03-04-2020, 12:49 PM
The findings, published today in Cell, hold promise as a treatment capable of stopping early infection of the novel coronavirus that, as of April 2, has affected more than 981,000 people and claimed the lives of 50,000 people worldwide.
The study provides new insights into key aspects of SARS-CoV-2, the virus that causes COVID-19, and its interactions on a cellular level, as well as how the virus can infect blood vessels and kidneys.
"We are hopeful our results have implications for the development of a novel drug for the treatment of this unprecedented pandemic," says Penninger, professor in UBC's faculty of medicine, director of the Life Sciences Institute and the Canada 150 Research Chair in Functional Genetics at UBC.
"This work stems from an amazing collaboration among academic researchers and companies, including Dr. Ryan Conder's gastrointestinal group at STEMCELL Technologies in Vancouver, Nuria Montserrat in Spain, Drs. Haibo Zhang and Art Slutsky from Toronto and especially Ali Mirazimi's infectious biology team in Sweden, who have been working tirelessly day and night for weeks to better understand the pathology of this disease and to provide breakthrough therapeutic options."
ACE2 -- a protein on the surface of the cell membrane -- is now at centre-stage in this outbreak as the key receptor for the spike glycoprotein of SARS-CoV-2. In earlier work, Penninger and colleagues at the University of Toronto and the Institute of Molecular Biology in Vienna first identified ACE2, and found that in living organisms, ACE2 is the key receptor for SARS, the viral respiratory illness recognized as a global threat in 2003. His laboratory also went on to link the protein to both cardiovascular disease and lung failure.
While the COVID-19 outbreak continues to spread around the globe, the absence of a clinically proven antiviral therapy or a treatment specifically targeting the critical SARS-CoV-2 receptor ACE2 on a molecular level has meant an empty arsenal for health care providers struggling to treat severe cases of COVID-19.
"Our new study provides very much needed direct evidence that a drug -- called APN01 (human recombinant soluble angiotensin-converting enzyme 2 -- hrsACE2) -- soon to be tested in clinical trials by the European biotech company Apeiron Biologics, is useful as an antiviral therapy for COVID-19," says Dr. Art Slutsky, a scientist at the Keenan Research Centre for Biomedical Science of St. Michael's Hospital and professor at the University of Toronto who is a collaborator on the study.
In cell cultures analyzed in the current study, hrsACE2 inhibited the coronavirus load by a factor of 1,000-5,000. In engineered replicas of human blood vessel and kidneys -- organoids grown from human stem cells -- the researchers demonstrated that the virus can directly infect and duplicate itself in these tissues. This provides important information on the development of the disease and the fact that severe cases of COVID-19 present with multi-organ failure and evidence of cardiovascular damage. Clinical grade hrsACE2 also reduced the SARS-CoV-2 infection in these engineered human tissues.
"Using organoids allows us to test in a very agile way treatments that are already being used for other diseases, or that are close to being validated. In these moments in which time is short, human organoids save the time that we would spend to test a new drug in the human setting," says Núria Montserrat, ICREA professor at the Institute for Bioengineering of Catalonia in Spain.
"The virus causing COVID-19 is a close sibling to the first SARS virus," adds Penninger. "Our previous work has helped to rapidly identify ACE2 as the entry gate for SARS-CoV-2, which explains a lot about the disease. Now we know that a soluble form of ACE2 that catches the virus away, could be indeed a very rational therapy that specifically targets the gate the virus must take to infect us. There is hope for this horrible pandemic."
This research was supported in part by the Canadian federal government through emergency funding focused on accelerating the development, testing, and implementation of measures to deal with the COVID-19 outbreak.
Journal Reference:
The paper appeared today in EBioMedicine, which is published by The Lancet, and is the first study to be published after critique from fellow scientists at outside institutions that describes a candidate vaccine for COVID-19. The researchers were able to act quickly because they had already laid the groundwork during earlier coronavirus epidemics.
"We had previous experience on SARS-CoV in 2003 and MERS-CoV in 2014. These two viruses, which are closely related to SARS-CoV-2, teach us that a particular protein, called a spike protein, is important for inducing immunity against the virus. We knew exactly where to fight this new virus," said co-senior author Andrea Gambotto, M.D., associate professor of surgery at the Pitt School of Medicine. "That's why it's important to fund vaccine research. You never know where the next pandemic will come from."
"Our ability to rapidly develop this vaccine was a result of scientists with expertise in diverse areas of research working together with a common goal," said co-senior author Louis Falo, M.D., Ph.D., professor and chair of dermatology at Pitt's School of Medicine and UPMC.
Compared to the experimental mRNA vaccine candidate that just entered clinical trials, the vaccine described in this paper -- which the authors are calling PittCoVacc, short for Pittsburgh Coronavirus Vaccine -- follows a more established approach, using lab-made pieces of viral protein to build immunity. It's the same way the current flu shots work.
The researchers also used a novel approach to deliver the drug, called a microneedle array, to increase potency. This array is a fingertip-sized patch of 400 tiny needles that delivers the spike protein pieces into the skin, where the immune reaction is strongest. The patch goes on like a Band-Aid and then the needles -- which are made entirely of sugar and the protein pieces -- simply dissolve into the skin.
"We developed this to build on the original scratch method used to deliver the smallpox vaccine to the skin, but as a high-tech version that is more efficient and reproducible patient to patient," Falo said. "And it's actually pretty painless -- it feels kind of like Velcro."
The system also is highly scalable. The protein pieces are manufactured by a "cell factory" -- layers upon layers of cultured cells engineered to express the SARS-CoV-2 spike protein -- that can be stacked further to multiply yield. Purifying the protein also can be done at industrial scale. Mass-producing the microneedle array involves spinning down the protein-sugar mixture into a mold using a centrifuge. Once manufactured, the vaccine can sit at room temperature until it's needed, eliminating the need for refrigeration during transport or storage.
"For most vaccines, you don't need to address scalability to begin with," Gambotto said. "But when you try to develop a vaccine quickly against a pandemic that's the first requirement."
When tested in mice, PittCoVacc generated a surge of antibodies against SARS-CoV-2 within two weeks of the microneedle prick.
Those animals haven't been tracked long term yet, but the researchers point out that mice who got their MERS-CoV vaccine produced a sufficient level of antibodies to neutralize the virus for at least a year, and so far the antibody levels of the SARS-CoV-2 vaccinated animals seem to be following the same trend.
Importantly, the SARS-CoV-2 microneedle vaccine maintains its potency even after being thoroughly sterilized with gamma radiation -- a key step toward making a product that's suitable for use in humans.
The authors are now in the process of applying for an investigational new drug approval from the U.S. Food and Drug Administration in anticipation of starting a phase I human clinical trial in the next few months.
"Testing in patients would typically require at least a year and probably longer," Falo said. "This particular situation is different from anything we've ever seen, so we don't know how long the clinical development process will take. Recently announced revisions to the normal processes suggest we may be able to advance this faster."
Additional authors on the study are Eun Kim, Geza Erdos, Ph.D., Shaohua Huang, Thomas Kenniston, Stephen Balmert, Ph.D., Cara Donahue Carey, Michael Epperly, Ph.D., William Klimstra, Ph.D., and Emrullah Korkmaz, Ph.D., all of Pitt; and Bart Haagmans, of Erasmus Medical Center.
Funding for this study was provided by National Institute of Allergy and Infectious Diseases grant R21-AI114264, National Institute of Arthritis and Musculoskeletal and Skin Diseases grants R01-AR074285, R01-AR071277 and R01-AR068249, and National Cancer Institute grant T32-CA175294.
The study provides new insights into key aspects of SARS-CoV-2, the virus that causes COVID-19, and its interactions on a cellular level, as well as how the virus can infect blood vessels and kidneys.
"We are hopeful our results have implications for the development of a novel drug for the treatment of this unprecedented pandemic," says Penninger, professor in UBC's faculty of medicine, director of the Life Sciences Institute and the Canada 150 Research Chair in Functional Genetics at UBC.
"This work stems from an amazing collaboration among academic researchers and companies, including Dr. Ryan Conder's gastrointestinal group at STEMCELL Technologies in Vancouver, Nuria Montserrat in Spain, Drs. Haibo Zhang and Art Slutsky from Toronto and especially Ali Mirazimi's infectious biology team in Sweden, who have been working tirelessly day and night for weeks to better understand the pathology of this disease and to provide breakthrough therapeutic options."
ACE2 -- a protein on the surface of the cell membrane -- is now at centre-stage in this outbreak as the key receptor for the spike glycoprotein of SARS-CoV-2. In earlier work, Penninger and colleagues at the University of Toronto and the Institute of Molecular Biology in Vienna first identified ACE2, and found that in living organisms, ACE2 is the key receptor for SARS, the viral respiratory illness recognized as a global threat in 2003. His laboratory also went on to link the protein to both cardiovascular disease and lung failure.
While the COVID-19 outbreak continues to spread around the globe, the absence of a clinically proven antiviral therapy or a treatment specifically targeting the critical SARS-CoV-2 receptor ACE2 on a molecular level has meant an empty arsenal for health care providers struggling to treat severe cases of COVID-19.
"Our new study provides very much needed direct evidence that a drug -- called APN01 (human recombinant soluble angiotensin-converting enzyme 2 -- hrsACE2) -- soon to be tested in clinical trials by the European biotech company Apeiron Biologics, is useful as an antiviral therapy for COVID-19," says Dr. Art Slutsky, a scientist at the Keenan Research Centre for Biomedical Science of St. Michael's Hospital and professor at the University of Toronto who is a collaborator on the study.
In cell cultures analyzed in the current study, hrsACE2 inhibited the coronavirus load by a factor of 1,000-5,000. In engineered replicas of human blood vessel and kidneys -- organoids grown from human stem cells -- the researchers demonstrated that the virus can directly infect and duplicate itself in these tissues. This provides important information on the development of the disease and the fact that severe cases of COVID-19 present with multi-organ failure and evidence of cardiovascular damage. Clinical grade hrsACE2 also reduced the SARS-CoV-2 infection in these engineered human tissues.
"Using organoids allows us to test in a very agile way treatments that are already being used for other diseases, or that are close to being validated. In these moments in which time is short, human organoids save the time that we would spend to test a new drug in the human setting," says Núria Montserrat, ICREA professor at the Institute for Bioengineering of Catalonia in Spain.
"The virus causing COVID-19 is a close sibling to the first SARS virus," adds Penninger. "Our previous work has helped to rapidly identify ACE2 as the entry gate for SARS-CoV-2, which explains a lot about the disease. Now we know that a soluble form of ACE2 that catches the virus away, could be indeed a very rational therapy that specifically targets the gate the virus must take to infect us. There is hope for this horrible pandemic."
This research was supported in part by the Canadian federal government through emergency funding focused on accelerating the development, testing, and implementation of measures to deal with the COVID-19 outbreak.
Journal Reference:
- Vanessa Monteil, Hyesoo Kwon, Patricia Prado, Astrid Hagelkrüys, Reiner A. Wimmer, Martin Stahl, Alexandra Leopoldi, Elena Garreta, Carmen Hurtado Del Pozo, Felipe Prosper, J.p. Romero, Gerald Wirnsberger, Haibo Zhang, Arthur S. Slutsky, Ryan Conder, Nuria Montserrat, Ali Mirazimi, Josef M. Penninger. Inhibition of SARS-CoV-2 infections in engineered human tissues using clinical-grade soluble human ACE2. Submitted to Cell, 2020 DOI: 10.1016/j.cell.2020.04.004
The paper appeared today in EBioMedicine, which is published by The Lancet, and is the first study to be published after critique from fellow scientists at outside institutions that describes a candidate vaccine for COVID-19. The researchers were able to act quickly because they had already laid the groundwork during earlier coronavirus epidemics.
"We had previous experience on SARS-CoV in 2003 and MERS-CoV in 2014. These two viruses, which are closely related to SARS-CoV-2, teach us that a particular protein, called a spike protein, is important for inducing immunity against the virus. We knew exactly where to fight this new virus," said co-senior author Andrea Gambotto, M.D., associate professor of surgery at the Pitt School of Medicine. "That's why it's important to fund vaccine research. You never know where the next pandemic will come from."
"Our ability to rapidly develop this vaccine was a result of scientists with expertise in diverse areas of research working together with a common goal," said co-senior author Louis Falo, M.D., Ph.D., professor and chair of dermatology at Pitt's School of Medicine and UPMC.
Compared to the experimental mRNA vaccine candidate that just entered clinical trials, the vaccine described in this paper -- which the authors are calling PittCoVacc, short for Pittsburgh Coronavirus Vaccine -- follows a more established approach, using lab-made pieces of viral protein to build immunity. It's the same way the current flu shots work.
The researchers also used a novel approach to deliver the drug, called a microneedle array, to increase potency. This array is a fingertip-sized patch of 400 tiny needles that delivers the spike protein pieces into the skin, where the immune reaction is strongest. The patch goes on like a Band-Aid and then the needles -- which are made entirely of sugar and the protein pieces -- simply dissolve into the skin.
"We developed this to build on the original scratch method used to deliver the smallpox vaccine to the skin, but as a high-tech version that is more efficient and reproducible patient to patient," Falo said. "And it's actually pretty painless -- it feels kind of like Velcro."
The system also is highly scalable. The protein pieces are manufactured by a "cell factory" -- layers upon layers of cultured cells engineered to express the SARS-CoV-2 spike protein -- that can be stacked further to multiply yield. Purifying the protein also can be done at industrial scale. Mass-producing the microneedle array involves spinning down the protein-sugar mixture into a mold using a centrifuge. Once manufactured, the vaccine can sit at room temperature until it's needed, eliminating the need for refrigeration during transport or storage.
"For most vaccines, you don't need to address scalability to begin with," Gambotto said. "But when you try to develop a vaccine quickly against a pandemic that's the first requirement."
When tested in mice, PittCoVacc generated a surge of antibodies against SARS-CoV-2 within two weeks of the microneedle prick.
Those animals haven't been tracked long term yet, but the researchers point out that mice who got their MERS-CoV vaccine produced a sufficient level of antibodies to neutralize the virus for at least a year, and so far the antibody levels of the SARS-CoV-2 vaccinated animals seem to be following the same trend.
Importantly, the SARS-CoV-2 microneedle vaccine maintains its potency even after being thoroughly sterilized with gamma radiation -- a key step toward making a product that's suitable for use in humans.
The authors are now in the process of applying for an investigational new drug approval from the U.S. Food and Drug Administration in anticipation of starting a phase I human clinical trial in the next few months.
"Testing in patients would typically require at least a year and probably longer," Falo said. "This particular situation is different from anything we've ever seen, so we don't know how long the clinical development process will take. Recently announced revisions to the normal processes suggest we may be able to advance this faster."
Additional authors on the study are Eun Kim, Geza Erdos, Ph.D., Shaohua Huang, Thomas Kenniston, Stephen Balmert, Ph.D., Cara Donahue Carey, Michael Epperly, Ph.D., William Klimstra, Ph.D., and Emrullah Korkmaz, Ph.D., all of Pitt; and Bart Haagmans, of Erasmus Medical Center.
Funding for this study was provided by National Institute of Allergy and Infectious Diseases grant R21-AI114264, National Institute of Arthritis and Musculoskeletal and Skin Diseases grants R01-AR074285, R01-AR071277 and R01-AR068249, and National Cancer Institute grant T32-CA175294.
"Let me issue and control a nation's money and I care not who writes the laws. - Mayer Rothschild
"Civil disobedience is not our problem. Our problem is civil obedience! People are obedient in the face of poverty, starvation, stupidity, war, and cruelty. Our problem is that grand thieves are running the country. That's our problem!" - Howard Zinn
"If there is no struggle there is no progress. Power concedes nothing without a demand. It never did and never will" - Frederick Douglass
"Civil disobedience is not our problem. Our problem is civil obedience! People are obedient in the face of poverty, starvation, stupidity, war, and cruelty. Our problem is that grand thieves are running the country. That's our problem!" - Howard Zinn
"If there is no struggle there is no progress. Power concedes nothing without a demand. It never did and never will" - Frederick Douglass