Free versus total ligand-binding assays: points to consider in biotherapeutic drug development
2011; Future Science Ltd; Volume: 3; Issue: 11 Linguagem: Inglês
10.4155/bio.11.73
ISSN1757-6199
Autores Tópico(s)Protein purification and stability
ResumoBioanalysisVol. 3, No. 11 EditorialFree AccessFree versus total ligand-binding assays: points to consider in biotherapeutic drug developmentJihong Yang & Valerie QuarmbyJihong Yang† Author for correspondenceBioAnalytical Sciences, Genentech, South San Francisco, CA 94080, USA. & Valerie QuarmbyBioAnalytical Sciences, Genentech, South San Francisco, CA, USAPublished Online:7 Jun 2011https://doi.org/10.4155/bio.11.73AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareShare onFacebookTwitterLinkedInRedditEmail Keywords: biotherapeutic drug developmentELISAligand-binding assaymonoclonal antibodypharmacodynamicpharmacokineticsAccurate quantitative information on biotherapeutic drug concentrations from both preclinical and clinical studies is critical to support drug development. These data reveal the relationship between drug concentrations in circulation and drug exposure; help characterize pharmacokinetics (PKs) of the drug candidates; facilitate projection of doses for human Phase I–III studies; and provide the foundation of PK/pharmacodynamic (PD) modeling. In addition, drug concentration data are important in revealing potential relationships between drug exposure and efficacy or safety. Information about circulating concentrations of soluble drug targets is also useful in understanding PK/PD relationships in situations where a biotherapeutic has a soluble circulating target. While there are many different techniques that can be used to quantify drug and target concentrations in a biological matrix such as serum or plasma, ligand-binding assays (LBAs) such as ELISA are commonly used. Recently, there has been an increasing interest among bioanalytical and PK scientists in understanding how the design and format of an LBA can impact apparent drug and/or target concentration data, and what the potential impact of this may be on interpretation of PK/PD and safety data. This is especially the case with monoclonal antibody (mAb)-based therapeutics.Since the introduction of the first mAb-based drug, muromonab-CD3 (orthoclone OKT3) [1], over 20 mAb therapeutics have been approved by the US FDA and over 100 additional mAb-based therapeutics are currently under development [2]. Typically, a therapeutic mAb needs to bind to its target antigen in order to exert its effect. When the mAb and the target both circulate, various molecular species of mAb and target co-exist in a dynamic equilibrium that is based upon the law of mass action, first described by Guldberg and Waage in 1864. These molecular species include: free drug, free circulating target, total drug and total target. In situations where there is more than one target-binding region on the drug (e.g., a full-length bivalent mAb) or more than one drug-binding region on the target (e.g., VEGF and IgE), more molecular species, such as partially complexed drug–target, may also be present. These partially complexed species are often considered to be part of the free fraction of drug or target.It has long been acknowledged that data on different drug and target species (e.g., free vs total levels of drug target as two possible biomarkers) may satisfy different needs. Recently, there has been a high level of interest in this topic and its potential impact on drug development in the biopharmaceutical industry. This has partially been driven by bioanalytical scientists, who are increasingly involved in helping to create PK, PD and safety strategies during drug development. In addition, advances in analytical technologies have helped bioanalytical scientists to better understand the limits of their own methods, and in some cases, have enabled them to develop more specific assays that enable the measurement of either free or total molecular species.A discussion group on free versus total PK/PD assays was formed by the AAPS LBA Bioanalytical Focus Group (LBABFG) following a hot topic session on this subject at the AAPS National Biotechnology Conference in 2008. Since then, multiple scientific presentations on this topic have been given at many different conferences. In addition, a growing number of publications have appeared on this topic in recent years, including detailed case studies and review articles as well as a consensus-based 'AAPS White Paper' that resulted from discussions of the LBABFG [3,4]. However, due to the complexity of the issue, it may not be realistic to have a 'one-size fits-all' bioanalytical strategy for all PK/PD and safety evaluations. This partially results from the technical challenges in designing and developing assays that measure only one molecular species (e.g., free or bound or total therapeutic species). Assay data are easily confounded by sample handling, dilutions, reagent limitations and so on. In addition, the lack of information on what specific data are actually needed to inform PK/PD and safety evaluations for a particular program also poses another challenge in formulating bioanalytical strategies [5]. For example, when only one PK assay (free or total assay) can be developed to measure drug concentrations due to limited resources, data from a free PK assay may be more informative for one project, while data from a total assay may be more informative for another project. Going into a new biotherapeutic development program, we do not typically know a priori if there is likely to be a difference between data from a free versus a total biotherapeutic drug assay. This is because the information on the target concentrations, especially after drug treatment, is not available prior to the development of a bioanalytical assay [6]. From the overall bioanalytical assay strategy point of view, it is also important to consider the development stage of a particular program because bioanalytical data usage may differ for preclinical and clinical studies and in support of a second-generation drug. As a result, the development stage of a molecule may affect the overall bioanalytical assay strategy. Given limited resources, it is often challenging to know what assay may be the most appropriate for a particular drug-development program. Information in the literature is scant on such assay comparisons and on the impact of the analyte selected for analysis on PK/PD and safety evaluation conclusions, which makes it hard to generalize from one particular case study.It is therefore very helpful for end users of bioanalytical data such as PK scientists and toxicologists to work together with bioanalytical scientists to assess the needs, technical feasibility and challenges of developing suitable bioanalytical assays in order to form a sound bioanalytical strategy. In addition, such discussions will facilitate data interpretation and help our PK/PD and safety assessment colleagues understand the limitations/caveats that are associated with this type of bioanalytical data. Thus, bioanalytical support of drug development will become more context driven and bioanalytical data should enable more informed decision making. It is worth pointing out that although there have been some significant advances in our knowledge regarding free versus total LBAs, it is still every bioanalytical scientist's dream that one day, sensitive and specific assays that are 'immune' from various interferences (e.g., those caused by sample handling and reagent limitation) could be readily developed. This means that we will need to significantly improve (or disruptively innovate beyond) current bioanalytical technologies. This discussion on free versus total LBAs is also expected to motivate analytical technology vendors to work more closely with their drug-development customers. Finally, we hope that a constructive and robust dialog on this topic will help to foster a collaborative environment for drug-discovery and -development scientists, regulatory agencies and instrument/technology manufacturers. This should help us all to speak the same (bioanalytical) language, understand each others needs and challenges, and, ultimately, develop new, efficacious and safe drugs that benefit the society that we are all a part of.Financial & competing interests disclosureThe authors are employees of Genentech, a member of the Roche Group. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.No writing assistance was utilized in the production of this manuscript.Bibliography1 Smith SL. Ten years of orthoclone OKT3 (muromonab-CD3): a review. J. Transpl. Coord.6(3),109–119, quiz 120–121 (1996).Crossref, Medline, CAS, Google Scholar2 Wang W, Wang EQ, Balthasar JP. Monoclonal antibody pharmacokinetics and pharmacodynamics. Clin. Pharmacol. Ther.84(5),548–558 (2008).Crossref, Medline, CAS, Google Scholar3 Lee JW, Kelley M, King LE et al. Bioanalytical approaches to quantify 'total' and 'free' therapeutic antibodies and their targets: technical challenges and PK/PD applications over the course of drug development. AAPS J.13(1),99–110 (2011).Crossref, Medline, CAS, Google Scholar4 Kuang B, King L, Wang HF. Therapeutic monoclonal antibody concentration monitoring: free or total? Bioanalysis2(6),1125–1140 (2010).Link, CAS, Google Scholar5 Lowe PJ, Tannenbaum S, Gautier A, Jimenez P. Relationship between omalizumab pharmacokinetics, IgE pharmacodynamics and symptoms in patients with severe persistent allergic (IgE-mediated) asthma. Br. J. Clin. Pharmacol.68(1),61–76, (2009).Crossref, Medline, CAS, Google Scholar6 Stefanini MO, Wu FT, Mac Gabhann F, Popel AS. Increase of plasma VEGF after intravenous administration of bevacizumab is predicted by a pharmacokinetic model. Cancer Res.70(23),9886–9894 (2010).Crossref, Medline, CAS, Google ScholarFiguresReferencesRelatedDetailsCited ByBest Practices in mAb and Soluble Target Assay Selection for Quantitative Modelling and Qualitative Interpretation9 February 2023 | The AAPS Journal, Vol. 25, No. 1Development of an immunoassay for aglycosylated murine IgG1 in mouse serum via generation of a specific tool antibodyCarrie Poon-Andersen, Hamsini Dhoolypala, Diana Wong, Marcus Soto, Hossein Salimi-Moosavi, Brian Chan, Agnieszka Kielczewska & Kevin D Cook12 May 2022 | Bioanalysis, Vol. 14, No. 9Assay pH and critical reagent optimization in measuring concentrations of a monoclonal antibody and its targetJihua Chen, Kimberly Kendra, Stacey Shank, Ashique Rafique, George Ehrlich, Kuan-Ju Lin, Lisa DeStefano, Matthew D Andisik, Michael A Partridge, Albert Torri & Giane Sumner17 March 2022 | Bioanalysis, Vol. 14, No. 8Development of a Meso Scale Discovery ligand-binding assay for measurement of free (drug-unbound) target in nonhuman primate serumYun Liu, Ronald Robinson, Thao Ung, Chrysanthe Spais, Justin Schreiber, Jacquelyn Lyons, Jean Husten, Hussein Hallak & Thelma Angeles22 March 2021 | Bioanalysis, Vol. 13, No. 7Modeling Approaches to Characterize Target‐Mediated Pharmacokinetics and Pharmacodynamics for Therapeutic Proteins15 February 2019Ligand Binding Assays in the Regulated Bioanalytical Laboratory26 April 2017Ligand-Binding Assay Development: What Do You Want to Measure Versus What You Are Measuring?15 December 2015 | The AAPS Journal, Vol. 18, No. 2The integration of ligand binding and LC-MS-based assays into bioanalytical strategies for protein analysisYan J Zhang, Timothy V Olah & Jianing Zeng26 August 2014 | Bioanalysis, Vol. 6, No. 13Free analyte QC concept: a novel approach to prove correct quantification of free therapeutic protein drug/biomarker concentrationsRoland F Staack, Gregor Jordan, Uwe Dahl & Julia Heinrich25 February 2014 | Bioanalysis, Vol. 6, No. 4Bioanalysis of therapeutic peptides: Differentiating between total and anti-drug antibody bound drug using liquid chromatography–tandem mass spectrometry quantitationJournal of Chromatography A, Vol. 1316Minimizing target interference in PK immunoassays: new approaches for low-pH-sample treatmentMichael A Partridge, John Pham, Olena Dziadiv, Onson Luong, Ashique Rafique, Giane Sumner & Albert Torri1 August 2013 | Bioanalysis, Vol. 5, No. 15BiopharmaceuticalsThe Development and Implementation of LC/MS-Based Bioanalytical Methods for the Quantification of Protein Therapeutics in Drug Discovery7 May 2013Mathematical simulations for bioanalytical assay development: the (un-)necessity and (im-)possibility of free drug quantificationRoland F Staack, Gregor Jordan & Julia Heinrich7 March 2012 | Bioanalysis, Vol. 4, No. 4 Vol. 3, No. 11 STAY CONNECTED Metrics History Published online 7 June 2011 Published in print June 2011 Information© Future Science LtdKeywordsbiotherapeutic drug developmentELISAligand-binding assaymonoclonal antibodypharmacodynamicpharmacokineticsFinancial & competing interests disclosureThe authors are employees of Genentech, a member of the Roche Group. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.No writing assistance was utilized in the production of this manuscript.PDF download
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