Rabies-virus inspired delivery of gold nanoparticles to treat brain tumors
When material science meets biology.
The best innovations in science and technology are often interdisciplinary.
What makes science so difficult and sometimes frustrating is the multidimensionality of problems we would like to solve. Another way of phrasing that is that life tends to be messy and complicated; its challenges do not care about our arbitrary distinctions between fields like statistics, physics or biology, they encompass them all.
Scientists have to consider it all. The physics of electron density and Van-der-Waals interactions underlie most biochemistry; the billions of billions of molecules which act in concert in a cell can only be understood with probability and statistics. Evolution has to be considered not as directed process, but as history record. Often, even in our most elaborate model systems, our results get screwed up by something we did not think about, or worse, we cannot even measure. That’s why science can be genuinely hard.
Once in a while, however, the very multidimensionality of the problem can be leveraged to come up with unique solutions.
Since some time already, biological scientists are not the only ones trying to find effective cancer treatments; physicists joined them in the quest. About a decade ago, physicists brought a creatively different idea to cancer research, called photothermal therapy (PTT).
In contrast to biological scientists (including physicians, doctors etc), who would majorly rely on drugs/biologically active compounds or surgery to target cancer cells for destruction, photothermal therapy is based on the interaction of electromagnetic waves (infrared light) with artificial nanoparticles (e.g. gold particles). Once taken up by a cell and hit with infrared light, these particles produce enough heat to destroy the cell and its surroundings. Sounds neat. But how do those particles get into cancer cells and avoid normal tissue?
Two strategies can be exploited.
A) Unique tumor biology. Usually, when a tumor forms, it requires new blood vessels in order to fuel its growth; these new blood vessels in/near tumors have different properties as compared to regular blood vessels, such as poor lymphatic drainage and a disorganized, leaky vasculature. These factors can lead to a significantly higher concentration of certain particles in a tumor as compared to the rest of the body.
B) Actively guided transport. Antibody-antigen interactions are the lock-key principle behind biological processes that require identification. For examples, antibodies are the class of proteins that our immune system uses to target defined “foreign” proteins displayed on the cell surface of invading parasites. Some cancer cells are known to be targetable by antibodies (immunotherapy).
While PTT only started quite recently to be explored, it did not come out of nowhere. Like many fields in science, there is a ideological predecessor, in this case it is called photodynamic therapy (PDT), an approach involving light and a photosensitizing chemical substance, used in conjunction with molecular oxygen to elicit cell death (phototoxicity). This has been widely used in treating acne and psoriasis, as well as approved by the FDA for treatment of macular degeneration. PDT is also being investigated in clinical trials against prostate cancer.
One prominent difference between PTT and PDT is the requirement of PDT for molecular oxygen to interact with the target cells, which might not always be a given. Furthermore, PDT requires higher energy electromagnetic waves, which can be harmful for surrounding tissue.
Before we can continue with PTT, we have to switch gears for a moment to virology.
Viruses come in many shapes and forms, vary widely in size (10nm to >1µm) and infect different hosts and tissues. The surface proteins of the viral capsid facilitate the cellular uptake of the virus cargo (RNA or DNA) into cells, and the strategies employed are as manifold as the viruses themselves.
For example, the HIV capsid protein called “envelope” (Env) binds to the primary cellular receptor CD4 of immune cells and subsequently to a cellular coreceptor. This sequential binding is shown to trigger fusion of the viral and host cell membranes, initiating infection.
For the very common influenza virus, viral entry is incompletely understood and reads like a sci-fi tale. The main actor is the capsid protein “hemagglutinin” (HA), a trimeric glycoprotein. The first step of influenza virus entry is the recognition of the host cell receptor molecule, terminal α-sialic acid, by HA. Attachment by multiple copies of trimetric HA triggers then the uptake (endocytosis) of influenza virus as endosome (imagine virus inside soap bubble). The endosome-trapped virus traffics to near the nucleus (imagine deep dive into the center of a cell). At this location, the interior pH of the endosome becomes acidic that induces a dramatic conformational change in HA, which induces a repositioning of the two membranes, and form a fusion pore (imagine docking to submarine) that allows the release of the genome segments of influenza virus… Pretty crazy and elaborate process for just a physical principle (viruses are not considered lifeforms), one might agree with me.
Anyway, to make my point: Viruses are crazy capable invaders because of their unique biological and surface properties.
This is why modified viruses have been used for more than 3 decades now by researchers to deliver a genetic cargo to a tissue of interest, and we have gotten quite good at it. Lentiviral based gene delivery systems today are ubiquitous in everyday lab work, to genetically modify cells in vitro, and more and more in vivo as well.
Now let’s return to why interdisciplinary thinking is great for innovation.
Recently, researchers from Sungkyunkwan University in South-Korea spearheaded a fantastic innovative study together with other major Korean universities, published in the scientific journal Advanced Materials.
The authors set themselves an ambitious goal; they wanted to deliver gold nanoparticles across the blood-brain barrier (BBB) to target glioblastoma (brain cancer) with PTT.
The blood-brain barrier is a highly selective membrane that separates the circulating blood from the extracellular fluid of our central nervous system, a gatekeeper that guards what molecules can and cannot reach our brain. Anybody who has ever had contact with pharmacology 101 can tell you that delivering therapeutic drugs to the brain is incredibly hard because of the BBB’s tight gatekeeping. Usually, only very small molecules like water and some gases can slip through, the rest needs some active transport (biological equivalent of an V.I.P pass) that lets them pass the BBB.
The only other alternative is to cheat the system, like invasion-specialist viruses do. One expert when it comes to infecting neurons in the brain is the rabies virus (RABV). The RABV is enveloped by five proteins, one of which is a rabies-virus glycoprotein (RVG) that interacts specifically with acetylcholine receptor (AchR) proteins, which are abundantly expressed on neuronal cells. One specific, 29 amino acid long (RVG-29) part of that glycoprotein is shown to bind so strongly to AchR on neuronal cells, that it can even outcompete the binding snake-venom neurotoxin a-bungarotoxin, which causes paralysis and respiratory failure by AchR-signaling inhibition.
Lee C. and his colleagues reasoned that if they could cover their gold-nanoparticles with this rabies-virus derived peptide RVG-29, they might camouflage and enable them to pass the BBB and reach their target, the brain tumor. To inform their design blue prints, they tried to match the size, shape and biological features of their gold-nanorods as closely as possible to the native shape of the rabies virus.
To test whether their RABV-Mimicry could actually enter neuronal cells, and do so better than random diffusion, Lee C. et al. went on to investigate cellular uptake of their construct compared to a similar gold-nanorod construct without the RAB-29 peptide on its surface.
While in vitro delivery of gold-nanoparticles had been reported previously, the real challenge for Lee C. et al. was crossing that pesky blood-brain barrier. In a mouse model of glioblastoma, they injected their RABV-inspired gold nanorods into the tail vein of mice (a technique commonly used in rodents to delivery drugs to the bloodstream) and imaged the brains and spines of mice at several time points after injection of the particles. With this, the authors could show that their RABV-inspired delivery method was able to transport the gold-nanorods necessary for photothermal therapy to the brain in a highly localized manner.
Finally, to also address the question of PTT feasibility with their RABV-inspired nanoparticle construct, the researchers used a conventional xenograft tumor model, based on inoculating tumor-bearing neuronal cells into the dorsal flanks of immunosuppressed mice. As expected, their RABV-inspired gold-nanorods accumulated in the xenograft tumors and produced high heat after being irradiated with an infrared laser, nicely visualized by imaging with a thermal camera.
In summary, we successfully developed gold nanorods that mimic the rabies virus in terms of size, shape, surface glycoprotein property, and in vivo behavior. […] Considering our data, the rod shape of our RVG-PEG-AuNRs@SiO2 appeared to play a crucial role in facilitating cellular uptake into neuronal cells in vitro and inducing a hyperthermal effect in response to NIR laser irradiation. The surface-modified RVG29 peptide substantially improved the in vivo distribution of the nanorods in the central nervous system. Importantly, it is of great interest that these nanorods not only resembled the appearance of the live rabies virus but targeted the brain through the neuronal pathway bypassing the blood–brain barrier. […]
Together, these results support that the rabies virus mimetic gold nanorods are a potential prototype delivery platform for treating brain tumors. — Lee C. et al. Advanced Materials. 2017
I could not have summarized it more nicely. Great, creative and inspiring interdisciplinary work. Science is a collaborative effort, it is in fact the biggest, generation-spanning quest for knowledge the world has ever seen.
So next time someone from a completely different field want to pitch an idea to you, I encourage you to hear him out.
Source: Lee C. et al. Advanced Materials. 2017
additional sources: Cook RL. et al. Journal of controlled release. 2015 , Peer D. et al, Nature Nanotechnology, 2007 (review)
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