Abstract
Antibody–drug conjugates (ADCs) combine the characteristics of large-molecule biologics and small-molecule drugs. Although a number of assays have been reported to characterize the ADCs, new and comprehensive bioanalytical strategy to overcome the limitations of the current methods is required for better characterization of ADCs for clinical use. An appropriate bioanalytical assay is essential for characterization of ADCs including physiochemical and pharmacokinetic properties due to their complex and heterogeneous structures. The commonly used bioanalytical methods that are typically implemented to characterize various ADCs are summarized in this chapter. The challenges and perspectives of the assays are also discussed.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Advani A, Coiffier B, Czuczman MS, Dreyling M, Foran J, Gine E et al (2010) Safety, pharmacokinetics, and preliminary clinical activity of inotuzumab ozogamicin, a novel immunoconjugate for the treatment of B-cell non-Hodgkin’s lymphoma: results of a phase I study. J Clin Oncol 28(12):2085–2093. doi:10.1200/JCO.2009.25.1900
Alley SC, Anderson KE (2013) Analytical and bioanalytical technologies for characterizing antibody–drug conjugates. Curr Opin Chem Biol 17(3):406–411. doi:http://dx.doi.org/10.1016/j.cbpa.2013.03.022
Alley SC, Benjamin DR, Jeffrey SC, Okeley NM, Meyer DL, Sanderson RJ et al (2008) Contribution of linker stability to the activities of anticancer immunoconjugates. Bioconjug Chem 19(3):759–765. doi:10.1021/bc7004329
Beck A (2014) Review of antibody-drug conjugates. Methods in molecular biology series: a book edited by Laurent Ducry. mAbs 6(1):30–33
Boghaert ER, Khandke KM, Sridharan L, Dougher M, DiJoseph JF, Kunz A et al (2008) Determination of pharmacokinetic values of calicheamicin-antibody conjugates in mice by plasmon resonance analysis of small (5 microl) blood samples. Cancer Chemother Pharmacol 61(6):1027–1035. doi:10.1007/s00280-007-0560-2
Buckwalter M, Dowell JA, Korth-Bradley J, Gorovits B, Mayer PR (2004) Pharmacokinetics of gemtuzumab ozogamicin as a single-agent treatment of pediatric patients with refractory or relapsed acute myeloid leukemia. J Clin Pharmacol 44(8):873–880. doi:10.1177/0091270004267595
Carter PJ, Senter PD (2008) Antibody-drug conjugates for cancer therapy. Cancer J 14(3):154–169. doi:10.1097/PPO.0b013e318172d704
Chari RV, Martell BA, Gross JL, Cook SB, Shah SA, Blattler WA et al (1992) Immunoconjugates containing novel maytansinoids: promising anticancer drugs. Cancer Res 52(1):127–131
Chen Y (2013) Drug-to-Antibody Ratio (DAR) by UV/Vis spectroscopy. In: Ducry L (ed) Antibody-drug conjugates. Humana Press, New York City, NY, USA, pp 267–273
Chen Y, Clark S, Wong T, Dennis MS, Luis E, Zhong F et al (2007) Armed antibodies targeting the mucin repeats of the ovarian cancer antigen, MUC16, are highly efficacious in animal tumor models. Cancer Res 67(10):4924–4932. doi:10.1158/0008-5472.CAN-06-4512
Clark T, Han X, King L, Barletta F (2013) Insights into antibody–drug conjugates: bioanalysis and biomeasures in discovery. Bioanalysis 5(9):985–987. doi:10.4155/bio.13.65
Cordoba AJ, Shyong BJ, Breen D, Harris RJ (2005) Non-enzymatic hinge region fragmentation of antibodies in solution. J chromatogr B, Analyt Technol Biomed Life Sci 818(2):115–121. doi:10.1016/j.jchromb.2004.12.033
Dere R, Yi JH, Lei C, Saad OM, Huang C, Li Y et al (2013) PK assays for antibody-drug conjugates: case study with ado-trastuzumab emtansine. Bioanalysis 5(9):1025–1040. doi:10.4155/bio.13.72
DeSilva B, Smith W, Weiner R, Kelley M, Smolec J, Lee B et al (2003) Recommendations for the bioanalytical method validation of ligand-binding assays to support pharmacokinetic assessments of macromolecules. Pharm Res 20(11):1885–1900
DiJoseph JF, Armellino DC, Boghaert ER, Khandke K, Dougher MM, Sridharan L et al (2004) Antibody-targeted chemotherapy with CMC-544: a CD22-targeted immunoconjugate of calicheamicin for the treatment of B-lymphoid malignancies. Blood 103(5):1807–1814. doi:10.1182/blood -2003-07-2466
Dowell JA, Korth-Bradley J, Liu H, King SP, Berger MS (2001) Pharmacokinetics of gemtuzumab ozogamicin, an antibody-targeted chemotherapy agent for the treatment of patients with acute myeloid leukemia in first relapse. J Clin Pharmacol 41(11):1206–1214
Francisco JA, Cerveny CG, Meyer DL, Mixan BJ, Klussman K, Chace DF et al (2003) cAC10-vcMMAE, an anti-CD30–monomethyl auristatin E conjugate with potent and selective antitumor activity. Blood 102(4):1458–1465. doi:10.1182/blood-2003-01-0039
Gorovits B, Alley SC, Bilic S, Booth B, Kaur S, Oldfield P et al (2013) Bioanalysis of antibody-drug conjugates: American association of pharmaceutical scientists antibody-drug conjugate working group position paper. Bioanalysis 5(9):997–1006. doi:10.4155/bio.13.38
Hamblett KJ, Senter PD, Chace DF, Sun MM, Lenox J, Cerveny CG et al (2004) Effects of drug loading on the antitumor activity of a monoclonal antibody drug conjugate. Clin Cancer Res 10(20):7063–7070. doi:10.1158/1078-0432.CCR-04-0789
Hinman LM, Hamann PR, Wallace R, Menendez AT, Durr FE, Upeslacis J (1993) Preparation and characterization of monoclonal antibody conjugates of the calicheamicins: a novel and potent family of antitumor antibiotics. Cancer Res 53(14):3336–3342
Junutula JR, Raab H, Clark S, Bhakta S, Leipold DD, Weir S et al (2008) Site-specific conjugation of a cytotoxic drug to an antibody improves the therapeutic index. Nature Biotechnol 26(8):925–932. doi:10.1038/nbt.1480
Junutula JR, Flagella KM, Graham RA, Parsons KL, Ha E, Raab H et al (2010) Engineered thio-trastuzumab-dm1 conjugate with an improved therapeutic index to target human epidermal growth factor receptor 2–positive breast cancer. Clin Cancer Res 16(19):4769–4778. doi:10.1158/1078-0432.ccr-10-0987
Kaur S, Xu K, Saad OM, Dere RC, Carrasco-Triguero M (2013) Bioanalytical assay strategies for the development of antibody-drug conjugate biotherapeutics. Bioanalysis 5(2):201–226. doi:10.4155/bio.12.299
King HD, Dubowchik GM, Mastalerz H, Willner D, Hofstead SJ, Firestone RA et al (2002) Monoclonal antibody conjugates of doxorubicin prepared with branched peptide linkers: inhibition of aggregation by methoxytriethyleneglycol chains. J Med Chem 45(19):4336–4343
Kovtun YV, Audette CA, Ye Y, Xie H, Ruberti MF, Phinney SJ et al (2006) Antibody-drug conjugates designed to eradicate tumors with homogeneous and heterogeneous expression of the target antigen. Cancer Res 66(6):3214–3221. doi:10.1158/0008-5472.CAN-05-3973
Kozak K, Raab H (2013) Assay methodologies and challenges. In: Phillips GL (ed) Antibody-drug conjugates and immunotoxins: Springer, New York, pp 41–56
Lazar AC, Wang L, Blättler WA, Amphlett G, Lambert JM, Zhang W (2005) Analysis of the composition of immunoconjugates using size-exclusion chromatography coupled to mass spectrometry. Rapid Commun Mass Spectrom 19(13):1806–1814. doi:10.1002/rcm.1987
Lewis Phillips GD, Li G, Dugger DL, Crocker LM, Parsons KL, Mai E et al (2008) Targeting HER2-positive breast cancer with trastuzumab-DM1, an antibody-cytotoxic drug conjugate. Cancer Res 68(22):9280–9290. doi:10.1158/0008-5472.CAN-08-1776
Luckey JA, Drossman H, Kostichka T, Smith LM (1993) High-speed DNA sequencing by capillary gel electrophoresis. In: Ray W (ed) Methods in enzymology. Academic Press, San Diego, CA, USA, pp 154–172
McDonagh CF, Turcott E, Westendorf L, Webster JB, Alley SC, Kim K et al (2006) Engineered antibody–drug conjugates with defined sites and stoichiometries of drug attachment. Protein Eng Des Sel 19(7):299–307. doi:10.1093/protein/gzl013
Ouyang J (2013) Drug-to-Antibody Ratio (DAR) and drug load distribution by hydrophobic interaction chromatography and reversed phase high-performance liquid chromatography. In: Ducry L (ed) Antibody-drug conjugates. Humana Press, New York City, NY, USA, pp 275–283
Polakis P (2005) Arming antibodies for cancer therapy. Curr Opin Pharmacol 5(4):382–387. doi:10.1016/j.coph.2005.04.008
Pollack VA, Alvarez E, Tse KF, Torgov MY, Xie S, Shenoy SG et al (2007) Treatment parameters modulating regression of human melanoma xenografts by an antibody-drug conjugate (CR011-vcMMAE) targeting GPNMB. Cancer Chemother Pharmacol 60(3):423–435. doi:10.1007/s00280-007-0490-z
Quiles S, Raisch KP, Sanford LL, Bonner JA, Safavy A (2009) Synthesis and preliminary biological evaluation of high-drug-load paclitaxel-antibody conjugates for tumor-targeted chemotherapy. J Med Chem 53(2):586–594. doi:10.1021/jm900899 g
Ramakrishnan NA, Drescher MJ, Sheikhali SA, Khan KM, Hatfield JS, Dickson MJ et al (2006) Molecular identification of an N-type Ca2+ channel in saccular hair cells. Neuroscience 139(4):1417–1434. doi:http://dx.doi.org/10.1016/j.neuroscience.2006.01.064
Safavy A, Bonner JA, Waksal HW, Buchsbaum DJ, Gillespie GY, Khazaeli MB et al (2003) Synthesis and biological evaluation of paclitaxel-C225 conjugate as a model for targeted drug delivery. Bioconjug Chem 14(2):302–310. doi:10.1021/bc020033z
Sanderson RJ, Hering MA, James SF, Sun MM, Doronina SO, Siadak AW et al (2005) In vivo drug-linker stability of an anti-CD30 dipeptide-linked auristatin immunoconjugate. Clin Cancer Res 11(2 Pt 1):843–852
Sapra P, Stein R, Pickett J, Qu Z, Govindan SV, Cardillo TM et al (2005) Anti-CD74 antibody-doxorubicin conjugate, IMMU-110, in a human multiple myeloma xenograft and in monkeys. Clin Cancer Res 11(14):5257–5264. doi:10.1158/1078-0432.CCR-05-0204
Schuck P (1997) Use of surface plasmon resonance to probe the equilibrium and dynamic aspects of interactions between biological macromolecules1. Annu Rev Biophys Biomol Struct 26(1):541–566. doi:10.1146/annurev.biophys.26.1.541
Shen BQ, Xu K, Liu L, Raab H, Bhakta S, Kenrick M et al (2012) Conjugation site modulates the in vivo stability and therapeutic activity of antibody-drug conjugates. Nature Biotechnol 30(2):184–189. doi:10.1038/nbt.2108
Siegel MM, Tabei K, Kunz A, Hollander IJ, Hamann RR, Bell DH et al (1997) Calicheamicin derivatives conjugated to monoclonal antibodies: determination of loading values and distributions by infrared and UV matrix-assisted laser desorption/ionization mass spectrometry and electrospray ionization mass spectrometry. Anal Chem 69(14):2716–2726
Stephan JP, Chan P, Lee C, Nelson C, Elliott JM, Bechtel C et al (2008) Anti-CD22-MCC-DM1 and MC-MMAF conjugates: impact of assay format on pharmacokinetic parameters determination. Bioconjug Chem 19(8):1673–1683. doi:10.1021/bc800059t
Stephan JP, Kozak KR, Wong WL (2011) Challenges in developing bioanalytical assays for characterization of antibody-drug conjugates. Bioanalysis 3(6):677–700. doi:10.4155/bio.11.30
Sun MMC, Beam KS, Cerveny CG, Hamblett KJ, Blackmore RS, Torgov MY et al (2005) Reduction–alkylation strategies for the modification of specific monoclonal antibody disulfides. Bioconjug Chem 16(5):1282–1290. doi:10.1021/bc050201y
Teicher BA, Chari RV (2011) Antibody conjugate therapeutics: challenges and potential. Clin Cancer Res 17(20):6389–6397. doi:10.1158/1078-0432.CCR-11-1417
Tolcher AW, Ochoa L, Hammond LA, Patnaik A, Edwards T, Takimoto C et al (2003) Cantuzumab mertansine, a maytansinoid immunoconjugate directed to the CanAg antigen: a phase I, pharmacokinetic, and biologic correlative study. J Clin Oncol 21(2):211–222
US FDA (2013) Guidance for industry: bioanalytical method validation. U.S. Food and Drug Administration, Washington, DC
Vater CA, Reid K, Bartle LM, Goldmacher VS (1995) Characterization of antibody binding to cell surface antigens using a plasma membrane-bound plate assay. Anal Biochem 224(1):39–50. doi:http://dx.doi.org/10.1006/abio.1995.1006
Wakankar A, Chen Y, Gokarn Y, Jacobson FS (2011) Analytical methods for physicochemical characterization of antibody drug conjugates. mAbs 3(2):161–172
Wang L, Amphlett G, Blättler WA, Lambert JM, Zhang W (2005) Structural characterization of the maytansinoid–monoclonal antibody immunoconjugate, huN901-DM1, by mass spectrometry. Protein Sci 14(9):2436–2446. doi:10.1110/ps.051478705
Xie H, Audette C, Hoffee M, Lambert JM, Blattler WA (2004) Pharmacokinetics and biodistribution of the antitumor immunoconjugate, cantuzumab mertansine (huC242-DM1), and its two components in mice. J Pharmacol Exp Therap 308(3):1073–1082. doi:10.1124/jpet.103.060533
Xu K, Liu L, Saad OM, Baudys J, Williams L, Leipold D et al (2011) Characterization of intact antibody–drug conjugates from plasma/serum in vivo by affinity capture capillary liquid chromatography–mass spectrometry. Anal Biochem 412(1):56–66. doi:http://dx.doi.org/10.1016/j.ab.2011.01.004
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 American Association of Pharmaceutical Scientists
About this chapter
Cite this chapter
Xie, C., Wang, Z. (2015). Bioanalytical Assay for Characterization of Antibody-Drug Conjugates (ADCs). In: Wang, J., Shen, WC., Zaro, J. (eds) Antibody-Drug Conjugates. AAPS Advances in the Pharmaceutical Sciences Series, vol 17. Springer, Cham. https://doi.org/10.1007/978-3-319-13081-1_6
Download citation
DOI: https://doi.org/10.1007/978-3-319-13081-1_6
Published:
Publisher Name: Springer, Cham
Print ISBN: 978-3-319-13080-4
Online ISBN: 978-3-319-13081-1
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)