Use of multicellular human liver models to investigate clearance of therapeutic antibodies

James, Bethany Hope ORCID: 0009-0005-1937-6184 (2024). Use of multicellular human liver models to investigate clearance of therapeutic antibodies. University of Birmingham. Ph.D.

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Abstract

Background and Aims: Therapeutic antibodies make up the majority share of the drug industry, estimated to be worth $495 billion by 2030. Often, inadequate understanding of the tissue distribution and pharmacokinetics has resulted in many biotherapeutics being unable to attain clinical success. The liver plays a crucial role in the internalisation and catabolism of these antibodies due to their vast vasculature system and high expression of scavenger and Fc-receptors. Liver sinusoidal endothelial cells (LSEC) express many of these receptors and are known to contribute in the clearance of immune complexes. The liver is often susceptible to injury and disease, possibly leading to changes in key receptors involved in antibody clearance. Therefore, we wanted to explore the role of LSEC in health and disease and assess how they impact upon non-specific clearance of therapeutic antibodies.

Method: LSEC were isolated from diseased and normal human liver tissue to ascertain changes in receptor expression by qPCR, western blotting, and immunohistochemistry. A novel recycling assay was adapted to quantify the binding and clearance of 11 therapeutic antibodies at 4- and 8-hours by ELISA, flow cytometry and confocal microscopy. Pharmaceutical inhibitors were utilised to interrogate the internalisation mechanism of both healthy and cirrhotic LSEC and colocalisation with lysosomal markers were also performed to establish the end fate of these therapeutics.

Results and Conclusions: Our results confirm that LSEC express an extensive portfolio of receptors able to bind biotherapeutics, and the distribution of these receptors are significantly altered in disease. We have also demonstrated that LSEC not only internalise antibodies, but they can also partition these for lysosomal degradation. The rate at which this happens differs between healthy and diseased cells. Most importantly, we describe how cirrhotic LSEC favour internalisation via a receptor-mediated clathrin-dependent pathway, whereas healthy cells favour a clathrin-independent pinocytic mechanism. Therefore, we conclude cirrhosis alters pharmacokinetics and that the role of LSEC (both in health and disease) should be considered during antibody development.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):