AP Biology class blog for discussing current research in Biology

Tag: clinical trials

A Life Saving Treatment: CRISPR Gene Editing

A proud, hard-working father is what Paddy Doherty looked up to all of his life until a sudden heart attack that took the life of his dad. What would you do if someone you love is unexpectedly gone without a goodbye?

His father had a career in construction and various home improvement projects which kept him active until his 60s until Doherty first caught glimpses of a worrying decline in his dad’s health. “I noticed him getting breathless on walks. He’d stop for a while and maybe make an excuse for stopping, saying, ‘Oh, isn’t that a lovely tree’ or whatever,” said Doherty, who lives in Ireland. Doctors chalked it up to angina, or chest pain caused by reduced blood flow to the heart, symptomatic of an underlying heart problem.

After his dad died, the true cause was discovered: a rare disease called transthyretin (ATTR) amyloidosis, characterized by a misfolded protein that builds up in the heart and interferes with normal function. As learned in AP Biology, misfolded proteins are caused by the lack of chaperonins that are present in cells to provide a secure hydrophilic environment. The misfolded proteins cannot achieve their native state and are contorted into shapes that are unfavorable to the environment it’s in. The formation of oligomers and aggregates occurs in the cell when a critical concentration of misfolded protein is reached. Aggregated proteins inside the cell often lead to the formation of an amyloid-like structure, which eventually causes different types of degenerative disorders and ultimately cell death.


Structure of Wild Type Human Transthyretin in Complex with Tafamidis, PDB 6E6Z

“Patients left untreated with this type of amyloidosis develop heart failure, low blood pressure, horrible bowel disturbance, and eventually become incontinent of urine and feces,” said Julian Gillmore, nephrologist and head of the National Amyloidosis Centre at University College London. “It’s a truly awful, gradually progressive disease that is ultimately fatal.”

In February last year, Doherty began to experience the same early breathing symptoms his father had had. As an avid hiker who had trekked the Himalayas, he was surprised to find himself getting winded on local hill walks. Testing confirmed that Doherty had a hereditary form of ATTR amyloidosis.

But there was one bit of good news: If Doherty had been diagnosed even a year earlier, no treatment options would have been available to him — an all-too-common situation for over 30 million U.S. patients with rare diseases. But Gillmore, Doherty’s doctor, offered him the chance to participate in an early-stage clinical trial using CRISPR, a groundbreaking genome editing therapy with the potential to cure his ATTR amyloidosis in a single dose.


“I had no side effects and left the facility after two days,” Doherty said. “The walk that I felt breathless on, which is a steep kind of mountain walk through a forest, I’m doing that every Sunday now.” CRISPR-Cas9 allows researchers to alter the DNA of living things at will. It works like genetic scissors that can insert, repair or edit individual genes to rewrite the code of life. The system itself consists of two molecules — a protein known as Cas9 that works like scissors and a guide RNA that takes Cas9 to the right place in the genome — that can be inserted into cells or the bloodstream.

In the case of the clinical trial on patients with ATTR amyloidosis, Gillmore and his colleagues aimed to edit the malfunctioning gene itself and demonstrate for the first time that direct infusion of CRISPR molecules into the bloodstream is safe effective.

The hereditary form of ATTR amyloidosis affects roughly 50,000 people worldwide with a large cluster of patients like Doherty with roots in Donegal County, Ireland. Because circulating transthyretin is made almost entirely in the liver — and everything that enters the bloodstream is carried to the liver to metabolize — the researchers realized they could simply inject patients with the CRISPR-based therapy.

The therapy, called NTLA-2001, appeared to knock out the mutated gene as intended. Only six patients were tested in total, but the three who received the higher of two doses — including Doherty — saw their transthyretin levels drop by an average of 87 percent after 28 days. The results remain preliminary, and several more patients will need to be tested before the trial is complete.

Doherty said he hopes his family members and fellow Donegal residents will be able to benefit from CRISPR as much as he has. Fortunately, testing shows his two daughters did not inherit ATTR amyloidosis. And along with his father, Paddy’s uncle and cousin both died of the disease.

“When the trial is over, I hope that CRISPR is available and affordable for all amyloidosis patients,” Doherty said. “If a pharmaceutical company can mass-produce something like that and sell it at a good price, it would be a godsend.”

That’s a Foul: “Interfere”-ons and the Fight Against Covid-19

Great news to those infected with Covid-19 – scientists have discovered a new (if risky) way to ameliorate the disease’s advanced symptoms.

According to Scientific American, several studies have concluded that immune proteins could suppress the potentially fatal “viral replication” that results from Covid-19. The proteins in question, called “interferons,” are released in “response to virus infection” and “induce numerous molecular changes that ‘interfere’ with cellular functions in order to prevent inflammation (according to ScienceDirect) .” Such a function could be likened to the way that enzymes in the lysosome break down foreign material, viruses, and bacteria. 

However, similar to lysosomes, the overpopulation or malfunctioning of interferons can have undesired consequences. Described by immunologist Eui-Cheol Shin as “a double-edged sword,” interferons may inadvertently worsen a patient’s deadly respiratory problems or lead to a dangerous disease called “children’s interferonopathies.”

Furthermore, several health organizations have warned against the use of interferons in treatments for the coronavirus. Citing “insufficient data to recommend either for or against the use of” the protein, the NIH does not endorse the proposal being practiced outside of extensive clinical trials. With major powers in the public health domain forbidding interferon-based treatments, it appeared that in-depth studies were needed to make a case for the idea.

Luckily, Shin and his fellow scientists were prepared to do just that. Research teams in France and South Korea conducted blood tests and RNA sequencing, respectively. The results of both tests were, to put it bluntly, not very conclusive. In the former experiment, patients with advanced Covid symptoms were surprisingly discovered to have a rather low count of interferons; conversely, the findings of the latter study showed that the interferon count of patients affected by severe symptoms was actually quite high compared to patients with mild symptoms. 

At first, this clash in results suggested that determining the full effect of interferons in cells would be extremely difficult. However, a clinical trial conducted by Eleanor Fish (an immunologist at the University of Toronto) showed that “[interferons] helped clear viral infections almost seven days sooner on average than people given arbidol hydrochloride, a drug thought to block viral entry to cells.” This conclusion would be a huge win in the rapidly escalating fight against Covid-19. Despite the notion in sports that an interference is an unfair foul, the interferons might just be the interference needed to save countless lives.

Gut Microbes Help to Advance Flu Vaccines

Beneficial Gut Bacteria

This September, a potentially monumental study was published in the scientific journal, Cell, reporting that researchers have confirmed that microbes present in the gut can change, lower, or jumpstart our immune response.  Previously research has only been done with other mammals such as mice, and this was the first study that linked the results to human subjects. Since most previous trials were conducted on other animals, researchers such as Dan Littman who studies microbiota at NYU School Of Medicine, emphasized there are likely to be large differences in the results for humans versus other animals.   

Specifically, researchers found that people who have not received a flu shot or had the flu within the past 3 years and then were administered broad spectrum antibiotics, produced lower levels of antibodies to the influenza virus. Those subjects who did not receive the antibiotics produced more antibodies to the flu virus. This publication is so noteworthy because previously so little actual human clinical trials were performed to understand the role of the human gut microbiome and its relationship to the strength of our immune response.  

Previous research on how the flu vaccine works and its varying efficiency among many people has been done.  In 2011, Bali Pulendran, an immunologist at Stanford University, found that increased activity in the gene receptor that recognizes the bacterial protein flagellin, the core part flagella, seemed to stand out as the one major change among how well the flu shot was working in varying groups of people.  This underscores the connection between the immune system’s recognition of bacteria (especially gut microbes) and  how well people may respond to the flu vaccine.  

In 2014, this research was followed by gene knockouts being given to mice for the receptor for bacterial flagellin in the flu shot.  The results showed that the mice who received the knockouts made were antibodies than the control mice in the trial.  The researchers suspected this reduction was controlled by the absence or presence of gut microbes and their ability to sense flagellin.  To confirm this, researchers followed up with separate trial in which mice’s microbiota were reduced by the administration of antibiotics before receiving the flu vaccine and control mice who did not receive the antibiotics so their microbiomes remained present.  The results again showed a link that gut microbiota play a role in levels of antibodies produced against their flu shot.  Because of these results, it seemed obvious to test the same situation with humans. 

The current study did just that and was designed as a Phase 1 clinical trial to determine if gut microbes are connected to the efficiency of flu vaccine immunity.   11 adults received broad spectrum antibiotics for 5 days and 11 served as the control and did not receive antibodies.  All subjects receive the influenza vaccine on day 4. The people who received the antibiotics had reduced levels of gut microbes.  However, no major difference was observed in response to the vaccine. These results prompted researchers to dig deeper and they next investigated people who had not had the flu shot or suffered from the flu virus within the last 3 years.  They wanted subjects that would be relatively clear of flu antibodies to begin with. They repeat a very similar study with 11 people, 5 receiving the antibiotics and 6 serving as controls. Everyone got the flu vaccine, but this time the results showed a marked difference in vaccine induced immunity.  Subjects who received antibiotics and had fewer microbes presents, made far fewer flu-specific antibodies.   

This research is very promising not only in the field of flu vaccination, but could reveal that changes to microbiota can have profound impacts on future vaccine development for a variety of pathogens.  Because the results were so tiring, Pulendran is continuing to research deeper into the relationship between gut bacteria and vaccines, for viruses that may affect us in the future. This holds promise for development of vaccines for a wide range of pathogens that attack the human race.  


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