An industry perspective on the monitoring of subvisible particles as a quality attribute for protein therapeutics. Chapter metal particles in ophthalmic ointments has been removed since the new chapter provides general particle control guidance. Significant advances in protein chemistry, protein folding, and biologics formulation were made in the past two decades 1521 to study the mechanism of many chemical and physical degradation pathways of protein based biologics. As a result, the requirement for drug product appearance for. Moreover, reallife protein formulations can differ in optical properties such as turbidity, color, refractive index which may potentially influence sbvp detection. Methods and protocols, second edition aims to cover each of these key aspects of protein drug production through the contributions of authors from highly esteemed industrial and academic institutions around the world. The uncertainties around subvisible particles svps may warrant their being controlled as a critical quality attribute, fda office of biotechnology products division of therapeutic proteins dtp deputy director barry cherney stressed at a casssfda cmc strategy forum in washington, d.
Cleanroom requirements for liquid particle testing. Carpenter jf1, randolph tw, jiskoot w, crommelin dj, middaugh cr, winter g, fan yx, kirshner s, verthelyi d, kozlowski s, clouse ka, swann pg, rosenberg a, cherney b. They can confound the analysis of the total subvisible particle population, and also have the potential to interact with the protein depending on formulation conditions14. Methods to asses these particles include dynamic or static light scattering 39. Matter in therapeutic protein injections linda narhi, amgen satish k. Therapeutic protein products used for the differentiation and characterization of subvisible particulates this application note provides an example of how archimedes can be used to detect and quantify the formation of protein subvisible particles and the introduction of silicone oil droplets, in. An industry perspective on the monitoring of subvisible. Overlooking subvisible particles in therapeutic protein. The characterization of protein aggregates is complex and requires the use of many different analytical techniques 3,4,12. Therapeutic protein products used for the differentiation. Initially, much of the discussion and debate in the field has centred on the utility and need for quantifying particles smaller than 10 micron.
Indeed, industry experience has demonstrated that therapeutic proteins such as monoclonal antibodies can exhibit a propensity for selfassociation leading to the formation of aggregates that range in size from nanometres oligomers to microns subvisible and visible particles. Blue 5 danny chou 6 mary cromwell 7 hansjuergen krause 8 hannschristian mahler 9 brian k. Subvisible particulates have historically been monitored through methods such as. Gaps that may compromise product quality carpenter jf, randolph tw, jiskoot w, crommelin dj, middaugh cr, winter g, fan yx, kirshner s, verthelyi d, kozlowski s, clouse ka, swann pg, rosenberg a, cherney b. Immunogenicity assessment for therapeutic protein products. Differentiation and characterization of subvisible particulates in therapeutic protein products we hope the information below is useful, but as always, if you have any further queries about the archimedes system and its capabilities just drop. Subvisible particle analysis the potential immunogenicity of particles ranging from nanometer to micron sizes in biotherapeutics has been a growing area of discussion between regulators and industry. A biopharmaceutical industry perspective on the control of. Usp 787 subvisible particulate matter in therapeutic. Engineering of therapeutic proteins 157 posttranslational modification, protein fusions, and genetic engineering, as illustrated in figure 7. Clayton gough identification of asp isomerization in proteins by o labeling and tandem mass spectrometry jennifer zhang and viswanatham katta monitoring of subvisible particles in therapeutic proteins satish k.
Usp, therapeutic protein injections beckman coulter. It has been proposed that aggregates and particles may lead to unwanted increases in the immune response with a possible impact on safety or efficacy. Protein aggregates are commonly measured using liquid chromatography with either uv detection hplcuv or fluorescence detection fld, which utilizes the native fluorescence of some therapeutic proteins e. The use of flow cytometry for the detection of subvisible. As an example, whereas only approximately 15 protein particles of 50. Among these, flow imaging also called flow microscopy has been widely employed. John carpenter professor of pharmaceutical sciences and codirector of the center for pharmaceutical biotechnology, university of colorado usa. Comparisons of two different analytical methodologies for the. Impacts on aggregation pathways, adverse immunogenicity and regulatory expectations for therapeutic proteins. As protein subvisible particles vary widely in shape and size, it is difficult to quantitatively describe the limitations of mdi and lo for actual samples of a protein therapeutic drug. Analyzing protein aggregation in biopharmaceuticals.
Division of therapeutic proteins, center for drug evaluation and research, us food and drug administration, rockville, maryland 20857 search. Understanding triggers of these phenomena and their complex effects are important for the development and optimization of protein formulations. The unintended presence of particulate matter in injectable products is an indicator of the quality of the product. Distinction between proteinaceous and nonproteinaceous svps is vital in monitoring formulation stability.
Pdf monitoring of subvisible particles in therapeutic. The presence of subvisible particles svps in parenteral formulations of biologics is a major challenge in the development of therapeutic protein formulations. After the identification of a potential protein drug, the next critical step is the production of sufficient authentic material for testing, characterization, and clinical trials, which, when successful, leads to. Rapid detection and characterization of protein aggregates by. Fh tobias frommknecht submitted to the graduate degree program in pharmaceutical chemistry and the graduate faculty of the. Phaseappropriate application of analytical methods to. Monitoring of subvisible particles in therapeutic proteins. Insights into the roles of particles in the protein aggregation pathway. Characterization of particles in protein solutions. Comparisons of two different analytical methodologies for the characterization of sub visible particles in therapeutic protein formulations by dipl. Hplcuv or fluorescence detection fld, which utilizes the native fluorescence of some therapeutic proteins e.
Evans, the use of flow cytometry for the detection of subvisible particles in therapeutic protein formulations, journal of pharmaceutical sciences, 100, 5, 16711678, 2010. Concern around the lack of monitoring of proteinaceous subvisible particulates in the 0. Contaminating particles in injectable or infusion drug therapies can be microbiological resulting in infection, allergic reaction or, in extreme cases, anaphylactic shock. The new usp subvisible particulate matter in therapeutic protein injections test can be used as an alternative to usp. Monitoring of subvisible particles in therapeutic proteins satish singh k. Recommendation is made that the range and levels of subvisible particles 210 microns present in therapeutic protein products, initially and over the course of the shelflife, should be assessed. The growing realization of the importance of monitoring all foreign as well as inherent particulates in therapeutic protein products has led to a number of. Usp chapter, subvisible particulate matter in therapeutic protein injections, became official august 1, 2014 and provides specific guidance for proteinbased formulations. Emphasizing the newest developments in the field, this second. Further, new chapters subvisible particulate matter in therapeutic protein injections and measurement of subvisible particulate matter in therapeutic protein injections provide.
Up to now, it had only been possible to fulfill the pharmaceutical standards 788 particulate matter in injections and 789. Comparisons of two different analytical methodologies for. You should also check out our latest application note on the topic of monitoring subvisible particles. The guidance goes on to indicate that as more analytical methods become available, there should be a move to characterize particles in smaller 0. With the apparent deficiencies in the analysis of the subvisible particles especially those smaller than 10. To explore the potential sensitivity of monitoring subvisible particle formation as a means of measuring aggregated protein, the estimated mass of protein in particles was calculated eq. This can occur with the first introduction of the drug or through repeated administration. Current pharmacopeial guidelines for monitoring subvisible particulates svps in therapeutic protein injections have expanded into new areas as usp is provided as an alternative to. The ability to detect and quantitate these aggregates particles in protein solutions is a function of their size, amount number, and the capability of the technique being employed. These preliminary results encourage further evaluation of fc as a practical tool that may facilitate development of safer. Characterization of subvisible particles in biotherapeutic. Detecting and quantifying the formation of protein sub. A multicompany assessment of submicron particle levels in.
The number and size range of subvisible particles in parenteral products have traditionally been measured by light obscuration, but when applied to monitoring protein particles, accuracy may be. A second method for monitoring subvisible particles is the membrane microscopy method also referred to as method 2, as outlined in the pharmacopeias 4 6. The rmm was used to measure sub visible particles before and after the application of shear stress and the results are presented in figure 2. Furthermore, unlike the concern regarding extraneous particles in small molecule parenteral products, protein particles can accumulate over time during storage of the final presentation. An industry perspective on the monitoring of subvisible particles as a. This educational session will focus on some of the new demands in counting particles in liquids, with a focus on the challenges of preparing and sampling protein based materials. Protein aggregate and subvisible particles analysis information.
The laboratory instruments svss and sbss are now compatible to the pharmacopoeia usp 787 subvisible particulate matter in therapeutic protein injections. The coulter principle can be used for analysis of subvisible particles in protein formulations. Characterization of sub visible particles in protein therapeutic formulations biologic formulations are complex and under certain circumstances can experience protein aggregation or poor stability. Aggregation and immunogenicity of therapeutic proteins. Special attenti the source of particles extrinsic, intrinsic, or inherent can be understood better and particlesize distribution and other characteristics can be studied and used to differentiate them if methods based on different measurement principles are used. Visible and subvisible protein particle inspection within.
In spite of these advances, the understanding of pathways leading to aggregateparticulate formation and prevention remains relatively poor. The aaps product attribute and biological consequences pabc focus group which has since transformed into an aaps community conducted a survey where participating labs rated their method of choice to. We describe the underlying theory, benefits, shortcomings, and. Monitoring of subvisible particles in therapeutic proteins springerlink. Subvisible particle counting provides a sensitive method of detecting and quantifying aggregation of monoclonal antibody caused by freeze. Immunogenicity assessment for therapeutic protein products this guidance represent s the food and drug administration s fdas current thinking on this topic. Usp chapter, subvisible particulate matter in therapeutic protein injections, became official august 1, 2014 and provides specific guidance for protein based formulations. Pdf overlooking subvisible particles in therapeutic. Inherent particles are particles which originate from the drug product, either the protein therapeutic itself or formulation components. Characterization of subvisible particles in protein. View enhanced pdf access article on wiley online library html view download pdf for offline viewing. The united states pharmacopoeia chapter on therapeutic proteins, usp 6, suggests that sources of particles found in parenteral products can be grouped into three sources. Pollo,2 muppalla sukumar2 1brightwell technologies inc. Singh 1 nataliya afonina 2 michel awwad 3 karoline bechtoldpeters 4 jeffrey t.
The coulter principle for analysis of subvisible particles. The aim of this study was thus to evaluate the ability of subvisible particles of a therapeutic. Monitoring of subvisible particles in therapeutic proteins article pdf available in methods in molecular biology clifton, n. Immunogenicity can be considered the ability of a material to induce an immune response in the body. This stimuli article presents a modified lightobscuration method that can be used in procedures to analyze and control subvisible particles in protein therapeutics. Usp is meant for therapeutic protein injections, making changes for smaller test product volumes and smaller test aliquots. The risk of of aggregates and particulates in biopharmaceutical formulation continues to be one of the major quality concerns in biotherapeutics development. The potential impact of subvisible particles svps in protein therapeutic products has received a great deal of attention recently. Changes to subvisible particle regulations for parenteral.
Request pdf an industry perspective on the monitoring of subvisible particles as a quality attribute for protein therapeutics concern around the lack of. The phaseappropriate application of analytical methods to characterize, monitor, and control particles is an important aspect of the development of safe and efficacious biotherapeutics. May 08, 2012 with the increasing number of organizations and academic institutions involved in biotherapeutics research, understanding of biochemical and biophysical profile of many classes of biological molecules such as antibodies, fusion proteins, and drug conjugates has improved significantly over the past decade. In this context, subvisible proteinaceous particles refer to aggregates that have grown in size to fall in the subvisible size range. Over the past several years there has been greatly increasing attention on subvisible particles in therapeutic protein products. Subvisible particle counting provides a sensitive method. Protein particulates can form in a wide range of sizes and shapes. Building on the valuable first edition, therapeutic proteins.
With the development and commercialization of protein therapeutics, the application of these. Particles in therapeutic protein formulations, part 1. Characterization of subvisible particles in biotherapeutic prefilled. All therapeutic protein products contain intrinsic particles formed by the aggregation. Subvisible particles sbvps are found in virtually all parental drug products dps and are currently considered a critical quality attribute. Therapeutic proteins methods and protocols vladimir. Uncertainties in subvisible particulates need to be. The presence of large quantities of aggregates is believed to be one of the causes of unwanted immunogenic responses. As a result, new analytical methods have emerged to characterize and quantify svps.
Extensive chemical modifications in the primary protein. Uncertainties in subvisible particulates need to be understood, fda tells biotech cmc strategy forum the uncertainties around subvisible particles svps may warrant their being controlled as a critical quality attribute, fda office of biotechnology products division of therapeutic proteins dtp deputy director barry cherney stressed at a. This educational session will focus on some of the new demands in counting particles in liquids, with a focus on the challenges of preparing and sampling proteinbased materials. Recent topics of research in the characterization and quality. Despite recent developments regarding sizing and counting of particulate matter, nosingle existing analytical techniques can coverthe entire size span of protein aggregates. Evaluation of protein aggregatessubvisible particles in.
Due to the aforementioned limitations, accurate determination, and characterization of protein subvisible particles remains a challenging and difficult task. Subvisible particulates have historically been monitored through methods such as light obscuration and membrane microscopy, as outlined in the united states pharmacopeia usp general chapter or the equivalent ph eur 2. Pdf monitoring of subvisible particles in therapeutic proteins. An industry perspective on the monitoring of subvisible particles as. The method includes appropriate sample handling, the ability to analyze volumes as small as 1 ml, and reporting of subvisible particles. An industry perspective on the monitoring of subvisible particles as a quality attribute for protein therapeutics author links open overlay panel satish k. Separation, characterization and discriminant analysis of.
Subvisible particulate matter in therapeutic protein. A comprehensive evaluation of nanoparticle tracking analysis. Protein particulate detection issues in biotherapeutics. Where the nature of the contents or the containerclosure system permits only limited capability for inspection of the total. An industry perspective on the monitoring of subvisible particles as a quality attribute for protein therapeutics august 2010 journal of pharmaceutical sciences 998. Sp scientific has announced a new lyolearn webinar, due to run on 27th june 20, entitled sub visible particles. Intrinsic particulate contamination is usually contamination from the vial or filling process due to inefficient cleaning, whereas extrinsic. When examining injections and parenteral infusions for subvisible particles.
Subvisible particle analysis protein antibody integrity bio. For the determination of particulate matter, two procedures, method 1 light obscuration particle count test and method 2 microscopic particle count test, are specified hereinafter. Before the syringeinduced stress, the archimedes system detected only very few particles, meaning that the solution was reasonably pure with a less number of large protein aggregates. Therapeutic proteins have made an immense contribution to treatment.
Thus, even though large protein aggregates that are classified as subvisible particles are potentially the most immunogenic class of protein aggregates, subvisible particles smaller than 10. Therapeutic protein aggregates have been linked to issues such as loss of efficacy and immunogenicity. Abstract usp is a new informational chapter intended to. Techniques to improve the characterization of protein. Abstract the unintended presence of particulate matter in injectable products is an indicator of the. Usp 787 subvisible particulate matter in therapeutic protein. Overlooking subvisible particles in therapeutic protein products. New methods or procedures need to be developed to help us better understand the issue of subvisible particles in protein pharmaceuticals. Characterization of pegylated biopharmaceutical products by lcms and lcmsms lihua huang and p. Detection and characterization of subvisible aggregates of.
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