Pathological crystallization of human immunoglobulins

Wang, Ying; Lomakin, Aleksey; Hideshima, Teru; Laubach, Jacob P.; Ogun, Olutayo; Richardson, Paul G.; Munshi, Nikhil C.; Anderson, Kenneth C.; Benedek, George B.

doi:10.1073/pnas.1211723109

Cited by 37 publications

(33 citation statements)

References 23 publications

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“…The gelation is reversible upon changing temperature and concentration. 6,18 The gel formed by one particular cryoglobulin antibody was proposed to be a kinetically arrested state that forms in a supersaturated solution at low temperature. If this model is correct for C4, variations in concentration, formulation pH, or rate of temperature change might result in aggregation or crystallization.…”

Section: Discussionmentioning

confidence: 99%

Resolving self-association of a therapeutic antibody by formulation optimization and molecular approaches

Casaz

Boucher²,

Wollacott

et al. 2014

mAbs

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(2014) Resolving selfassociation of a therapeutic antibody by formulation optimization and molecular approaches, mAbs, 6:6, 1533-1539, DOI: 10.4161/19420862.2014 To link to this article: https://doi.org/10. 4161/19420862.2014 A common challenge encountered during development of high concentration monoclonal antibody formulations is preventing self-association. Depending on the antibody and its formulation, self-association can be seen as aggregation, precipitation, opalescence or phase separation. Here we report on an unusual manifestation of selfassociation, formation of a semi-solid gel or "gelation." Therapeutic monoclonal antibody C4 was isolated from human B cells based on its strong potency in neutralizing bacterial toxin in animal models. The purified antibody possessed the unusual property of forming a firm, opaque white gel when it was formulated at concentrations >30 mg/mL and the temperature was <6 C. Gel formation was reversible with temperature. Gelation was affected by salt concentration or pH, suggesting an electrostatic interaction between IgG monomers. A comparison of the C4 amino acid sequences to consensus germline sequences revealed differences in framework regions. A C4 variant in which the framework sequence was restored to the consensus germline sequence did not gel at 100 mg/mL at temperatures as low as 1 C. Additional genetic analysis was used to predict the key residue(s) involved in the gelation. Strikingly, a single substitution in the native antibody, replacing heavy chain glutamate 23 with lysine (E23K), was sufficient to prevent gelation. These results indicate that the framework region is involved in intermolecular interactions. The temperature dependence of gelation may be related to conformational changes near glutamate 23 or the regions it interacts with. Molecular engineering of the framework can be an effective approach to resolve the solubility issues of therapeutic antibodies.

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Section: Discussionmentioning

confidence: 99%

Resolving self-association of a therapeutic antibody by formulation optimization and molecular approaches

Casaz

Boucher²,

Wollacott

et al. 2014

mAbs

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“…For instance, optimum pH range for cryoprecipitation, inhibitory pH range, critical concentration, temperature threshold, and the kinetics of precipitation can all vary from clone to clone even among cryoglobulins [177, 181–185]. There is one cryo-IgG clone that was shown to crystallize even at 37°C when the pH was shifted to a range between 5.0 and 6.5 [126].…”

Section: Roles Of Differential Physicochemical Properties In the Ementioning

confidence: 99%

Aggregates, Crystals, Gels, and Amyloids: Intracellular and Extracellular Phenotypes at the Crossroads of Immunoglobulin Physicochemical Property and Cell Physiology

Hasegawa

2013

International Journal of Cell Biology

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Recombinant immunoglobulins comprise an important class of human therapeutics. Although specific immunoglobulins can be purposefully raised against desired antigen targets by various methods, identifying an immunoglobulin clone that simultaneously possesses potent therapeutic activities and desirable manufacturing-related attributes often turns out to be challenging. The variable domains of individual immunoglobulins primarily define the unique antigen specificities and binding affinities inherent to each clone. The primary sequence of the variable domains also specifies the unique physicochemical properties that modulate various aspects of individual immunoglobulin life cycle, starting from the biosynthetic steps in the endoplasmic reticulum, secretory pathway trafficking, secretion, and the fate in the extracellular space and in the endosome-lysosome system. Because of the diverse repertoire of immunoglobulin physicochemical properties, some immunoglobulin clones' intrinsic properties may manifest as intriguing cellular phenotypes, unusual solution behaviors, and serious pathologic outcomes that are of scientific and clinical importance. To gain renewed insights into identifying manufacturable therapeutic antibodies, this paper catalogs important intracellular and extracellular phenotypes induced by various subsets of immunoglobulin clones occupying different niches of diverse physicochemical repertoire space. Both intrinsic and extrinsic factors that make certain immunoglobulin clones desirable or undesirable for large-scale manufacturing and therapeutic use are summarized.

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“…In type I, the monoclonal component undergoes crystallization and aggregation, which is dependent on temperature and concentration. 9 Although the definition of cryoglobulins is precipitation at cold temperatures, this process can occur at room temperature at high cryoglobulin concentrations. 2 This probably explains why distal extremities (lower temperatures) and kidneys (increase in concentration as a result of ultrafiltration) are major sites of pathology.…”

Section: Pathogenesis Cryoprecipitationmentioning

confidence: 99%

How I treat cryoglobulinemia

Muchtar

Magen

Gertz

2017

Blood

144

146

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Cryoglobulinemia is a distinct entity characterized by the presence of cryoglobulins in the serum. Cryoglobulins differ in their composition, which has an impact on the clinical presentation and the underlying disease that triggers cryoglobulin formation. Cryoglobulinemia is categorized into two main subgroups: type I, which is seen exclusively in clonal hematologic diseases, and type II/III, which is called mixed cryoglobulinemia and is seen in hepatitis C virus infection and systemic diseases such as B-cell lineage hematologic malignancies and connective tissue disorders. Clinical presentation is broad and varies between types but includes arthralgia, purpura, skin ulcers, glomerulonephritis, and peripheral neuropathy. Life-threatening manifestations can develop in a small proportion of patients. A full evaluation for the underlying cause is required, because each type requires a different kind of treatment, which should be tailored on the basis of disease severity, underlying disease, and prior therapies. Relapses can be frequent and can result in significant morbidity and cumulative organ impairment. We explore the spectrum of this heterogeneous disease by discussing the disease characteristics of 5 different patients.

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Pathological crystallization of human immunoglobulins

Cited by 37 publications

References 23 publications

Resolving self-association of a therapeutic antibody by formulation optimization and molecular approaches

Resolving self-association of a therapeutic antibody by formulation optimization and molecular approaches

Aggregates, Crystals, Gels, and Amyloids: Intracellular and Extracellular Phenotypes at the Crossroads of Immunoglobulin Physicochemical Property and Cell Physiology

How I treat cryoglobulinemia

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