Posted on by Dr. Francis Collins
If used to make non-heritable genetic changes, CRISPR gene-editing technology holds tremendous promise for treating or curing a wide range of devastating disorders, including sickle cell disease, vision loss, and muscular dystrophy. Early efforts to deliver CRISPR-based therapies to affected tissues in a patient’s body typically have involved packing the gene-editing tools into viral vectors, which may cause unwanted immune reactions and other adverse effects.
Now, NIH-supported researchers have developed an alternative CRISPR delivery system: nanocapsules. Not only do these tiny, synthetic capsules appear to pose a lower risk of side effects, they can be precisely customized to deliver their gene-editing payloads to many different types of cells or tissues in the body, which can be extremely tough to do with a virus. Another advantage of these gene-editing nanocapsules is that they can be freeze-dried into a powder that’s easier than viral systems to transport, store, and administer at different doses.
In findings published in Nature Nanotechnology , researchers, led by Shaoqin Gong and Krishanu Saha, University of Wisconsin-Madison, developed the nanocapsules with specific design criteria in mind. They would need to be extremely small, about the size of a small virus, for easy entry into cells. Their surface would need to be adaptable for targeting different cell types. They also had to be highly stable in the bloodstream and yet easily degraded to release their contents once inside a cell.
After much hard work in the lab, they created their prototype. It features a thin polymer shell that’s easily decorated with peptides or other ingredients to target the nanocapsule to a predetermined cell type.
At just 25 nanometers in diameter, each nanocapsule still has room to carry cargo. That cargo includes a single CRISPR/Cas9 scissor-like enzyme for snipping DNA and a guide RNA that directs it to the right spot in the genome for editing.
In the bloodstream, the nanocapsules remain fully intact. But, once inside a cell, their polymer shells quickly disintegrate and release the gene-editing payload. How is this possible? The crosslinking molecules that hold the polymer together immediately degrade in the presence of another molecule, called glutathione, which is found at high levels inside cells.
The studies showed that human cells grown in the lab readily engulf and take the gene-editing nanocapsules into bubble-like endosomes. Their gene-editing contents are then released into the cytoplasm where they can begin making their way to a cell’s nucleus within a few hours.
Further study in lab dishes showed that nanocapsule delivery of CRISPR led to precise gene editing of up to about 80 percent of human cells with little sign of toxicity. The gene-editing nanocapsules also retained their potency even after they were freeze-dried and reconstituted.
But would the nanocapsules work in a living system? To find out, the researchers turned to mice, targeting their nanocapsules to skeletal muscle and tissue in the retina at the back of eye. Their studies showed that nanocapsules injected into muscle or the tight subretinal space led to efficient gene editing. In the eye, the nanocapsules worked especially well in editing retinal cells when they were decorated with a chemical ingredient known to bind an important retinal protein.
Based on their initial results, the researchers anticipate that their delivery system could reach most cells and tissues for virtually any gene-editing application. In fact, they are now exploring the potential of their nanocapsules for editing genes within brain tissue.
I’m also pleased to note that Gong and Saha’s team is part of a nationwide consortium on genome editing supported by NIH’s recently launched Somatic Cell Genome Editing program. This program is dedicated to translating breakthroughs in gene editing into treatments for as many genetic diseases as possible. So, we can all look forward to many more advances like this one.
 A biodegradable nanocapsule delivers a Cas9 ribonucleoprotein complex for in vivo genome editing. Chen G, Abdeen AA, Wang Y, Shahi PK, Robertson S, Xie R, Suzuki M, Pattnaik BR, Saha K, Gong S. Nat Nanotechnol. 2019 Sep 9.
Saha Lab (University of Wisconsin-Madison)
Shaoqin (Sarah) Gong (University of Wisconsin-Madison)
NIH Support: National Eye Institute; National Institute of General Medical Sciences; National Institute of Neurological Disorders and Stroke; National Heart, Lung, and Blood Institute; Common Fund
Posted on by Dr. Francis Collins
Recently, I’ve highlighted just a few of the many amazing advances coming out of the NIH-led Brain Research through Advancing Innovative Neurotechnologies® (BRAIN) Initiative. And for our grand finale, I’d like to share a cool video that reveals how this revolutionary effort to map the human brain is opening up potential plans to help people with disabilities, such as vision loss, that were once unimaginable.
This video, produced by Jordi Chanovas and narrated by Stephen Macknik, State University of New York Downstate Health Sciences University, Brooklyn, outlines a new strategy aimed at restoring loss of central vision in people with age-related macular degeneration (AMD), a leading cause of vision loss among people age 50 and older. The researchers’ ultimate goal is to give such people the ability to see the faces of their loved ones or possibly even read again.
In the innovative approach you see here, neuroscientists aren’t even trying to repair the part of the eye destroyed by AMD: the light-sensitive retina. Instead, they are attempting to recreate the light-recording function of the retina within the brain itself.
How is that possible? Normally, the retina streams visual information continuously to the brain’s primary visual cortex, which receives the information and processes it into the vision that allows you to read these words. In folks with AMD-related vision loss, even though many cells in the center of the retina have stopped streaming, the primary visual cortex remains fully functional to receive and process visual information.
About five years ago, Macknik and his collaborator Susana Martinez-Conde, also at Downstate, wondered whether it might be possible to circumvent the eyes and stream an alternative source of visual information to the brain’s primary visual cortex, thereby restoring vision in people with AMD. They sketched out some possibilities and settled on an innovative system that they call OBServ.
Among the vital components of this experimental system are tiny, implantable neuro-prosthetic recording devices. Created in the Macknik and Martinez-Conde labs, this 1-centimeter device is powered by induction coils similar to those in the cochlear implants used to help people with profound hearing loss. The researchers propose to surgically implant two of these devices in the rear of the brain, where they will orchestrate the visual process.
For technical reasons, the restoration of central vision will likely be partial, with the window of vision spanning only about the size of one-third of an adult thumbnail held at arm’s length. But researchers think that would be enough central vision for people with AMD to regain some of their lost independence.
As demonstrated in this video from the BRAIN Initiative’s “Show Us Your Brain!” contest, here’s how researchers envision the system would ultimately work:
• A person with vision loss puts on a specially designed set of glasses. Each lens contains two cameras: one to record visual information in the person’s field of vision; the other to track that person’s eye movements enabled by residual peripheral vision.
• The eyeglass cameras wirelessly stream the visual information they have recorded to two neuro-prosthetic devices implanted in the rear of the brain.
• The neuro-prosthetic devices process and project this information onto a specific set of excitatory neurons in the brain’s hard-wired visual pathway. Researchers have previously used genetic engineering to turn these neurons into surrogate photoreceptor cells, which function much like those in the eye’s retina.
• The surrogate photoreceptor cells in the brain relay visual information to the primary visual cortex for processing.
• All the while, the neuro-prosthetic devices perform quality control of the visual signals, calibrating them to optimize their contrast and clarity.
While this might sound like the stuff of science-fiction (and this actual application still lies several years in the future), the OBServ project is now actually conceivable thanks to decades of advances in the fields of neuroscience, vision, bioengineering, and bioinformatics research. All this hard work has made the primary visual cortex, with its switchboard-like wiring system, among the brain’s best-understood regions.
OBServ also has implications that extend far beyond vision loss. This project provides hope that once other parts of the brain are fully mapped, it may be possible to design equally innovative systems to help make life easier for people with other disabilities and conditions.
Age-Related Macular Degeneration (National Eye Institute/NIH)
Macknik Lab (SUNY Downstate Health Sciences University, Brooklyn)
Martinez-Conde Laboratory (SUNY Downstate Health Sciences University)
Show Us Your Brain! (BRAIN Initiative/NIH)
NIH Support: BRAIN Initiative
Posted on by Dr. Francis Collins
Infrared vision often brings to mind night-vision goggles that allow soldiers to see in the dark, like you might have seen in the movie Zero Dark Thirty. But those bulky goggles may not be needed one day to scope out enemy territory or just the usual things that go bump in the night. In a dramatic advance that brings together material science and the mammalian vision system, researchers have just shown that specialized lab-made nanoparticles applied to the retina, the thin tissue lining the back of the eye, can extend natural vision to see in infrared light.
The researchers showed in mouse studies that their specially crafted nanoparticles bind to the retina’s light-sensing cells, where they act like “nanoantennae” for the animals to see and recognize shapes in infrared—day or night—for at least 10 weeks. Even better, the mice maintained their normal vision the whole time and showed no adverse health effects. In fact, some of the mice are still alive and well in the lab, although their ability to see in infrared may have worn off.
When light enters the eyes of mice, humans, or any mammal, light-sensing cells in the retina absorb wavelengths within the range of visible light. (That’s roughly from 400 to 700 nanometers.) While visible light includes all the colors of the rainbow, it actually accounts for only a fraction of the full electromagnetic spectrum. Left out are the longer wavelengths of infrared light. That makes infrared light invisible to the naked eye.
In the study reported in the journal Cell, an international research team including Gang Han, University of Massachusetts Medical School, Worcester, wanted to find a way for mammalian light-sensing cells to absorb and respond to the longer wavelengths of infrared . It turns out Han’s team had just the thing to do it.
His NIH-funded team was already working on the nanoparticles now under study for application in a field called optogenetics—the use of light to control living brain cells . Optogenetics normally involves the stimulation of genetically modified brain cells with blue light. The trouble is that blue light doesn’t penetrate brain tissue well.
That’s where Han’s so-called upconversion nanoparticles (UCNPs) came in. They attempt to get around the normal limitations of optogenetic tools by incorporating certain rare earth metals. Those metals have a natural ability to absorb lower energy infrared light and convert it into higher energy visible light (hence the term upconversion).
But could those UCNPs also serve as miniature antennae in the eye, receiving infrared light and emitting readily detected visible light? To find out in mouse studies, the researchers injected a dilute solution containing UCNPs into the back of eye. Such sub-retinal injections are used routinely by ophthalmologists to treat people with various eye problems.
These UCNPs were modified with a protein that allowed them to stick to light-sensing cells. Because of the way that UCNPs absorb and emit wavelengths of light energy, they should to stick to the light-sensing cells and make otherwise invisible infrared light visible as green light.
Their hunch proved correct, as mice treated with the UCNP solution began seeing in infrared! How could the researchers tell? First, they shined infrared light into the eyes of the mice. Their pupils constricted in response just as they would with visible light. Then the treated mice aced a series of maneuvers in the dark that their untreated counterparts couldn’t manage. The treated animals also could rely on infrared signals to make out shapes.
The research is not only fascinating, but its findings may also have a wide range of intriguing applications. One could imagine taking advantage of the technology for use in hiding encrypted messages in infrared or enabling people to acquire a temporary, built-in ability to see in complete darkness.
With some tweaks and continued research to confirm the safety of these nanoparticles, the system might also find use in medicine. For instance, the nanoparticles could potentially improve vision in those who can’t see certain colors. While such infrared vision technologies will take time to become more widely available, it’s a great example of how one area of science can cross-fertilize another.
 Mammalian Near-Infrared Image Vision through Injectable and Self-Powered Retinal Nanoantennae. Ma Y, Bao J, Zhang Y, Li Z, Zhou X, Wan C, Huang L, Zhao Y, Han G, Xue T. Cell. 2019 Feb 27. [Epub ahead of print]
 Near-Infrared-Light Activatable Nanoparticles for Deep-Tissue-Penetrating Wireless Optogenetics. Yu N, Huang L, Zhou Y, Xue T, Chen Z, Han G. Adv Healthc Mater. 2019 Jan 11:e1801132.
Diagram of the Eye (National Eye Institute/NIH)
Infrared Waves (NASA)
Visible Light (NASA)
Han Lab (University of Massachusetts, Worcester)
NIH Support: National Institute of Mental Health; National Institute of General Medical Sciences