Achieving drug delivery across the blood-brain barrier is a major challenge in the development of drugs to treat diseases of the central nervous system (CNS), especially when using biological drugs such as monoclonal antibodies and enzyme replacement therapies.
Blood-Brain Barrier (BBB)
The blood-brain barrier (BBB) is a protective barrier between the blood vessels, cells, and other tissues that make up the brain, providing a defense mechanism for the brain against foreign pathogens and toxins in the bloodstream. The development of therapies for central nervous system (CNS) diseases is complicated by the presence of the BBB. While the BBB maintains CNS homeostasis by tightly controlling specific nutrients and restricting the passage of harmful xenobiotic molecules, the BBB also prevents drugs and large molecule therapies such as biologics from entering the brain, thereby greatly reducing their efficacy.
Over the past two decades, biologics such as monoclonal antibodies have become increasingly popular in the field of drug development. Seven of the top 10 best-selling drugs in the world in 2019 are biologics. However, due to the presence of BBB, biologics are present in the brain at a level equivalent to only 0.01-0.1% of plasma. There are no biologics for CNS diseases, and therefore efficient delivery technologies are needed to transport biologics to the CNS.
Biological Delivery Technologies Crossing the BBB
Biomolecules are transported across the blood-brain barrier by two routes: the uncontrolled nonspecific transport system and the controlled endogenous blood-brain barrier transport system. Uptake-mediated nonspecific transport greatly limits the therapeutic potential of therapeutic proteins and antibodies because their indiscriminate cellular uptake is not only a major reason in terms of off-target effects but may lead to poor pharmacokinetic properties. Exploring endogenous transport systems across the blood-brain barrier is more appropriate for CNS protein and antibody therapy. Receptor-mediated transcellular transport (RMT) and carrier-mediated transport (CMT) are the main endogenous transport systems for BBB. Substrate-selective CMT is responsible for the delivery of small-molecule nutrients, including glucose, amino acids, hormones, ions, vitamins, etc. RMT delivers larger molecules, such as transferrin, insulin, TNF-α, and epidermal growth factor, through vesicles of the ligand-receptor complex.
Most of the studies on delivering biomolecules have been on RMT receptors such as transferrin receptor (TfR), insulin receptor (InsR), CD98hc, low-density lipoprotein receptor-related protein 1 (LRP1), and folate receptor (FR). The most studied are TfR and InsR RMT receptors.
Transferrin Receptor (TfR)
There are two known transferrin receptors, TfR1 and TfR2. TfR1 has been extensively studied for protein and antibody therapeutics across the blood-brain barrier due to its high level of expression in brain endothelial cells (BEC). TfR is a type II transmembrane receptor, a homodimer linked by disulfide bonds on Cys89 and Cys98(97). TfR binds to iron through a helical structural domain and a protease-like structural domain to iron-bound Holo-Tf (containing iron). At neutral pH, iron binds tightly to transferrin, and the iron/transferrin complex is transported intracellularly via transferrin. In low-pH endosomes, iron is released from transferrin. TfR-associated apo-Tf (which does not contain iron) is transported back to the cell surface. Under neutral pH conditions, the low affinity of apo-Tf for TfR results in the release of apo-Tf. TfR is universally expressed, with erythrocytes and proliferating cells expressing it at high levels due to the metabolic demand for iron. BEC also expresses TfR. Transferrin is transported through the blood-brain barrier, suggesting that the TfR pathway can naturally transport macromolecules across the blood-brain barrier. Direct use of transferrin as a carrier is impractical for some reasons, including circulating endogenous transferrin concentrations as high as 3 mg/mL. Therefore, it is more feasible to use antibody-targeted TfR as a delivery vehicle across the blood-brain barrier. To avoid interference with the normal transferrin receptor function of iron/transferrin and apo-transferrin across cell membranes and the blood-brain barrier, antibodies should not compete with transferrin binding to the transferrin receptor. For example, vectors currently found to target TfR typically bind to the apical region of the TfR, which is far from where Tf binds TfR.
Insulin Receptor (InsR)
The insulin receptor is a single type I transmembrane receptor with two subunits (alpha and beta chains) connected by a disulfide bond. Insulin binds to two different sites on each subunit of the receptor, cross-linking the two receptors, and resulting in high affinity. Insulin receptor binding activates the catalytic structural domain of the intracellular tyrosine kinase, which then triggers a downstream signaling cascade that regulates various biological functions such as glucose uptake. Insulin binding to InsR can be internalized. Insulin receptors are highly expressed in BEC, and since insulin is indispensable for glucose metabolism in the brain, it transports insulin across the blood-brain barrier through an active, energy-dependent process. However, insulin is unsuitable as a transport carrier due to underlying carbohydrate metabolism dysfunction. Therefore, it is more feasible to use InsR antibody as a transport carrier across the blood-brain barrier.
InsR antibodies have been extensively studied as carriers for human insulin receptor (HIR)-mediated brain delivery. Antibodies should bind InsR noncompetitively with insulin.InsR antibodies are usually designed at the N-terminus enzymes or proteins are designed at the C-terminus, and the whole structure is a bivalent antibody-enzyme/protein fusion.
TfR and InsR have now been extensively studied as BBB delivery platforms, but they are not perfect RTM receptor systems because their broad peripheral distribution can lead to rapid clearance of the drug from the bloodstream, thereby limiting access to the brain. While strategies such as low-affinity TfR-conjugated antibodies and monovalent carrier design can minimize rapid clearance from the blood, higher and more frequent doses may also be required to compensate for clearance to achieve the desired therapeutic concentration in the brain. Therefore, there is a need to investigate new RMT receptors that need to be more specific to the brain.
References
- Tosi G.; et al. Nanoparticles as carriers for drug delivery of macromolecules across the blood-brain barrier. Expert Opinion on Drug Delivery. 2020, 17(1): 23-32.
- Patel M. M.; et al. Crossing the blood–brain barrier: recent advances in drug delivery to the brain. CNS Drugs. 2017, 31: 109-133.
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