The objective of this study was to explore the fouling behavior of a glycoconjugate vaccine drug product consisting of four different polysaccharide serotypes. Sterile filtration data were obtained with 0.22 µm Durapore® membranes at both constant flux and constant pressure for both the individual serotypes and the drug product containing multiple serotypes. Fouled membranes were examined by confocal microscopy, demonstrating that all four serotypes deposit in a narrow band near the filter inlet. The different ionic composition of the formulation buffer (compared to the buffers used with the drug substance) had a large effect on the fouling behavior. In addition, the fouling resistance associated with the drug product was greater than the sum of the resistances of the individual serotypes. These results provide important insights into the sterile filtration behavior of these multivalent glycoconjugate vaccines.

Characterizing Silicone Oil-Induced Protein Aggregation with Stimulated Raman Scattering Imaging.

Particles in biopharmaceutical products present high risks due to their detrimental impacts on product quality and safety. Identification and quantification of particles in drug products are important to understand particle formation mechanisms, which can help develop control strategies for particle formation during the formulation development and manufacturing process. However, existing analytical techniques such as microflow imaging and light obscuration measurement lack the sensitivity and resolution to detect particles with sizes smaller than 2 µm. More importantly, these techniques are not able to provide chemical information to determine particle composition. In this work, we overcome these challenges by applying the stimulated Raman scattering (SRS) microscopy technique to monitor the C-H Raman stretching modes of the proteinaceous particles and silicone oil droplets formed in the prefilled syringe barrel. By comparing the relative signal intensity and spectral features of each component, most particles can be classified as protein-silicone oil aggregates. We further show that morphological features are poor indicators of particle composition. Our method has the capability to quantify aggregation in protein therapeutics with chemical and spatial information in a label-free manner, potentially allowing high throughput screening or investigation of aggregation mechanisms.

Current State and Opportunities with Long-acting Injectables: Industry Perspectives from the Innovation and Quality Consortium "Long-Acting Injectables" Working Group.

Long-acting injectable (LAI) formulations can provide several advantages over the more traditional oral formulation as drug product opportunities. LAI formulations can achieve sustained drug release for extended periods of time, which results in less frequent dosing requirements leading to higher patient adherence and more optimal therapeutic outcomes. This review article will provide an industry perspective on the development and associated challenges of long-acting injectable formulations. The LAIs described herein include polymer-based formulations, oil-based formulations, and crystalline drug suspensions. The review discusses manufacturing processes, including quality controls, considerations of the Active Pharmaceutical Ingredient (API), biopharmaceutical properties and clinical requirements pertaining to LAI technology selection, and characterization of LAIs through in vitro, in vivo and in silico approaches. Lastly, the article includes a discussion around the current lack of suitable compendial and biorelevant in vitro models for the evaluation of LAIs and its subsequent impact on LAI product development and approval.

The Effect of the Drug Life Cycle Price on Cost-Effectiveness: Case Studies Using Real-World Pricing Data.

OBJECTIVES: Cost-effectiveness analyses (CEAs) generally assume constant drug prices throughout the model time horizon, yet it is known that prices are not constant, often with price decreases near loss of exclusivity (LOE). This study explores the impact of using dynamic drug-specific prices on the incremental cost-effectiveness ratio (ICER) using selected reproduced case studies. METHODS: Case studies were selected following explicit criteria to reflect a variety of drug characteristics. For each drug, a published CEA model was identified, replicated, and modified with dynamic real-world pricing data, to compare ICERs based on constant drug prices with estimates obtained when including drug life cycle pricing. The impact of dynamic real-world pricing-inclusive LOE-was analyzed using a single patient cohort and multiple cohorts over time. RESULTS: Fluvastatin, alendronic acid + colecalciferol combination therapy, letrozole and clopidogrel were selected as case studies. Inclusion of real-world pricing data compared with applying constant prices reduced the ICER in a single-cohort setting up to 43%. In the multicohort analyses, further reductions of the ICERs were observed of up to 113%. The ICERs were sensitive to the period of drug usage relative to the models' time horizons, the relative proportions of drug costs in the overall treatment costs, and timing of LOE compared with the cost year of the original analysis. CONCLUSIONS: Assuming dynamic drug prices may lead to more representative ICER estimates. Future CEAs for drugs could account for predicted and disaggregated life cycle price developments based on retrospective data.

Stability and safety key factors of the oncolytic protoparvovirus H-1 from manufacturing to human application.

The oncolytic rodent protoparvovirus H-1PV has been successfully used in phase I/II clinical trials to treat recurrent glioblastoma multiforme and pancreatic cancer. The present work focuses on the stability and environmental safety of the H-1PV drug product from production up to its use in patients. We identified hold-steps in manufacturing for up to 3 months and showed 7-years stability for the optimal product formulation. Stress testing via UV, temperature, and pH also determined that the drug product is stable. De- and rehydration for lyophilization simulation are possible without infectious virus loss. Furthermore, we prove in-use stability for 4 days at room temperature and show no virus adsorption to injection devices, guaranteeing the correct administration dose. Iodixanol in the formulation, resulting in high viscosity, protects H-1PV against UV and some disinfectants. Nonetheless, H-1PV is depleted with rapid heat deactivation, autoclavation, and nanofiltration. Assessment of chemical disinfectants that are currently recommended by the Robert Koch-Institute demonstrated that ethanol-based hand disinfectants are not effective; however, aldehyde-based disinfectants for surfaces and instruments demonstrate sufficient H-1PV deactivation in aqueous formulations by 4 to 6 log10. With these results, we could establish a specific hygiene plan for all involved facilities from manufacturing to patient application. Overall, using 48% Iodixanol in Visipaque/Ringer as a drug formulation stabilizes H-1PV infectivity over years and protects against virus loss from short-term UV, low pH, and temperature exposure. KEY POINTS: • Optimal formulation of drug product protects the H-1PV protoparvovirus against UV, temperatures up to 50 °C, and low pH (> 1.25), stabilizing the virus during manufacturing, storage, transport, and application. • H-1PV is stable during in-use and does not adsorb to injection devices during patient administration. • Hygiene plan for H-1PV with physicochemical methods has been established.

Development and Validation of a Stability-indicating UPLC-DAD Method for the Simultaneous Determination of Ivermectin and Praziquantel in Pharmaceutical Tablets and Dissolution Media.

Currently, there is no single rapid and accurate stability-indicating quantitative method that can simultaneously determine both ivermectin and praziquantel and their related compounds. Thus, the goal of this research is to develop and validate a new rapid, accurate, and stability-indicating ultra-performance liquid chromatography (UPLC) method. The method uses a water, acetonitrile, and methanol gradient. The chromatographic separation was achieved on a C18 (1.7 µm, 2.1 × 50 mm) column with a flow rate of 0.7 mL/min, and the column temperature was maintained at 40°C. Analytes are detected at 245 nm. The method was validated in accordance with ICH Q2R1 guidelines. The linearity (R2) was >0.9987 and 0.9997 for praziquantel and ivermectin, respectively. The corresponding accuracy ranged between 98.0 and 102.0%. Intermediate precision (assessed as inter-day precision) was determined by calculating the cumulative %CV of eighteen assay preparations and was less than 2.0% for both praziquantel and ivermectin. The specificity of the method was shown by the resolution of the two active pharmaceutical ingredients (APIs) from any interfering excipients, impurities, or degradation products. The limit of detection and quantitation for ivermectin was 26.80 ng/mL and 81.22 ng/mL, respectively. The limit of detection and quantitation for praziquantel was 1.39 µg/mL and 4.22 µg/mL, respectively. The robustness study proved that method performance is stable against small variations in sample processing parameters (shaking, sonication time, and acetonitrile % in solvent solution) and also against small variations in the initial % of mobile phase components and gradient slope. Using ICH Q2R2 criteria, the method was demonstrated to be specific, accurate, stability indicating, and robust to small variations of chromatographic variables.

The journey of CAR-T therapy in hematological malignancies.

Chimeric antigen receptor T (CAR-T) cells therapy has revolutionized the treatment paradigms for hematological malignancies, with multi-line therapy-refractory patients achieving durable complete remissions (CR) and relatively high objective response rate (ORR). So far, many CAR-T products, such as Kymriah, Yescarta and Tecartus, have been developed and got the unprecedented results. However, some patients may relapse afterwards, driving intense investigations into promoting the development of novel strategies to overcome resistance and mechanisms of relapse. Notable technical progress, such as nanobodies and CRISPR-Case9, has also taken place to ensure CAR-T cell therapy fully satisfies its medical potential. In this review, we outline the basic principles for the development and manufacturing processes of CAR-T cell therapy, summarize the similarities and differences in efficacy of different products as well as their corresponding clinical results, and discuss CAR-T immunotherapy combined with other clinical effects of drug therapy.

Proteomics Analysis of Antitumor Activity of Agrimonia pilosa Ledeb. in Human Oral Squamous Cell Carcinoma Cells.

Oral cancer is a malignant neoplasm of oral cavity. It accounts for approximately 5% of all malignant tumors. Approximately 97% of all oral cancers are squamous cell carcinomas, followed by adenocarcinomas, and rarely malignant melanomas. It occurs particularly in males (twice as common in males than in females) of middle age (above 40 years). Agrimonia pilosa Ledeb. has traditionally been known for its effective antitumor activity and is currently used in China for cancer therapy. A. pilosa Ledeb. has been traditionally used for the treatment of abdominal pain, sore throat, headache, blood discharge, parasitic infections, and eczema in Korea and other Asian countries. Most studies on A. pilosa Ledeb. are related to the leaves and a few investigated the roots of the plant. However, detailed mechanisms of antitumor activity of A. pilosa Ledeb. have not been fully elucidated. Furthermore, to date, there have been no reports on the antitumor effect of A. pilosa Ledeb. in oral squamous cells. In this study, we used proteomic technology to observe changes in proteins related to anticancer activity of A. pilosa Ledeb. and identified target proteins among altered proteins to reveal the underlying mechanism of action.

Development of an induced pluripotent stem cell-specific microRNA assay for detection of residual undifferentiated cells in natural killer cell therapy products.

Most clinically evaluated chimeric antigen receptor (CAR)-based cell therapies are generated from autologous immune cells. However, there are several limitations to autologous cell therapy, including low availability, poor quality of starting cellular material and limited expansion capability. Recently, induced pluripotent stem cell (iPSC)-derived allogeneic cell therapy platforms have gained popularity, as they seek to overcome many of the challenges inherent to current autologous cell therapies. However, teratoma risk associated with residual undifferentiated cells (i.e., iPSCs) in the drug product may restrict potential clinical applications if left unaddressed. To ensure the safety of the final cell therapy product, there is a need to develop quality control assays to detect residual iPSCs. Combining microRNA (miRNA) sequencing data with publicly archived miRNA microarray datasets, we demonstrated that miRNAs belonging to the 300 family (miR-302a-5p, miR-302c-3p and miR-302d-5p) and 500 family (miR-518f-5p and miR-519-3p) were highly expressed in iPSCs (both periperal blood mononuclear cell- and T cell-derived iPSCs) compared with a number of differentiated cell types. We developed and validated a sensitive digital droplet polymerase chain reaction (ddPCR) assay targeting these miRNAs to detect low levels of residual iPSCs in differentiated cell samples. The miRNA ddPCR-based method with primers for miR-302a-5p, miR-302c-3p and miR-302d-5p detected as few as 5, 3 and 10 undifferentiated iPSCs, respectively, in the background of 106 iPSC-derived natural killer (iNK) cells. These results suggest that our method targeting identified iPSC-specific miRNA transcripts is specific and sensitive for the quality assessment of NK cell therapy products derived from iPSCs.

Vehicles for Drug Administration to Children: Results and Learnings from an In-Depth Screening of FDA-Recommended Liquids and Soft Foods for Product Quality Assessment.

PURPOSE: Mixing with liquids or soft foods is a common procedure to improve acceptability of oral medicines in children but may affect drug stability and the in vivo performance of the administered drug product. The aim of the present study was to obtain an overview of the variability of critical attributes of commonly used vehicles and to identify which vehicle characteristics need to be considered when developing in vitro methods for evaluating product quality. METHODS: One product of each vehicle listed in the FDA draft guidance "Use of Liquids and/or Soft Foods as Vehicles for Drug Administration" was analyzed with regard to composition, calorific content and physicochemical properties. RESULTS: The studied vehicles show wide variability, both in composition and physicochemical properties. No correlation was observed between vehicle composition and physicochemical properties. Comparison of results of the present study with previously published data also provided variability in physicochemical properties within individual vehicle types. CONCLUSIONS: To identify acceptable (qualified) vehicles for global drug product labeling, it is important that the vehicles selected for in vitro compatibility screening reflect the variability in composition and essential physicochemical properties of the vehicles recommended on the product label, rather than relying on results obtained with a single vehicle of each type. Future activities will focus on the development of standardized dosing vehicles that can represent key vehicle characteristics in all their variability to ensure reliable risk assessment.

Color, Scent and Size: Exploring Women's Preferences Around Design Characteristics of Drug-Releasing Vaginal Rings.

Steroid-releasing vaginal rings are available for contraception and estrogen replacement therapy, and a new antiretroviral-releasing ring was recently approved for HIV prevention. Marketed rings are white or transparent in appearance, non-scented, and supplied as one-size-fits-all devices with diameters ranging from 54 to 56 mm. In this study, drug-free silicone elastomer rings were manufactured in different sizes, colors and scents, and the opinions/preferences of 16 women (eThekwini District, South Africa; 20-34 years) assessed through focus group discussions and thematic analysis. Opinions varied on ring color and scent, with some women preferring specific colors or scent intensities, while for others these attributes were unimportant. Concerns about color and scent were linked to perceptions around vaginal health and safety related to chemical composition. There was greater agreement on preferred ring size; flexibility and width were considered important factors for insertion and comfort. Greater choice with ring products could facilitate acceptability and overall uptake.

Direct Compaction Drug Product Process Modeling.

Most challenges during the development of solid dosage forms are related to the impact of any variations in raw material properties, batch size, or equipment scales on the product quality and the control of the manufacturing process. With the ever pertinent restrictions on time and resource availability versus heightened expectations to develop, optimize, and troubleshoot manufacturing processes, targeted and robust science-based process modeling platforms are essential. This review focuses on the modeling of unit operations and practices involved in batch manufacturing of solid dosage forms by direct compaction. An effort is made to highlight the key advances in the past five years, and to propose potentially beneficial future study directions.