Categories
Neuroscience

Transparent Zebrafish Study Reveals How Sleep Repairs Damaged Neuronal DNA

Using transparent zebrafish, Israeli researchers were able to confirm neuronal DNA repair as a function of sleep, also identifying a protein that triggers both DNA repair and sleep.

Background

The functions of sleep, though widely researched, have largely remained a mystery. It is known that sleep influences cognition, benefitting learning and memory, but proposed physiological functions have had little strong evidence. Proposed functions of sleep include removing toxic byproducts in the brain caused by wakefulness, replenishing energy and supplies for cells, and remediating neural damage and cellular stress.

Prior research showed that wakefulness and neuronal activity causes DNA double-strand breaks. These lesions are accumulated during wakefulness, contributing to homeostatic sleep drive (the pressure to sleep that builds up as time awake increases). Sleep has been demonstrated to decrease this DNA damage.

Researchers from Israel’s Bar-Ilan University and Tel Aviv University used zebrafish to study neuronal DNA repair as a potential novel function of sleep. The zebrafish is a tiny freshwater fish that has been widely used as a model organism due to their 70% genetic homology (similarity) to humans. Among other parallel physiologies, zebrafish exhibit a diurnal sleep cycle with states closely resembling mammalian slow-wave sleep (SWS) and rapid eye movement (REM) sleep. A mutated type of zebrafish is transparent, enabling researchers to observe the previously unobservable.

Confocal microscopy image showing the developing face of a 6 day old zebrafish larva. / Oscar Ruiz and George Eisenhoffer, University of Texas MD Anderson Cancer Center

By inducing neuronal DNA damage in zebrafish, they were able to determine its relation to sleep as well as causal proteins.

Results

First, the researchers confirmed a significant positive correlation between levels of neuronal DNA damage and total sleep time (R = 0.76). During wakefulness, zebrafish larvae were treated with pentylenetetrazol, which stimulated their neuronal activity. Consequently, the larvae had increased neuronal DNA damage and a 5-fold increase in total sleep time.

Because neuronal DNA damage is not only caused by cell activity, the researchers then exposed the larvae to UV radiation, which damaged their DNA without increasing their neuronal activity. Their subsequent increased sleep further confirmed that DNA damage was the cause.

By testing the rates at which genes involved in the DNA damage response (DDR) were expressed during sleep, they found that the RAD52 and Ku80 proteins were responsible for repairing double-strand breaks during sleep.

Further analysis uncovered that the PARP1 protein, a DNA damage detector that organizes the DDR, was immediately recruited and activated upon neuronal DNA damage.

When the researchers provoked greater expression of PARP1, total sleep time and depth increased—demonstrating that the protein is what connects the DNA damage response to homeostatic sleep drive. PARP1 was also shown to promote sleep regardless of damage.

Discussion

The study confirms in multiple experiments that neuronal DNA repair is a function of sleep regulated by the PARP1 enzyme and carried out by the DNA repair proteins RAD52 and Ku80.

Triggering neuronal DNA damage via cellular excitation and UV light both caused the expected result of increased sleep, with PARP1 as the protein responsible for detecting the damage, provoking a repair response, and causing increased sleep drive.

The study authors noted their intrigue that FDA-approved PARP1 inhibitors used as antitumor agents all caused fatigue as the prominent side effect, suggesting that inhibiting PARP1 masks its sleep-promoting signals. The use of PARP1 inhibitors was also found to cause increased DNA damage. These results, separate from their study, align with their findings regarding the functions of PARP1 as observed in zebrafish.

References
  • Frank, M. G. (2006, August 1). The mystery of sleep function: Current perspectives and future directions. De Gruyter. https://doi.org/10.1515/revneuro.2006.17.4.375
  • Yourgenome. (2021, July 21). Why use the zebrafish in research? https://www.yourgenome.org/facts/why-use-the-zebrafish-in-research
  • Zada, D., et al. (2021, December 16). Parp1 promotes sleep, which enhances DNA repair in neurons. Molecular Cell. https://doi.org/10.1016/j.molcel.2021.10.026
Categories
Neuroscience

New Stem Cell Mechanism Behind Down Syndrome Discovered, Paves Way for Treatment

Human chromosomes. / Hesed Padilla-Nash and Thomas Ried, National Cancer Institute, National Institutes of Health

A team of researchers from MIT’s Alana Down Syndrome Center published results in Cell Stem Cell that pointed to a mechanistic link between Down syndrome (DS) and genome-wide transcriptional disruption. The team, led by Hiruy Meharena, revealed that senescence (the state when cells stop dividing) may play an important role in the progression of down syndrome and could lead to novel therapeutic approaches for treating individuals with DS.

Down syndrome, also known as trisomy 21 (T21), is a congenital disorder that is caused by the triplication of  the 21st chromosome. Previous studies have linked the cognitive deficits of the disorder to a lack of dysfunctional neural progenitor cells (NPCs). These are the stem cells that differentiate into glial and neuronal cells which make up the central nervous system. Neurogenesis, the process in which new neurons are formed in the brain, was found to be significantly reduced in the brains of mice with DS because the lack of NPCs ultimately led to a decrease in cortical matter.

Another study linked T21 to transcriptional disruption in human-derived induced pluripotent stem cells (or iPSCs). His research team found that the disruption to those cells was also consistent with other papers that examined related aneuploidies. Nonetheless, while both studies did reveal a connection between T21 and the cells, they did not delve into the factors that caused this relationship.

As a result, Meharena’s study explores a potential mechanism that may explain how this transcriptional disruption may take place. His team examined T21’s effect on DS-related iPSCs and NPCs at the genetic, epigenetic and mRNA levels. Their finding revealed that although the iPSC were not significantly affected, the NPCs that were derived from these iPSCs revealed significant chromosomal introversion and disrupted lamina-associated domains. This made it so that the squished chromosomes would have increased genetic interactions within itself but decreased interactions with the other chromosomes. Additionally, they found decreased NPC’s chromatin accessibility to the A-compartment, which is associated with transcriptional downregulation, and increased accessibility to the B-compartment, which is associated with transcriptional upregulation.

What was notable was that these changes of differentially expressed genes of NPCs shared many qualities with cells that became senescent via oxidative stress conditions. Furthermore, senolytic pharmaceutical treatments of dasatinib and quercetin was found to ease some of the transcriptional disruption and deficiencies that arose from T21 affected NPCs. The use of these drugs not only improved gene accessibility and transcription but also helped with cellular migration and proliferation.

Although this treatment is not practical due to the neutropenic (neutrophil-decreasing) side effects of dasatinib, it stills elucidates a previously unknown avenue for research into potential treatments to be conducted. With Down syndrome being one the most common intellectual disabilities, having a stronger understanding of the mechanisms behind its progression may provide an opportunity to restore or prevent some of the dysfunctions of the disease. Although this research is only a small piece to the larger puzzle that is DS, it still provides a step to better understanding the condition, hopefully one day assisting individuals with the disorder and their families.

References
Categories
Cardiology Immunotherapy

CAR T Cells Produced by mRNA Injection Reduce Cardiac Fibrosis, Restore Function to Heart

Two individual CAR T cells attacking fibroblast activation protein (FAP)-producing cells. / Rurik et al., 2022

Researchers at the University of Pennsylvania’s Perelman School of Medicine have published a method to treat cardiac fibrosis using an mRNA injection that enables an individual’s own CAR T cells to fight the disease.

Background

Cardiac fibrosis is a medical condition caused by many different types of heart disease that can lead to scarring and stiffening in the muscle wall of the heart. Normally, cells in the heart called cardiac fibroblasts help to develop the heart and maintain its homeostasis (that is, it helps the heart stay in a stable condition). However, in a patient with cardiac fibrosis, these cells no longer perform their normal function. Following a cardiac injury, fibrosis can progress from scarring to complete heart failure.

T cells are a type of white blood cell that play a key role in immune response, killing cells that they recognize to be infected with viruses, cancers, or certain other pathogens. Chimeric antigen receptor (CAR) T cells are T cells that have been engineered to recognize specific proteins as harmful. This enables them to target and kill cells that have proteins from diseases that they otherwise would not recognize as harmful.

Innovation

The Penn researchers developed a CAR T-cell therapy that works by engineering T cells to recognize and kill cells that express (create) the fibroblast activation protein (FAP), a protein key to the pathology of cardiac fibrosis. Killing FAP-expressing cells consequently treats cardiac fibrosis.

By encoding a messenger RNA (mRNA) strand that results in the creation of CAR T cells that target FAP, the researchers had the idea to deliver them to a patient’s cells through an injection containing the mRNA within a lipid nanoparticle.

Lipid nanoparticles (LNP) are a relatively new technology discovered in the 1990s. To deliver an mRNA strand into cells to provoke a protein-expressing response, the mRNA is inserted into a sphere made of lipids that is injected into a patient. This then allows cells to uptake the LNP through endocytosis (bringing material into the cell). The mRNA then exits the LNP, causing the cell to read the mRNA instructions to create the desired protein.

09908-feature1-nanoparticle.jpg
Structure of the LNP. / Genevant Sciences via ACS.org

Without the LNP, mRNA would be unable to enter cells. mRNA vaccines for COVID-19 are a prominent use of this technology, as the mRNA that gives cells instructions to create the spike protein is protected and brought into cells by LNP.

Results

In rodents with cardiac fibrosis, the Penn researchers revealed that their mRNA injection successfully resulted in the creation of FAP-targeting CAR T cells. Observing the hearts of rodents before and after treatment showed notable improvements in cardiac function. This means that as the CAR T cells killed cells that expressed FAP, fibrosis was reduced.

Video of rodent echocardiograph recorded two weeks after treatment with CAR T-cell therapy that was given after an injury that caused cardiac fibrosis. / Rurik et al., 2022

In rodents with injuries causing cardiac fibrosis, the CAR T-cell treatment halved the percentage of fibrosis in the ventricles.

Discussion

The implications of this new treatment are of great significance. Reduction of fibrosis and restoration of cardiac function in rodents with cardiac fibrosis reveals a promising new form of treatment for human patients with the potentially fatal disease.

According to the CDC, about 659,000 people in the United States die from heart disease each year, accounting for 1 in every 4 deaths–all costing the country hundreds of billions of dollars each year. Thus, biotechnological innovations in treatment of cardiac disease can have a great impact.

Earlier CAR T-cell therapies have required a patient’s T cells to be extracted from blood, sent to a lab, engineered to find and kill certain targets, then returned intravenously to the patient. This is an extremely time-consuming and cost-prohibitive process, potentially costing patients hundreds of thousands of dollars.

The innovation of using mRNA injections to create CAR T cells within a patient’s own body instead of a lab may greatly reduce the time and financial burdens associated with CAR T-cell therapies. Rather than extracting, modifying, and replacing T cells from each patient, mRNA shots that provoke the creation of CAR T cells can be mass-produced and given to any patient.

The scope of this innovation reaches far beyond cardiac fibrosis, as it can potentially be applied to CAR T-cell therapies for cancer and other diseases.

References
Categories
COVID-19 Immunology

As Antibodies Wane in Quantity and Efficacy, T Cells Remain Effective Against Omicron

Killer T cells surround a cancer cell // NIH Image Gallery

Background

As the Omicron variant of COVID-19 becomes increasingly dominant among skyrocketing cases, including in vaccinated individuals, concerns of the variant’s immune escape abilities have grown.

Vaccines provoke important responses in the immune system to prevent disease, including creation of T cells and antibodies specific to the pathogen they introduce. In the case of mRNA vaccines for COVID-19, genetic code (mRNA) for the spike protein enters our cells, causing them to manufacture spike proteins. Our immune system then recognizes these proteins as foreign to our bodies, promptly destroying them while creating T cells and antibodies that can work against them in the future.

Antibodies and T cells play different roles in the event of an infection. Antibodies work by creating sites that bind to certain parts of a pathogen. As they pertain to mRNA COVID-19 vaccines, the antibodies will bind strongly to the spike protein. Since the spike protein is what enables SARS-CoV-2 to enter cells, binding antibodies to them will prevent infection.

On the other hand, T cells help fight infection by injecting poison into cells that are already infected, killing both the cell and the pathogen. mRNA vaccines help T cells recognize when a cell is infected with SARS-CoV-2. This means that antibodies are more useful for preventing infection via neutralization while T cells are better at stopping an infection that has already infected some cells.

These figures model T cell and antibody responses to viral infection. In an average SARS-CoV-2 infection, T cells have a greater response than antibodies, and this response effectively decreases viral load. In a severe infection, antibodies are far more prominent than T cells, and this response is ineffective at decreasing the viral load. T cells are more effective at managing instead of preventing an infection, so they would be more useful than antibodies in an already severe infection. (Sette, Crotty 2021)

Much of the focus surrounding COVID-19 vaccines and their efficacy has related to antibody quantity and binding affinity to the changed spike proteins of new variants in order to prevent infection instead of the role of T cells in managing infection. Researchers sought to quantify both antibody and T cell counts and efficacy in unvaccinated, twice vaccinated, and three-times vaccinated patients.

Results

A preprint study from Erasmus University Medical Center in the Netherlands detected high antibody levels against the original SARS-CoV-2 spike protein following receipt of the Pfizer or Moderna mRNA vaccines. Lower (but still significant) antibody levels were detected from the Johnson & Johnson viral vector vaccine. However, antibody levels from the mRNA vaccines decreased significantly within 6 months, while those from the viral vector vaccine did not. Though, even if neutralizing antibody levels remained high, researchers from Beijing’s Peking University found that Omicron escapes most SARS-CoV-2 neutralizing antibodies.

Studies from the Icahn School of Medicine at Mount Sinai in New York supported earlier reports that convalescent (unvaccinated, previously infected) and twice-vaccinated individuals had nonexistent protection against symptomatic infection from the Omicron variant. Boosted (three-times vaccinated) individuals had about 75% protection against symptomatic disease from Omicron, though it is unknown how long this protection will last.

Importantly, the Erasmus study found that unlike neutralizing antibodies, SARS-CoV-2-specific T cells were still detected in the blood 6 months after mRNA and viral vector vaccination as well as natural infection.

In contrast to findings that most neutralizing antibodies are largely ineffective against Omicron, data from Pfizer and BioNTech showed that 80% of spike-specific T cells in vaccinated individuals retained function. The Erasmus researchers corroborate this, finding that vaccinated individuals retain T cell immunity to the Omicron variant.

Discussion

Results from multiple studies now support a consensus that naturally infected and twice-vaccinated individuals have nonexistent protection against symptomatic infection due to depleted and ineffective neutralizing antibodies.

However, both populations can reattain significant protection against symptomatic infection by receiving initial vaccinations or a booster–though it is still unknown how long this protection will last.

These data indicate that convalescent individuals greatly benefit from vaccination, an observation that is of significant public health importance.

Carreño et al., 2021

Even though it has become significantly more difficult to prevent symptomatic infection due to the waning quantity and efficacy of neutralizing antibodies in convalescent and vaccinated individuals, T cells have been shown to remain active and are expected to still help prevent severe infection.

This is supported by new data that has shown that SARS-CoV-2-specific T cells remain present in the long-term and are still mostly effective against the Omicron variant in convalescent and vaccinated individuals.

Well-preserved T cell immunity to Omicron is likely to contribute to protection from severe COVID-19, supporting early clinical observations from South Africa.

Keeton et al., 2021
References
  • BioNTech. “Update – Omicron Variant (B.1.1.529).” BioNTech Investors & Media, 8 Dec. 2021, https://investors.biontech.de/static-files/47b4131a-0545-4a0b-a353-49b3a1d01789.
  • Cao, Yunlong, et al. “Omicron Escapes the Majority of Existing SARS-COV-2 Neutralizing Antibodies.” Nature, 23 Dec. 2021, https://doi.org/10.1038/d41586-021-03796-6.
  • Carreño, Juan Manuel, et al. “Activity of Convalescent and Vaccine Serum against SARS-COV-2 Omicron.” Nature, 31 Dec. 2021, https://doi.org/10.1038/d41586-021-03846-z.
  • Geurts van Kessel, Corine H., et al. “Divergent Sars Cov-2 Omicron-Specific T- and B-Cell Responses in COVID-19 Vaccine Recipients.” MedRxiv, 29 Dec. 2021, https://doi.org/10.1101/2021.12.27.21268416.
  • Keeton, Roanne, et al. “SARS-COV-2 Spike T Cell Responses Induced upon Vaccination or Infection Remain Robust against Omicron.” MedRxiv, 28 Dec. 2021, https://doi.org/10.1101/2021.12.26.21268380.
  • Sette, Alessandro, and Shane Crotty. “Adaptive Immunity to SARS-COV-2 and COVID-19.” Cell, 18 Feb. 2021, https://doi.org/10.1016/j.cell.2021.01.007.
  • Wu, Katherine J. “T Cells Might Be Our Bodies’ Best Shot against Omicron.” The Atlantic, 14 Dec. 2021, https://www.theatlantic.com/science/archive/2021/12/t-cells-omicron-vaccine-immunity/620995/.
Categories
COVID-19

Researchers Reveal Portable COVID Testing Method, Gives Results Within One Second

person holding test tubes
Photo by Polina Tankilevitch on Pexels.com

Researchers from the University of Florida, along with collaborators from the National Chiao Tung University, recently created the world’s fastest COVID detection test to date using a new method with antibody-infused test strips and a small circuit board.

Since the beginning of the COVID-19 pandemic, RT-PCR tests, commonly referred to as PCR, have been regarded as the gold standard for COVID-19 testing.

Reverse Transcription Polymerase Chain Reaction (RT-PCR) works by first converting RNA into DNA, followed by copying small segments of this DNA over and over, primarily using temperature to denature and bind DNA, along with “primers” to make new copies. The process takes about two hours and uses expensive machinery. Such amplification of DNA makes it easy for machines to detect the small amounts of viral particles present in infected patient samples, but difficult to apply to large populations during a pandemic.

Denaturation is when double stranded DNA splits in two and occurs at high temperatures. The green and purple segments in the image are “primers” which bind to DNA at lower temperatures and elongate it. / microbenotes.com

One of the defining features of the coronavirus is the spike proteins, which enable the virus to penetrate host cells due to their geometry and location. Rather than having to convert RNA to DNA, copy the DNA, and read a signal as is done with RT-PCR tests, a new study described a system which uses the spike protein-antibody bond and circuitry for detection.

Antibodies are Y-shaped proteins our immune system produces to fight and prevent future infection. They work by creating sites to which infectious particles bind, effectively blocking those particles from infecting cells. These sites can include binding locations for viruses such as SARS-CoV-2, which researchers have found to be quite useful for detection.

COVID-19 surrounded by Y-shaped antibodies / cdc.gov


As our need for fast, cheap, and portable detection grows, researchers have been searching for new methods. The researchers from the University of Florida ingeniously combined knowledge of antibodies and circuitry to detect presence of COVID in one second.

First, they modeled commercially available glucose testing strips commonly used for testing blood sugar levels in diabetic patients. If you were to dissect a glucose test strip, you would find several electrodes, coated and made of different materials.

Most commonly, glucose test strips are coated with an enzyme that reacts with glucose to steal its available electrons. These electrons are then transported to the electrode which can detect and quantify their presence, indicating how much glucose was in the blood sample.

A glucose strip, mainly used for diabetes patients, is dissected to reveal electrode and adhesive layers. / electrochem.org

In the study, researchers worked to transform the electrodes using different biological and chemical materials. One of the electrodes was plated with gold then “biofunctionalized” with coronavirus antibodies.  An electrode in the middle was connected to an electronic component called a metal-oxide-semiconductor field-effect transistor (MOSFET), which is used to control and amplify electrical signals.

When spike proteins from a sample interact with the surface, the antibody-antigen complex will spring up and down, causing an electrical signal to be sent to the gate of the MOSFET. The device’s circuit board can then quickly convert and read the signal. 

The MOSFET is especially important as it can convert electrical activity from the interaction of a very small amount of coronavirus with the antibodies into a very large signal, similar to how RT-PCR tests amplify the small amount of genetic material into a much larger and easier-to-detect sample.

Figure (a) shows a completed rendition of the detection test. The microfluidic channel is where saliva is inserted for COVID-19 detection. Figure (b) reveals the inner electronics, which connect the test strips to a MOSFET. / che.ufl.edu

The accuracy and acute sensitivity of this method are a direct result of combining electrical and biological tools of detection. Not only does this allow for the detection of extremely low quantities of virus particles, but it can be accomplished in merely 1 second. Furthermore, the device is inexpensive and portable, paving the way for fast, economical, and highly sensitive at-home diagnostic kits.

Notably, Minghan Xian, first author of the study, remarked that by simply altering the type of antibody used, this detection kit could be reapplied to a multitude of other infectious diseases. The electronic components can also be reused with new electrodes.

References
Categories
COVID-19 Public Health

Recovered Patients of Severe COVID-19 Infection 233% More Likely To Die Within Year Than Negative Counterparts

a grieving man kneeling on a grave with red flower
Photo by RODNAE Productions on Pexels.com

Research published by University of Florida scientists in Frontiers in Medicine reported that patients (aged 18-65) who recovered from severe COVID-19 infection were 233% more likely to die within 12 months than COVID-19-negative counterparts.

Methodology

The study analyzed 13,638 patients in the University of Florida Health system over a 12-month period, including positive (mild, severe) and negative cases. A severe case was defined as one requiring hospitalization within 30 days of a positive COVID-19 test. The 12-month risk of mortality was adjusted for age, sex, race, and comorbidities–meaning these factors did not affect the data.

Results

Survival curve showing probability of survival over time following mild, severe, and lack of COVID-19 illness. / Mainous 2021

Patients aged 18 to 65 who recovered from an initial episode of severe COVID-19 had a 233% increased incidence of mortality in a 12-month period compared to negative counterparts. Recovered patients aged over 65 also had increased mortality compared to negative counterparts, but to a lesser extent.

The difference in 12-month mortality between COVID-negative and mild COVID patients was not statistically significant.

Only 20% of the deaths in the 12-month period were attributed to cardiovascular or respiratory conditions.

Discussion

These results show that those who recover from severe COVID-19 infections are much more likely to die within 12 months of recovery compared to those with mild or no infection. This reveals that the increased risk of death from COVID-19 is not limited to the initial episode of infection, indicating that the biological and physiological insult from severe infection is significant. This is further demonstrated by the unexpectedly low portion of deaths caused by cardiovascular or respiratory conditions.

Arch G. Mainous III, Ph.D., first author of the study and University of Florida College of Medicine faculty member, said in a statement to the University of Florida Health Newsroom that “patients may feel that if they are hospitalized and recover from COVID-19 then they have beaten COVID-19. Unfortunately, having a substantially increased [risk] of death in the next year after recovery from a severe episode of COVID-19 shows that this is not the case. Preventing severe COVID-19 should be our primary focus.”

The study mentions that nearly all hospitalizations and severe infections are preventable. Pfizer and Moderna’s COVID-19 vaccines prevent severe infection in more than 95% of cases.

Mainous hopes that the data, which he described as devastating, will “make everyone rethink the impact of COVID-19.”

References
Categories
COVID-19 Pharmacology

Combining a Protein Found in Milk with Benadryl Reduces SARS-CoV-2 Replication in Lung Cells by 99%

Benadryl tablets. // Castorly Stock via Pexels

Researchers looking for prevention and treatment strategies for COVID-19 that are not impacted by SARS-CoV-2 mutations published findings in Pathogens that showed that a combination of diphenhydramine (the active ingredient antihistamine in Benadryl) with lactoferrin (an immunologically active protein found in human and cow milk) reduced SARS-CoV-2 replication by 99% in human cells.

Background

The key to the researchers’ findings related to proteins called the sigma receptors. These receptor proteins are located in the endoplasmic reticulum (ER), an organelle responsible for protein folding and transportation. Sigma receptors have multiple functions, including regulation of the ER stress response.

The ER stress response occurs when the ER is overwhelmed with unfolded or misfolded proteins. This triggers the unfolded protein response (UPR), which seeks to return the cell to a normal state by increasing protein folding, autophagy (destruction of damaged proteins), and in the case of prolonged UPR, apoptosis (cell suicide).

ER stress usually occurs when the ER is overwhelmed with unfolded or misfolded proteins. Cells mitigate ER stress by provoking the unfolded protein response (UPR), which includes increased protein folding, autophagy (destruction of damaged proteins) and, in prolonged cases, apoptosis (cell suicide).

When the UPR causes autophagy, it does so by forming sites near the ER called autophagosomes. Coronaviruses (CoV) have been found to bind directly to the sigma-2 receptor to cause ER stress, enabling them to hijack autophagosomes for use as virus replication sites.

Implication

Researchers found that by binding a drug molecule to the sigma-2 receptor, SARS-CoV-2 would no longer be able to bind to it to cause ER stress (and ultimately virus replication). This is made even more effective by also binding to and activating the function of the sigma-1 receptor.

Results

The team identified a ligand called AZ66 as being able to bind to both sigma-1 and sigma-2 receptors. In experiments with human lung cells infected with SARS-CoV-2, AZ66 completely blocked virus production. However, the safety of AZ66 is unknown, as the drug candidate has not been tested in clinical trials.

Molecular docking model of human sigma-2 receptor (orange) bound to AZ66 (yellow).

Searching for common compounds with proven records of safety, the researchers analyzed electronic medical records to identify diphenhydramine (DPH), the active ingredient antihistamine in Benadryl, as being associated with higher survival rates for COVID-19 patients. This is due to DPH having effects on the sigma-1 receptor. DPH was found to reduce replication of SARS-CoV-2 in the infected human lung cells by about 30%.

Lactoferrin is an antimicrobial and immunostimulatory iron-sequestering protein found in human and cow milk that was brought to a researcher’s attention by the Global Virus Network’s COVID-19 task force due to its antiviral effects on SARS-CoV-2. When tested, it was also found to reduce virus replication by about 30%. The milk protein has a proven safety record as a supplement widely used to treat stomach ulcers.

When a diphenhydramine/lactoferrin combination was tested in human and monkey epithelial lung cells, they found that a synergistic effect occurred, reducing virus replication by 99%.

Commentary

The study’s first author, David A. Ostrov, Ph.D. of the University of Florida, hailed diphenhydramine and lactoferrin as “effective, economical,” and unlike AZ66, “[having] a long history of safety.” The combination could be used to prevent infection as well as decrease recovery time from COVID-19.

While the researchers await potential interest from pharmaceutical companies, Ostrov told the University of Florida Health Newsroom that he cautions against self-medicating with diphenhydramine or lactoferrin as a COVID-19 prevention or treatment. He said that any off-label use of medication should follow a consultation with a physician. Further, commercially available lactoferrin used for treatment of stomach ulcers is not exactly the same as the lactoferrin used in the study.

Lactovid™ is a combination of diphenhydramine and lactoferrin

Would you be interested in purchasing Lactovid™ as a non-FDA approved over-the-counter product?(required)

Warning against off-label self-medication

This article does not offer medical advice. University of Florida researcher, David A. Ostrov, Ph.D., said that any off-label use of medication should follow a consultation with a physician. Off-label use is when a medication is used for anything other than its approved purpose.

This article is based on the following sources

– Bennett, D. (2020, December 3). Existing antihistamine drugs show effectiveness against COVID-19 virus in cell testing. University of Florida Health Newsroom. https://ufhealth.org/news/2020/existing-antihistamine-drugs-show-effectiveness-against-covid-19-virus-cell-testing
– Bennett, D. (2021, November 22). Two common compounds show effectiveness against COVID-19 virus in early testing. University of Florida Health Newsroom. https://ufhealth.org/news/2021/two-common-compounds-show-effectiveness-against-covid-19-virus-early-testing
– Ostrov, D. A., Bluhm, A. P., Li, D., Norris, M. H., et al. (2021, November 20). Highly specific sigma receptor ligands exhibit anti-viral properties in SARS-Cov-2 infected cells. Pathogens. https://doi.org/10.3390/pathogens10111514
– Vela, J. M. (2020). Repurposing sigma-1 receptor ligands for COVID-19 therapy? Frontiers in Pharmacology. https://doi.org/10.3389/fphar.2020.582310

Categories
Cardiology COVID-19

American Heart Association Quells Vaccine Myocarditis Fears Amid Growing Public Concern

Model heart. / Ali Hajiluyi on Unsplash

As concerns regarding mRNA vaccine-caused myocarditis skyrocket on social media and news outlets, researchers have published a study in the American Heart Association’s Circulation journal with statistics regarding the potential side effect.

Google Search interest in “COVID vaccine myocarditis” from December 6, 2020 to December 6, 2021. / trends.google.com

Myocarditis is a condition that causes inflammation in the heart, which can weaken its ability to regularly pump blood throughout the body. It can lead to heart failure, abnormal heartbeat, and sudden death. Most cases of myocarditis are caused by viruses, but the rare heart condition has been noticed as a potential side effect of mRNA vaccines for COVID-19.

The study published in Circulation statistically analyzed cases in patients younger than 21 years old who had received an mRNA vaccine within 30 days of showing symptoms of myocarditis. Researchers found that, in general, young people who experience myocarditis as a side effect of the COVID-19 vaccine recover quickly and completely.

The Circulation study found that 90.6% of adolescent and young adult patients who experienced myocarditis after vaccination were male. In most cases, symptoms presented 2 days after vaccination. The most common symptom was chest pain, which presented in 99.3% of the patients. 18.7% of the patients had low left ventricular ejection fraction (LVEF), meaning that not enough blood was pumping out of their hearts. However, all patients with low LVEF who followed up had fully recovered with normalized function.

Another study published in the New England Journal of Medicine reported that only 2.13 in 100,000 people who received the mRNA vaccine experienced myocarditis. This is much lower than the 150 in 100,000 rate of myocarditis in unvaccinated patients infected with COVID-19 as reported by the Morbidity and Mortality Weekly Report. This means that unvaccinated people infected with COVID-19 had about 70 times greater incidence of myocarditis than any person receiving the mRNA vaccine.

The Circulation study’s first author, Dr. Dongngan T. Truong, told the American Heart Association Newsroom that the data showed that “most cases of suspected COVID-19 vaccine-related myocarditis in people younger than 21 are mild and resolve quickly.”

As the data shows that myocarditis as a side effect of COVID-19 vaccination is extremely rare (2.13 in 100,000) and that almost all of those patients recovered quickly and completely, the American Heart Association continues holding its position that COVID-19 vaccines are safe and highly effective (preventing hospitalization and death in 91% of severe infections). Dr. Donald M. Lloyd-Jones, president of the AHA, said that COVID-19 vaccines were “fundamental to saving lives, protecting our families and communities against COVID-19, and ending the pandemic,” then urging parents to vaccinate their children as soon as possible.

References

Note from the Editors

This article does not offer medical advice. It is a review of statements and data offered by the American Heart Association. Consult with a doctor regarding concerns related to health effects from the COVID-19 vaccine.

Categories
COVID-19

Mutation of Nucleocapsid, Not Spike Protein, Responsible for Delta Variant’s Increased Transmissibility

Single SARS-CoV-2 virion with all parts labeled. / Maya Peters Kostman for the Innovative Genomics Institute

Since the emergence of the Delta variant as the world’s predominant variant of SARS-CoV-2, little has been discovered regarding its mechanism for increased transmissibility. Researchers from the Gladstone Institute of Data Science and Biotechnology (San Francisco, CA) and the Innovative Genomics Institute at UC Berkeley (Berkeley, CA) believe they have found a key site of genetic mutations in the nucleocapsid that could be responsible for the Delta variant’s increased transmissibility.

One author of the study, Abdullah Syed, described in a press release that the life cycle of a virus can be divided into three parts:

  • Entry: the virus is enters a cell
  • Replication: the virus hijacks the cell, causing it to create more copies of proteins and genetic material that comprise the virus
  • Assembly: the copied proteins and genetic material are packaged into new virus particles

The nucleocapsid, a multifunctional structural protein, is critical to the efficiency of the assembly stage in the coronavirus. Though it was previously hypothesized that mutations in the spike protein were causing increased efficiency during the entry stage, the researchers found that mutations in the nucleocapsid were the most significant contributor to Delta’s higher infectivity.

Typically, researching the nucleocapsid would require real viruses, because unlike the spike protein, it is located inside of the virus. This has caused nucleocapsid research to be overlooked, as working with real viruses is inherently dangerous due to the possibility of infecting researchers.

Taha Taha of the Gladstone Institute of Virology described the method that enabled the researchers to study virus replication without using live SARS-CoV-2 virions. They used virus-like particles (VLPs), which have the same structure as the virus, but lack any genetic material for replication. This means that researchers do not risk infection, as the VLPs are unable to replicate. VLPs are also easier to mutate than live viruses.

The researchers genetically engineered the VLPs to express luciferase (the enzyme that causes fireflies to glow), as the light it gives off can be used to gauge activity of a protein-expressing gene. By mutating the nucleocapsid with the mutations found in the Delta variant, an increase in luciferase expression (measured by light) would signal that the mutations increase the functionality of the nucleocapsid.

This was confirmed as one single amino acid mutation (the most basic mutation possible) in the nucleocapsid was found to cause a tenfold increase of luciferase expression, meaning that the mutated virus-assembly protein was ten times as active. Syed noted that this matched the tenfold increase in viral load observed in patients infected with the Delta variant.

The researchers further proved that the mutations increased the activity of the nucleocapsid by infecting cells with real SARS-CoV-2 virions in a highly controlled lab setting, finding that the real mutated virus also demonstrated faster reproduction.

This research has the potential to completely redirect the focus of scientists searching for culprit mutations in new variants of SARS-CoV-2, as research has largely focused on the spike protein until now. An improved understanding of the mechanisms of improved infectivity is important to researchers developing new therapies.

The novel virus-like particle method used could also prove to change virus research forever, as Taha states that this faster and safer alternative to using real viruses could also be used to test existing therapeutics (like vaccines) on new variants. Syed mentioned that the VLP method could even be used to find out if certain animal coronaviruses are capable of infecting humans, to develop new methods of therapy for COVID-19, or to do research on newly emerging viruses that are potentially too dangerous to work with.

This article is based on the following sources

– A. M. Syed et al., Science. (2021, November 4). Rapid assessment of SARS-Cov-2 evolved variants using virus-like particles. https://doi.org/10.1126/science.abl6184
– Henderson, H. (2021, November 4). New method sheds light on why some SARS-Cov-2 variants are more infectious. Innovative Genomics Institute (IGI). https://innovativegenomics.org/news/sars-cov-2-variants-infection/
– Peters Kostman, M. (2021, May 17). Free COVID-19 (SARS-Cov-2) illustrations. Innovative Genomics Institute (IGI). https://innovativegenomics.org/free-covid-19-illustrations

Categories
Immunotherapy Oncology

Mass-Producible Specialized T Cells Exhibit High Cancer-Killing Efficacy, Minimized Complications

An engineered HSC-iNKT cell (blue) attacking a human tumor cell. / Yang lab, UCLA

UCLA researchers have shown in preclinical studies that their mass-producible engineered invariant natural killer T (iNKT) cells demonstrate promising antitumor efficacy and low immunogenicity (unwanted immune response) compared to current cell-based immunotherapy for cancer treatment.

Invariant natural killer T (iNKT) cells are specialized T cells that are notable for their speedy response to danger signals and activation of macrophages (white blood cells that destroy cancer cells, microbes, cellular debris, and foreign substances).

Dr. Lili Yang’s UCLA lab generated iNKT cells by engineering hematopoietic stem cells (HSCs, precursors to all types of blood cells). These cells are allogeneic—they are not genetically specific to patients. Normally, in the realm of cell-based immunotherapy, this would be expected to cause an immune response in the form of graft-versus-host disease (GvHD), a condition in which donor stem cells attack the recipient. The study mentioned that such immunogenicity can also decrease efficacy of therapeutic cells. Therefore, allogeneic cells have not been widely used for T-cell-based therapies, with most therapies using autologous (from the patient) cells instead.

Generally, autologous T-cell therapy requires a patient’s T cells to be extracted from blood, sent to a lab, engineered to find and kill cancer cells, then returned intravenously to the patient—all costing hundreds of thousands of dollars.

Unexpectedly, when tested on mice, the Yang Engineering Immunity Lab’s allogeneic HSC-iNKT cells did not cause the negative effects associated with allogeneic cells. The researchers found that while other types of allogeneic T cells killed mice by GvHD after 2 months of cell transfer, the mice that received HSC-iNKT cells sustained long-term survival.

Figures showing (G) experimental design, (H) mouse tumor imaging, (I) quantification of tumor size based on imaging, (J) survival curves of mice over 4 months following tumor challenge.

Following irradiation of mice, those without cell therapy (labeled as vehicle) died of tumors within 45 days. Those treated with allogeneic BCAR-T cells were tumor-free but died of GvHD. Only those treated with allogeneic HSC-iNKT were tumor-free and survived long term.

This important development means that cell-based cancer therapies would no longer have to rely only on autologous cells extracted from each individual patient. Instead, with the advent of the Yang lab’s one-size-fits-all allogeneic solution, therapeutic cells could be mass-produced and given to any patient, significantly bringing down treatment costs.

The reason why allogeneic HSC-iNKT cells do not cause GvHD is currently unknown to researchers.

Graphs showing tumor load of irradiated mice over time. The Yang lab’s HSC-iNKT cells are shown to have decreased tumor load to near-zero levels (p < 0.001), a more significant decrease than was shown by the other tested therapy (PBMC-NK).
Frozen and fresh allogeneic HSC-iNKT cells were shown to kill more live lung cancer cells (H292-FG) than PBMC-NK cell therapy.

The study also showed that both frozen and fresh allogeneic HSC-iNKT cells killed live leukemia, melanoma, lung cancer, prostate cancer, and multiple myeloma cells in vitro. Compared to PBMC-NK cells, the Yang lab’s cells displayed greater tumor-killing efficacy. Importantly, allogeneic HSC-iNKT cells were also found to remain functional following freezing and thawing, which is crucial for their viability as a widespread, mass-produced treatment.

Factors that support allogeneic HSC-iNKT cells’ prospects as a future widespread cancer therapy include remaining functional following freezing and thawing, high tumor-killing efficacy, and mass-producible by virtue of low immunogenicity.

Dr. Yang told the UCLA Newsroom that one peripheral blood donation could yield 300,000 doses. The researchers are now focused on streamlining manufacturing processes, hoping to better enable mass-production, potentially bringing it to clinical and commercial development more quickly. The Newsroom noted that clinical trials have not yet occurred—this therapy has yet to be tested in humans or evaluated by the FDA. The UCLA Technology Development Group has filed a patent application for this method.

This article is based on the following sources

– UCLA scientists make strides toward an ‘off-the-shelf’ immune cell therapy for cancer. (2021, November 16). UCLA Newsroom. https://newsroom.ucla.edu/releases/off-the-shelf-immune-cell-therapy-for-cancer
– Yang, L., et al. (2021, November 16). Development of allogeneic HSC-engineered iNKT cells for off-the-shelf cancer immunotherapy. Cell Reports Medicine. https://doi.org/10.1016/j.xcrm.2021.100449