Reed Berkowitz

Reed is a Health Science student and published virology researcher at the University of Florida. His areas of interest are immunology and general biomedical research. Reed founded OneResearch.org as a free online source to highlight biomedical research and combat medical disinformation.

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

Categories
Public Health

Study Shows Reddit’s Potential as an Early Warning System for New Designer Drugs

In a new study, researchers demonstrated the use of data mining of social networks like Reddit to predict increased use of novel psychoactive substances (NPS).

Reddit is a social network and media aggregator divided into subreddits pertaining to certain topics, like college football, art, movies, and world news. Niche subreddits exist for many topics, including specific drugs.

Users on subreddits like r/ResearchChemicals discuss designer drugs (including analogs) and other newly discovered substances. Analogs are two drug compounds that have similar molecular structures and/or effects. Sometimes, an analog of an illegal drug has similar effects and properties to its illegal counterpart but has yet to be made illegal.

Under the Federal Analogue Act, compounds “substantially similar” to controlled substances are meant to be treated as if they were the controlled substances themselves. However, designer drugs are often overlooked or may take years for federal authorities to address.

Reddit post on “research chemicals” subreddit discussing novel psychoactive substances.

Researchers from Florida Atlantic University, the University of Florida, and New York University gathered and analyzed data on mentions of newly discovered NPS on Reddit. They found that users mentioned certain NPS months to years before the substances became prevalent in toxicology reports. Such reports are conducted to identify a culprit substance when a patient has overdosed or is subjected to a toxic exposure.

The study mentioned that NPS mentioned on Reddit before any exposures were recorded include:

  • Carfentanil
  • U-47700
  • Eutylone
  • Flualprazolam
  • N-ethylpentylone
  • Isotonitazene
  • Brorphine

Carfentanil, for example, was first mentioned on Reddit in February 2013–almost 4 years before its first reported exposure in October 2016.

Reddit mentions of N-ethylpentylone peaked 5 months before the number of reported cases. (Barenholtz et al, 2021)

The study found that seven of the eight analyzed NPS were mentioned on Reddit before there were any reported cases of the substances in toxicology reports.

Five of the eight analyzed NPS peaked in Reddit mentions months before the substances peaked in reported exposures.

The researchers posit that these data mining methods could prove to be a useful tool for early detection of NPS trends by public health authorities and legislatures.

This article is based on the following works:

– Barenholtz et al. (2021, August 5). Online surveillance of novel psychoactive substances (NPS): Monitoring Reddit discussions as a predictor of increased NPS-related exposures. https://doi.org/10.1016/j.drugpo.2021.103393

Categories
Pharmacology

How “Docking Software” Helped Discover Compounds That Could Boost Stroke, TBI Recovery

Example of molecular docking of a ligand in the main protease of SARS-CoV-2. Yellow: ligand position determined by computer simulation. Black: ligand position determined by experiment. Source: Joshua Vermaas/ORNL, US Department of Energy.

Using the University of Florida’s HiPerGator supercomputer with the University of California San Francisco’s DOCK software, researchers have identified two dipeptides as small molecule activators of the peptidase neurolysin enzyme.

Render of peptidase neurolysin enzyme.

Peptidase neurolysin is an enzyme that has been found to be “one of the brain’s potent, self-protective mechanisms promoting preservation and recovery of the brain after acute injury” (Karamyan). Karamyan’s research posited that in the case of these injuries, neurolysin is responsible for:

  • reducing fluid buildup around the brain (cerebral edema)
  • reducing neural cell death caused by excess neurotransmitters (excitotoxicity)
  • reducing inflammatory response in the brain (neuroinflammation)

Searching for ways to make neurolysin more effective, lead researcher Vardan Karamyan, PhD (Texas Tech University Health Sciences Center Department of Pharmaceutical Sciences) enlisted the help of David Ostrov, PhD (University of Florida College of Medicine Department of Pathology, Immunology and Laboratory Medicine).

Neurolysin is similar in structure to a molecule called ACE2 (angiotensin converting enzyme-2), the main receptor for the virus that causes COVID. Dr. Ostrov previously identified drugs that bind ACE2, and he used the same strategy to identify drug candidates that bind neurolysin.

Superposition of neurolysin (copper) on ACE2 (teal).

Ostrov used UF’s HiPerGator supercomputer to screen 139,725 compounds from a repository at the National Cancer Institute Developmental Therapeutics Program to identify candidates that could fit a site on neurolysin that would facilitate an increase in the enzyme’s function. This was achieved using the DOCK program package, developed by UCSF.

DOCK’s molecular docking simulations were used to identify if and how a ligand (drug-like small molecule) can bind to the active site of a macromolecule (in this case, neurolysin). This is usually done to find ways to promote or inhibit a macromolecule’s function, making molecular docking software a key component in the ever-expanding world of computational drug discovery.

According to the UFHealth Newsroom, UF’s HiPerGator supercomputer completed the query in 15 hours–only 1.4% of the time it would take a desktop computer to complete the same task.

In the lab, the researchers tested the function of DOCK’s best matches on rat neurolysin. They found that the dipeptides L-histidlyl-L-tyrosine and L-histidlyl-L-histidine enhanced the activity of neurolysin.

According to Ostrov, human clinical trials with the mentioned drug candidates could begin within 2 years if FDA approved drugs are found to enhance neurolysin activity. An approved drug could assist or facilitate recovery in stroke and traumatic brain injury (TBI) patients.

This article is based on the following sources

– Bennett, D. (2021, September 7). UF health researcher, collaborators discover compounds that might boost stroke recovery. University of Florida Health. https://ufhealth.org/news/2021/uf-health-researcher-collaborators-discover-compounds-might-boost-stroke-recovery
– Karamyan, V. T. (2019, October). Peptidase neurolysin is an endogenous cerebroprotective mechanism in acute neurodegenerative disorders. https://doi.org/10.1016/j.mehy.2019.109309
– Statements from David A. Ostrov, PhD (University of Florida College of Medicine, Department of Pathology, Immunology and Laboratory Medicine)