The blood-brain barrier plays a vital role in ensuring that the network of cells that surrounds the blood vessels in the brain maintain function by limiting the entry of harmful substances, such as toxins, pathogens and large molecules – all while still allowing the necessary nutrients to pass through. In practical terms, this means the vast majority of blood vessels in the brain have a different structure to those found in other areas of the body. The endothelial cells that line every blood vessel are fused together to form tight junctions that prevent entry to the aforementioned invaders.

Embedded around the surface of the blood vessel in what’s known as the basal lamina are chemical detector cells called pericytes, which pick up on anything in the bloodstream that shouldn’t be there and contract to prevent entry so that nearby macrophages can dispose of it. The final element is the end-feet of astrocytes – specialised glial cells that supply nutrients from blood vessels to neurons – which wrap around the entire blood vessel and control the defensive functions of the other cells mentioned. Despite the crucial function the barrier provides, it also prevents the delivery of the majority of drugs to the brain.

“It is estimated that only 2% of developed drugs are able to cross the blood-brain barrier, and for this reason, the delivery of drugs to the brain is an unmet clinical need,” says Vera Neves, whose work centres around finding drug delivery systems to bypass the barrier. Research teams have been searching for ways to overcome the blood-brain barrier for some time, but with advances in the field of peptides – short chains of amino acids that serve as building blocks for proteins – some believe we can use ‘peptide shuttles’ to sneak active pharmaceutical ingredients past it, effectively tricking the cells that form the blood-brain barrier into letting them use the same pathway that ferries nutrients to where they’re needed.

Peptide shuttles

Proponents of peptide shuttles believe they could be used to improve the treatment of a wide range of neurological conditions, such as Alzheimer’s, Parkinson’s and brain tumours, by facilitating the delivery of therapeutic agents to the brain. Neves explains that peptides serve an important role in transporting drugs across the blood-brain barrier. “In simple terms, it’s like having a key that recognises a blood-brain barrier property (the lock) in the cells. For instance, peptides (the key) can be used to enable transport using overexpressed receptors, such as transferring receptor (TfR), in a mechanism designated Receptor Mediated Transport (RMT).”

Peptides can mediate the delivery of drugs using the unique properties of cell components (lipids, glycolipids, or glycoproteins), in a mechanism called adsorptive mediated transport (AMT), says Neves. “In AMT, positively-charged peptides induce interactions with the blood-brain barrier cell membrane, which will culminate with vesicular transport across it,” she adds. “These types of mediated transports are particularly important for the delivery of large therapeutics, such as antibodies, that have been successfully used to treat diseases like cancer in other parts of the body, but have very limited application in the brain.”

As well as enabling a treatment pathway that could open the door for new therapeutics, advantages to using peptide shuttles over traditional drug delivery methods include them being more cost-effective and scalable, since they’re small and can easily be synthesised. They can also be designed to specifically target certain areas, reducing the need for unnecessary off-target therapy and the side-effects that can result. However, as with any potential drug delivery method, there are challenges associated with developing peptide shuttles for the brain, as Stacy Blain, chief science officer of Concarlo Therapeutics and professor at SUNY Downstate Medical Center, explains: “There are many safety concerns or even cost issues, but neurological diseases are so difficult to treat that cost is less of an issue. The bar is quite low. Any drugs that we can get across the blood-brain barrier would be a benefit, regardless of price.”

In terms of safety, it’s important to remember that the peptide shuttles are just that: shuttles. “They are not the drug themselves, they’re just a modality to get another medicine across the barrier,” says Blain. “So, we will have to take into account the inherent safety to your cost of what the peptide shuttles are transporting.”

“There are many safety concerns or even cost issues, but neurological diseases are so difficult to treat that cost is less of an issue. The bar is quite low.”
Stacy Blain, chief science officer of Concarlo Therapeutics

Beyond cost and safety, there’s also efficacy to consider – and here, too, there are many other potential complications. “The major challenge associated with peptide shuttles is their short lifespan in vivo, due to the abundance of peptidases in the digestive system (which limits oral administration), blood, liver, the blood-brain barrier,” says Neves. “As a consequence, the use of peptide shuttles might result in lower effectiveness.” Nevertheless, it has been proposed that modifications to peptides might increase their time in the blood. Such modifications include the use of unnatural amino acids, as well as cyclisation and glycosylation – two chemical modulation techniques that can improve bioavailability and reduce the rate at which the body clears the peptide from the blood. “Another important aspect for trans-blood-brain-barrier delivery is the prerequisite to maximise transcytosis – meaning that peptide shuttles need to avoid the endosomal/ lysosomal degradation pathway that would reduce the amount that effectively reaches the brain,” says Neves. “Finally, apart from the strong evidence that peptide shuttles dramatically improve brain delivery – the amount reaching the brain remains low. In the end, it is important to evaluate the efficacy of the payload to effectively assess that the amount ‘delivered’ is enough for treatment.”

“In the end, it is important to evaluate the efficacy of the payload to effectively assess that the amount ‘delivered’ is enough for treatment.”
Vera Neves, senior staff researcher at the University of Lisbon

The future of drug delivery

While the effects of peptide shuttles to bypass the blood-brain barrier are yet to be tested on humans, studies have shown promising results in animal models of brain diseases, including Alzheimer’s disease and glioblastoma. Peptide shuttles have been intensively investigated for the delivery of different drugs, some examples being: chemotherapeutics for the treatment of brain cancer (glioma), such as Paclitaxel, Doxorubicin, Temozolomide; and the biologics like peptise, proteins and antibodies, for the treatment of cerebral Ischemia, Parkinson’s, Alzheimer’s, Hunter Syndrome and other brain diseases, explains Neves.

Among a handful of companies who are on the cutting edge of developing this technology is Denali Therapeutics, whose large molecule transport vehicle (TV) platform technology uses an engineered Fc fragment that exploits receptormediated transcytosis for CNS delivery of biotherapeutics. Denali Therapeutics is one of many pharmaceutical companies who believe that the development of the peptide shuttle will play a critical role in enabling effective neurological treatments.

Despite the development of peptide shuttles and other strategies to delivery therapeutic agencies across the blood-brain barrier being a promising area of research, there is still the risk of an immune response. Many of these peptides can induce an immune response and cause toxicity, which has the potential to increase the risk of adverse side effects. As well as this, the pharmacokinetics of blood-brain-barrier-targeting peptides can pose a challenge due to their short half-life, which can limit their overall effectiveness. These are all areas for which researchers will need to find a solution before they can move peptide shuttles towards first-in-human trials with therapeutics. Currently, there are over 150 peptides in the process of being developed for clinical development. Evidently, the field of peptide shuttles for neurological diseases is rapidly evolving and the potential for it is vast.

Successful shuttles

In a paper for the journal Pharmaceutics, researchers Macarena Sanchez-Navarro and Ernest Giralt, discuss the different types of peptides that have effectively been used as shuttles, including cell-penetrating peptides, receptor-specific peptides, and brain-targeting peptides. Examples of successful drug delivery peptide shuttles include:

■ Delivery of insulin-like growth factor-1 (IGF-1) to the brain: A receptor-specific peptide shuttle was developed that could bind to the insulin receptor on the blood-brain barrier and transport IGF-1 across the barrier. The peptide shuttle successfully delivery IGF-1 to the brain and protected against ischemic damage in a mouse model.

■ Delivery of neuropeptide Y (NPY) to the brain: A cell-penetrating peptide shuttle was developed that could transport NPY across the blood-brain barrier. The peptide shuttle successfully delivered NPY to the brain and reduced brain injury in a rat model of traumatic brain injury.

■ Delivery of doxorubicin to brain tumours: A brain-targeting peptide shuttle was developed that could bind to receptors on the surface of brain tumour cells and transport doxorubicin across the blood-brain barrier. The peptide shuttle successfully delivered doxorubicin to brain tumours and improved survival in a mouse model of brain cancer.