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Advanced Neurology Brain bioavailability of targeted protein degraders

0.1% to 0.4% of their respective serum concentrations. 78-80 endocytosis regulates insulin transport across the BBB in
Confocal microscopy of the brain reveals that the neonatal the hypothalamus, while clathrin-mediated mechanisms
Fc receptor is the primary Fc receptor on brain capillaries. drive insulin binding in isolated brain microvessels.
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However, the transfer of IgG molecules from blood to the The major facilitator superfamily domain containing
brain is not mediated by the BBB. Instead, IgG molecules 2a protein in the endothelial cells of the BBB modulates
undergo asymmetrical transcytosis through Fc receptors the lipid composition of the plasma membrane, thus
in the brain-to-blood direction. 82 limiting the formation of caveolae vesicles and restricting
The molecular weight of TPDs generally ranges between transcytosis. 97,98
700 and 1000 Da, which is a significant limiting factor for Antibody conjugates are also being explored as a
their use in CNS indications. Despite this, several TPDs are means of achieving the desired CNS concentration of
in the advanced pre-clinical development stage for CNS piggyback molecules for intended therapeutic efficacy.
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indications, with a few listed in Table 2. Antibody variable domain binding has been used in RMT-
It is important to note that for some of the TPDs based delivery techniques to target brain endothelial cell
mentioned in the table above, the reported plasma and receptors. 100-103 Some BBB receptors facilitate endocytosis
brain concentrations were so disparate (double-digit µM into endothelial cells but do not mediate transcytosis
versus sub-nM, respectively) that it potentially indicates across the endothelial barrier.
vascular contamination rather than meaningful BBB The surface of brain endothelial cells contains
permeation. Estimating the vascular-restricted atenolol approximately 10% of the total receptor pool for the
could help rule out the possibility of blood contamination. recycling receptor TfR. Anti-TfR antibodies have been
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In addition, washout or perfusion technique could be shown to traverse the BBB. 105-107 These studies suggest that
employed to flush out residual drug within the vascular if drugs intended for CNS indications are tagged with TfR
volume. ligands, such as antibody fragments, their brain delivery is
Macromolecules, such as proteins, can enter the brain enhanced. 108-110
through vesicular transport or transcytosis. However, To harness RMT, protein therapies have been designed
this process is highly selective and is actively repressed by to bind these receptors, which are abundantly expressed on
recently discovered homeostatic mechanisms. It has been brain endothelial cells, thereby delivering the therapeutic
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reported that transcytosis of macromolecules primarily payload to the CNS. A few examples of such therapies are
occurs through endocytic mechanisms, including receptor- listed in Table 3.
mediated transcytosis (RMT), adsorptive-mediated
transcytosis (AMT), or carrier-mediated transport RMT of biotherapeutics conjugated to a receptor-
(CMT). 89 binding ligand has shown promise, although the net
quantity transported remains relatively low. The same
In RMT, ligands bind to specific receptors, such as applies to drugs conjugated to Angiopep-2 and activatable
the transferrin receptor (TfR), insulin receptor, or low- cell-penetrating peptides. Antibodies or endogenous
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density lipoprotein receptor-related protein 1, on the protein receptor ligands have been shown to be transported
luminal surface of endothelial cells. This binding triggers across endothelium at the capillary level. Brain vasculature
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an endocytic cascade, followed by vesicle-mediated hemodynamics vary across arteries, arterioles, capillaries,
transport across the BBB. These “Trojan horses” transport venules, and veins, 110,112 with the TfRs being most abundant
large biomolecules such as insulin (5734 Da), transferrin in capillaries. It is therefore conceivable that the slowest
(80,000 Da), insulin-like growth factor 1 (7,649 Da), or blood velocity in capillaries could prolong the time TfR
antidiuretic hormone (1,084.24 Da). Lipoparticles or substrates interact with the endothelial surface, thereby
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iron-bound transferrin bind to specific receptors on brain increasing the likelihood of internalization. However, it is
endothelial cells and are transported across the BBB. important to note that red blood cells (RBCs) also express
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RMT at the BBB endothelium primarily occurs at clathrin-
coated pits, with a marginal contribution from caveolae. TfR1, and the high affinity of TfR1 may lead to drug
accumulation in RBCs, potentially causing hemolysis in
The BBB endothelium contains a large number of clathrin- some instances.
coated pits, most of which are located on the luminal
membrane, indicating that clathrin-dependent transport Adsorption-mediated transcytosis is a saturable
primarily occurs in the blood-to-brain direction. 92,93 A mechanism of transport. 94,113 AMT requires the molecule
well-known example of clathrin-dependent endocytosis to be positively charged at physiological pH 7.4 for
is the transfer of iron across the BBB, which is mediated electrostatic interactions with cell surface binding sites.
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by transferrin and its receptor. 94,95 Caveolin-mediated The positive charge is necessary due to the highly charged

Volume 4 Issue 2 (2025) 60 doi: 10.36922/an.5140