Unexpected Thrombocytopenia and Anemia in Cynomolgus Monkeys Induced by a Therapeutic Human Monoclonal Antibody

Everds, Nancy E.; Li, Nianyu; Bailey, Keith; Fort, Madeline M.; Stevenson, Riki; Jawando, Remi; Salyers, Kevin L.; Jawa, Vibha; Narayanan, Padma K.; Stevens, Erin; He, Ching; Nguyen, Mai Phuong; Tran, Sam M; Doyle, Nancy; Poitout-Belissent, Florence; Jolette, Jacquelin; Xu, Cen; Sprugel, K. H.

doi:10.1177/0192623312474727

Cited by 22 publications

(12 citation statements)

References 39 publications

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“…Our results further support that some of these off-target effects require Fc-Fc␥R binding (10), and that the elimination of the effector function of a mAb may eliminate off-target reactions as well (as shown for mAbX and mAbY (9, 10)). The generation of a SEFL mAb construct offers a path forward for development of IgG1-based mAbs when cytotoxicity is not desired, by eliminating the Fc-mediated events.…”

Section: Discussionsupporting

confidence: 81%

“…Cynomolgus Monkey Platelet Phagocytosis-mAbY is an IgG2 mAb that has previously been shown to mediate non-target dependent phagocytosis of platelets by cynomolgus monocyte/ macrophages both in vivo and in vitro in a process requiring Fc function (10). In vitro, parental mAbY.IgG2 and IgG1 constructs, mAbY.IgG1 and mAbW.IgG1, caused phagocy- tosis of cynomolgus monkey platelets at concentrations Ն0.1 mg/ml (Fig.…”

Section: Journal Of Biological Chemistry 1877mentioning

confidence: 99%

“…In addition, IgG2 can also bind to Fc␥ receptors to induce non-cytotoxic effector functions. For example, IgG2 isotypes can mediate phagocytosis via monocytes/macrophages and neutrophils through interaction with the Fc␥RIIa on platelets (8), and binding to the Fc␥RIIa along with a cell surface antigen has been implicated in off-target effects of mAbs on platelets (9,10).…”

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confidence: 99%

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Biological Characterization of a Stable Effector Functionless (SEFL) Monoclonal Antibody Scaffold in Vitro

Liu¹,

Jacobsen²,

Everds

et al. 2017

Journal of Biological Chemistry

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Edited by Peter CresswellThe stable effector functionLess (SEFL) antibody was designed as an IgG1 antibody with a constant region that lacks the ability to interact with Fc␥ receptors. The engineering and stability and pharmacokinetic assessments of the SEFL scaffold is described in the accompanying article (Jacobsen, F. W., Stevenson, R., Li, C., Salimi-Moosavi, H., Liu, L., Wen, J., Luo, Q., Daris, K., Buck, L., Miller, S., Ho, S-Y., Wang, W., Chen, Q., Walker, K., Wypych, J., Narhi, L., and Gunasekaran, K. (2017) J. Biol. Chem. 292). The biological properties of these SEFL antibodies were assessed in a variety of human and cynomolgus monkey in vitro assays. Binding of parent molecules and their SEFL variants to human and cynomolgus monkey Fc␥Rs were evaluated using flow cytometry-based binding assays. The SEFL variants tested showed decreased binding affinity to human and cynomolgus Fc␥Rs compared with the wild-type IgG1 antibody. In addition, SEFL variants demonstrated no antibody-dependent cell-mediated cytotoxicity in vitro against Daudi cells with cynomolgus monkey peripheral blood mononuclear cells, and had minimal complement-dependent cytotoxicity activity similar to that of the negative control IgG2 in a CD20؉ human Raji lymphoma cell line. SEFL mutations eliminated off-target antibody-dependent monocyte phagocytosis of cynomolgus monkey platelets, and cynomolgus platelet activation in vitro. These experiments demonstrate that the SEFL modifications successfully eliminated Fc-associated effector binding and functions.Monoclonal antibodies (mAbs) are the largest class of biopharmaceuticals and have diverse clinical applications (1). The choice of therapeutic mAb isotype to develop (IgG1, IgG2, or IgG4) is dependent on the target (cell surface versus soluble), desired biology, safety (risk of immunogenicity and undesired immunological effects), and manufacturability (expression, formulation, and stability). More than 80% of approved therapeutic mAbs are IgG1 isotypes that target cell surface receptors and are effective for oncology indications (2). For these therapeutic approaches, mAb isotypes that can induce cell killing such as complement-dependent cytotoxicity (CDC) 2 and antibody-dependent cell-mediated cytotoxicity (ADCC) are often desirable (3, 4). However, for non-oncologic indications, therapeutic mAbs without cytotoxic effector function may be more appropriate because cell killing may not be a goal of therapy.The Fc portion of IgG has interaction sites for the effector ligands, including Fc␥ receptors (Fc␥RI, Fc␥RII, and Fc␥RIII), C1q complement, and the neonatal Fc receptor (FcRn). IgG isotypes differentially engage Fc␥ receptors and C1q binding to recruit immune effector functions and initiate cytotoxic effector functions, (either ADCC or CDC (5)). Historically, IgG2 or IgG4 isotypes were thought to have minimal cytotoxic effector function, and have been selected for applications where cytotoxic effector function is not required or desirable (5). However, recent evidence suggests that IgG2 is...

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Section: Discussionsupporting

confidence: 81%

Section: Journal Of Biological Chemistry 1877mentioning

confidence: 99%

mentioning

confidence: 99%

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Biological Characterization of a Stable Effector Functionless (SEFL) Monoclonal Antibody Scaffold in Vitro

Liu¹,

Jacobsen²,

Everds

et al. 2017

Journal of Biological Chemistry

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“…Subsequently it was shown that the functional differences between these antibodies had a structural basis: The unexpected toxicity was shown to be driven by one to three amino acid differences in the light chain. 27 Obviously, such structural differences are not within the scope of a biosimilar development. Although the observed toxicity came unexpectedly for the company developing the drug, it is not an example that could be applied to biosimilars when structural and functional similarity have already been shown by analytical and in vitro methods.…”

Section: Safety Evaluationmentioning

confidence: 99%

Biosimilars entering the clinic without animal studies

Aerts¹,

Smet²,

Reichmann

et al. 2014

mAbs

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“…Re-gating was used to successfully enumerate and classify RBCs and platelets. Study conditions causing erroneous results: A nonclinical safety study was conducted in male and female cynomolgus monkeys at doses of 10, 50, and 300 mg/kg subcutaneously and 300 mg/kg intravenously (Everds et al 2013). Animals dosed at !50 mg/kg had unexpected acute profound thrombocytopenia (nadir *3.0 Â 10 3 platelets/ml), sometimes with mild to marked decreases in red cell mass.…”

Section: Nancy Everds and David Zelmanovicmentioning

confidence: 99%

Nontraditional Applications in Clinical Pathology

Jordan

Tripathi

et al. 2014

Toxicol Pathol

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Most published reviews of preclinical toxicological clinical pathology focus on the fundamental aspects of hematology, clinical chemistry, coagulation, and urinalysis in routine toxicology animal species, for example, rats, mice, dogs, and nonhuman primates. The objective of this continuing education course was to present and discuss contemporary examples of nonroutine applications of clinical pathology endpoints used in the drug development setting. Area experts discussed bone turnover markers of laboratory animal species, clinical pathology of pregnant and growing laboratory animals, clinical pathology of nonroutine laboratory animal species, and unique applications of the Siemens Advia 1 hematology analyzer. This article is a summary based on a presentation given at the 31st Annual Symposium of the Society of Toxicologic Pathology, during the Continuing Education Course titled ''Nontraditional Applications of Clinical Pathology in Drug Discovery and Preclinical Toxicology.''

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Unexpected Thrombocytopenia and Anemia in Cynomolgus Monkeys Induced by a Therapeutic Human Monoclonal Antibody

Abstract: Cynomolgus monkeys dosed with a therapeutic monoclonal antibody (mAbY

Cited by 22 publications

References 39 publications

Biological Characterization of a Stable Effector Functionless (SEFL) Monoclonal Antibody Scaffold in Vitro

Biological Characterization of a Stable Effector Functionless (SEFL) Monoclonal Antibody Scaffold in Vitro

Biosimilars entering the clinic without animal studies

Nontraditional Applications in Clinical Pathology

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