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CRISPR/Cas9

Biomedical Research Leads Science’s 2021 Breakthroughs

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Artificial Antibody Therapies, AI-Powered Predictions of Protein Structures, Antiviral Pills for COVID-19, and CRISPR Fixes Genes Inside the Body

Hi everyone, I’m Larry Tabak. I’ve served as NIH’s Principal Deputy Director for over 11 years, and I will be the acting NIH director until a new permanent director is named. In my new role, my day-to-day responsibilities will certainly increase, but I promise to carve out time to blog about some of the latest research progress on COVID-19 and any other areas of science that catch my eye.

I’ve also invited the directors of NIH’s Institutes and Centers (ICs) to join me in the blogosphere and write about some of the cool science in their research portfolios. I will publish a couple of posts to start, then turn the blog over to our first IC director. From there, I envision alternating between posts from me and from various IC directors. That way, we’ll cover a broad array of NIH science and the tremendous opportunities now being pursued in biomedical research.

Since I’m up first, let’s start where the NIH Director’s Blog usually begins each year: by taking a look back at Science’s Breakthroughs of 2021. The breakthroughs were formally announced in December near the height of the holiday bustle. In case you missed the announcement, the biomedical sciences accounted for six of the journal Science’s 10 breakthroughs. Here, I’ll focus on four biomedical breakthroughs, the ones that NIH has played some role in advancing, starting with Science’s editorial and People’s Choice top-prize winner:

Breakthrough of the Year: AI-Powered Predictions of Protein Structure

The biochemist Christian Anfinsen, who had a distinguished career at NIH, shared the 1972 Nobel Prize in Chemistry, for work suggesting that the biochemical interactions among the amino acid building blocks of proteins were responsible for pulling them into the final shapes that are essential to their functions. In his Nobel acceptance speech, Anfinsen also made a bold prediction: one day it would be possible to determine the three-dimensional structure of any protein based on its amino acid sequence alone. Now, with advances in applying artificial intelligence to solve biological problems—Anfinsen’s bold prediction has been realized.

But getting there wasn’t easy. Every two years since 1994, research teams from around the world have gathered to compete against each other in developing computational methods for predicting protein structures from sequences alone. A score of 90 or above means that a predicted structure is extremely close to what’s known from more time-consuming work in the lab. In the early days, teams more often finished under 60.

In 2020, a London-based company called DeepMind made a leap with their entry called AlphaFold. Their deep learning approach—which took advantage of 170,000 proteins with known structures—most often scored above 90, meaning it could solve most protein structures about as well as more time-consuming and costly experimental protein-mapping techniques. (AlphaFold was one of Science’s runner-up breakthroughs last year.)

This year, the NIH-funded lab of David Baker and Minkyung Baek, University of Washington, Seattle, Institute for Protein Design, published that their artificial intelligence approach, dubbed RoseTTAFold, could accurately predict 3D protein structures from amino acid sequences with only a fraction of the computational processing power and time that AlphaFold required [1]. They immediately applied it to solve hundreds of new protein structures, including many poorly known human proteins with important implications for human health.

The DeepMind and RoseTTAFold scientists continue to solve more and more proteins [1,2], both alone and in complex with other proteins. The code is now freely available for use by researchers anywhere in the world. In one timely example, AlphaFold helped to predict the structural changes in spike proteins of SARS-CoV-2 variants Delta and Omicron [3]. This ability to predict protein structures, first envisioned all those years ago, now promises to speed fundamental new discoveries and the development of new ways to treat and prevent any number of diseases, making it this year’s Breakthrough of the Year.

Anti-Viral Pills for COVID-19

The development of the first vaccines to protect against COVID-19 topped Science’s 2020 breakthroughs. This year, we’ve also seen important progress in treating COVID-19, including the development of anti-viral pills.

First, there was the announcement in October of interim data from Merck, Kenilworth, NJ, and Ridgeback Biotherapeutics, Miami, FL, of a significant reduction in hospitalizations for those taking the anti-viral drug molnupiravir [4] (originally developed with an NIH grant to Emory University, Atlanta). Soon after came reports of a Pfizer anti-viral pill that might target SARS-CoV-2, the novel coronavirus that causes COVID-19, even more effectively. Trial results show that, when taken within three days of developing COVID-19 symptoms, the pill reduced the risk of hospitalization or death in adults at high risk of progressing to severe illness by 89 percent [5].

On December 22, the Food and Drug Administration (FDA) granted Emergency Use Authorization (EUA) for Pfizer’s Paxlovid to treat mild-to-moderate COVID-19 in people age 12 and up at high risk for progressing to severe illness, making it the first available pill to treat COVID-19 [6]. The following day, the FDA granted an EUA for Merck’s molnupiravir to treat mild-to-moderate COVID-19 in unvaccinated, high-risk adults for whom other treatment options aren’t accessible or recommended, based on a final analysis showing a 30 percent reduction in hospitalization or death [7].

Additional promising anti-viral pills for COVID-19 are currently in development. For example, a recent NIH-funded preclinical study suggests that a drug related to molnupiravir, known as 4’-fluorouridine, might serve as a broad spectrum anti-viral with potential to treat infections with SARS-CoV-2 as well as respiratory syncytial virus (RSV) [8].

Artificial Antibody Therapies

Before anti-viral pills came on the scene, there’d been progress in treating COVID-19, including the development of monoclonal antibody infusions. Three monoclonal antibodies now have received an EUA for treating mild-to-moderate COVID-19, though not all are effective against the Omicron variant [9]. This is also an area in which NIH’s Accelerating COVID-19 Therapeutic Interventions and Vaccines (ACTIV) public-private partnership has made big contributions.

Monoclonal antibodies are artificially produced versions of the most powerful antibodies found in animal or human immune systems, made in large quantities for therapeutic use in the lab. Until recently, this approach had primarily been put to work in the fight against conditions including cancer, asthma, and autoimmune diseases. That changed in 2021 with success using monoclonal antibodies against infections with SARS-CoV-2 as well as respiratory syncytial virus (RSV), human immunodeficiency virus (HIV), and other infectious diseases. This earned them a prominent spot among Science’s breakthroughs of 2021.

Monoclonal antibodies delivered via intravenous infusions continue to play an important role in saving lives during the pandemic. But, there’s still room for improvement, including new formulations highlighted on the blog last year that might be much easier to deliver.

CRISPR Fixes Genes Inside the Body

One of the most promising areas of research in recent years has been gene editing, including CRISPR/Cas9, for fixing misspellings in genes to treat or even cure many conditions. This year has certainly been no exception.

CRISPR is a highly precise gene-editing system that uses guide RNA molecules to direct a scissor-like Cas9 enzyme to just the right spot in the genome to cut out or correct disease-causing misspellings. Science highlights a small study reported in The New England Journal of Medicine by researchers at Intellia Therapeutics, Cambridge, MA, and Regeneron Pharmaceuticals, Tarrytown, NY, in which six people with hereditary transthyretin (TTR) amyloidosis, a condition in which TTR proteins build up and damage the heart and nerves, received an infusion of guide RNA and CRISPR RNA encased in tiny balls of fat [10]. The goal was for the liver to take them up, allowing Cas9 to cut and disable the TTR gene. Four weeks later, blood levels of TTR had dropped by at least half.

In another study not yet published, researchers at Editas Medicine, Cambridge, MA, injected a benign virus carrying a CRISPR gene-editing system into the eyes of six people with an inherited vision disorder called Leber congenital amaurosis 10. The goal was to remove extra DNA responsible for disrupting a critical gene expressed in the eye. A few months later, two of the six patients could sense more light, enabling one of them to navigate a dimly lit obstacle course [11]. This work builds on earlier gene transfer studies begun more than a decade ago at NIH’s National Eye Institute.

Last year, in a research collaboration that included former NIH Director Francis Collins’s lab at the National Human Genome Research Institute (NHGRI), we also saw encouraging early evidence in mice that another type of gene editing, called DNA base editing, might one day correct Hutchinson-Gilford Progeria Syndrome, a rare genetic condition that causes rapid premature aging. Preclinical work has even suggested that gene-editing tools might help deliver long-lasting pain relief. The technology keeps getting better, too. This isn’t the first time that gene-editing advances have landed on Science’s annual Breakthrough of the Year list, and it surely won’t be the last.

The year 2021 was a difficult one as the pandemic continued in the U.S. and across the globe, taking far too many lives far too soon. But through it all, science has been relentless in seeking and finding life-saving answers, from the rapid development of highly effective COVID-19 vaccines to the breakthroughs highlighted above.

As this list also attests, the search for answers has progressed impressively in other research areas during these difficult times. These groundbreaking discoveries are something in which we can all take pride—even as they encourage us to look forward to even bigger breakthroughs in 2022. Happy New Year!

References:

[1] Accurate prediction of protein structures and interactions using a three-track neural network. Baek M, DiMaio F, Anishchenko I, Dauparas J, Grishin NV, Adams PD, Read RJ, Baker D., et al. Science. 2021 Jul 15:eabj8754.

[2] Highly accurate protein structure prediction with AlphaFold. Jumper J, Evans R, Pritzel A, Green T, Senior AW, Kavukcuoglu K, Kohli P, Hassabis D. et al. Nature. 2021 Jul 15.

[3] Structural insights of SARS-CoV-2 spike protein from Delta and Omicron variants. Sadek A, Zaha D, Ahmed MS. preprint bioRxiv. 2021 Dec 9.

[4] Merck and Ridgeback’s investigational oral antiviral molnupiravir reduced the risk of hospitalization or death by approximately 50 Percent compared to placebo for patients with mild or moderate COVID-19 in positive interim analysis of phase 3 study. Merck. 1 Oct 2021.

[5] Pfizer’s novel COVID-19 oral antiviral treatment candidate reduced risk of hospitalization or death by 89% in interim analysis of phase 2/3 EPIC-HR Study. Pfizer. 5 November 52021.

[6] Coronavirus (COVID-19) Update: FDA authorizes first oral antiviral for treatment of COVID-19. Food and Drug Administration. 22 Dec 2021.

[7] Coronavirus (COVID-19) Update: FDA authorizes additional oral antiviral for treatment of COVID-19 in certain adults. Food and Drug Administration. 23 Dec 2021.

[8] 4′-Fluorouridine is an oral antiviral that blocks respiratory syncytial virus and SARS-CoV-2 replication. Sourimant J, Lieber CM, Aggarwal M, Cox RM, Wolf JD, Yoon JJ, Toots M, Ye C, Sticher Z, Kolykhalov AA, Martinez-Sobrido L, Bluemling GR, Natchus MG, Painter GR, Plemper RK. Science. 2021 Dec 2.

[9] Anti-SARS-CoV-2 monoclonal antibodies. NIH COVID-19 Treatment Guidelines. 16 Dec 2021.

[10] CRISPR-Cas9 in vivo gene editing for transthyretin amyloidosis. Gillmore JD, Gane E, Taubel J, Kao J, Fontana M, Maitland ML, Seitzer J, O’Connell D, Walsh KR, Wood K, Phillips J, Xu Y, Amaral A, Boyd AP, Cehelsky JE, McKee MD, Schiermeier A, Harari O, Murphy A, Kyratsous CA, Zambrowicz B, Soltys R, Gutstein DE, Leonard J, Sepp-Lorenzino L, Lebwohl D. N Engl J Med. 2021 Aug 5;385(6):493-502.

[11] Editas Medicine announces positive initial clinical data from ongoing phase 1/2 BRILLIANCE clinical trial of EDIT-101 For LCA10. Editas Medicine. 29 Sept 2021.

Links:

Structural Biology (National Institute of General Medical Sciences/NIH)

The Structures of Life (NIGMS)

COVID-19 Research (NIH)

2021 Science Breakthrough of the Year (American Association for the Advancement of Science, Washington, D.C)


Engineering a Better Way to Deliver Therapeutic Genes to Muscles

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Green adenovirus delivers therapeutic genes to muscles which glow green

Amid all the progress toward ending the COVID-19 pandemic, it’s worth remembering that researchers here and around the world continue to make important advances in tackling many other serious health conditions. As an inspiring NIH-supported example, I’d like to share an advance on the use of gene therapy for treating genetic diseases that progressively degenerate muscle, such as Duchenne muscular dystrophy (DMD).

As published recently in the journal Cell, researchers have developed a promising approach to deliver therapeutic genes and gene editing tools to muscle more efficiently, thus requiring lower doses [1]. In animal studies, the new approach has targeted muscle far more effectively than existing strategies. It offers an exciting way forward to reduce unwanted side effects from off-target delivery, which has hampered the development of gene therapy for many conditions.

In boys born with DMD (it’s an X-linked disease and therefore affects males), skeletal and heart muscles progressively weaken due to mutations in a gene encoding a critical muscle protein called dystrophin. By age 10, most boys require a wheelchair. Sadly, their life expectancy remains less than 30 years.

The hope is gene therapies will one day treat or even cure DMD and allow people with the disease to live longer, high-quality lives. Unfortunately, the benign adeno-associated viruses (AAVs) traditionally used to deliver the healthy intact dystrophin gene into cells mostly end up in the liver—not in muscles. It’s also the case for gene therapy of many other muscle-wasting genetic diseases.

The heavy dose of viral vector to the liver is not without concern. Recently and tragically, there have been deaths in a high-dose AAV gene therapy trial for X-linked myotubular myopathy (XLMTM), a different disorder of skeletal muscle in which there may already be underlying liver disease, potentially increasing susceptibility to toxicity.

To correct this concerning routing error, researchers led by Mohammadsharif Tabebordbar in the lab of Pardis Sabeti, Broad Institute of MIT and Harvard and Harvard University, Cambridge, MA, have now assembled an optimized collection of AAVs. They have been refined to be about 10 times better at reaching muscle fibers than those now used in laboratory studies and clinical trials. In fact, researchers call them myotube AAVs, or MyoAAVs.

MyoAAVs can deliver therapeutic genes to muscle at much lower doses—up to 250 times lower than what’s needed with traditional AAVs. While this approach hasn’t yet been tried in people, animal studies show that MyoAAVs also largely avoid the liver, raising the prospect for more effective gene therapies without the risk of liver damage and other serious side effects.

In the Cell paper, the researchers demonstrate how they generated MyoAAVs, starting out with the commonly used AAV9. Their goal was to modify the outer protein shell, or capsid, to create an AAV that would be much better at specifically targeting muscle. To do so, they turned to their capsid engineering platform known as, appropriately enough, DELIVER. It’s short for Directed Evolution of AAV capsids Leveraging In Vivo Expression of transgene RNA.

Here’s how DELIVER works. The researchers generate millions of different AAV capsids by adding random strings of amino acids to the portion of the AAV9 capsid that binds to cells. They inject those modified AAVs into mice and then sequence the RNA from cells in muscle tissue throughout the body. The researchers want to identify AAVs that not only enter muscle cells but that also successfully deliver therapeutic genes into the nucleus to compensate for the damaged version of the gene.

This search delivered not just one AAV—it produced several related ones, all bearing a unique surface structure that enabled them specifically to target muscle cells. Then, in collaboration with Amy Wagers, Harvard University, Cambridge, MA, the team tested their MyoAAV toolset in animal studies.

The first cargo, however, wasn’t a gene. It was the gene-editing system CRISPR-Cas9. The team found the MyoAAVs correctly delivered the gene-editing system to muscle cells and also repaired dysfunctional copies of the dystrophin gene better than the CRISPR cargo carried by conventional AAVs. Importantly, the muscles of MyoAAV-treated animals also showed greater strength and function.

Next, the researchers teamed up with Alan Beggs, Boston Children’s Hospital, and found that MyoAAV was effective in treating mouse models of XLMTM. This is the very condition mentioned above, in which very high dose gene therapy with a current AAV vector has led to tragic outcomes. XLMTM mice normally die in 10 weeks. But, after receiving MyoAAV carrying a corrective gene, all six mice had a normal lifespan. By comparison, mice treated in the same way with traditional AAV lived only up to 21 weeks of age. What’s more, the researchers used MyoAAV at a dose 100 times lower than that currently used in clinical trials.

While further study is needed before this approach can be tested in people, MyoAAV was also used to successfully introduce therapeutic genes into human cells in the lab. This suggests that the early success in animals might hold up in people. The approach also has promise for developing AAVs with potential for targeting other organs, thereby possibly providing treatment for a wide range of genetic conditions.

The new findings are the result of a decade of work from Tabebordbar, the study’s first author. His tireless work is also personal. His father has a rare genetic muscle disease that has put him in a wheelchair. With this latest advance, the hope is that the next generation of promising gene therapies might soon make its way to the clinic to help Tabebordbar’s father and so many other people.

Reference:

[1] Directed evolution of a family of AAV capsid variants enabling potent muscle-directed gene delivery across species. Tabebordbar M, Lagerborg KA, Stanton A, King EM, Ye S, Tellez L, Krunnfusz A, Tavakoli S, Widrick JJ, Messemer KA, Troiano EC, Moghadaszadeh B, Peacker BL, Leacock KA, Horwitz N, Beggs AH, Wagers AJ, Sabeti PC. Cell. 2021 Sep 4:S0092-8674(21)01002-3.

Links:

Muscular Dystrophy Information Page (National Institute of Neurological Disorders and Stroke/NIH)

X-linked myotubular myopathy (Genetic and Rare Diseases Information Center/National Center for Advancing Translational Sciences/NIH)

Somatic Cell Genome Editing (Common Fund/NIH)

Mohammadsharif Tabebordbar (Broad Institute of MIT and Harvard and Harvard University, Cambridge, MA)

Sabeti Lab (Broad Institute of MIT and Harvard and Harvard University)

NIH Support: Eunice Kennedy Shriver National Institute of Child Health and Human Development; Common Fund


Could CRISPR Gene-Editing Technology Be an Answer to Chronic Pain?

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Active Neurons
Credit: iStock/Firstsignal

Gene editing has shown great promise as a non-heritable way to treat a wide range of conditions, including many genetic diseases and more recently, even COVID-19. But could a version of the CRISPR gene-editing tool also help deliver long-lasting pain relief without the risk of addiction associated with prescription opioid drugs?

In work recently published in the journal Science Translational Medicine, researchers demonstrated in mice that a modified version of the CRISPR system can be used to “turn off” a gene in critical neurons to block the transmission of pain signals [1]. While much more study is needed and the approach is still far from being tested in people, the findings suggest that this new CRISPR-based strategy could form the basis for a whole new way to manage chronic pain.

This novel approach to treating chronic pain occurred to Ana Moreno, the study’s first author, when she was a Ph.D. student in the NIH-supported lab of Prashant Mali, University of California, San Diego. Mali had been studying a wide range of novel gene- and cell-based therapeutics. While reading up on both, Moreno landed on a paper about a mutation in a gene that encodes a pain-enhancing protein in spinal neurons called NaV1.7.

Moreno read that kids born with a loss-of-function mutation in this gene have a rare condition known as congenital insensitivity to pain (CIP). They literally don’t sense and respond to pain. Although these children often fail to recognize serious injuries because of the absence of pain to alert them, they have no other noticeable physical effects of the condition.

For Moreno, something clicked. What if it were possible to engineer a new kind of treatment—one designed to turn this gene down or fully off and stop people from feeling chronic pain?

Moreno also had an idea about how to do it. She’d been working on repressing or “turning off” genes using a version of CRISPR known as “dead” Cas9 [2]. In CRISPR systems designed to edit DNA, the Cas9 enzyme is often likened to a pair of scissors. Its job is to cut DNA in just the right spot with the help of an RNA guide. However, CRISPR-dead Cas9 no longer has any ability to cut DNA. It simply sticks to its gene target and blocks its expression. Another advantage is that the system won’t lead to any permanent DNA changes, since any treatment based on CRISPR-dead Cas9 might be safely reversed.

After establishing that the technique worked in cells, Moreno and colleagues moved to studies of laboratory mice. They injected viral vectors carrying the CRISPR treatment into mice with different types of chronic pain, including inflammatory and chemotherapy-induced pain.

Moreno and colleagues determined that all the mice showed evidence of durable pain relief. Remarkably, the treatment also lasted for three months or more and, importantly, without any signs of side effects. The researchers are also exploring another approach to do the same thing using a different set of editing tools called zinc finger nucleases (ZFNs).

The researchers say that one of these approaches might one day work for people with a large number of chronic pain conditions that involve transmission of the pain signal through NaV1.7. That includes diabetic polyneuropathy, sciatica, and osteoarthritis. It also could provide relief for patients undergoing chemotherapy, along with those suffering from many other conditions. Moreno and Mali have co-founded the spinoff company Navega Therapeutics, San Diego, CA, to work on the preclinical steps necessary to help move their approach closer to the clinic.

Chronic pain is a devastating public health problem. While opioids are effective for acute pain, they can do more harm than good for many chronic pain conditions, and they are responsible for a nationwide crisis of addiction and drug overdose deaths [3]. We cannot solve any of these problems without finding new ways to treat chronic pain. As we look to the future, it’s hopeful that innovative new therapeutics such as this gene-editing system could one day help to bring much needed relief.

References:

[1] Long-lasting analgesia via targeted in situ repression of NaV1.7 in mice. Moreno AM, Alemán F, Catroli GF, Hunt M, Hu M, Dailamy A, Pla A, Woller SA, Palmer N, Parekh U, McDonald D, Roberts AJ, Goodwill V, Dryden I, Hevner RF, Delay L, Gonçalves Dos Santos G, Yaksh TL, Mali P. Sci Transl Med. 2021 Mar 10;13(584):eaay9056.

[2] Nuclease dead Cas9 is a programmable roadblock for DNA replication. Whinn KS, Kaur G, Lewis JS, Schauer GD, Mueller SH, Jergic S, Maynard H, Gan ZY, Naganbabu M, Bruchez MP, O’Donnell ME, Dixon NE, van Oijen AM, Ghodke H. Sci Rep. 2019 Sep 16;9(1):13292.

[3] Drug Overdose Deaths. Centers for Disease Control and Prevention.

Links:

Congenital insensitivity to pain (National Center for Advancing Translational Sciences/NIH)

Opioids (National Institute on Drug Abuse/NIH)

Mali Lab (University of California, San Diego)

Navega Therapeutics (San Diego, CA)

NIH Support: National Human Genome Research Institute; National Cancer Institute; National Institute of General Medical Sciences; National Institute of Neurological Disorders and Stroke


What A Year It Was for Science Advances!

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Science Breakthroughs of the Year 2020

At the close of every year, editors and writers at the journal Science review the progress that’s been made in all fields of science—from anthropology to zoology—to select the biggest advance of the past 12 months. In most cases, this Breakthrough of the Year is as tough to predict as the Oscar for Best Picture. Not in 2020. In a year filled with a multitude of challenges posed by the emergence of the deadly coronavirus disease 2019 (COVID-2019), the breakthrough was the development of the first vaccines to protect against this pandemic that’s already claimed the lives of more than 360,000 Americans.

In keeping with its annual tradition, Science also selected nine runner-up breakthroughs. This impressive list includes at least three areas that involved efforts supported by NIH: therapeutic applications of gene editing, basic research understanding HIV, and scientists speaking up for diversity. Here’s a quick rundown of all the pioneering advances in biomedical research, both NIH and non-NIH funded:

Shots of Hope. A lot of things happened in 2020 that were unprecedented. At the top of the list was the rapid development of COVID-19 vaccines. Public and private researchers accomplished in 10 months what normally takes about 8 years to produce two vaccines for public use, with more on the way in 2021. In my more than 25 years at NIH, I’ve never encountered such a willingness among researchers to set aside their other concerns and gather around the same table to get the job done fast, safely, and efficiently for the world.

It’s also pretty amazing that the first two conditionally approved vaccines from Pfizer and Moderna were found to be more than 90 percent effective at protecting people from infection with SARS-CoV-2, the coronavirus that causes COVID-19. Both are innovative messenger RNA (mRNA) vaccines, a new approach to vaccination.

For this type of vaccine, the centerpiece is a small, non-infectious snippet of mRNA that encodes the instructions to make the spike protein that crowns the outer surface of SARS-CoV-2. When the mRNA is injected into a shoulder muscle, cells there will follow the encoded instructions and temporarily make copies of this signature viral protein. As the immune system detects these copies, it spurs the production of antibodies and helps the body remember how to fend off SARS-CoV-2 should the real thing be encountered.

It also can’t be understated that both mRNA vaccines—one developed by Pfizer and the other by Moderna in conjunction with NIH’s National Institute of Allergy and Infectious Diseases—were rigorously evaluated in clinical trials. Detailed data were posted online and discussed in all-day meetings of an FDA Advisory Committee, open to the public. In fact, given the high stakes, the level of review probably was more scientifically rigorous than ever.

First CRISPR Cures: One of the most promising areas of research now underway involves gene editing. These tools, still relatively new, hold the potential to fix gene misspellings—and potentially cure—a wide range of genetic diseases that were once to be out of reach. Much of the research focus has centered on CRISPR/Cas9. This highly precise gene-editing system relies on guide RNA molecules to direct a scissor-like Cas9 enzyme to just the right spot in the genome to cut out or correct a disease-causing misspelling.

In late 2020, a team of researchers in the United States and Europe succeeded for the first time in using CRISPR to treat 10 people with sickle cell disease and transfusion-dependent beta thalassemia. As published in the New England Journal of Medicine, several months after this non-heritable treatment, all patients no longer needed frequent blood transfusions and are living pain free [1].

The researchers tested a one-time treatment in which they removed bone marrow from each patient, modified the blood-forming hematopoietic stem cells outside the body using CRISPR, and then reinfused them into the body. To prepare for receiving the corrected cells, patients were given toxic bone marrow ablation therapy, in order to make room for the corrected cells. The result: the modified stem cells were reprogrammed to switch back to making ample amounts of a healthy form of hemoglobin that their bodies produced in the womb. While the treatment is still risky, complex, and prohibitively expensive, this work is an impressive start for more breakthroughs to come using gene editing technologies. NIH, including its Somatic Cell Genome Editing program, continues to push the technology to accelerate progress and make gene editing cures for many disorders simpler and less toxic.

Scientists Speak Up for Diversity: The year 2020 will be remembered not only for COVID-19, but also for the very public and inescapable evidence of the persistence of racial discrimination in the United States. Triggered by the killing of George Floyd and other similar events, Americans were forced to come to grips with the fact that our society does not provide equal opportunity and justice for all. And that applies to the scientific community as well.

Science thrives in safe, diverse, and inclusive research environments. It suffers when racism and bigotry find a home to stifle diversity—and community for all—in the sciences. For the nation’s leading science institutions, there is a place and a calling to encourage diversity in the scientific workplace and provide the resources to let it flourish to everyone’s benefit.

For those of us at NIH, last year’s peaceful protests and hashtags were noticed and taken to heart. That’s one of the many reasons why we will continue to strengthen our commitment to building a culturally diverse, inclusive workplace. For example, we have established the NIH Equity Committee. It allows for the systematic tracking and evaluation of diversity and inclusion metrics for the intramural research program for each NIH institute and center. There is also the recently founded Distinguished Scholars Program, which aims to increase the diversity of tenure track investigators at NIH. Recently, NIH also announced that it will provide support to institutions to recruit diverse groups or “cohorts” of early-stage research faculty and prepare them to thrive as NIH-funded researchers.

AI Disentangles Protein Folding: Proteins, which are the workhorses of the cell, are made up of long, interconnected strings of amino acids that fold into a wide variety of 3D shapes. Understanding the precise shape of a protein facilitates efforts to figure out its function, its potential role in a disease, and even how to target it with therapies. To gain such understanding, researchers often try to predict a protein’s precise 3D chemical structure using basic principles of physics—including quantum mechanics. But while nature does this in real time zillions of times a day, computational approaches have not been able to do this—until now.

Of the roughly 170,000 proteins mapped so far, most have had their structures deciphered using powerful imaging techniques such as x-ray crystallography and cryo–electron microscopy (cryo-EM). But researchers estimate that there are at least 200 million proteins in nature, and, as amazing as these imaging techniques are, they are laborious, and it can take many months or years to solve 3D structure of a single protein. So, a breakthrough certainly was needed!

In 2020, researchers with the company Deep Mind, London, developed an artificial intelligence (AI) program that rapidly predicts most protein structures as accurately as x-ray crystallography and cryo-EM can map them [2]. The AI program, called AlphaFold, predicts a protein’s structure by computationally modeling the amino acid interactions that govern its 3D shape.

Getting there wasn’t easy. While a complete de novo calculation of protein structure still seemed out of reach, investigators reasoned that they could kick start the modeling if known structures were provided as a training set to the AI program. Utilizing a computer network built around 128 machine learning processors, the AlphaFold system was created by first focusing on the 170,000 proteins with known structures in a reiterative process called deep learning. The process, which is inspired by the way neural networks in the human brain process information, enables computers to look for patterns in large collections of data. In this case, AlphaFold learned to predict the underlying physical structure of a protein within a matter of days. This breakthrough has the potential to accelerate the fields of structural biology and protein research, fueling progress throughout the sciences.

How Elite Controllers Keep HIV at Bay: The term “elite controller” might make some people think of video game whizzes. But here, it refers to the less than 1 percent of people living with human immunodeficiency virus (HIV) who’ve somehow stayed healthy for years without taking antiretroviral drugs. In 2020, a team of NIH-supported researchers figured out why this is so.

In a study of 64 elite controllers, published in the journal Nature, the team discovered a link between their good health and where the virus has inserted itself in their genomes [3]. When a cell transcribes a gene where HIV has settled, this so-called “provirus,” can produce more virus to infect other cells. But if it settles in a part of a chromosome that rarely gets transcribed, sometimes called a gene desert, the provirus is stuck with no way to replicate. Although this discovery won’t cure HIV/AIDS, it points to a new direction for developing better treatment strategies.

In closing, 2020 presented more than its share of personal and social challenges. Among those challenges was a flood of misinformation about COVID-19 that confused and divided many communities and even families. That’s why the editors and writers at Science singled out “a second pandemic of misinformation” as its Breakdown of the Year. This divisiveness should concern all of us greatly, as COVID-19 cases continue to soar around the country and our healthcare gets stretched to the breaking point. I hope and pray that we will all find a way to come together, both in science and in society, as we move forward in 2021.

References:

[1] CRISPR-Cas9 gene editing for sickle cell disease and β-thalassemia. Frangoul H et al. N Engl J Med. 2020 Dec 5.

[2] ‘The game has changed.’ AI triumphs at protein folding. Service RF. Science. 04 Dec 2020.

[3] Distinct viral reservoirs in individuals with spontaneous control of HIV-1. Jiang C et al. Nature. 2020 Sep;585(7824):261-267.

Links:

COVID-19 Research (NIH)

2020 Science Breakthrough of the Year (American Association for the Advancement of Science, Washington, D.C)


Discussing the Long Arc of Discovery with NIH’s Newest Nobelist

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Discussion with Dr. Harvey Alter

It’s been a tough year for our whole world because of everything that’s happening as a result of the coronavirus disease 2019 (COVID-19) pandemic. Yet there are bright spots that still shine through, and this week brought some fantastic news about NIH-supported researchers being named 2020 Nobel Prize Laureates for their pioneering work in two important fields: Chemistry and Physiology or Medicine.

In the wee hours of Wednesday morning, NIH grantee Jennifer A. Doudna, a biochemist at the University of California, Berkeley, got word that she and Emmanuelle Charpentier, a microbiologist at the Max Planck Institute for Infection Biology, Berlin, Germany, had won the 2020 Nobel Prize in Chemistry for developing the CRISPR/cas approach to genome editing. Doudna has received continuous NIH funding since 1997, mainly from the National Institute of General Medical Sciences and National Human Genome Research Institute.

The CRISPR/cas system, which consists of a short segment of RNA attached to the cas enzyme, provides the ability to make very precise changes in the sequence, or spelling, of the genetic instruction books of humans and other species. If used to make non-heritable edits in relevant tissues, such technology holds enormous potential to treat or even cure a wide range of devastating diseases, including thousands of genetic disorders where the DNA misspelling is precisely known.

Just two days before Doudna learned of her big award, a scientist who’s spent almost his entire career at the NIH campus in Bethesda, MD, received news that he too was getting a Nobel—the 2020 Nobel Prize in Physiology or Medicine. Harvey Alter, a senior scholar in the NIH Clinical Center’s Transfusion Medicine Department, was recognized for his contributions in identifying the potentially deadly hepatitis C virus. He shares this year’s prize with Michael Houghton, now with University of Alberta, Edmonton, and Charles M. Rice, The Rockefeller University, New York, who’s received continuous NIH funding since 1987, mainly from the National Institute of Allergy and Infectious Diseases.

In a long arc of discovery rooted in basic, translational, and clinical research that spanned several decades, Alter and his colleagues doggedly pursued biological clues that at first led to tests, then life-saving treatments, and, today, the very real hope of eradicating the global health threat posed by hepatitis C infections.

We at NIH are particularly proud of the fact that Alter is the sixth Nobel Prize winner—and the first in 26 years—to have done the entirety of his award-winning research in our Intramural Research Program. So, I jumped at the opportunity to talk with Harvey on NIH’s Facebook Live and Twitter chats just hours after he got the good news on Monday. Here’s a condensed version of our conversation, which took place on the NIH campus, but at a safe physical distance to minimize the risk of COVID-19 spread.

Collins: Harvey, let me start off by asking, how did you find out you’d won the Nobel Prize?

Alter: At 4:15 this morning. I was asleep and heard the telephone ringing. I ignored it. Five minutes later, I got another call. Now, I’m getting kind of perturbed. But I ignored it, thinking the call must be some kind of solicitation. Then, the phone rang a third time. I answered it, prepared to tell the person on the other end not to call me anymore. I heard a man’s voice say, “I’m the Secretary General of the Nobel Prize, calling you from Stockholm.” At that point, I just froze.

Collins: Did you think it might be a hoax?

Alter: No, I didn’t think it was a hoax. But I wasn’t expecting to win the prize. I knew about three years ago that I’d been on a Nobel list. But it didn’t happen, and I just forgot about it. Truthfully, I didn’t know that today was the day that the announcement was being made. The news came as a complete shock.

Collins: Please say a few words about viral hepatitis. What is it?

Alter: Sure. Viral hepatitis is an infection of the liver that causes inflammation and can lead to scarring, or cirrhosis. Early in my career, two viruses were known to cause the disease. One was the hepatitis A virus. You got it from consuming contaminated water or food. The second was the hepatitis B virus, which has a blood-borne transmission, typically from blood transfusions. In the 1970s, we realized that some other agent was causing most of the hepatitis from blood transfusions. Since it wasn’t A and it wasn’t B, we cleverly decided to call it: non-A, non-B. We did that because we hadn’t yet proven that the causative agent was a virus.

Collins: So, even though you screened donor units for the hepatitis B virus to eliminate tainted blood, people were still getting hepatitis from blood transfusions. How did you go about trying to solve this mystery?

Alter: The main thing was to follow patients prospectively, meaning forward in time. We drew a blood sample before they were transfused, and then serially afterwards. We saved those samples and also the donor samples to compare them. Using a liver function test, we found that 30 percent of patients who had open heart surgery at NIH prior to 1970 developed liver abnormalities indicative of hepatitis. That’s 1 in 3 people.

We then looked for the reasons. We found the main one was our source of blood. We were buying blood, which was then in short supply, from commercial laboratories. It turned out that their paid donors were engaging in high-risk behaviors [Note: like IV drug users sharing hypodermic needles]. We immediately stopped using these laboratories, and, through various other measures, we got the rate down to around 4 percent in 1987.

That’s when Michael Houghton, then at Chiron Corp. and a co-recipient of this year’s prize, cloned the virus. Think about it, he and his colleagues looked at 6 million clones and found just one that reacted with the convalescent serum of a patient with non-A, non-B. In other words, having contracted the virus, the patient already made antibodies against it that were present in the serum. If that one clone came from the virus, the antibodies in the serum would recognize it. They did, and Chiron then developed an assay to detect antibodies to the virus.

Collins: And that’s when they contacted you.

Alter: Yes, they wanted to use our panel of patient blood samples that had fooled a lot of people who claimed to have developed a non-A, non-B assay. Nobody else had “broken” this panel, but the Chiron Corp. did. We found that every case of non-A, non-B was really hepatitis C, the agent that they had cloned. Hepatitis C was the missing piece. As far as we could tell, there were no other agents beside hepatitis B and C that would result in transfusion transmission of the disease.

Collins: This story is clearly one of persistence. So, say something about persistence as an important characteristic of a scientist. You’re a great example of someone who was always looking out for opportunities that might not have seemed so promising at first.

Alter: I first learned persistence from Dr. Baruch Blumberg, my first NIH mentor who discovered the hepatitis B virus in 1967. [Note: Other NIH researchers identified the hepatitis A virus in 1977] The discovery started when we found this “Australian antigen,” a molecular structure that the immune system recognizes as foreign and attacks. It was a serendipitous finding that could have been easily just dropped. But he just kept at it, kept at it, kept at it. He had this famous wall where he diagrammed his hypotheses with all the contingencies if one worked or failed. Then, all of a sudden, the antigen was associated with hepatitis B. It became the basis of the hepatitis B vaccine, which is highly effective and used throughout the world. Dr. Blumberg won the Nobel Prize for his work on the hepatitis B virus in 1976.

Collins: Sometimes people look at NIH and ask why we don’t focus all of our efforts on curing a particular disease. I keep answering, ‘Wait a moment, we don’t know enough to know how to do that.’ What’s the balance that we ought to be seeking between basic research and clinical applications?

Alter: There is this tendency now to pursue highly directed research to solve a problem. That’s certainly how biopharma works. They want a payoff. The NIH is different. It’s a place where you can pursue your scientific interests, wherever they lead. The NIH leadership understands that the details of a problem often aren’t obvious at first. Researchers need to be allowed to observe things and then to pursue their leads as far as possible, with the understanding that not everything will work out. I think it’s very important to keep this basic research component in parallel with the more clinical applications. In the case of hepatitis C, it started as a clinical problem that led to a basic research investigation, which led back to a clinical problem. It was bedside-to-bench-to-bedside.

Collins: Are people still getting infected with hepatitis C?

Alter: Yes, hepatitis C remains a global problem. Seventy million people have contracted the virus, though the majority are generally asymptomatic, meaning they don’t get sick from it. Instead, they carry around the virus for decades without knowing it. That’s because the hepatitis C virus likes to persist, and our immune system doesn’t seem to be able to get rid of it easily.

However, some of those infected will have bad outcomes, such as cirrhosis or cancer of the liver. But there’s no way of knowing who will and who won’t get sick over time. The trick now is to identify people when they’re asymptomatic and without obvious disease.

That involves testing. We’re in a unique position with hepatitis C, where we have great tests that are highly sensitive and very specific to the virus. We also have great treatments. We can cure everybody who is tested and found to be positive.

Collins: People may be surprised to hear that. Here is a chronic viral illness, for which we actually have a cure. That’s come along fairly recently. Say a bit more about that—it’s such a great story of success.

Alter: For many years, the only treatment for hepatitis C was interferon, a very difficult treatment that initially had only about a 6 percent cure rate. With further progress, it got up to around 50 percent. But the big breakthrough came in the late 1990s when Gilead Corp., having the sequenced genome of the hepatitis C virus, deduced what it needs to replicate. If we know what it needs and we interfere with that, we can stop the replication. Gilead came out with a blockbuster drug that, now in combination with another drug, aims at two different sites on the virus and cures at least 98 percent of people. It’s an oral therapy taken for only 12 weeks, sometimes as little as 8 weeks, and with virtually no side-effects. It’s like a miracle drug.

Collins: What would you say to somebody who is thinking about becoming a scientist? How do you pick an area of research that will be right for you?

Alter: It’s a tough question. Medical research is very difficult, but there’s nothing more rewarding than doing something for patients and to see a good outcome like we had with hepatitis C.

The best path forward is to work for somebody who’s already an established investigator and a good teacher. Work in his or her lab for a few years and get involved in a project. I’ve learned not get into a lot of projects. Get into something where you can become the expert and pursue it.
The other thing is to collaborate. There’s no way that one person can do everything these days. You need too much technology and lots of different areas of expertise.

Collins: You took on a high-risk project in which you didn’t know that you’d find the answer. What’s the right balance between a project that you know will be productive, and something that might be risky, but, boy, if it works, could be transformative? How did you decide which of those paths to go?

Alter: I don’t think I decided. I just went! But there were interim rewards. Finding that the paid donors were bad was a reward and it had a big impact. And the different donor testing, decreasing the amount of blood [transfused], there were all kinds of steps along the way that gave you a reward. Now, did I think that there would be a treatment, an eradication of post-transfusion hepatitis at the end of my line? No, I didn’t.

And it wouldn’t have happened if it was only me. I just got the ball rolling. But it needed Houghton’s group. It needed the technology of Charlie Rice, a co-recipient of this year’s Nobel Prize. It needed joint company involvement. So, it required massive cooperation, and I have to say that here at NIH, Bob Purcell did most of the really basic work in his lab. Patrizia Farci, my closest collaborator, does things that I can’t do. You just need people who have a different expertise.

Collins: Harvey, it’s been maybe six hours since you found out that you won the Nobel Prize. How are you going to spend the rest of your day?

Alter: Well, I have to tell you a story that just happened. We had a press conference earlier today at NIH. Afterwards, I wanted to return to my NIH office and the easiest route was through the parking garage across the street from where we held the press conference. When I entered the garage, a security guard said, “You can’t come in, you haven’t been screened for COVID.” I assured him that I had been screened when I drove onto the NIH campus. He repeated that I had to go around to the front of the building to get screened.

Finally, I said to him, “Would it make any difference if I told you that I won the Nobel Prize today?” He replied, ‘That’s nice, but you must go around to the front of the building.’” So, winning the Nobel doesn’t give you immediate rewards!

Collins: Let me find that security guard and give him a bonus for doing a good job. Well, Harvey, will there be that trip to Stockholm coming up in December?

Alter: Not this year. I’ve heard that they will invite us to Stockholm next year to receive the award. But there’s going to be something in the US. I don’t know what it will be. I’ll invite you.

Collins: I will be glad to take part in the celebration. Well, Harvey, I really want to thank you for taking some time on this special day to reflect on your career and how the Nobel Committee came calling at 4:30 this morning. We’re really proud of you!

Alter: Thank you.

Links:

Hepatitis C (National Institute of Diabetes and Digestive and Kidney Diseases/NIH)

The Nobel Assembly at Karolinska Institutet has today decided to award the 2020 Nobel Prize in Physiology or Medicine jointly to Harvey J. Alter, Michael Houghton and Charles M. Rice for the discovery of Hepatitis C virus,” Nobel Prize announcement, October 5,2020.

Harvey Alter (Clinical Center/NIH)

The Road Not Taken, or How I Learned to Love the Liver: A Personal Perspective on Hepatitis History” Alter HJ, Hepatology. 2014 Jan;59(1):4-12.

Reflections on the History of HCV: A Posthumous Examination.” Alter HJ, Farci P, Bukh J, Purcell RH. Clinical Liver Disease, 15:1, Feb 2020.

Is Elimination of Hepatitis B and C a Pipe Dream or Reality?” Alter HJ, Chisari FV. Gastroenterology. 2019 Jan;156(2):294-296.

Michael Houghton (University of Alberta, Edmonton)

Charles Rice (The Rockefeller University, New York)

What is genome editing? (National Human Genome Research Institute/NIH)

Jennifer Doudna (University of California, Berkeley)

Emmanuelle Charpentier (Max Planck Institute for Infection Biology, Berlin, Germany)


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