Protein engineering approaches for regulating blood–brain barrier transcytosis

Goulatis, Loukas I.; Shusta, Eric V.

doi:10.1016/j.sbi.2016.12.005

Cited by 56 publications

(53 citation statements)

References 49 publications

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“…Interestingly, comparing the permeabilities of the sdAb and larger macromolecules revealed a marginal, but significantly higher, permeability for the sdAb (Table 1). Enhanced permeability of certain sdAbs across the BBB in vitro has been reported previously and was attributed to highly positively charged isoelectric points (pI>9) 35 , which is hypothesized to foster adsorptive interactions with the negatively charged luminal surface of the BBB to facilitate transcytosis 7 . We performed isoelectric focusing (IEF) of the sdAb as well as 10-and 155-kDa dextrans to explore whether there were disparities in charge that might explain their enhanced permeability.…”

Section: Igg Transcytosis Across Ibecs Is Not Receptor-mediated Givementioning

confidence: 71%

Antibody transcytosis across brain endothelial-like cells occurs nonspecifically and independent of FcRn

Ruano-Salguero

Lee

2020

Sci Rep

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The blood-brain barrier (BBB) hinders the brain delivery of therapeutic immunoglobulin γ (igG) antibodies. Evidence suggests that IgG-specific processing occurs within the endothelium of the BBB, but any influence on transcytosis remains unclear. Here, involvement of the neonatal Fc receptor (FcRn), which mediates IgG recycling and transcytosis in peripheral endothelium, was investigated by evaluating the transcytosis of IgGs with native or reduced FcRn engagement across human induced pluripotent stem cell-derived brain endothelial-like cells. Despite differential trafficking, the permeability of all tested IgGs were comparable and remained constant irrespective of concentration or competition with excess IgG, suggesting IgG transcytosis occurs nonspecifically and originates from fluid-phase endocytosis. Comparison with the receptor-enhanced permeability of transferrin indicates that the phenomena observed for IgG is ubiquitous for most macromolecules. However, increased permeability was observed for macromolecules with biophysical properties known to engage alternative endocytosis mechanisms, highlighting the importance of biophysical characterizations in assessing transcytosis mechanisms.The brain endothelial cells (BECs) that form the main structural component of the blood-brain barrier (BBB) are central to the protection of brain parenchyma. Unique from peripheral endothelium, BECs exhibit substantially reduced permeability of most bloodborne molecules 1 . Accordingly, this restrictive physiology also poses a formidable obstacle in the brain delivery of therapeutic molecules 2 . In particular, conventional immunoglobulin γ (IgG) antibody-based passive immunotherapies, which are structurally and functionally similar to endogenous IgGs, only demonstrate brain uptake of 0.1-0.3% of the injected dose 3,4 . Despite the need to improve the brain delivery of conventional therapeutic IgGs 5 , progress is hindered by the current lack of understanding regarding their interactions with BECs.IgG-endothelium interactions in the periphery are dominated by the neonatal fragment crystallizable (Fc) receptor (FcRn). Following internalization of circulating IgG, the acidic microenvironment of endosomal compartments enables FcRn to bind and recycle IgG back to the lumen in a pH-dependent manner 6 . FcRn-mediated recycling therefore limits lysosomal degradation and contributes to the extended serum half-life of IgGs. However, FcRn can also mediate the abluminal transcytosis of IgG, which is exemplified in the transfer of maternal IgG across the placental endothelium. In this regard, FcRn is a unique transcytosis receptor, e.g. compared to the transferrin receptor (TfR) 7 , as it can shuttle its ligand bidirectionally to either cell surface (i.e. luminal recycling or abluminal transcytosis) 8 . Traditional transcytosis pathways are categorized as fluid-phase (e.g. macropinocytosis) or adsorptive-mediated, which occur via specific (e.g. receptor-mediated transcytosis (RMT)) or nonspecific (e.g. electrostatic adsorption) processes....

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Section: Igg Transcytosis Across Ibecs Is Not Receptor-mediated Givementioning

confidence: 71%

Antibody transcytosis across brain endothelial-like cells occurs nonspecifically and independent of FcRn

Ruano-Salguero

Lee

2020

Sci Rep

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“…However, a discussion of the mechanisms of transcytosis across this specialized endothelial bed is beyond the scope of this review. The reader is referred to a recent authoritative review on this subject …”

Section: Epithelial Transcytosis—immunoglobulins Lead the Waymentioning

confidence: 99%

Transcellular vesicular transport in epithelial and endothelial cells: Challenges and opportunities

Fung

Fairn

Lee

2017

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Vesicle‐mediated transcellular transport or simply “transcytosis” is a cellular process used to shuttle macromolecules such as lipoproteins, antibodies, and albumin from one surface of a polarized cell to the other. This mechanism is in contrast to the transit of small molecules such as anions, cations and amino acids that occur via uptake, diffusion through the cytosol and release and is also distinct from paracellular leak between cells. Importantly, transcytosis has evolved as a process to selectively move macromolecules between 2 neighboring yet unique microenvironments within a multicellular organism. Examples include the movement of lipoproteins out of the circulatory system and into tissues and the delivery of immunoglobulins to mucosal surfaces. Regardless of whether the transport is conducted by endothelial or epithelial cells, the process often involves receptor‐mediated uptake of a ligand into an endocytic vesicle, regulated transit of the carrier through the cytoplasm and release of the cargo via an exocytic event. While transcytosis has been examined in detail in epithelial cells, for both historical and technical reasons, the process is less understood in endothelial cells. Here, we spotlight aspects of epithelial transcytosis including recent findings and review the comparative dearth of knowledge regarding the process in endothelial cells highlighting the opportunity for further study.

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“…5 These systems, while mediating transport across the BBB, are somewhat inefficient and nonspecific, and can result in deleterious off-target effects. [6][7][8] While it is possible to mitigate these effects by antibody engineering strategies, 9 there remains a significant need for the discovery of new BBB RMT-targeting antibody pairs that may address these challenges. Several approaches have been implemented to identify new antibody-RMT pairs.…”

Section: Introductionmentioning

confidence: 99%

Antibody screening using a human iPSC‐based blood‐brain barrier model identifies antibodies that accumulate in the CNS

et al. 2020

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Drug delivery across the blood‐brain barrier (BBB) remains a significant obstacle for the development of neurological disease therapies. The low penetration of blood‐borne therapeutics into the brain can oftentimes be attributed to the restrictive nature of the brain microvascular endothelial cells (BMECs) that comprise the BBB. One strategy beginning to be successfully leveraged is the use of endogenous receptor‐mediated transcytosis (RMT) systems as a means to shuttle a targeted therapeutic into the brain. Limitations of known RMT targets and their cognate targeting reagents include brain specificity, brain uptake levels, and off‐target effects, driving the search for new and potentially improved brain targeting reagent‐RMT pairs. To this end, we deployed human‐induced pluripotent stem cell (iPSC)‐derived BMEC‐like cells as a model BBB substrate on which to mine for new RMT‐targeting antibody pairs. A nonimmune, human single‐chain variable fragment (scFv) phage display library was screened for binding, internalization, and transcytosis across iPSC‐derived BMECs. Lead candidates exhibited binding and internalization into BMECs as well as binding to both human and mouse BBB in brain tissue sections. Antibodies targeted the murine BBB after intravenous administration with one particular clone, 46.1‐scFv, exhibiting a 26‐fold increase in brain accumulation (8.1 nM). Moreover, clone 46.1‐scFv was found to associate with postvascular, parenchymal cells, indicating its successful receptor‐mediated transport across the BBB. Such a new BBB targeting ligand could enhance the transport of therapeutic molecules into the brain.

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Protein engineering approaches for regulating blood–brain barrier transcytosis

Cited by 56 publications

References 49 publications

Antibody transcytosis across brain endothelial-like cells occurs nonspecifically and independent of FcRn

Antibody transcytosis across brain endothelial-like cells occurs nonspecifically and independent of FcRn

Transcellular vesicular transport in epithelial and endothelial cells: Challenges and opportunities

Antibody screening using a human iPSC‐based blood‐brain barrier model identifies antibodies that accumulate in the CNS

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