Novel Technologies Could Make Coronavirus Vaccines More Stable for Worldwide Shipping
Ssendi Bosco has long known to fear the rainy season. As deputy health officer of Mubende District, a region in Central Uganda, she is only too aware of the threat that heavy storms can pose to her area's fragile healthcare facilities.
In early October, persistent rain overwhelmed the power generator that supplies electricity to most of the region, causing a blackout for three weeks. The result was that most of Mubende's vaccine supplies against diseases such as tuberculosis, diphtheria, and polio went to waste. "The vaccines need to be constantly refrigerated, so the generator failing means that most of them are now unusable," she says.
This week, the global fight against the coronavirus pandemic received a major boost when Pfizer and their German partner BioNTech released interim results showing that their vaccine has proved more than 90 percent effective at preventing participants in their clinical trial from getting COVID-19.
But while Pfizer has already signed deals to supply the vaccine to the U.S., U.K., Canada, Japan and the European Union, Mubende's recent plight provides an indication of the challenges that distributors will face when attempting to ship a coronavirus vaccine around the globe, particularly to low-income nations.
Experts have estimated that somewhere between 12 billion and 15 billion doses will be needed to immunize the world's population against COVID-19, a staggering scale, and one that has never been attempted before. "The logistics of distributing COVID-19 vaccines have been described as one of the biggest challenges in the history of mankind," says Göran Conradson, managing director of Swedish vaccine manufacturer Ziccum.
But even these estimates do not take into account the potential for vaccine spoilage. Every year, the World Health Organization estimates that over half of the world's vaccines end up being wasted. This happens because vaccines are fragile products. From the moment they are made, to the moment they are administered, they have to be kept within a tightly controlled temperature range. Throughout the entire supply chain – transportation to an airport, the flight to another country, unloaded, distribution via trucks to healthcare facilities, and storage – they must be refrigerated at all times. This is known as the cold chain, and one tiny slip along the way means the vaccines are ruined.
"It's a chain, and any chain is only as strong as its weakest link," says Asel Sartbaeva, a chemist working on vaccine technologies at the University of Bath in the U.K.
For COVID-19, the challenge is even greater because some of the leading vaccine candidates need to be kept at ultracold temperatures. Pfizer's vaccine, for example, must be kept at -70 degrees Celsius, the kind of freezer capabilities rarely found outsides of specialized laboratories. Transporting such a vaccine across North America and Europe will be difficult enough, but supplying it to some of the world's poorest nations in Asia, Africa and South America -- where only 10 percent of healthcare facilities have reliable electricity -- might appear virtually impossible.
But technology may be able to come to the rescue.
Making Vaccines Less Fragile
Just as the world's pharmaceutical companies have been racing against the clock to develop viable COVID-19 vaccine candidates, scientists around the globe have been hastily developing new technologies to try and make vaccines less fragile. Some approaches involve various chemicals that can be added to the vaccine to make them far more resilient to temperature fluctuations during transit, while others focus on insulated storage units that can maintain the vaccine at a certain temperature even if there is a power outage.
Some of these concepts have already been considered for several years, but before COVID-19 there was less of a commercial incentive to bring them to market. "We never felt that there is a need for an investment in this area," explains Sam Kosari, a pharmacist at the University of Canberra, who researches the vaccine cold chain. "Some technologies were developed then to assist with vaccine transport in Africa during Ebola, but since that outbreak was contained, there hasn't been any serious initiative or reward to develop this technology further."
In her laboratory at the University of Bath, Sartbaeva is using silica - the main constituent of sand – to encase the molecular components within a vaccine. Conventional vaccines typically contain protein targets from the virus, which the immune system learns to recognize. However, when they are exposed to temperature changes, these protein structures degrade, and lose their shape, making the vaccine useless. Sartbaeva compares this to how an egg changes its shape and consistency when it is boiled.
When silica is added to a vaccine, it molds to each protein, forming little protective cages around them, and thus preventing them from being affected by temperature changes. "The whole idea is that if we can create a shell around each protein, we can protect it from physically unravelling which is what causes the deactivation of the vaccine," she says.
Other scientists are exploring similar methods of making vaccines more resilient. Researchers at the Jenner Institute at the University of Oxford recently conducted a clinical trial in which they added carbohydrates to a dengue vaccine, to assess whether it became easier to transport.
Both research groups are now hoping to collaborate with the COVID-19 vaccine candidates being developed by AstraZeneca and Imperial College, assuming they become available in 2021.
"It's good we're all working on this big problem, as different methods could work better for different types of COVID-19 vaccines," says Sartbaeva. "I think it will be needed."
Next-Generation Vaccine Technology
While these different technologies could be utilized to try and protect the first wave of COVID-19 vaccines, efforts are also underway to develop completely new methods of vaccination. Much of this research is still in its earliest stages, but it could yield a second generation of COVID-19 vaccine candidates in 2022 and beyond.
"After the first round of mass vaccination, we could well observe regional outbreaks of the disease appearing from time to time in the coming years," says Kosari. "This is the time where new types of vaccines could be helpful."
One novel method being explored by Ziccum and others is dry powder vaccines. The idea is to spray dry the final vaccine into a powder form, where it is more easily preserved and does not require any special cooling while being transported or stored. People then receive the vaccine by inhaling it, rather than having it injected into their bloodstream.
Conradson explains that the concept of dry powder vaccines works on the same principle as dried food products. Because there is no water involved, the vaccine's components are far less affected by temperature changes. "It is actually the water that leads to the destruction of potency of a vaccine when it gets heated," he says. "We're looking to develop a dry powder vaccine for COVID-19 but this will be a second-generation vaccine. At the moment there are more than 200 first-generation candidates, all of which are using conventional technologies due to the timeframe pressures, which I think was the correct decision."
Dry powder COVID-19 vaccines could also be combined with microneedle patches, to allow people to self-administer the vaccine themselves in their own home. Microneedles are miniature needles – measured in millionths of a meter – which are designed to deliver medicines through the skin with minimal pain. So far, they have been used mainly in cosmetic products, but many scientists are working to use them to deliver drugs or vaccines.
At Georgia Institute of Technology in Atlanta, Mark Prausnitz is leading a couple of projects looking at incorporating COVID-19 vaccines into microneedle patches with the hope of running some early-stage clinical trials over the next couple of years. "The advantage is that they maintain the vaccine in a stable, dry state until it dissolves in the skin," he explains.
Prausnitz and others believe that once the first generation of COVID-19 vaccines become available, biotech and pharmaceutical companies will show more interest in adapting their products so they can be used in a dried form or with a microneedle patch. "There is so much pressure to get the COVID vaccine out that right now, vaccine developers are not interested in incorporating a novel delivery method," he says. "That will have to come later, once the pressure is lessened."
The Struggle of Low-Income Nations
For low-income nations, time will only tell whether technological advancements can enable them to access the first wave of licensed COVID-19 vaccines. But reports already suggest that they are in danger of becoming an afterthought in the race to procure vaccine supplies.
While initiatives such as COVAX are attempting to make sure that vaccine access is equitable, high and middle-income countries have already inked deals to secure 3.8 billion doses, with options for another 5 billion. One particularly sobering study by the Duke Global Health Innovation Center has suggested that such hoarding means many low-income nations may not receive a vaccine until 2024.
For Bosco and the residents of Mubende District in Uganda, all they can do is wait. In the meantime, there is a more pressing problem: fixing their generators. "We hope that we can receive a vaccine," she says. "But the biggest problem will be finding ways to safely store it. Right now we cannot keep any medicines or vaccines in the conditions they need, because we don't have the funds to repair our power generators."
From infections with no symptoms to why men are more likely to be hospitalized in the ICU and die of COVID-19, new research shows that your genes play a significant role
Early in the pandemic, genetic research focused on the virus because it was readily available. Plus, the virus contains only 30,000 bases in a dozen functional genes, so it's relatively easy and affordable to sequence. Additionally, the rapid mutation of the virus and its ability to escape antibody control fueled waves of different variants and provided a reason to follow viral genetics.
In comparison, there are many more genes of the human immune system and cellular functions that affect viral replication, with about 3.2 billion base pairs. Human studies require samples from large numbers of people, the analysis of each sample is vastly more complex, and sophisticated computer analysis often is required to make sense of the raw data. All of this takes time and large amounts of money, but important findings are beginning to emerge.
Asymptomatics
About half the people exposed to SARS-CoV-2, the virus that causes the COVID-19 disease, never develop symptoms of this disease, or their symptoms are so mild they often go unnoticed. One piece of understanding the phenomena came when researchers showed that exposure to OC43, a common coronavirus that results in symptoms of a cold, generates immune system T cells that also help protect against SARS-CoV-2.
Jill Hollenbach, an immunologist at the University of California at San Francisco, sought to identify the gene behind that immune protection. Most COVID-19 genetic studies are done with the most seriously ill patients because they are hospitalized and thus available. “But 99 percent of people who get it will never see the inside of a hospital for COVID-19,” she says. “They are home, they are not interacting with the health care system.”
Early in the pandemic, when most labs were shut down, she tapped into the National Bone Marrow Donor Program database. It contains detailed information on donor human leukocyte antigens (HLAs), key genes in the immune system that must match up between donor and recipient for successful transplants of marrow or organs. Each HLA can contain alleles, slight molecular differences in the DNA of the HLA, which can affect its function. Potential HLA combinations can number in the tens of thousands across the world, says Hollenbach, but each person has a smaller number of those possible variants.
She teamed up with the COVID-19 Citizen Science Study a smartphone-based study to track COVID-19 symptoms and outcomes, to ask persons in the bone marrow donor registry about COVID-19. The study enlisted more than 30,000 volunteers. Those volunteers already had their HLAs annotated by the registry, and 1,428 tested positive for the virus.
Analyzing five key HLAs, she found an allele in the gene HLA-B*15:01 that was significantly overrepresented in people who didn’t have any symptoms. The effect was even stronger if a person had inherited the allele from both parents; these persons were “more than eight times more likely to remain asymptomatic than persons who did not carry the genetic variant,” she says. Altogether this HLA was present in about 10 percent of the general European population but double that percentage in the asymptomatic group. Hollenbach and her colleagues were able confirm this in other different groups of patients.
What made the allele so potent against SARS-CoV-2? Part of the answer came from x-ray crystallography. A key element was the molecular shape of parts of the cold virus OC43 and SARS-CoV-2. They were virtually identical, and the allele could bind very tightly to them, present their molecular antigens to T cells, and generate an extremely potent T cell response to the viruses. And “for whatever reasons that generated a lot of memory T cells that are going to stick around for a long time,” says Hollenbach. “This T cell response is very early in infection and ramps up very quickly, even before the antibody response.”
Understanding the genetics of the immune response to SARS-CoV-2 is important because it provides clues into the conditions of T cells and antigens that support a response without any symptoms, she says. “It gives us an opportunity to think about whether this might be a vaccine design strategy.”
Dead men
A researcher at the Leibniz Institute of Virology in Hamburg Germany, Guelsah Gabriel, was drawn to a question at the other end of the COVID-19 spectrum: why men more likely to be hospitalized and die from the infection. It wasn't that men were any more likely to be exposed to the virus but more likely, how their immune system reacted to it
Several studies had noted that testosterone levels were significantly lower in men hospitalized with COVID-19. And, in general, the lower the testosterone, the worse the prognosis. A year after recovery, about 30 percent of men still had lower than normal levels of testosterone, a condition known as hypogonadism. Most of the men also had elevated levels of estradiol, a female hormone (https://pubmed.ncbi.nlm.nih.gov/34402750/).
Every cell has a sex, expressing receptors for male and female hormones on their surface. Hormones docking with these receptors affect the cells' internal function and the signals they send to other cells. The number and role of these receptors varies from tissue to tissue.
Gabriel began her search by examining whole exome sequences, the protein-coding part of the genome, for key enzymes involved in the metabolism of sex hormones. The research team quickly zeroed in on CYP19A1, an enzyme that converts testosterone to estradiol. The gene that produces this enzyme has a number of different alleles, the molecular variants that affect the enzyme's rate of metabolizing the sex hormones. One genetic variant, CYP19A1 (Thr201Met), is typically found in 6.2 percent of all people, both men and women, but remarkably, they found it in 68.7 percent of men who were hospitalized with COVID-19.
Lung surprise
Lungs are the tissue most affected in COVID-19 disease. Gabriel wondered if the virus might be affecting expression of their target gene in the lung so that it produces more of the enzyme that converts testosterone to estradiol. Studying cells in a petri dish, they saw no change in gene expression when they infected cells of lung tissue with influenza and the original SARS-CoV viruses that caused the SARS outbreak in 2002. But exposure to SARS-CoV-2, the virus responsible for COVID-19, increased gene expression up to 40-fold, Gabriel says.
Did the same thing happen in humans? Autopsy examination of patients in three different cites found that “CYP19A1 was abundantly expressed in the lungs of COVID-19 males but not those who died of other respiratory infections,” says Gabriel. This increased enzyme production led likely to higher levels of estradiol in the lungs of men, which “is highly inflammatory, damages the tissue, and can result in fibrosis or scarring that inhibits lung function and repair long after the virus itself has disappeared.” Somehow the virus had acquired the capacity to upregulate expression of CYP19A1.
Only two COVID-19 positive females showed increased expression of this gene. The menopause status of these women, or whether they were on hormone replacement therapy was not known. That could be important because female hormones have a protective effect for cardiovascular disease, which women often lose after going through menopause, especially if they don’t start hormone replacement therapy. That sex-specific protection might also extend to COVID-19 and merits further study.
The team was able to confirm their findings in golden hamsters, the animal model of choice for studying COVID-19. Testosterone levels in male animals dropped 5-fold three days after infection and began to recover as viral levels declined. CYP19A1 transcription increased up to 15-fold in the lungs of the male but not the females. The study authors wrote, “Virus replication in the male lungs was negatively associated with testosterone levels.”
The medical community studying COVID-19 has slowly come to recognize the importance of adipose tissue, or fat cells. They are known to express abundant levels of CYP19A1 and play a significant role as metabolic tissue in COVID-19. Gabriel adds, “One of the key findings of our study is that upon SARS-CoV-2 infection, the lung suddenly turns into a metabolic organ by highly expressing” CYP19A1.
She also found evidence that SARS-CoV-2 can infect the gonads of hamsters, thereby likely depressing circulating levels of sex hormones. The researchers did not have autopsy samples to confirm this in humans, but others have shown that the virus can replicate in those tissues.
A possible treatment
Back in the lab, substituting low and high doses of testosterone in SARS-COV-2 infected male hamsters had opposite effects depending on testosterone dosage used. Gabriel says that hormone levels can vary so much, depending on health status and age and even may change throughout the day, that “it probably is much better to inhibit the enzyme” produced by CYP19A1 than try to balance the hormones.
Results were better with letrozole, a drug approved to treat hypogonadism in males, which reduces estradiol levels. The drug also showed benefit in male hamsters in terms of less severe disease and faster recovery. She says more details need to be worked out in using letrozole to treat COVID-19, but they are talking with hospitals about clinical trials of the drug.
Gabriel has proposed a four hit explanation of how COVID-19 can be so deadly for men: the metabolic quartet. First is the genetic risk factor of CYP19A1 (Thr201Met), then comes SARS-CoV-2 infection that induces even greater expression of this gene and the deleterious increase of estradiol in the lung. Age-related hypogonadism and the heightened inflammation of obesity, known to affect CYP19A1 activity, are contributing factors in this deadly perfect storm of events.
Studying host genetics, says Gabriel, can reveal new mechanisms that yield promising avenues for further study. It’s also uniting different fields of science into a new, collaborative approach they’re calling “infection endocrinology,” she says.
New device finds breast cancer like earthquake detection
Mammograms are necessary breast cancer checks for women as they reach the recommended screening age between 40 and 50 years. Yet, many find the procedure uncomfortable. “I have large breasts, and to be able to image the full breast, the radiographer had to manipulate my breast within the machine, which took time and was quite uncomfortable,” recalls Angela, who preferred not to disclose her last name.
Breast cancer is the most widespread cancer in the world, affecting 2.3 million women in 2020. Screening exams such as mammograms can help find breast cancer early, leading to timely diagnosis and treatment. If this type of cancer is detected before the disease has spread, the 5-year survival rate is 99 percent. But some women forgo mammograms due to concerns about radiation or painful compression of breasts. Other issues, such as low income and a lack of access to healthcare, can also serve as barriers, especially for underserved populations.
Researchers at the University of Canterbury and startup Tiro Medical in Christchurch, New Zealand are hoping their new device—which doesn’t involve any radiation or compression of the breasts—could increase the accuracy of breast cancer screening, broaden access and encourage more women to get checked. They’re digging into clues from the way buildings move in an earthquake to help detect more cases of this disease.
Earthquake engineering inspires new breast cancer screening tech
What’s underneath a surface affects how it vibrates. Earthquake engineers look at the vibrations of swaying buildings to identify the underlying soil and tissue properties. “As the vibration wave travels, it reflects the stiffness of the material between that wave and the surface,” says Geoff Chase, professor of engineering at the University of Canterbury in Christchurch, New Zealand.
Chase is applying this same concept to breasts. Analyzing the surface motion of the breast as it vibrates could reveal the stiffness of the tissues underneath. Regions of high stiffness could point to cancer, given that cancerous breast tissue can be up to 20 times stiffer than normal tissue. “If in essence every woman’s breast is soft soil, then if you have some granite rocks in there, we’re going to see that on the surface,” explains Chase.
The earthquake-inspired device exceeds the 87 percent sensitivity of a 3D mammogram.
That notion underpins a new breast screening device, the brainchild of Chase. Women lie face down, with their breast being screened inside a circular hole and the nipple resting on a small disc called an actuator. The actuator moves up and down, between one and two millimeters, so there’s a small vibration, “almost like having your phone vibrate on your nipple,” says Jessica Fitzjohn, a postdoctoral fellow at the University of Canterbury who collaborated on the device design with Chase.
Cameras surrounding the device take photos of the breast surface motion as it vibrates. The photos are fed into image processing algorithms that convert them into data points. Then, diagnostic algorithms analyze those data points to find any differences in the breast tissue. “We’re looking for that stiffness contrast which could indicate a tumor,” Fitzjohn says.
A nascent yet promising technology
The device has been tested in a clinical trial of 14 women: one with healthy breasts and 13 with a tumor in one breast. The cohort was small but diverse, varying in age, breast volume and tumor size.
Results from the trial yielded a sensitivity rate, or the likelihood of correctly detecting breast cancer, of 85 percent. Meanwhile, the device’s specificity rate, or the probability of diagnosing healthy breasts, was 77 percent. By combining and optimizing certain diagnostic algorithms, the device reached between 92 and 100 percent sensitivity and between 80 and 86 percent specificity, which is comparable to the latest 3D mammogram technology. Called tomosynthesis, these 3D mammograms take a number of sharper, clearer and more detailed 3D images compared to the single 2D image of a conventional mammogram, and have a specificity score of 92 percent. Although the earthquake-inspired device’s specificity is lower, it exceeds the 87 percent sensitivity of a 3D mammogram.
The team hopes that cameras with better resolution can help improve the numbers. And with a limited amount of data in the first trial, the researchers are looking into funding for another clinical trial to validate their results on a larger cohort size.
Additionally, during the trial, the device correctly identified one woman’s breast as healthy, while her prior mammogram gave a false positive. The device correctly identified it as being healthy tissue. It was also able to capture the tiniest tumor at 7 millimeters—around a third of an inch or half as long as an aspirin tablet.
Diagnostic findings from the device are immediate.
When using the earthquake-inspired device, women lie face down, with their breast being screened inside circular holes.
University of Canterbury.
But more testing is needed to “prove the device’s ability to pick up small breast cancers less than 10 to 15 millimeters in size, as we know that finding cancers when they are small is the best way of improving outcomes,” says Richard Annand, a radiologist at Pacific Radiology in New Zealand. He explains that mammography already detects most precancerous lesions, so if the device will only be able to find large masses or lumps it won’t be particularly useful. While not directly involved in administering the clinical trial for the device, Annand was a director at the time for Canterbury Breastcare, where the trial occurred.
Meanwhile, Monique Gary, a breast surgical oncologist and medical director of the Grand View Health Cancer program in Pennsylvania, U.S., is excited to see new technologies advancing breast cancer screening and early detection. But she notes that the device may be challenging for “patients who are unable to lay prone, such as pregnant women as well as those who are differently abled, and this machine might exclude them.” She adds that it would also be interesting to explore how breast implants would impact the device’s vibrational frequency.
Diagnostic findings from the device are immediate, with the results available “before you put your clothes back on,” Chase says. The absence of any radiation is another benefit, though Annand considers it a minor edge “as we know the radiation dose used in mammography is minimal, and the advantages of having a mammogram far outweigh the potential risk of radiation.”
The researchers also conducted a separate ergonomic trial with 40 women to assess the device’s comfort, safety and ease of use. Angela was part of that trial and described the experience as “easy, quick, painless and required no manual intervention from an operator.” And if a person is uncomfortable being topless or having their breasts touched by someone else, “this type of device would make them more comfortable and less exposed,” she says.
While mammograms remain “the ‘gold standard’ in breast imaging, particularly screening, physicians need an option that can be used in combination with mammography.
Fitzjohn acknowledges that “at the moment, it’s quite a crude prototype—it’s just a block that you lie on.” The team prioritized function over form initially, but they’re now planning a few design improvements, including more cushioning for the breasts and the surface where the women lie on.
While mammograms remains “the ‘gold standard’ in breast imaging, particularly screening, physicians need an option that is good at excluding breast cancer when used in combination with mammography, has good availability, is easy to use and is affordable. There is the possibility that the device could fill this role,” Annand says.
Indeed, the researchers envision their new breast screening device as complementary to mammograms—a prescreening tool that could make breast cancer checks widely available. As the device is portable and doesn’t require specialized knowledge to operate, it can be used in clinics, pop-up screening facilities and rural communities. “If it was easily accessible, particularly as part of a checkup with a [general practitioner] or done in a practice the patient is familiar with, it may encourage more women to access this service,” Angela says. For those who find regular mammograms uncomfortable or can’t afford them, the earthquake-inspired device may be an option—and an even better one.
Broadening access could prompt more women to go for screenings, particularly younger women at higher risk of getting breast cancer because of a family history of the disease or specific gene mutations. “If we can provide an option for them then we can catch those cancers earlier,” Fitzjohn syas. “By taking screening to people, we’re increasing patient-centric care.”
With the team aiming to lower the device’s cost to somewhere between five and eight times less than mammography equipment, it would also be valuable for low-to-middle-income nations that are challenged to afford the infrastructure for mammograms or may not have enough skilled radiologists.
For Fitzjohn, the ultimate goal is to “increase equity in breast screening and catch cancer early so we have better outcomes for women who are diagnosed with breast cancer.”