Enhanced human blood-brain barrier chip provides in vivo transport of drugs and antibodies

Like airport security gates that allow either authorized travelers to prevent unauthorized travelers and their baggage from accessing central areas of operations, the blood-brain barrier tightly controls transportation essential nutrients and energy metabolites in the brain and prevents the circulation of unwanted substances into the bloodstream. It is important to note that its highly organized structure of thin blood vessels and supporting cells is also the main obstacle preventing drugs from saving lives reaching the brain in order to effectively treat cancer, neurodegeneration and dementia. 39, other diseases of the central nervous system. In a number of brain diseases, BBB can also decompose locally, leading to the penetration of neurotoxicants, blood cells and pathogens into the brain, causing irreparable damage.

To study BBB and the transport of the drug through it, the researchers mainly used animal models such as the mouse. However, the precise composition and transport of BBBs in these models may differ considerably from those in human patients, making them unreliable in predicting drug delivery and therapeutic efficacy. In addition, in vitro models attempting to recreate human BBB with the aid of primary cells derived from brain tissue have so far not been able to mimic the physical barrier, the transport functions and drug and antibody shuttle activities of the BBB sufficiently to be useful as therapeutic development tools.

Now, a team led by Donald Ingber, MD, Ph.D. The Harvard Wyss Institute for Biologically Inspired Engineers has overcome these limitations by using its microfluidic organ technology on chips (microarray chips). organs) in combination with an approach mimicking development-induced hypoxia to differentiate human pluripotent stem cells (iPS) into microvascular endothelial brain cells (BMVEC). The resulting "hypoxia enhanced BBB chip" summarizes the cellular organization, barrier functions and transport capabilities of human BBB; and it allows the transport of therapeutic drugs and antibodies in a manner that more closely mimics the transport through the BBB in vivo compared to existing in vitro systems. Their study is reported in Nature Communications.

"Our approach to modeling the circulation of drugs and antibodies in human BBB in vitro with such high and unprecedented fidelity represents a significant advance over existing capabilities in this extremely challenging area of ​​research," said Ingber, director founder of Wyss Institute. "It responds to a critical need for drug development programs in the pharmaceutical and biotechnology world, which we are now aiming to solve with a Wyss Institute's" Blood-brain barrier transport program ", which uses our talent and unique resources. " Ingber is also a Judah Folkman Professor of Vascular Biology at HMS and the Vascular Biology Program at Boston Children's Hospital, as well as a professor of bioengineering at SEAS.

The BBB consists of thin capillary blood vessels formed by BMVEC, multifunctional cells called pericytes that wrap around the outside of the vessels and star-shaped astrocytes, brain cells non-neurons that also come into contact with the blood vessels with processes similar to those of the foot. . In the presence of pericytes and astrocytes, endothelial cells can generate the tightly sealed vascular wall barrier typical of human BBB.

The Ingber team first differentiated human iPS cells into brain endothelial cells in the culture dish using a method previously developed by co-author Eric Shusta, Ph.D. Chemical and biological engineering at the University of Wisconsin-Madison, but the added power of bioinspiration. "Because in the embryo, the BBB is formed under conditions of lack of oxygen (hypoxia), we have differentiated iPS cells for a prolonged period in an atmosphere with only 5% instead of concentration Normal oxygen in 20%, "said the co-first author, Tae-Eun Park, Ph.D." As a result, iPS cells have initiated a development program very similar to that of the embryo, producing BMVECs having a higher functionality than BMVECs generated under normal oxygen conditions. " Park was a postdoctoral member of the Ingber team and currently holds the position of badistant professor at the Ulsan National Institute of Science and Technology in the Republic of Korea.

Building on a previous human BBB model, the researchers then transferred hypoxia-induced human BMVECs into one of two parallel channels of a microfluidic organ-on-chip device, divided by a porous membrane and perfused continuously with a medium. The other channel was populated with a mixture of human brain pericytes and astrocytes. After an additional day of hypoxia treatment, the human BBB chip could be stably maintained for at least 14 days at normal oxygen levels, which is much longer than the in vitro models of human models of BBB tested in the laboratory. past.

Under the shear stress of fluids infusing the BBB chip, BMVECs form a blood vessel and develop a dense interface with pericytes aligning with them on the other side of the porous membrane, as well as on the other hand. with astrocytes extending towards them. through small openings in the membrane. "The distinct morphology of the modified BBB goes hand in hand with the formation of a tighter barrier containing a high number of selective transport systems and drug shuttles compared to the control BBBs that we generated without hypoxia or shear stress. fluid, or with endothelium derived from the adult brain instead of. iPS cells, "said Nur Mustafaoglu, Ph.D., co-lead author of the study and postdoctoral fellow working on the subject. Ingber team. "In addition, we could mimic the effects of treatment strategies in patients in the clinic.For example, we have reversibly opened the BBB plate for a short time by increasing the concentration of a mannitol solute. [osmolarity] allow the pbadage of large drugs such as the anti-cancer antibody Cetuximab ".

To provide further evidence that the hypoxia-enhanced human BBB chip can be used as an effective tool for studying drug delivery to the brain, the team investigated a range of transport mechanisms that prevent drugs to reach their targets in the brain by pumping them back. in the bloodstream (efflux), or that, on the contrary, allow the selective transport of nutrients and drugs through the BBB (transcytosis).

"When we specifically blocked the function of P-gp, a key pump for endothelial efflux, we were able to dramatically increase the transport of doxorubicin, an anticancer drug, from the vascular cbad to the brain cbad, in a very similar to that seen in human patients. "said Park. "Thus, our in vitro system could be used to identify new approaches to reduce efflux and thus facilitate the transport of drugs into the brain in the future."

On another site, drug developers are trying to leverage "receptor-mediated transcytosis" as a means of transporting drug-laden nanoparticles, larger drugs and protein-based drugs, as well as therapeutic antibodies to through the BBB. "The hypoxia-enhanced human BBB chip recapitulates the function of critical transcytosis pathways, such as those used by LRP-1 and transferrin receptors, that are responsible for the capture of vital lipoproteins and iron in circulating blood and to release them. in the brain on the other side of the body, exploiting these receptors using different preclinical strategies, we can faithfully mimic the previously demonstrated multiplication of therapeutic antibodies that target transferrin receptors in vivo, while now the integrity of the BBB in vitro, "said Mustafaoglu.

Based on these findings, the Wyss Institute has launched a "Blood-Brain Barrier Transport Program". "Initially, BBB's transport program was aimed at discovering new shuttle targets enriched on the vascular surface of BMVEC, using novel approaches to transcriptomics, proteomics and iPS cells, and at the same time developing fully human antibody shuttles. directed against known shuttle targets brain targeting capabilities, "said James Gorman, MD, Ph.D., responsible for BBB's transportation program working with Ingber. "Our goal is to collaborate with several biopharmaceutical partners in a precompetitive relationship to develop shuttles offering exceptional technical efficiency and flexibility for incorporation into antibody-based drugs and protein, because patients and everyone else need it. "

The authors believe that in addition to drug development studies, the hypoxia-enhanced human BBB chip can also be used to model aspects of brain disease that affect the disease, such as Alzheimer's disease and Parkinson's disease. that advanced approaches to personalized medicine using methods derived from the patient. iPS cells.