No Effectiveness of Hydroxychloroquine in Randomized Control Trial of Patients with COVID-19

It has been a month since Donald Trump tweeted that hydroxychloroquine may be a big game changer for the treatment of COVID-19. Since that time, the FDA granted emergency use authorization for this medication in the case of hospitalized patients who do not have access to a randomized clinical trial. Unfortunately there have not been many robust clinical trials to date and people are still unsure if this medication works.

A preprint was recently posted that described the results of an open-labeled, randomized controlled trial for 150 hospitalized patients. In this post, I’ll take a deeper look at this study.

Study Design

Hospitalized patients in Ruijin Hospital, China were randomized either to standard of care or standard of care with hydroxychloroquine.

Hydroxychloroquine was provided the same day of randomization. 1,200 mg daily were given for three days followed by 800 mg for the remaining 2-3 weeks. Just for reference, this is 2-3 times greater than the dose provided in France and in the United States.

Patient Demographics

These patients were relatively healthy, with an average age in the 40s and most had moderate COVID-19, although the authors did not describe what constitute moderate disease. One potential confounder is that more patients in the hydroxychloroquine arm had preexisting comorbidities, shortness of breath, and higher CRP and IL-6 levels upon initial presentation.

Tang et al. 2020.
Tang et al. 2020
Tang et al. 2020

Adverse Effects

It is no surprise that patients with hydroxychloroquine had more adverse effects. The most dangerous complication is arrhythmia from QT prolongation, but that is not reported here.

Tang et al. 2020

Results

Hydroxychloroquine did not lead to greater viral clearance of SARS-CoV-2 as detected by PCR compared to standard of care. There was no significant improvement in clinical symptoms with this medication as well compared to standard of care treatment. This pretty much kills the idea that hydroxychloroquine is a cure for COVID-19. The authors do report there is some reduction in IL-6 and CRP levels, but at the end of the day, we care about clinical improvement and the results clearly show that that hydroxychloroquine is not some sort of magic cure.

Tang et al. 2020
Tang et al. 2020

Conclusions

This likely is a practice changing trial. Every hospital in the world has been empirically providing hydroxychloroquine for treatment given a few in vitro results and two small randomized trials. However, this medication does come with side effects and there is no clear evidence that hydroxychloroquine significantly improves symptoms or decreases viral load.

I suspect every hospital will start cutting back on their use of hydroxychloroquine.

Compassionate Use of Remdesivir for Patients with Severe COVID-19

The biggest update about investigational therapies to combat the COVID-19 pandemic is this recent NEJM article, which showed that remdesivir is associated with clinical improvement in 36/53 patients who received this medication for compassionate use. In this post, I’ll take a deeper dive into this specific paper and provide my own personal opinions. For more background about remdesivir, please see a previous post. This was not a randomized clinical trial, so we cannot make an efficacy claim meaning that we cannot tell if remdesivir was the specific cause that these patients improved.

Background

Remdesivir is a nucleotide analog that is metabolized to create a dummy adenosine triphosphate group. Viruses normally need nucleotides like adenosine to replicate their viral genome, so adding these dummy group limits viral expansion.

In January to February, there were no widespread clinical trials in the United States for remdesivir so clinicians had to send these request approvals to get this drug. Gilead only approved requests for patients with RT-PCR confirmed COVID, good kidney function, and no failed liver. Once approved, patients received a 10 day course of remdesivir (200 mg IV day 1 and 100 mg daily afterwards).

Patient Demographics

Below are the baseline patient demographics for the patients included in this study. 2/3 were on a ventilator and 1/3 just had noninvasive oxygen support (i.e. nasal cannula or face mask). The big takeaway for me was that the patients’ kidney function was really good (Cr of 1) and liver function was really good (ALT, AST < 40). This means they did not want to take a chance on having really bad adverse effects if this drug is poorly cleared.

Grein et al. NEJM 2020.

Clinical Improvement

The paper then had three figures showing clinical improvement with remdesivir.

In the first figure, they stratified every patient’s baseline condition based on oxygen requirement along columns and described their resulting medical status in rows. In patients with invasive ventilation, 56% improved with 8 patients getting discharged and another 8 now without any oxygen requirement. The results were even better for those with noninvasive ventilation, in which 5/7 patients were discharged.

Grein et al. NEJM 2020.

The next figure took a deeper dive at the oxygen status of every single patient who received therapy. There are several interesting stories from this figure.

There are 3 patients who were on ECMO (extracorporeal membrane oxygen support) requiring an external device to oxygenate their blood, who made near miraculous recoveries.

Of the patients who were intubated, 1/2 recovered, 1/4 remained the same, and 1/4 died. There was greater clinical recovery with remdesivir in patients with less severe oxygen requirements, in which all but two patients recovered.

Grein et al. NEJM 2020

The next figure plotted the course of clinical recovery with remdesivir. The authors claimed that it took a few weeks to lead to full clinical improvement with this medication, patients with noninvasive ventilation did better, and younger patients recovered faster. None of these are big surprises.

Grein et al. NEJM 2020.

Adverse Effects

Finally the authors listed all the adverse effects. Honestly, we cannot tell if these adverse effects are from the medication or from worsening clinical status from the virus. This table was not too helpful from my assessment. We know hepatotoxicity is associated with remdesivir, but we do not have an idea of how much hepatic enzymes increased with this medication. Presenting the raw numbers would have been way more helpful.

Conclusions

From this trial, it seems like remdesivir is associated with clinical improvement severe COVID-19.

However, it is difficult to make any large claims about remdesivir efficacy. We do not know the natural history and clinical improvement for patients with COVID-19. Are patients recovering because they have built enough antibodies (which takes weeks to mount) or are they recovering because of the medication?

We can speculate that remdesivir won’t work well for patients who have large viral load causing multiorgan failure. Perhaps this drug works better when given earlier.

Ventec VOCSN Ventilator

The world is in a global shortage of ventilators. There are currently about 200,000 ventilators in the United States, but 1 million patients may need one of these life-saving technologies during the COVID-19 pandemic.

Every ventilator company is scaling up production to fulfill this dire need. To scale up production, ventilator medical device companies are partnering with large scale manufactures to overcome bottlenecks in supply chain. One such collaboration is between Ventec and General Motors.

General motors just converted their Kokomo facility from automotive to medical manufacturing and trained 1,000 employees to make Ventec’s VOCSN ventilator.

In this post, I’ll describe some of the features of the VOCSN ventilator. The main advantage of this medical device is that it can perform five distinct respiratory functions at the same time – ventilation, oxygenation, cough support, suction, and nebulzation.

A hands on demonstration is available here:

Hands on Demonstration of VOCSN Features

Ventilation

The VOCSN device is at its core a full-functioning ventilator, which has all the standard settings of a ventilator (i.e. mode of ventilation, respiration rate, end expiratory pressure, etc). A couple benefits of this ventilation device are that it is portable with a weight of only 18 pounds) and battery-powered with 9 hours of internal battery power.

Additional Features

The additional features of the VOCSN device are what make it so attractive.

This device has an internal oxygen concentrator that can deliver 6 L/min of oxygen, but can also be connected to an external high or low pressure source. I’m not sure how much we are in critical demand of oxygen tanks, but the VOCSN ventilator wastes less oxygen compared to other ventilators (Pangillnan et al. Respiratory Care 2019).

This device also has cough-support. This features simulates a cough in a patient by rapidly alternating a inspiratory air pressure followed by an expiratory air pressure. Cough reflexes help patients clear mucus and debris in their airways. Intubated patients with COVID-19 have a suppressed cough reflex, so including this feature will help someone breathe better.

The VOCSN device also has in-built suction. When someone has COVID-19 pneumonia, he or she has increased mucus in their airways. In the hospital, intubated patients have suctioning devices to suck up any mucus that may prevent air from going into the lungs. This in-built feature prevents the need for another device and will decrease the amount of viral particles that can become airborne and infect others.

Finally, the VOCSN device has an in-built nebulizer. Patients with COPD or asthma often need inhaled steroids and airway dilation medication to decrease inflammation and prevent their airways from collapsing.

VOCSN for COVID-19

Normal respiratory support for someone suffering from respiratory failure due to COVID-19 includes a ventilator, oxygen tank, cough-assist device, suction, and nebulizer.

These together are 55 pounds, require lots of nursing and physician time, and increase the risk of cross-infection. The biggest benefit of VOCSN is that it is one machine, easy-to-use, and light-weight.

Source: https://www.venteclife.com/assets/pdf/VOCSN_Multi-Function_Ventilator-COVID-19-Coronavirus-Response.pdf

Challenges

The biggest challenge for this device is that it includes so many different parts, as seen below. If any of these parts are missing or delayed, the entire ventilator production supply will also lag. For example, one of the parts in this device was made in a factory in India, but the Indian government quarantined the entire region, thus stalling production. This is where General Motors used its expertise in supply chain manufacturing to overcome this issue.

Source: https://www.venteclife.com/assets/pdf/VOCSN_Multi-Function_Ventilator-COVID-19-Coronavirus-Response.pdf

Conclusion

I am extremely hopeful that the Ventec-General Motors collaboration can ramp up ventilator production. These devices are life-saving and we need them now.

Public Non-Medical Masks Wearing to Quell the Pandemic

The United States has a shortage of personal protective equipment. Every surgical mask and N95 respirator should be prioritized to frontline healthcare professionals. However, should the general public be wearing non-medical masks (i.e. cloth masks, scarves, bandanas, etc)?

This topic is controversial. As of April 2, the CDC in the United States recommended to only wear face masks if sick.

US CDC Guidelines. Source

However, the CDC in China does recommend wearing face masks in public.

China CDC Guidelines. Source

Academic clinicians in the New England Journal of Medicine state that wearing a mask outside a hospital offers little protection (Klompas et al. NEJM 2020).

Klompas et al. NEJM 2020

However, many people on Twitter, including the former FDA chief Scott Gottlieb, suggest the countries that flattened the curve of disease all had public mask wearing policies.

News about #masksforall on Twitter
Source: Twitter

Who is right? In this post, I’ll talk about the evidence for public mask wearing and the evidence for non-medical masks to quell respiratory infections.

Public Masks during 2003 SARS

Our current COVID-19 coronavirus is similar in biology, viral transmission, and danger to the 2003 SARS virus that stemmed from Hong Kong.

Lau et al. showed that during the 2003 SARS outbreak, wearing a mask in public was a protective preventive measure in reducing the risk of infection (Odds ratio of 0.27, p value < 0.001).

Lau et al. Emerging Infectious Disease 2004.

Similarly, Wu et al. also showed that wearing a mask in public, either sometimes (Odds Ratio 0.5, p = 0.02) or always (Odds Ratio 0.3, p < 0.001) was protective in preventing SARS.

Wu et al. Emerging Infectious Disease 2004.

It is pretty clear from both studies that wearing masks in public prevents SARS infection.

Medical vs Non-Medical Masks

We also know from the 2003 SARS virus that the greatest at-risk population were frontline healthcare workers. Every respirator and surgical mask should be prioritized to these individuals. But how effective are non-medical masks for the general population to prevent community transmission?

Dr. Sui Huang answered this question in a beautiful Medium post, stating that we should all wear masks in public even if they are homemade. Briefly, Huang argued that public mask wearing can help flatten the curve by at least 50%, which can help our overburdened healthcare system and save us time to build respirators, diagnostic tests, therapeutic drugs, and protective equipment to send to our hospitals.

Sui Huang. Source

Cloth based masks prevent 67% of particles from entering your mouth from the environment and prevent 10% of your cough from leaking into the environment.

Sui Huang. Source

Historical Evidence for Public Non-Medical Masks

There is also historical evidence that publicly mandated non-medical masks are effective.

For example, during the Spanish flu of 1918, a range of interventions were tried to quell the pandemic including banning of mass gatherings, mandated mask wearing, isolation, and disinfection (Bootsma and Ferguson, PNAS 2007). Cities that implemented these programs earlier and longer had fewer deaths (Strochlic and Champine).

Dr. Capps also showed mandated non-medical masks to be effective to prevent respiratory illness in an Army military camp (Capps, JAMA 1918). After the mask mandate, they reduced infection from scarlet fever and measles by greater than 95%. These results were replicated by Dr. McLester to control a bronchopneumonia outbreak at a hospital in Alabama and Dr. Lichty to control another pneumonia outbreak at Mercy Hospital in Pittsburgh.

Conclusion

In a personal protective equipment shortage, all medical masks (surgical and N95) should be prioritized to healthcare frontline workers. However, there is good evidence to suggest that public non-medical masks are beneficial.

Update

As of April 3, the US CDC now recommends cloth masks for everyone when they go outside. This is a major development and a step in the right direction.

FDA Emergency Use Authorization for Cellex Serologic Antibody Test for COVID-19

In my last post, I discussed the value of serologic antibody tests for COVID-19. Briefly, these tests detect the presence of IgM and IgG antibodies that a person builds to fight an infection. These tests can be used at a global scale to find out how many people have been infected with COVID-19, who potentially can return to work, and perhaps who we can retrieve convalescent plasma to treat active infection.

Today, the FDA provided its first emergency use authorization for serologic antibody testing to Cellex, Inc. Cellex, Inc is a biotechnology company that specializes in point-of-care diagnostics technology. They have previously developed high sensitive and specific immunochromatographic assays for various infectious, tropical, gastrointestinal, and respiratory diseases. For their COVID-19 antibody test, they use a lateral flow immunochromatographic assay to detect the presence of IgM and IgG antibodies.

Lateral Flow Test

Here is a great video demonstration of how a lateral flow immunoassay test works.

A drop of analyte (i.e. blood, plasma, serum) is applied to the sample pad. This drop will flow down the cellulose membrane through capillary mass action. The substances in the analyte then bind to their matching antibodies tagged with a marker that can be visualized or with control line antibodies. Matched antibodies are then captured and displayed at the test line of the immunoassay.

Source: Youtube

Cellex Test for COVID-19 Antibodies

Here is the Cellex brochure for their antibody test. The instructions are simple: add 10 microLiters of sample (blood, plasma, or serum), add two drops of sample diluent, and then wait 15 minutes for the result.

Instructions for Cellex Test

A negative result shows the presence of the C band without any color in the G or M band. There are several possibilities of positive results, shown below. If only G band is present, this suggests a late stage primary, early secondary, or past infection with COVID-19. If only the M band is active, this suggests a fresh primary coronavirus infection. If there are both M and G bands, this means there is a current primary or early secondary infection.

Providers are recommended that a negative test does not rule out COVID-19 infection and should not be used as a sole basis for treatment decisions.

Interpretation of Antibody Results

Efficacy of Test

Cellex, Inc. tested the efficacy of their serological antibody test against 128 samples with SARS-CoV-2 infection confirmed with RT-PCR and 250 negative control samples. The sensitivity was 93.75% and specificity was 96.4%.

Source

There was no cross reactivity with other viruses like HIV, influenza, or Rhinovirus. There was also no false positive findings with elevations in hemoglobin, bilirubin, cholesterol, rheumatoid factor, human IgG/IgM, or common antibiotics.

FDA Guidance

The FDA provided emergency use authorization stating that it is reasonable to believe that the Cellex, Inc antibody test may be effective in diagnosing COVID-19.

FDA EUA Criteria for Issuance

Conclusion

There are a ton of products now in the market that provide serologic antibody tests. However, all those products are required to state that they are not reviewed by the FDA, negative results do not rule out infection, and positive results can be contaminated from a past non COVID-19 related infection.

This makes it difficult for any healthcare provider to determine which test to purchase. This is especially important because Spain had to recall 58,000 ordered tests after learning they had poor accuracy.

However, FDA EUA authorization gives more credence and confidence to purchase Cellex products..

Serologic Antibody Tests for COVID-19

In order to diagnose someone with an active COVID-19 infection, we take a nasal swab and run an RT-PCR test to identify if there is elevated coronavirus RNA in the specimen. But how do we tell if someone is mounting an immune response to a virus? How do we tell if someone contracted the virus, recovered, and now immune? How do we tell if someone has antibodies in plasma that can be donated to someone else for treatment?

A serological antibody test can answer all those questions. There have been many small biotechnology companies developing these serological tests. In this post, I’ll outline a few of the ones I found online.

Background

When someone first contracts an infection from a virus, there is first an asymptomatic phase of about one week before he or she has her first symptom like fever or cough. Generally, this coincides with increased virus replication and increased viral RNA, which can be identified through a polymerase chain reaction test. However, our body then starts to mount an immune response, first with IgM antibodies which last for months and then with IgG antibodies which last for years.

Source

We do not know the exact length of time it takes someone to build these IgM/IgG antibodies. An early report from Guo et al, suggested it takes 5 days for IgM to be detected and 14 days for IgG to be detected after symptom onset. If this current SARS-CoV-2 virus is similar to SARS-CoV1, then we should expect it to take a few weeks after the onset of symptoms to build a robust immune response (Li et al. NEJM, 2003).

Li et al. NEJM 2003

Serological Tests in Literature

There have been a few wonderful research reports detailing the application of serological tests.

For example, Amanat et al. developed an assay that detects antibodies to the spike protein of SARS-CoV2. Reagents, plasmids, and proteins can be requested from their website. More details of their protocols are available here.

Protocol overview of ELISA test for SARS-CoV-2 antibodies
Source

Li et al. also developed a rapid IgM-IgG combined antibody test with a sensitivty of 88% and specificity of 90%.

Li et al. Journal of Medical Virology. 2020

There are now many companies building serological tests for detecting antibodies against COVID-19. Two that I found online are from Pinnacle BioLabs and BD. To be honest, these reports are a little confusing because they note the FDA approved their products, but no serologic tests were granted Emergency Use Authorization, as of March 31. More details here and here.

FDA Recommendations

The FDA made the following recommendation for use of serological tests:

Source: FDA

Serologic Testing Moving Forward

Different countries are using serologic testing for various purposes.

The United Kingdom recently bought 3.5 million tests and plans to deliver them to homes.

In Spain, serological tests were ordered from China, but withdrawn when discovered to have a 30% detection rate.

Germany will issue coronavirus antibody certificates to allow people to re-enter society based on their serology tests.

It seems like the CDC is developing serology tests to identify the full scope of the coronavirus outbreak. It is still unclear what is the United States strategy. I feel like we are still trying to figure out who is actively infected and recommending social distancing measures to make sure our healthcare system is not totally overwhelmed.

In my opinion, the biggest benefits of serology tests are for healthcare workers to allow them to return to work if they have developed immunity and to identify a cohort of people for which we can retrieve convalescent plasma.

CanSino Biologics Ad5-nCoV Vaccine

With a strong potential that we will see yearly coronavirus outbreaks every winter, we are in desperate need for a vaccine. In my previous post, I discussed the Moderna mRNA vaccine which is currently in Phase I clinical trials. In this post, I will talk about another promising vaccine technology and company also in Phase I clinical trials: CanSino Biologics and their adenovirus-vector vaccine.

Adenovirus-Vector Vaccines

There are three basic types of vaccines: 1. whole pathogen vaccines, 2. subunit vaccines, and 3. nucleic acid vaccines. In whole pathogen vaccines, the virus is killed or significantly weakened and then delivered to a person in order to stimulate an immune response against the virus. In subunit vaccines, only a component of the virus is delivered. Finally in nucleic acid vaccines, the virus DNA or RNA is delivered into a human cell which then get made into viral proteins and prompt an immune response.

An adenovirus-vector vaccine is somewhat like a combination of a whole pathogen vaccine and a subunit vaccine. In this case, we first identify the genetic sequence that codes for proteins on the virus surface. These surface viral proteins will be identified by our immune system and prompt an immune response to kill the virus. The next thing we do is insert this viral genetic sequence into an adenovirus, which are common, relatively benign viruses that cause the common cold, pink eye, etc. The final thing we do is inactivate the E1 gene of the recombined adenovirus, which prevents virus replication and amplification. A schematic of this process is below. A review article detailing the methods and clinical development of adenovirus-vectored vaccines is available from Afkhami et al.

Steps to build an adenovirus-vector vaccine
Source: Afkhami et al. Methods and Clinical Development 2016.

The end result of this process is that we have an inactivated adenovirus that displays a protein of a different virus for which we want to build memory immune cells to fight against (schematic below, more details from Rollier et al.).

Virus antigen presentation to immune cells from adenovirus-vector vaccine
Source: Rollier et al. Current Opinion in Immunology 2011.

The advantages for adenovirus-vector vaccines is that they are safe, create a long-lasting immune response, and create potent T cell responses. One disadvantage is that these vaccine types are less effective if a human has pre-existing immunity to adenoviruses.

CanSino Biologics Technology

CanSino Biologics is a Chinese biopharmaceutical company founded in 2009. To date, they have created 16 vaccine candidates using adenovirus-vector technologies. They have not been in the news as other much as other big biotechnology companies probably because they are based in China, but they have an impressive pipeline of products.

CanSino Biologics Pipeline.
Source: http://www.cansinotech.com/homes/article/plist/45.html

Their most recent vaccine is the Ad5-EBOV vaccine, which is a recombined adenovirus with an Ebola glycoprotein. CanSino Biologics was able to develop this vaccine and get it approved by the Chinese National Government in three years. This vaccine is part of the Chinese National Stockpile emergency use and is part of their national stockpile.

Zhu et al. showed that the Ad5-EBOV vaccine was safe and highly effective in building immune responses against the Ebola glycoprotein in a double-blinded phase II randomized clinical trial.

Ad5-EBOV vaccine creates robust glycoprotein-specific antibody responses.
Source: Zhu et al. Lancet 2016.

CanSino Biologics Ad5-nCoV Vaccine

CanSino Biologics used their same adenovirus-vector pipeline to build a vaccine candidate against COVID-19. In this case, they inserted the spike glycoprotein (the protein responsible for coronavirus entry into cells) into adenoviruses.

The company reported that they were able to trigger a strong response and satisfactory safety profile in preclinical animal studies. They were approved to begin a Phase I clinical trial in China on March 17.

Details of their clinical trial are here. Briefly, they are assessing the safety of three different doses of vaccine, and will measure adverse reactions for up to 6 months. They will also measure the amount of anti-spike protein antibodies, anti-SARS-CoV-2 neutralizing antibodies, and specific T cell responses against the virus for up to 6 months.

Conclusion

CanSino Biologics is another innovative biopharmaceutical company building a much-needed vaccine against coronavirus. They are an established company with many vaccine products and have already brought a vaccine to approval in a short amount of time. I am optimistic that they can succeed in their coronavirus vaccine as well.

FDA Grants Emergency Use Authorization for Chloroquine

On March 27, the FDA issued an Emergency Use Authorization for chloroquine phosphate and hydroxychloroquine sulfate for treatment of COVID-19.

These drugs have been the subject of fierce debate recently, with Donald Trump proclaiming that this can be “one of the biggest game changers int he history of medicine” and critics outcrying the lack of rigorous clinical trials providing evidence for this medication.

The evidence for chloroquine is still primarily anecdotal from case series experiences. Given the immediacy and debate, the FDA provided this emergency use authorization.

Chloroquine Updates

The French group that initially published a report stating that hydroxychloroquine and azithromycin together improve recovery from COVID-19 provided an update with their experience, available here.

Gautret et al. report that in 80 patients given this drug combination, 78 improved clinically with PCR levels of virus decreasing every day. Similarly, the number of patients with a positive viral culture dropped daily. In their experience, they had 1 death, 3 transfers to the ICU, and 12 patients requiring oxygen therapy.

PCR levels of Coronavirus Decreased with Hydroxychloroquine and Azithromycin
Gautret et al. 2020.
Number of Patients with Positive Coronavirus Culture
Gautret et al. 2020.

At present, the French group treated over 1,000 patients and only had one death with hydroxychloroquine and azithromycin.

Chen et al. posted the results of their clinical trial evaluating the efficacy and safety of hydroxychloroquine. In their experience, patients who received hydroxychloroquine had a 2 day faster recovery time from cough and fever. Additionally, a significantly greater proportion of patients had improved imaging findings with hydroxychloroquine (80%) compared to control (55%). All four patients who progressed to severe illness were in the control group.

Chen et al.
Source: https://www.medrxiv.org/content/10.1101/2020.03.22.20040758v1.full.pdf

Emergency Use Authorization

The FDA granted emergency use authorization for chloroquine and hydroxychloroquine for treatment of patients hospitalized with COVID-19 for whom a clinical trial is not available. The reasons for their recommendation are as follows:

Source: https://www.fda.gov/media/136534/download
Source: https://www.fda.gov/media/136535/download
Source: https://www.fda.gov/media/136537/download

Conclusions

This is the first therapy granted emergency use authorization from the FDA. 30 million doses of hydroxychloroquine sulfate and 1 million doses of chloroquine phosphate are added to the Strategic National Stockpile, provided by Sandoz and Bayer Pharmaceuticals.

We still need more evidence to help determine what is the optimal dosage and for how long therapy should be administered.

This authorization helps clear a lot of the hysteria about these medications, given that there has been at least one death from someone taking chloroquine phosphate used to clean fish tanks. People should not use chloroquine phosphate intended for fish, as the FDA noted. These aquarium products have not even been evaluated to see if they are safe for fish, let alone humans.

There is a lot of bickering on social media and news outlets about the efficacy of chloroquine and efficacy. I think the typical peer review publication process which takes months, requires volunteer reviewers, and still lets plenty of fake results through the pipeline is inappropriate to address our current crisis. We need something better.

Abbott Rapid Point of Care Detection of COVID-19

Yesterday, the FDA has issued Emergency Use Authorization for Abbott’s point of care detection of COVID-19.

Source: Abbott ID Now

Abbott’s technology can deliver positive results in as little as 5 minutes and negative results in 15 minutes. This test utilizes the ID NOW platform, which is currently used for rapid detection of influenza, respiratory syncytial virus, and streptococcus pneumoniae. Using this technology, Abbott plans to delivery 50,000 COVID-19 tests per day. Press releases from Abbott are here and here. A video summary of the device is below.

Abbott ID Now Demo Video

In this post, I’ll summarize what we know about this technology.

Background of Nucleic Acid Amplification

The gold standard of nucleic acid amplification is polymerase chain reaction. This is also our current gold standard for detecting COVID-19 in a patient.. In this process, template DNA is denatured and split into two separate strands (Step 1: denaturation), primers are added that bind to the split strands (Step 2: annealing), and new nucleic acids are added which elongate from the primers and bind to the template DNA (Step 3: elongation). Steps 1, 2, and 3 are repeated to continuously amplify nucleic acids. This process works well, but is laborious, technically demanding, expensive, and takes hours to complete. It is also limited in scenarios with increased patient volumes, such as what we are seeing in the current COVID-19 pandemic.

Steps for PCR.
Source: Wikipedia

The most laborious step of polymerase chain reaction is the constant denaturation and elongation steps which require different temperature ranges to function. To overcome this challenge, scientists have developed various isothermal nucleic acid amplification techniques to detect a specific nucleic acid sequence.

Abbott’s Technology

The company’s technology is based off a patent from Maples et al. in 2009 to provide isothermal amplification of nucleic acids using a nicking mechanism.

The mechanism is described in more detail in Bell et al. and Nie. et al, with a figure below. My understanding is that you first start with a template DNA strand, constructed using reverse transcriptase from viral RNA. Then specific nicking enzymes open the template strand which allow for DNA polymerase enzymes to create new DNA. However, each newly constructed DNA will also get nicked to allow for construction of another DNA strand. This process leads to exponential growth of DNA strands in minutes.

Source: Bell et al. Journal of Clinical Virology 2014.

Sensitivity and Specificity compared to PCR

Abbott’s ID Now technology was previously shown to have over 96% sensitivty and specificity to detect influenza when compared to RT-PCR as the gold standard.

Source: Bell et al. Journal of Clinical Virology 2014.

Results disclosed to the FDA show that the ID Now also is consistent with RT-PCR for detecting SARS-CoV-2. There was 100% agreement in detecting 30 samples of SARS-CoV-2 and not detecting the virus in 30 controls.

Source: https://www.fda.gov/media/136525/download

The limit of detection was 125 genome equivalents/mL which can be obtained >95% of the time with a standard nasopharyngeal swab.

Overall, this is a major breakthrough in our fight against COVID-19. We can quickly, safely, accurately detect COVID-19 in a massive scale.

Limitations

One caveat is that we do know that the sensitivity of RT-PCR for detecting COVID-19 is variable across test kits. The sensitivity can be as low as 71% compared to a clinical suspicion of COVID-19 with positive chest CT findings (Source Fang et al. Radiology 2020). This means that there may be cases where the Abbott technology provides a false negative result. This may be more prevalent during early signs of disease, and this is why some people need two or more tests to confirm or rule out COVID-19. However, this is less of a problem for Abbott’s technology because it only takes ~10 minutes to run.

Moderna mRNA Vaccine for COVID-19

In the last 20 years, we had three major pandemics from coronaviruses: SARS, MERS, and COVID-19. Now that COVID-19 is evolving and mutating, we are facing the threat that the world faces yearly coronavirus outbreaks, similar to influenza. We cannot afford that reality, so we are in urgent need of a vaccine now. The typical length of time it takes to develop a vaccine is 10-15 years, involving rigorous Phase I, II, III, and IV trials demonstrating safety and efficacy. However, we need something now.

Moderna, Inc was the first company to develop a potential mRNA-based vaccine and launch a Phase I human study in collaboration with the National Institute of Allergy and Infectious Diseases. In this post, I’ll describe what we know about Moderna, mRNA vaccination, and the Phase I study.

Moderna, Inc

Moderna was founded in 2010, based off the laboratory research of Derrick Rossi. At that time, Rossi’s lab made synthetic mRNA in order to convert somatic cells (i.e. skin cells) into pluripotent stem cells and then muscle cells (Warren et al. Cell Stem Cell 2010). Tucked away in their supplementary figures is how they make synthetic RNA.

Source: Warren et al. Cell Stem Cell 2010

Moderna first partnered with Flagship Ventures to make mRNA to design any protein of interest. Using this technology, they have made synthetic mRNA solutions to treat infectious diseases, cancers, and rare diseases.

They have built prophylactic mRNA vaccines against a variety of infectious diseases such as Cytomegalovirus, Zika virus, and Ebstein-Barr virus.

Source: Moderna, Inc

A full list of their preclinical publications showing the efficacy of the mRNA vaccine in animals is listed here. For example, they showed that their mRNA vaccine can protect both a pregnant mouse and its newborn from the Zika virus (Jagger et al. Journal of Infectious Diseases 2019).

Source: Jagger et al. Journal of Infectious Diseases 2019

mRNA Vaccines

There are three broad categories of vaccine types: whole-pathogens, subunit, and nucleic acid derived. More detail here. Briefly, whole pathogen vaccines are killed or weakened viruses which are then delivered to people, such as the MMR vaccine. Subunit vaccines include only the components of the virus that activate the immune system (i.e. Pertussis vaccine). Nucleic acid vaccines introduce virus DNA or RNA into a cell, which then create viral proteins, and then lead to an immune response against the newly created viral proteins.

There are no current mRNA vaccines on the market used for prophylaxis against infectious disease. These vaccines have been challenging to make in the past because they have been difficult to deliver in vivo and they are prone to degradation from ribonucleases. However, there have been several technologies developed in the last decade to address these technologies. More detail is in the following review article by Pardi et al.

To improve delivery in vivo, scientists have tried a variety of approaches, summarized in the visual below:

Source: Pardi et al. Nature Reviews Drug Discovery 2018

To improve stability, scientists have added modifications to synthetic mRNA to make it resemble more like native mRNA, summarized below:

Source: Pardi et al. Nature Reviews Drug Discovery 2018

From my understanding of the research literature, a popular delivery mechanism is the liquid nanoparticle, which enhances antigen expression dramatically (Zhang et al. Front Immunol 2019).

Using liquid nanoparticles and mRNA vaccines, scientists have created robust immune responses against many viruses, such as the Zika virus and Ebola virus. These responses were all in animals, but a phase I trial using a liquid nanoparticle mRNA vaccine did show a significant immune response against the H10N8 and H7N9 influenza viruses in human volunteers. Figures showing the antibody persistence are below.

Feldman et al. Vaccine 2019

Moderna Clinical Trial

The Moderna mRNA vaccine is a synthetically stabilized mRNA sequence coding for the full-length, prefusion stabilized spike protein of SARS-CoV-2, which is responsible for viral entry into the cell. They deliver this vaccine in a lipid nanoparticle.

It seems like Moderna based their vaccine off Wu et al.‘s first genomic sequence of SARS-CoV-2 and Wrapp et al.‘s characterization of the spike protein in the prefusion conformation (both figures below).

Genomic sequence of SARS-CoV-2
Wu et al. Nature 2020
Spike protein 3D structure
Wrapp et al. Science 2020

Moderna then incorporated this sequence into their preexisting mRNA vaccine pipeline and was able to get a vaccine in a remarkable 42 days after the genetic sequence was published. Moderna press release here. It normally takes years to get to this step.

Moderna then launched a phase I open labeled study, enrolling 45 people into three cohorts, testing three different doses of the vaccine. Each person will get an IM injection of the mRNA vaccine on Day 1 and Day 29 of the study. The primary objective is to assess safety of the vaccine but Moderna will also assess how well this vaccine creates antibodies against SARS-CoV-2 at around the 3 month mark.

This means we are still months away from knowing how well this vaccine works. We are still at least 12 months out from getting this vaccine past phase II and III trials so that it can be used in humans. However, after reading the literature I am pretty optimistic.

Summary

We desperately need a vaccine against coronaviruses. We already had 3 pandemics in the last 20 years, and we will probably get another coronavirus pandemic. There is a real risk that we get yearly outbreaks just like influenza.

The Moderna mRNA vaccine is a new technology that has not been proven. However, it is fast to develop and the preclinical data of the mRNA vaccine technology is promising. I am hopeful that this will work.