Are adenoviral vectors safe? | 117 Three People

Whether or not to use adenovirus vector vaccines ultimately comes down to a risk/benefit ratio issue that often needs to be considered during drug development.

Written by | Shi Jun

In recent weeks, there has been increasing news about the side effects of adenovirus-vectored COVID-19 vaccines.

On April 7, the European Medicines Regulatory Agency found a possible link between the chimpanzee adenovirus vector-based COVID-19 vaccine (ChAdOx1 nCoV-19, also called AZD1222) developed by AstraZeneca and Oxford University and rare blood clot problems. Abnormal blood clots and low platelets will be added to the vaccine's product information as "very rare" side effects. The British medicines regulator also said there may be a link between AstraZeneca's COVID-19 vaccine and rare blood clots.

On Tuesday morning, April 13, the U.S. federal health regulator recommended suspending the use of Johnson & Johnson's adenovirus-based COVID-19 vaccine (Ad26.COV2.S) because six women aged 18 to 48 had blood clots after receiving the vaccine. One of them died and another was hospitalized with severe illness.

On Wednesday (April 14), two more cases were confirmed: a seventh woman, and a man in a clinical trial.

In seven of the eight cases, the symptom was cerebral venous sinus blood clots, all of which occurred 6-14 days after vaccination.

After discussion, the Vaccine Advisory Committee of the Centers for Disease Control and Prevention (CDC) decided to suspend the use of Johnson & Johnson vaccines in the United States for a few weeks. During this period, data on the possible side effects of the Johnson & Johnson vaccine causing blood clots will continue to be collected before further decisions are made. This is because three million doses of the Johnson & Johnson vaccine were administered in the past two weeks, and side effects may take several weeks to appear.

Adenovirus is a weakened cold virus that was first discovered in human adenoids in 1953. In humans, adenovirus usually causes mild respiratory and gastrointestinal infections. However, adenovirus infection can be fatal in people with weakened immune systems or pre-existing respiratory or heart diseases. Adenoviruses are found in a variety of mammals, including monkeys, chimpanzees and humans. More than 50 adenoviruses have been isolated from humans, of which Ad5 is the most commonly used in the biomedical field.

Adenovirus vector vaccines are vaccines that insert a viral gene into an adenovirus gene. This modified virus is usually called a vector by scientists. The only purpose of a viral vector is to infect cells and express the useful genes it carries. When the adenovirus vaccine is injected into the body and infects cells, the viral gene will be expressed, and proteins will be produced as antigens, triggering an immune response. A popular metaphor is to imagine the adenovirus vector as a taxi, which carries important supplies to a designated location and then unloads the supplies.

Before the COVID-19 vaccine, the only approved adenovirus vaccine was the first dose of the Ebola vaccine developed by Johnson & Johnson (a second dose is required later), which uses Ad26 as a vector. The vaccine was approved only in Europe last July. There are also some adenovirus vector vaccines under development: adenovirus vector vaccines for HIV, Ebola virus, influenza virus, etc. are in human clinical trials; adenovirus vector vaccines for rabies virus, dengue virus and Middle East respiratory syndrome coronavirus are still in preclinical development.

The most widely studied HIV vaccine in the world uses the Ad5 vector to express three HIV antigens. However, this vaccine has failed to reduce viral load in clinical trials and has increased the risk of HIV infection in some men.

Adenovirus vectors have also been used in gene therapy, but they almost paralyzed the entire gene therapy field.

In the 1990s, there lived a family of six in Tucson, Arizona, USA. They were a father, a mother and four children, two boys and two girls. One of the boys was in high school, named Jesse Gelsinger. He was a kind and smart boy who loved riding motorcycles and professional wrestling, and worked part-time in a supermarket. Gelsinger was not a very ambitious child. He only had $10.10 in his bank account because he needed $10 to keep the account open. As he grew older, he became more and more independent and even started to rebel a little. Unfortunately, rebellion was not just a stage of growth for him, but also a threat to his health and life.

Figure 1: Jesse Gelsinger. (Source: Wikipedia)

Kissinger suffers from a rare metabolic disease called ornithine transcarbamylase deficiency syndrome (OTCD). OTCD patients completely or partially lack ornithine transcarbamylase (OTC). OTC is one of the five key enzymes in the human body's urea cycle and plays an important role in breaking down and removing nitrogen from the body. The lack of OTC enzyme causes excess nitrogen in the body to accumulate in the blood in the form of ammonia. Ammonia is a neurotoxin, and excess ammonia enters the central nervous system through the blood, causing vomiting, anorexia, drowsiness and coma. In some infants with severe OTCD, ammonia accumulates in the blood to lethal levels. These infants fall into a coma and suffer brain damage within 72 hours of birth. Half of the infants die within a month, and the other half of the survivors die before the age of 5.

Kissinger was diagnosed with OTCD at age two, but it was not a typical case: his mutation seemed to have arisen spontaneously in the womb, and it was what scientists call a "mosaic mutation"—not all cells had the mutation, but only a small number of cells were missing the OTC enzyme. As a result, his symptoms were not that severe. Although he still had occasional attacks, most of them could be controlled with a low-protein diet and 32 pills a day.

However, at the age of 17 (1998), he began to have rebellious thoughts, often disobeying doctor's orders and secretly stopping medication. On December 22, 1998, his father came home and found Kissinger curled up on the sofa, vomiting continuously. He was taken to the hospital, intubated and kept in a coma until the ammonia level in his body was under control. After being discharged from the hospital, he never dared to miss a pill again.

Kissinger's pediatric genetics doctor told him that researchers at the University of Pennsylvania in Philadelphia were developing a gene therapy to repair the missing OTC gene. This gene therapy uses an adenovirus vector Ad5 to express the OTC enzyme. This modified, theoretically non-pathogenic adenovirus is infused into the patient's body through the right hepatic artery, infecting the patient's liver cells and expressing the carried OTC enzyme.

Kissinger and his father were very interested in this. If the treatment was successful, Kissinger might be able to live like a normal person, without having to take so many pills or strictly control his diet - for him, even eating half a hot dog was a luxury. Kissinger wanted to sign up immediately, but he had to wait until he was 18 years old to be eligible to participate in clinical trials.

At the time, gene therapy had only been tried in a handful of patients with genetic diseases. The researchers' experimental treatment extended the lifespan of mice lacking the OTC enzyme in the lab, and scientists hope the gene repair method could eventually be used to treat liver disease in people.

When Kissinger signed up for the trial, he knew he wouldn’t benefit: The trial was a phase I study designed to investigate the safety, not the effectiveness, of the gene therapy in infants with the most severe symptoms—the goal was to find a “maximum tolerated dose”: a dose high enough for the gene to work but low enough that patients wouldn’t suffer severe side effects. Yet Kissinger was eager to contribute to the research of a new treatment. “What’s the worst thing that could happen to me? I die,” he told a friend. “But this is for babies with terminal illnesses.”

June 18, 1999 was Kissinger's 18th birthday. He and his family flew to Philadelphia to visit relatives on his father's side. They visited the Liberty Bell and the Rocky Statue, where Jesse raised his fist and took a photo with the statue (Figure 2). On the 22nd, they went to the University of Pennsylvania, where doctors explained the clinical trial to them and performed blood and liver function tests on Kissinger to see if he met the conditions for participating in the trial. He met the conditions! The trial was scheduled to start in the fall, and he would be the youngest volunteer.

Figure 2: 18-year-old Jesse Gelsinger (Jesse Gelsinger) next to a statue at the University of Pennsylvania. (Source: The Arizona Daily Star/AP Photo)

The clinical trial was planned to include 18 adults, and ultimately 19 volunteers signed up, with Kissinger being the 18th volunteer to be infused with the modified adenovirus.

Kissinger's treatment begins on Monday, September 13. He will receive the highest dose of the viral vector. One of the 17 volunteers who has already been treated is a woman who received the same dose of the virus as Jesse (although from a different production batch) and responded well.

Kissinger began receiving the viral infusion at 10:30 a.m. and finished at 12:30 p.m. In the evening, Kissinger felt sick and had a fever of 40.3 degrees (104.5 degrees Fahrenheit). The doctors were not particularly surprised because other patients had experienced the same reaction. His father, Paul Gelsinger, who was still at home in Arizona and was preparing to fly to the hospital in a few days, called him. After talking, they said I love you to each other and said goodbye. Those were the last words they said.

The next morning, Kissinger began to feel confused and showed signs of jaundice. Tests confirmed that Kissinger's bilirubin, a breakdown product of red blood cells, was four times higher than normal. The doctors began to worry, because this was not a good sign. Abnormally high bilirubin usually means one of two things: liver failure or a coagulopathy—red blood cells are breaking down faster than the liver can metabolize them. The scientists had seen the same symptoms when they tested an enhanced version of the adenovirus vector in monkeys. Such symptoms would be life-threatening in anyone, but they are particularly dangerous for people with OTCD because people with OTCD cannot process the nitrogen produced by the protein released when red blood cells break down well. At noon, Kissinger fell into a coma. By 11:30 p.m., the ammonia level in his blood had risen to more than 10 times the normal level. Doctors began dialysis.

Kissinger's father flew from home to Philadelphia overnight and arrived at the hospital on Wednesday morning. In the afternoon, Kissinger seemed to be stable. However, in the evening, his condition began to deteriorate again. He began to have strong inflammatory reactions and blood clots, and then his kidneys, liver and lungs failed one after another.

On the morning of September 17, 1999, five days into his drug treatment, Kissinger was declared brain dead. The team of doctors and nurses caring for him were shocked by his rapid deterioration and death. Due to Kissinger's death, the last volunteer was not treated.

There is a Wright Mountain in Kissinger's hometown of Tucson. The jagged peaks stand tall, overlooking a deep canyon. The desert at the bottom of the canyon is dotted with cacti and extends to lush yellow pine forests. It is said that this is the closest place to heaven in southern Arizona and it is also Kissinger's favorite place. In early November 1999, seven weeks after Kissinger's death, on a sunny Sunday afternoon, about 20 mourners, including Kissinger's father, mother, stepmother, two sisters, a brother, three attending doctors and some friends, carried a medicine bottle containing his ashes, trekked five miles along a steep path to the top of the mountain and scattered his ashes into the canyon.

Figure 3: Mount Wrightson (picture from the Internet).

Kissinger's death was the first case directly related to gene therapy using adenovirus vectors. The cause of death listed on his death certificate was adult respiratory distress syndrome. However, the actual cause of death was much more complicated.

Prior to the clinical trial, the laboratory of Professor James Wilson of the University of Pennsylvania conducted more than 20 efficacy tests in mice and completed 12 safety studies in mice, rhesus monkeys and baboons. Another volunteer was also infused with the viral vector, using the same dose as Kissinger received, and the expected flu-like side effects and some mild liver inflammation were observed, which disappeared on its own.

However, after Kissinger received the vector, a series of unpredictable chain reactions occurred - jaundice, coagulopathy, kidney failure, lung failure and brain death. In other words, multiple organ system failure. The real cause of death is still not completely clear. A popular theory is that the adenovirus triggered an inflammatory storm, but the specific reason is still not clear.

The news that a gene therapy under investigation had killed a generally healthy volunteer shocked the field of gene therapy, and even the entire field of biomedical research. News reports portrayed the researchers as eager and careless, taking shortcuts and ignoring the principle of protecting the volunteers' health first. Kissinger's family filed a lawsuit. The entire gene therapy field began to collapse, with investors reluctant to invest and many startups failing. Professor Wilson found himself at the center of multiple investigations. He was stripped of his title, his gene therapy center was disbanded, and he was banned from conducting any more clinical trials. This situation lasted until 2010.

Figure 4: Professor James Wilson (Source: https://gtp.med.upenn.edu/people/dr-jim-wilson)

Despite this, Wilson reduced the number of staff in his lab and focused on finding safer viral vectors. Ultimately, his career and the field of gene therapy staged a stunning comeback. His lab’s work ultimately led to the discovery of multiple serotypes of adeno-associated virus (AAV), which led to its widespread use in gene therapy.

Adeno-associated viruses are smaller than adenoviruses, do not cause disease on their own, and cannot replicate independently. In 2019, Novartis Pharmaceuticals' approved gene therapy Zolgensma uses an adeno-associated virus AAV9, which can save infants with fatal neurological diseases.

Ten years ago, everyone would take a detour when they saw Wilson. Now, gene therapy has become a new hot spot in the biomedical field. Wilson is the director of the Gene Therapy and Orphan Disease Center at the University of Pennsylvania and the founder of many biotech companies. His team at the University of Pennsylvania has more than 200 people, occupying several floors of multiple buildings, like a small biotechnology company. Several rooms have been converted into small AAV production workshops; there is also a large animal room that houses many animals (mice, monkeys, cats and dogs) to test the effectiveness and safety of AAV gene therapy.

In the biomedical community, some people praise Wilson for promoting the revival of gene therapy, while others believe that it was his mistakes 20 years ago that almost destroyed the entire field. In any case, there is no doubt that the resurgence of gene therapy is closely related to Wilson.

However, for Paul Kissinger, his beloved son Jesse Kissinger will never come back. He believes that if Wilson had strictly followed the clinical trial regulations and conducted an immune response screening test on Jesse Kissinger, Jesse Kissinger would not have met the conditions for participating in the clinical trial and might not have died as a result.

Figure 5: Paul Kissinger at a hearing on gene therapy on February 2, 2000 (Photo from C-SPAN).

As of April 15, 7.4 million people in the United States had been vaccinated with the Johnson & Johnson vaccine, and no serious adverse reactions other than blood clots had been reported.

Experts have not yet determined whether the vaccine is directly related to the occurrence of blood clots. The CDC/FDA has temporarily suspended the use of the Johnson & Johnson vaccine out of an abundance of caution while investigating this rare blood clot.

"Pause of use" is a common practice. During clinical trials and after the vaccine is widely used, experts will track any medical problems experienced by people who have received the vaccine. If certain symptoms appear unusually often, regulators may decide to suspend the trial or stop using the vaccine for further investigation. The discovery of such a rare side effect just shows that the tracking system works after the vaccine is used on a large scale.

What exactly causes this side effect still needs further investigation. The purpose of the investigation is to figure out whether these problems are just coincidental or directly related to the vaccine. If the results show that the vaccine does have risks, perhaps scientists can identify some common inherent characteristics of high-risk groups, so that new guidelines can be written to indicate who should not be vaccinated. The suspension also gives regulators time to advise doctors how to recognize and treat the symptoms of side effects.

According to the CDC, 300,000 to 600,000 people in the United States develop blood clots in their lungs or leg veins or other parts of the body each year. This means that about 1,000 to 2,000 people in the U.S. population will have blood clots every day. Now that millions of people are vaccinated every day, it is likely that some vaccinated people will randomly develop blood clots, which are not directly related to the vaccine.

However, the blood clot symptoms found in the Johnson & Johnson vaccine recipients so far are different and much less common than common blood clot symptoms. In addition to the formation of cerebral venous sinus thrombosis (CVST) in the brain, all of these recipients had very low levels of platelets, which made them prone to abnormal bleeding. This is very similar to the symptoms of some AstraZeneca vaccine recipients.

It is not yet completely confirmed that the adenovirus technology caused the problem. The New England Journal of Medicine (NEJM) published two articles in a row, explaining the possible mechanism of AstraZeneca's new crown vaccine causing blood clots, which seems to be directly related to the viral vector.

Both Johnson & Johnson and AstraZeneca's COVID-19 vaccines use adenovirus vectors, Ad26 and chimpanzee adenovirus vectors (ChAdOx1). Russia's COVID-19 vaccine Sputnik V (Ad26+Ad5) and China's CanSino vaccine (Ad5) also use similar technology.

Because adenoviruses are pathogens that can cause the common cold, many people have been naturally infected with adenoviruses in the past (especially as children) and have developed an immune response to the viral vectors, and already have antibodies against these adenoviruses in their bodies. Pre-existing immunity acquired from these previous infections may reduce the effectiveness of adenovirus vaccines. Similarly, the effectiveness of the second dose of adenovirus vaccine may be greatly reduced compared to the first dose. According to statistics from one paper, the rates of pre-existing immunity to Ad5 in South Africa, Kenya, Uganda, and Thailand were 87.9-89.5%, 90.5%, 86.4%, and 82.2%, respectively, and the rates for Ad26 were 43.1–53.2%, 66.2%, 67.8%, and 54.6% [3]. An earlier study showed that pre-existing immunity to the chimpanzee adenovirus ChAdOx1 was low in adults in the UK and Gambia [4]. However, the rates of pre-existing immunity to a particular adenovirus vary from region to region.

So far, the chance of CVST after receiving the Johnson & Johnson vaccine is one in a million, and the chance of CVST after receiving the AstraZeneca vaccine is five in a million. The chance of blood clots after infection with the coronavirus is 135,000 in a million. According to an analysis of data published on April 14 by researchers at the University of Oxford that has not yet been peer-reviewed, the chance of a patient infected with the coronavirus having symptoms similar to CVST is 39 in a million [5]. In other words, if you do not get vaccinated, the chance of getting CVST if you get infected with the coronavirus is much greater than the chance of getting CVST after getting vaccinated.

Ultimately, this is a risk/benefit ratio that needs to be considered frequently during drug development. The risk/benefit ratio of any drug/therapy is different for each person, and you need to make your own judgment. I hope that experts can quickly find a risk indicator to distinguish the high-risk groups for CVST after vaccination, so that the rest of the people can get the vaccine with confidence.

References

[1] https://www.nytimes.com/1999/11/28/magazine/the-biotech-death-of-jesse-gelsinger.html

[2] https://cen.acs.org/business/The-redemption-of-James-Wilson-gene-therapy-pioneer/97/i36

[3] DH Barouch et al., International seroepidemiology of adenovirus serotypes 5, 26, 35, and 48 in pediatric and adult populations. Vaccine 29, 5203-5209 (2011).

[4] MDJ Dicks et al., A Novel Chimpanzee Adenovirus Vector with Low Human Seroprevalence: Improved Systems for Vector Derivation and Comparative Immunogenicity. PLOS ONE 7, e40385 (2012).

[5] https://osf.io/a9jdq/