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Glycoform-resolved pharmacokinetic studies in a rat model employing glycoengineered variants of a therapeutic monoclonal antibody.

ABSTRACT: Good pharmacokinetic (PK) behavior is a key prerequisite for sufficient efficacy of therapeutic monoclonal antibodies (mAbs). Fc glycosylation is a critical quality attribute (CQA) of mAbs, due to its impact on stability and effector functions. However, the effects of various IgG Fc glycoforms on antibody PK remain unclear. We used a combination of glycoengineering and glycoform-resolved PK measurements by mass spectrometry (MS) to assess glycoform effects on PK. Four differently glycoengineered mAbs, each still containing multiple glycoforms, were separately injected into rats. Rat models have been shown to be predictive of human PK. At different time points, blood was taken, from which the mAbs were purified and analyzed with a liquid chromatography-MS-based bottom-up glycoproteomics approach. This allowed us to follow changes in the glycosylation profiles of each glycoengineered mAb over time. Enzyme-linked immunosorbent assay measurements provided an absolute concentration in the form of a sum value for all glycoforms. Information from both readouts was then combined to calculate PK parameters per glycoform. Thereby, multiple glycoform kinetics were resolved within one mAb preparation. We confirmed increased clearance of high-mannose (Man5) and hybrid-type (Man5G0) glycoforms. Specifically, Man5 showed a 1.8 to 2.6-fold higher clearance than agalactosylated, complex glycans (G0F). Unexpectedly, clearance was even higher (4.7-fold) for the hybrid-type glycan Man5G0. In contrast, clearance of agalactosylated, monoantennary glycoforms (G0F-N) was only slightly increased over G0F (1.2 to 1.4-fold). Thus, monoantennary, hybrid-type and high-mannose glycoforms should be distinguished in CQA assessments. Strikingly, α2,3-linked sialylation did not affect clearance, contradicting the involvement of the asialoglycoprotein receptor in mAb clearance.

SUBMITTER: Falck D

PROVIDER: S-EPMC7781607 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Autoimmune diseases of the peripheral nervous system have so far been treated mainly with exogenous high-dose intravenous immunoglobulins (IVIg), that act through several mechanisms, including neutralization of pathogenic autoantibodies, modulation of lymphocyte activity, interference with antigen presentation, and interaction with Fc receptors, cytokines, and the complement system. Other therapeutic strategies have recently been developed, in part to address the increasing shortage of IVIg, prime among which is the use of B cell depleting monoclonal antibodies, or small molecule inhibitors targeting the B-cell specific kinases. Rituximab, a chimeric monoclonal antibody against CD20 + B lymphocytes, is currently the most used, especially in anti-MAG antibody neuropathy and autoimmune neuropathies with antibodies to nodal/paranodal antigens that are unresponsive to IVIg. After several reports of its efficacy in chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), rituximab is currently under investigation in three Phase 2 trials in CIDP. In addition, the possible role of complement activation in the pathogenesis of chronic autoimmune neuropathies has brought into consideration drugs that can block the complement cascade, such as eculizumab, a monoclonal antibody already assessed in acute polyradiculoneuropathies, and approved for myasthenia gravis. Preliminary data on eculizumab in multifocal motor neuropathy have been published, but randomized controlled studies are pending. Moreover, the neonatal Fc receptor, that recycles IgGs by preventing their lysosome degradation, is an important and attractive pharmacological target. Antibodies against FcRn, which reduce circulating IgG (both pathogenic and non-pathogenic) have been developed. The FcRn blocker efgartigimod, a humanized IgG1-derived Fc fragment, which competitively inhibits the FcRn, has recently been approved for the treatment of myasthenia gravis and is currently under investigation in CIDP. In addition, the anti-human FcRn monoclonal antibody rozanolixizumab is currently being assessed in phase 2 trials in CIDP. However, none of the abovementioned monoclonal antibodies is currently approved for treatment of any immune-mediated neuropathies. While more specific and individualized therapies are being developed, the possibility of combined treatments targeting different pathogenic mechanisms deserves consideration as well.

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Glycoform-resolved pharmacokinetic studies in a rat model employing glycoengineered variants of a therapeutic monoclonal antibody.

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