Tag Archive for: Li

RNA molecule

New deep learning system helps scientists edit RNA

RNA molecule

The Children’s National team built DeepCas13 on a newer and less studied CRISPR platform, called CRISPR-Cas13d, which instead focuses on RNA.

Children’s National Hospital scientists have created a revolutionary machine-learning system that predicts the effects of changing ribonucleic acid (RNA) molecules using a gene-editing tool built on CRISPR technology.

Called DeepCas13, the system is among the world’s first deep-learning frameworks to recognize the challenges of editing RNA – and then applying data science and machine learning to solve the intricate problems that stem from modifying biological code. Details of the DeepCas13 system were published recently in Nature Communications.

Born from an international collaboration, DeepCas13 could provide the backbone for treatments for diseases based on errors in RNA, including debilitating neurodegenerative diseases such as Huntington’s disease and muscular dystrophy.

“I am an optimistic person, so I expect to have treatments within five to 10 years,” said Wei Li, Ph.D., principal investigator at the Center for Genetic Medicine Research at Children’s National. “Of course, there are going to be lots of obstacles. If we have a very good system, like DeepCas13, with very good performance that can generate treatments, the next problem is how we deliver the system to the right tissue in the patients.”

The big picture

Most research in this space has focused on a version of CRISPR – or Clustered Regularly Interspaced Short Palindromic Repeats – that edits DNA, called CRISPR-Cas9. The Children’s National team built DeepCas13 on a newer and less studied CRISPR platform, called CRISPR-Cas13d, which instead focuses on RNA. In doing so, researchers are opening the door to treating a host of disorders of RNA, given its biological role in coding, decoding, regulating and supporting gene expression.

DeepCas13 combines hundreds of thousands of data points with considerable computing power to help scientists target errant pieces of RNA, while minimizing any off-target changes that could damage the health of cells.

“We only want to target the RNA molecule that is causing diseases, and we don’t want the system to edit normal RNA,” said Xiaolong Cheng, Ph.D., a member of the Li lab and the first author of the study. “With DeepCas13, we can design highly efficient, and highly specific, rules.”

What’s ahead

The FDA has approved one method for delivering RNA treatments to cells, using a virus known as AAV or adeno-associated virus. So far, the gene therapy method has had limited applications. But Li and other researchers see the potential for life-changing treatments in the coming years, built on DeepCas13 and other advances.

The system was developed with partners from around the world, including the University of Illinois Urbana-Champaign and Northeastern University in Shenyang, China. It is open source and available for free to researchers looking for targets to treat RNA-related diseases.

Li says this international partnership is leading the way: “We tested our DeepCas13 model over other methods, and we confirmed that our method has the highest prediction accuracy.”

DeepCas13 was funded by grants from NIH and the Children’s National Research Institute.

HIV virus

CRISPR gene editing identifies possible drug targets for HIV

HIV virus

Working with researchers at Johns Hopkins University, the Children’s National team used CRISPR gene technology to test drug targets that find and attack latent HIV, paving the way for drug treatments that may someday completely cure the virus.

Researchers at Children’s National Hospital have identified several new drug targets that may enhance the elimination of latent HIV in patients, a major bottleneck to the full treatment of the virus, according to new findings published in Science Translational Medicine.

Working with researchers at Johns Hopkins University, the Children’s National team used CRISPR gene technology to test drug targets that find and attack latent HIV, paving the way for drug treatments that may someday completely cure the virus. Currently, anti-retroviral therapies (ARTs) can only slow its progress.

Why we’re excited

“In less than one month, we were able to use CRISPR to test 20,000 gene candidates in one single experiment. It was an amazing application of the technology,” said Wei Li, Ph.D., a co-author of the study and assistant professor at the Center for Genetic Medicine Research at Children’s National. “The CRISPR technology provides a global, unbiased approach to understanding molecular aspects of HIV-1 infection, including the ways that HIV-1 enters cells and replicates. This research could someday revolutionize how we treat the virus pharmaceutically.”

The big picture

More than 30 million people worldwide live with HIV-1, the most common form of the virus that can eventually lead to AIDS. But no single agent can entirely eliminate HIV-1 in these patients.

Researchers have sought ways to attack this elusiveness and turned to the CRISPR gene-editing tool, which can locate specific bits of DNA inside a cell. They trained CRISPR screens on the HIV-1 genome to identify critical factors that allow or prevent the virus from lying latent. In the latter case, these pieces of DNA will be the ideal targets of a drug that will push the virus out of the latent stage so it can be targeted by therapies.

What’s ahead

The findings of the Children’s National and Johns Hopkins scientists point to novel drug therapies and validation systems that could someday eradicate HIV.

Bicna Song, a postdoctoral researcher in Li’s laboratory at the Center for Genetic Medicine, said that reversing HIV-1 latency will allow for the killing of infected cells and give researchers opportunities to actually cure patients with HIV.

“So far, no single latency-reversing agent – alone or in combination with another drug – has been shown to effectively reduce the latent reservoir size in persons living with HIV-1,” said Song, who contributed to the study. “With this work, we are meeting the urgent need to identify factors that can lead to new drug targets.”

illustration of the brain

New research provides glimpse into landscape of the developing brain

illustration of the brain

Stem and progenitor cells exhibit diversity in early brain development that likely contributes to later neural complexity in the adult cerebral cortex, this according to a new study in Science Advances. This research expands on existing ideas about brain development, and could significantly impact the clinical care of neurodevelopmental diseases in the future.

Stem and progenitor cells exhibit diversity in early brain development that likely contributes to later neural complexity in the adult cerebral cortex, this according to a study published Nov. 6, 2020, in Science Advances. Researchers from the Center for Neuroscience Research (CNR) at Children’s National Hospital say this research expands on existing ideas about brain development, and could significantly impact the clinical care of neurodevelopmental diseases in the future. The study was done in collaboration with a research team at Yale University led by Nenad Sestan, M.D, Ph.D.

“Our study provides a new glimpse into the landscape of the developing brain. What we are seeing are new complex families of cells very early in development,” says Tarik Haydar, Ph.D., director of CNR at Children’s National, who led this study. “Understanding the role of these cells in forming the cerebral cortex is now possible in a way that wasn’t possible before.”

The cerebral cortex emerges early in development and is the seat of higher-order cognition, social behavior and motor control. While the rich neural diversity of the cerebral cortex and the brain in general is well-documented, how this variation arises is relatively poorly understood.

“We’ve shown in our previous work that neurons generated from different classes of cortical stem and progenitor cells have different functional properties,” says William Tyler, Ph.D., CNR research faculty member and co-first author of the study. “Part of the reason for doing this study was to go back and try to classify all the different progenitors that exist so that eventually we can figure out how each contributes to the diversity of neurons in the adult brain.”

Using a preclinical model, the researchers were able to identify numerous groups of cortical stem and precursor cells with distinct gene expression profiles. The team also found that these cells showed early signs of lineage diversification likely driven by transcriptional priming, a process by which a mother cell produces RNA for the sole purpose of passing it on to its daughter cells for later protein production.

Tarik Haydar

“Our study provides a new glimpse into the landscape of the developing brain. What we are seeing are new complex families of cells very early in development,” says Tarik Haydar, Ph.D., director of CNR at Children’s National, who led this study. “Understanding the role of these cells in forming the cerebral cortex is now possible in a way that wasn’t possible before.”

Using novel trajectory reconstruction methods, the team observed distinct developmental streams linking precursor cell types to particular excitatory neurons. After comparing the dataset of the preclinical model to a human cell database, notable similarities were found, such as the surprising cross-species presence of basal radial glial cells (bRGCs), an important type of progenitor cell previously thought to be found mainly in the primate brain.

“At a very high level, the study is important because we are directly testing a fundamental theory of brain development,” says Zhen Li, Ph.D., CNR research postdoctoral fellow and co-first author of the study. The results add support to the protomap theory, which posits that early stem and progenitor diversity paves the way for later neuronal diversity and cortical complexity. Furthermore, the results also hold exciting translational potential.

“There is evidence showing that neurodevelopmental diseases affect different populations of the neural stem cells differently,” says Dr. Li. “If we can have a better understanding of the complexity of these neural stem cells there is huge implication of disease prevention and treatment in the future.”

“If we can understand how this early landscape is affected in disorders, we can predict the resulting changes to the cortical architecture and then very narrowly define ways that groups of cells behave in these disorders,” adds Dr. Haydar. “If we can understand how the cortex normally achieves its complex architecture, then we have key entry points into improving the clinical course of a given disorder and improving quality of life.”

Future topics the researchers hope to study include the effects of developmental changes on brain function, the origin and operational importance of bRGCs, and the activity, connections and cognitive features enabled by different families of neurons.

Yuan Zhu

Study suggests glioblastoma tumors originate far from resulting tumors

Yuan Zhu

“The more we continue to learn about glioblastoma,” Yuan Zhu, Ph.D., says, “the more hope we can give to these patients who currently have few effective options.”

A pre-clinical model of glioblastoma, an aggressive type of cancer that can occur in the brain, suggests that this recalcitrant cancer originates from a pool of stem cells that can be a significant distance away from the resulting tumors. The findings of a new study, led by Children’s National Hospital researchers and published July 22 in the journal Nature Communications, suggest new ways to fight this deadly disease.

Despite decades of research, glioblastoma remains the most common and lethal primary brain tumor in adults, with a median survival of only 15 months from diagnosis, says study leader Yuan Zhu, Ph.D., the scientific director and endowed professor of the Gilbert Family Neurofibromatosis Institute at Children’s National. Unlike many cancers, which start out as low-grade tumors that are more treatable when they’re caught at an early stage, most glioblastomas are almost universally discovered as high-grade and aggressive lesions that are difficult to treat with the currently available modalities, including surgery, radiation and chemotherapy.

“Once the patient has neurological symptoms like headache, nausea, and vomiting, the tumor is already at an end state, and disease progression is very rapid,” Dr. Zhu says. “We know that the earlier you catch and treat cancers, the better the prognosis will be. But here, there’s no way to catch the disease early.”

However, some recent research in glioblastoma patients shows that the subventricular zone (SVZ) – an area that serves as the largest source of stem cells in the adult brain – contains cells with cancer-driving mutations that are shared with tumors found in other often far-distant brain regions.

To see if the SVZ might be the source for glioblastoma tumors, Dr. Zhu and his colleagues worked with pre-clinical models that carried a single genetic glitch: a mutation in a gene known as p53 that typically suppresses tumors. Mutations in p53 are known to be involved in glioblastoma and many other forms of cancer.

Using genetic tests and an approach akin to those used to study evolution, the researchers traced the cells that spurred both kinds of tumors back to the SVZ. Although both single and multiple tumors had spontaneously acquired mutations in a gene called Pten, another type of tumor suppressor, precursor cells for the single tumors appeared to acquire this mutation before they left the SVZ, while precursor cells for the multiple tumors developed this mutation after they left the stem cell niche. When the researchers genetically altered the animals to shut down the molecular pathway that loss of Pten activates, it didn’t stop cancer cells from forming. However, rather than migrate to distal areas of the brain, these malignant cells remained in the SVZ.

Dr. Zhu notes that these findings could help explain why glioblastoma is so difficult to identify the early precursor lesions and treat. This work may offer potential new options for attacking this cancer. If new glioblastoma tumors are seeded by cells from a repository in the SVZ, he explains, attacking those tumors won’t be enough to eradicate the cancer. Instead, new treatments might focus on this stem cell niche as target for treatment or even a zone for surveillance to prevent glioblastoma from developing in the first place.

Another option might be to silence the Pten-suppressed pathway through drugs, a strategy that’s currently being explored in various clinical trials. Although these agents haven’t shown yet that they can stop or reverse glioblastomas, they might be used to contain cancers in the SVZ as this strategy did in the pre-clinical model — a single location that might be easier to attack than tumors in multiple locations.

“The more we continue to learn about glioblastoma,” Dr. Zhu says, “the more hope we can give to these patients who currently have few effective options.”

Other Children’s National researchers who contributed to this study include Yinghua Li, Ph.D., Wei Li, Ph.D., Yuan Wang, Ph.D., Seckin Akgul, Ph.D., Daniel M. Treisman, Ph.D., Brianna R. Pierce, B.S., Cheng-Ying Ho, M.D. /Ph.D.

This work is supported by grants from the National Institutes of Health (2P01 CA085878-10A1, 1R01 NS053900 and R35CA197701).

tube labeled "CRISPR"

$2M from NIH to extract meaningful data from CRISPR screens

tube labeled "CRISPR"

Protein-coding genes comprise a mere 1% of DNA. While the other 99% of DNA was once derided as “junk,” it has become increasingly apparent that some non-coding genes enable essential cellular functions.

Wei Li, Ph.D., a principal investigator in the Center for Genetic Medicine Research at Children’s National in Washington, D.C., proposes to develop statistical and computational methods that sidestep existing hurdles that currently complicate genome-wide CRISPR/Cas9 screening. The National Institutes of Health has granted him $2.23 million in funding over five years to facilitate the systematic study of genes, non-coding elements and genetic interactions in various biological systems and disease types.

Right now, a large volume of screening data resides in the public domain, however it is difficult to compare data that is stored in one library with data stored at a different library. Over the course of the five-year project, Li aims to:

  • Improve functional gene identification from CRISPR screens.
  • Develop new analyses algorithms for screens targeting non-coding elements.
  • Study genetic interactions from CRISPR screens targeting gene pairs.

Ultimately, Li’s work will examine a range of disease types. Take cancer.

“There is abundant information already available in the public domain, like the Project Achilles  from the Broad Institute. However, no one is looking to see what is going in inside these tumors,” Li says. “Cancer is a disease of uncontrolled cell growth that makes tumors grow faster.”

Li and colleagues are going to ask which genes control this process by looking at genes that hit the brakes on cell growth as well as genes that pump the gas.

“You knock out one gene and then look: Does the cell grow faster or does it grow more slowly? If the cell grows more slowly, you know you are knocking out a gene that has the potential to stop tumor growth. If cells are growing faster, you know that you’re hitting genes that suppress cancer cell growth.”

In a nutshell, CRISPR (clustered regularly interspaced short palindromic repeats) screens knock out different genes and monitor changes in corresponding cell populations. When CRISPR first became popular, Li decided he wanted to do something with the technology. So, as a Postdoc at Harvard, he developed comprehensive computational algorithms for functional screens using CRISPR/Cas9.

To reach as many people as possible, he offered that MAGeCK/MAGeCK-VISPR software free to as many researchers as possible, providing source code and offering internet tutorials.

“So far, I think there are quite a lot of people using this. There have been more than 40,000 software downloads,” he adds. “It’s really exciting and revolutionary technology and, eventually, we hope the outcomes also will be exciting. We hope to find something really helpful for cancer patients.”

Research reported in this publication was supported by the National Human Genome Research Institute of the National Institutes of Health under award number R01HG010753.