8 tools that helped us tackle the coronavirus

Adenoviral vectors

Viruses might be nature’s most efficient gene-delivery vehicles. At its essence, a virus is simply a package that shuttles genes into a host cell, which it then hijacks to make more viruses. About 4 decades ago, scientists started hijacking the viruses themselves in an attempt to make novel vaccines. The fruits of that labor have led to some of the most advanced vaccines for fighting COVID-19.

In early 2020, scientists around the world—most prominently, those at the University of Oxford (later joined by AstraZeneca) in the UK, Johnson & Johnson in the US, CanSino Biologics in China, and the Gamaleya National Center of Epidemiology and Microbiology in Russia—began using an old but largely experimental vaccine technology to make vaccines for COVID-19.

The technology relies on adenoviruses, perhaps best known for causing the common cold. Scientists began tinkering with adenoviruses in the 1980s, first removing genes that the adenoviruses relied on for replication and then inserting those genes into special cells designed to grow the viruses in the lab.

The gutted adenoviruses, called adenoviral vectors, are versatile tools into which scientists can slip new genes at will. To make COVID-19 vaccines, researchers insert DNA encoding the SARS-CoV-2 spike protein into the vectors. The modified viruses are then grown, isolated, and packaged into vaccines, which contain about 50 billion vectors per shot. Once injected, the vectors slip into our cells and trick our bodies into producing coronavirus spike proteins, which trigger the immune system to develop antibodies and T cells that target the coronavirus.

Before last year, several groups had tried, and failed, to use adenoviral vectors to make vaccines for some of medicine’s toughest problems: HIV infection, malaria, and cancer. In July 2020, success for the technology finally arrived when Johnson & Johnson’s adenoviral vector vaccine for Ebola virus disease was approved in Europe. This year could bring several authorizations for COVID-19 vaccines based on the technology. Even if these vaccines turn out to be less effective than the messenger RNA vaccines, they might make a bigger global impact because they are cheaper to produce and easier to distribute.

CRISPR

Better known for its use as a gene-editing tool, CRISPR got attention amid the pandemic as a new option for diagnosing infectious diseases. Two start-ups, Sherlock Biosciences and Mammoth Biosciences, led the way in developing these tests.

CRISPR’s use as a gene-editing tool is based on its precision; it allows researchers to program in an exact genetic sequence, and some members of the Cas family of enzymes used in gene editing cut in that spot alone. But that cutting precision is not required for diagnosis. In fact, company officials from both Mammoth and Sherlock have told C&EN that the less-specific cutting ability of some Cas enzymes is what makes CRISPR work as a diagnostic.

The companies use Cas coupled to small bits of genetic code called guide RNA, which, like a pig rooting out truffles, digs through a person’s swab sample to find the genetic signature of SARS-CoV-2. If the particular Cas enzymes these companies use find any SARS-CoV-2 gene, the enzyme goes into overdrive and starts cutting any nucleic acid nearby, including bits of synthetic DNA added to the reaction mix. Those bits of DNA are tied to fluorescent reporters that, when cut by Cas, are free to shine.

A special machine runs this reaction in about an hour, giving a positive result for COVID-19 when it detects a certain threshold of the fluorescent molecule.

Sherlock Biosciences, cofounded by the Massachusetts Institute of Technology’s CRISPR guru Feng Zhang, received an emergency use authorization for its test from the US Food and Drug Administration in May, making it the first CRISPR-based product of any kind to get a nod from the government agency. The test is available in large diagnostic labs. Mammoth Biosciences has partnered with GlaxoSmithKline to produce a handheld test and with MilliporeSigma and Hamilton to produce a high-throughput test, which is expected to launch in early 2021.

Meanwhile, both Sherlock and Mammoth are working on CRISPR-based lateral-flow assays, which could potentially be taken at home. And a new test developed by CRISPR scientist Jennifer Doudna and her colleagues can spot the coronavirus in a mere 5 min.

Cryo-EM

Studying an emerging virus and designing vaccines and treatments are easier when you can visualize the virus’s intricate structural details. So when the first cases of COVID-19 surfaced, structural biologists got to work. By February, researchers had deposited multiple SARS-CoV-2-related protein structures in the open Protein Data Bank (PDB) repository. In the past year, protein structures became a vital part of the scientific study of SARS-CoV-2. These structures were made possible largely by an essential technique called cryogenic electron microscopy, or cryo-EM.

To date, researchers have deposited into the PDB over 150 SARS-CoV-2-related structures captured by the technique. Those structures include the first image of the virus’s spike protein, which became the centerpiece of COVID-19 vaccines; viral enzymes that could be targeted by small-molecule drugs; and neutralizing antibodies—made by human immune cells—that were developed as treatments for COVID-19.

Cryo-EM bounces electrons off individual proteins that are frozen—hence the “cryo”—onto a metal or carbon grid. A detector then captures the projected electrons, and heavy-duty computing power processes thousands of individual protein images, reconstructing the protein’s 3-D shape.

Over the past 10 years, improvements in detector technology, computing power, and access to cryo-EM machines mean that the technology was primed and ready for its star turn during the pandemic. Atomic-level imaging by cryo-EM is now possible, and researchers say that if you have access to the equipment and can make copies of your protein, you can move quickly to get useful structural information.

The high cost of purchasing and running cryo-EM equipment is still a barrier for some. Instrument maker Thermo Fisher Scientific recently announced it will soon offer a smaller, cheaper cryo-electron microscope, the Tundra, that uses a less powerful electron beam than the high-end machines that cost millions of dollars. And researchers at the Medical Research Council Laboratory of Molecular Biology in the UK are also developing a lower-cost alternative.

LAMP

Some COVID-19 diagnostic tests take several hours to perform. Others take a few minutes. Why the difference? The secret to many of the rapid molecular diagnostics for COVID-19 is LAMP—loop-mediated isothermal amplification­—a method of copying genetic material. The diagnostic technique is at the center of many molecular tests for COVID-19, including the first at-home product to receive an emergency use authorization from the US Food and Drug Administration.

In the haystack of human genetic material—any nose or throat sample from someone with COVID-19—is a tiny needle of viral genetic material.

For a diagnostic to accurately detect that tiny viral needle, the genetic material has to be amplified—copied several times—to reach the sensitivity of the machine doing the test. Traditionally, this copying is done through polymerase chain reaction (PCR), a technique that relies on several molecular reactions that have to occur at different temperatures. It’s a precise method, but the heating and cooling of the reaction can take a long time.

LAMP uses similar steps for its copying but can conduct them at the same temperature thanks to a special enzyme, often a fragment of a DNA polymerase isolated from a bacterium that thrives in extreme heat. If a sample contains any viral genetic material, it’s converted to a form that the LAMP polymerase amplifies. As one part of the enzyme is making a new copy of the genetic material, another part is ripping that copy open to be copied again. Overall, LAMP is faster and cheaper than PCR, making it a potentially useful tool for regular testing and in low-income countries. But critics have pointed to accuracy limitations that could lead to false negatives and positives.

mRNA

In nature, messenger RNA (mRNA) is the universal language. All living organisms use mRNA as an intermediary between the DNA code of their genomes and the amino acid sequences that compose proteins. For decades, scientists have dreamed of quickly and easily drafting blueprints of artificial mRNA to coax cells into producing therapeutic proteins to treat disease or making viral proteins for vaccination. And in December 2020, that dream became reality when the US Food and Drug Administration authorized the first two mRNA vaccines to prevent COVID-19.

The idea, so simple in theory, was hard to put to practice. Naked mRNA stimulates unwanted immune reactions when injected in the body. Researchers had to package mRNA in lipid nanoparticles to protect it from degradation and to shuttle it into cells, but early formulations of those nanoparticles were also highly toxic.

Companies like BioNTech, CureVac, and Moderna spent over a decade ironing out these problems and others that arose along the way. Doing so required raising billions of dollars for research, which drew ire or ardor from biotech insiders who debated whether the investments were wasteful or revolutionary.

Until last year, no one had ever tested an mRNA vaccine in a large clinical study. In March, the first volunteers were injected with Moderna’s vaccine. Pfizer and BioNTech started trials of their jointly developed vaccine in April. Both groups began large Phase 3 studies in late July; less than 4 months later, they reported that the vaccines are about 95% effective at preventing COVID-19. By December, the first vials were rolled out in Canada, the UK, the US, and other countries. This year, hundreds of millions could get the vaccines.

mRNA’s big breakthrough in COVID-19 is no guarantee that future mRNA vaccines and therapies will be successful. But firms are ready to show what mRNA can do for other infectious diseases, rare genetic disorders, and even cancer. mRNA vaccines are poised to disrupt the pandemic. Now drug and vaccine developers will be watching to see if mRNA can disrupt the biotech industry at large.

Rapid monoclonal antibody development

Drugs normally take years to design and even longer to test in clinical trials. But in November, two antibody drugs—both of which work by binding to the virus before it can infect cells—were granted emergency use authorizations by the US Food and Drug Administration to treat mild cases of COVID-19. It was a mere 9 months after the drugs’ discovery.

The race began in February 2020, when Regeneron Pharmaceuticals used its VelociMouse technology—a strain of mice genetically engineered to have human immune systems—to create human antibodies that recognize the spike protein, which SARS-CoV-2 uses to infect our cells. That same month, AbCellera Biologics obtained convalescent plasma from a person who had recovered from COVID-19. The Canadian start-up scoured the plasma for antibodies that target the spike protein. In March, Eli Lilly and Company began working with AbCellera. Meanwhile, Regeneron expanded its search and also looked for spike protein antibodies in human plasma.

Both groups relied on sophisticated tools to quickly identify and assess hundreds of potential antibodies that target SARS-CoV-2. Their success hinged on techniques for isolating individual B cells—the immune cells that make antibodies—and on DNA-sequencing and synthesis technologies to quickly isolate and study genes that encode the best antibodies against the virus. The companies then copy and paste those genes into cells, which are grown in large vats to manufacture the antibody therapies.

Lilly and Regeneron both started testing their antibodies in humans by early June. Lilly settled on a single antibody, while Regeneron opted for a cocktail of two antibodies—one from humans and one from the VelociMouse. By the end of September, both groups reported preliminary data from larger clinical trials, and in early October, US president Donald J. Trump got an infusion of Regeneron’s experimental therapy.

Both firms applied for US emergency use authorizations of their therapies in early October and were granted them in November. It’s pandemic-pace drug development. And it sets a record that’s unlikely to be broken anytime soon.

Single-cell genomics

When a new disease emerges, scientists devote much of their time to understanding the pathogen causing it. But sometimes, disease isn’t caused by only the invader; the immune system’s response can also be a culprit, making the illness worse. This is the case with COVID-19: for some people, the carefully regulated immune response goes off the rails, causing potentially deadly damage to lungs and other organs.

Historically, understanding that immune response—and identifying drug targets and antibodies to try to harness it—can be a tougher scientific challenge than understanding the pathogen itself. Enter single-cell genomics, a means of parsing which genes are active in individual cells. During the pandemic, scientists are using the technology to probe what individual cells and cell types are doing when a foreign invader like SARS-CoV-2 sets up shop.

Those analyses have led to key discoveries, such as which cells SARS-CoV-2 infects, how the immune response varies in people according to the severity of their disease, and the way antibodies produced against SARS-CoV-2 block the virus from entering another cell.

To interrogate a single cell, researchers first isolate it from its brethren in a human sample. In 10x Genomics’ system, that cell sits in a tiny drop of oil containing a bead that is covered with bits of synthetic, bar-coded RNA. There, the cell is split open, and all the RNA inside binds to the synthetic-RNA-coated bead. A reaction in the assay converts the bound RNA to a form that can be amplified, and a gauntlet of bioinformatic algorithms tells the scientists about the cell and its function.

That analysis can identify the kind of cell and, in the case of COVID-19, differentiate between a B cell, which produces antibodies, and a T cell, which recognizes bits of viral protein. The bioinformatic programs also quantify the genes that are active in the cell, giving scientists a better understanding of how that cell is responding to the invader.

Spike protein mutation

The spiky ball—you’ve seen it everywhere. That artistic rendering of the novel coronavirus, often depicted with menacing red spikes, has become the de facto symbol of the COVID-19 pandemic. And as any scientist familiar with the inner workings of the coronavirus can tell you, those spike proteins are also the key to ending the pandemic.

Coronavirus spike proteins are like grappling hooks. They lock on to receptors that jut from the surface of human cells and pull the virus inside, where it begins to multiply. Our immune systems can eventually develop antibodies that target the spikes and physically block infection.

Most COVID-19 vaccines rely on spike proteins—formulated with an adjuvant, encoded in messenger RNA, or encoded in DNA that is packaged in an adenoviral vector—to trigger antibody production. But this approach has a potential problem.

Although spike proteins look static in illustrations, they are wily shape-shifters. Upon binding to human receptors, the stubby spikes spring into elongated spears that forcefully fuse the virus with the cell. Thus, only antibodies that bind to the spike’s stubby prefusion form, not its elongated postfusion form, can block that crucial first contact and prevent infection. Unfortunately, isolated spike proteins used in vaccines are liable to spontaneously spring into the postfusion form.

Remarkably, scientists Jason McLellan, from the University of Texas at Austin, and Barney Graham, from the National Institute of Allergy and Infectious Diseases, devised a solution to this problem before the pandemic even began. While working on a vaccine for a different coronavirus, the one that causes Middle East respiratory syndrome, their labs discovered that the addition of two strategically placed prolines—the most rigid of amino acids—keeps that coronavirus’s spike protein locked in its prefusion conformation.

Last year, they found that the protein-engineering trick, dubbed the 2P mutation, works for the SARS-CoV-2 spike protein as well. Several companies, including Moderna, Pfizer and BioNTech, Johnson & Johnson, and Novavax, have used the 2P mutation in their COVID-19 vaccines.