Snapshot: What Does Success Mean in Clinical Trials with Antisense Oligonucleotides (ASO)?

Research is rapidly moving from the bench to the bedside to treat neurological inherited disorders of all types, including spinocerebellar ataxias. SCAsource has previously gone over the science behind ASO therapy. These diseases share a common theory that the DNA mutation leads to the formation of an altered protein that is toxic. ASO therapy is meant to stop the formation of the toxic protein by “shooting the messenger”.

What is involved in these clinical trials?

To see what might happen in ataxia trials, let’s look at ASO trials happening right now in related polyglutamine diseases. In Huntington’s disease (HD), there are two programs that are currently in clinical trials. Regulatory authorities view ASOs as drugs and require that the product be shown to be both safe and effective in patients.

ASOs cannot be given as pills and they are currently injected into the spinal fluid. This is called intrathecal administration to get the drug directly in the fluid space where it can circulate back to the brain. Patients in phase 1 studies in HD are asked to have up to 7 injections and one phase 3 program requires injections every second month for 2 years. This involves a large commitment to the study and is asking a lot from patients and their families.

The only published phase 1 double-blind, placebo-controlled study in HD (Tabrizi et al., New England Journal of Medicine, 2019) has identified that a series of 4 injections were safe. They measured changes of the “bad” protein in the spinal fluid as a proof of concept that ASOs could lower protein levels. The good news was that they found that there was a dose-related reduction in this protein of about 40%. Patients from this study were offered “open label” monthly injections and this has shown a 60% reduction in the abnormal protein according to a recent presentation. Open label extensions are when patients can continue taking a drug after the formal time of the clinical trial is over.

medical doctor in blue scrubs and a white lab coat holding a stethoscope. They are off to one side, so only have their body can be seen, not inclduing their face.
What will ataxia clinical trials involving ASOs look like in the future? What will success look like?

So, what does success mean?

The phase 3 studies that are currently ongoing in HD are designed to see if there is a slowing of disease progression. This is being measured by assessing motor, cognitive and behavioral symptom change over time. Changes occur slowly in HD and SCA. Therefore, large numbers of patients are required over a relatively long study time.

The bottom line is that a successful study that shows slowing disease progression is likely to mean that the patients may not experience any obvious improvement while receiving the treatment and that they will continue to have progressive symptoms over time. Hopefully, this will be at a slower rate compared to the placebo group. Since there are no treatments available for SCA or HD, this will be welcome. It is by no means considered to be a cure or likely to stop the progression. True cures in medicine are rare, where a cure is defined as a drug ending disease.

Graphs of symptoms vs time. The "typical progression" line has more symptoms more quickly. The "delayed progression after potential treatment" line has fewer symptoms, but still increases over time.
Graph explaining how a potential ASO treatment might work in the future. Although it might not make symptoms go away completely, it could reduce how severe symptoms are, the number of symptoms, and/or delay when symptoms first appear. Illustration by Celeste Suart.

In the HD research community, we are asking questions that include:

  1. Is it a good idea to reduce the good protein that is part of our normal brain chemistry? In the current phase 3 study, the ASO reduces both the “good” and the “bad” HD protein. Another program in phase 1 uses an ASO that only reduces the “bad” protein.
  2. When is the best time to use ASO therapy? Since these conditions are associated with nerve cell damage and loss, it makes sense to use these types of therapy very early, even before damage occurs. This will mean that patients with moderate or advanced symptoms may not be good candidates for ASO therapy.
  3. Should we consider treatment in people who have had predictive genetic testing before symptoms start? This is being actively discussed but it is too early to consider this. We have to show that ASOs are safe and effective in symptomatic patients. We need to have good measures to determine if treatments are working. Regulatory authorities have required evidence that treatments have a positive effect on patients lives. This may be difficult to show in a short study. We must consider that it takes patients decades to get these diseases: slowing or stopping this could take just as long.

We can only figure out the answers to these questions in clinical trials. The goals of these trials are to improve people’s quality of life. To do this we need information from real people with these diseases, and not just models of disease. This is a process that will take time but will tell us which approach has the most promise and is worth pursuing faster. Thus, the patients and families at this point are just as important as the researchers in lab coats working together to treat these diseases.

If you would like to learn more about clinical trials, take a look at this resource by the FDA or our previous Snapshot on the subject.

Snapshot written by Dr. Mark Guttman and edited by Dr. Ray Truant.

Snapshot: What is recessive ataxia?

What is a recessive disorder?

A recessive disorder is one that has a specific disease mechanism. For a recessive disorder to occur, both copies of the causative gene must be mutated for a patient to show symptoms.  Ataxias that follow this disease mechanism are known as recessive ataxia. However, having a mutation in only one copy of the gene does not lead to a disorder. As people with only one mutated copy of the gene can pass on the defective gene, these people are known as an unaffected carrier. Recessive ataxias range in symptoms and severity but are linked by their disease mechanism. While none of the Spinocerebellar Ataxias (SCAs) are recessive, there are many types of recessive ataxias, including Autosomal Recessive Cerebellar Ataxia Type 1 and 2 (ARCA1 and ARCA2), Autosomal Recessive Spastic Ataxia of Charlevoix-Saguenay (ARSACS), Friedreich’s Ataxia, and Ataxia Telangiectasia. For example, Friedreich’s Ataxia is caused by a trinucleotide repeat expansion in the frataxin (FXN) gene. People with only one expanded copy of the FXN gene do not show any symptoms, while people with two expanded copies of the FXN gene are affected by Friedreich’s Ataxia.

How are recessive ataxias inherited?

For every gene in our body, we have two copies, one that is inherited from our mother and one from our father. Both parents of an affected individual have to have at least one copy of the mutation for a child to be born with a recessive disorder. If both parents are unaffected carriers, each child will have a 1 in 4 chance of getting the disorder.

For a patient affected with a recessive ataxia, the chances of having a child affected by the same disorder are low. For a patient to pass on the disease, their spouse must have at least one mutated copy of the causative gene. In the case where a patient’s spouse is a carrier, children have an equal chance of being an unaffected carrier or being affected by the disease. However, carrier rates for ataxias are low in the population, which makes it unlikely that a patient’s spouse is also a carrier for the ataxia mutation.

Map showing the statistical chance of two unnafected carrier parents passing on a mutated gene (25% unaffected child, 50% carrier child, 25% affected child) or an affected parrent and unaffected carrier (50% carrier child, 50% affected child)
How recessive disorders are inherited. Image by Eder Xhako, created with BioRender

How can a patient prevent passing on a recessive disorder to their children?

Generally, when a patient with recessive ataxia passes on the disorder to their children, their spouse is an unaffected carrier. If you are a patient with a form of recessive ataxia and are thinking about having children, your spouse can undergo carrier testing to find out if they are a carrier for the same recessive ataxia. This will determine the likelihood that the recessive ataxia is passed on to your children. If it is determined that the spouse is a carrier, options like IVF with embryo screening can help patients prevent passing on recessive ataxia to their children.

If you would like to know more trinucleotide repeat expansions, you can look at our past Snapshot on Polyglutamine Expansion.

If you would like to learn more about carrier and embryo screening, take a look at these resources by the American College of Obstetricians & Gynecologists and Integrated Genetics.

Snapshot written by Eder Xhako and edited by Larissa Nitschke.

Snapshot: What is Magnetic Resonance Imaging (MRI)?

What is it?

Magnetic resonance imaging (MRI) is a type of technology used to take detailed pictures of the body. It is commonly used to detect abnormalities in the body, diagnose diseases, and to regularly monitor patients who are undergoing treatments. It can generate three-dimensional images of non-bony tissues, such as the brain. MRI procedures are non-invasive, require minimal preparation, and are not associated with health risks, as it does not use harmful types of radiation such as X-rays.

How does it work?

Human tissues contain water, which contain very small particles known as protons that behave like tiny magnets. An MRI machine uses large, powerful magnets to generate a magnetic field that can change how these particles rotate in your body, making them align with the magnetic field. Non-harmful radio waves are then pulsed through the patient, changing the direction of these particles, such that they are no longer aligned with the magnetic field. The radio waves are then turned off, and the particles can then re-align with the magnetic field. Different types of tissue and structures in the body will have particles that re-align differently, which can be detected by the machine to generate a detailed black and white image of the scanned area of the body. In addition to such structural information, MRI scans can provide information about how the brain is wired, levels of important chemicals, blood flow, metabolism, and brain function by acquiring information differently with the same machine.

3D view of an entire human brain taken by MRI, shown from two angles.
3D view of an entire human brain taken by 7 Tesla MRI. Photo courtesy of  B.L. Edlow et al, bioRxiv, 2019.

How do you prepare for an MRI scan?

Since an MRI scan uses a large magnet, electronic devices and metal objects, such as glasses and jewelry, must be removed. There is usually no other preparation required for the scan. Patients must lie very still to generate a clear image. Patients do not need to be sedated, unless they have trouble lying still for the procedure. MRI scans that are obtained for research do not use anaesthesia to avoid unnecessary risk to research participants.

What happens during an MRI scan?

The patient lies down on a table that will move into the tunnel-shaped chamber. The patient is usually awake and will remain in the chamber as several scans are taken during the procedure (about 30-60 minutes). As the scan proceeds, there are often loud mechanical sounds, so earplugs are provided for protection. Some patients may experience claustrophobia, or are bothered by the noises. Becoming more familiar with the procedure, or listening to music or closing your eyes can help alleviate discomfort during the scan.

What do doctors look for in patients with SCAs?

MRI scans are often used to image the brain to detect signs of spinocerebellar ataxia (SCA), especially in a region of the brain known as the cerebellum. SCA is associated with brain cell loss, and appears as reduced volume of brain tissue in the MRI image.

If you would like to learn more about Magnetic Resonance Imaging (MRI), take a look at these resources by the National Institutes of Health and the Mayo Clinic.

Snapshot written by Dr. Claudia Hung and edited by Dr. Gülin Öz.

Snapshot: What is RNAi?

RNA interference, or RNAi, is a natural biological process that inhibits the expression of a specific gene. In medicine, targeted RNAi therapies can be used to silence the expression of a disease-causing gene. To understand RNAi, you first have to understand RNA.

DNA is transcribed to make mRNA, which is tranlated by the ribosome to make protein.

An overview of  RNA is the messager between the DNA (the instructions) and the protein (the product). RNA is transcribed from the DNA. The ribosome translates the mRNA into protein. Graphic designed by Colleen Stoyas and illustrated by Celeste Suart.

Genes encode the instruction manual of our biology, but this material cannot leave the nucleus of your cells. Think of genes as a lecturer that provides instruction for your homework, which you must copy and take home to use later. The equivalent of copying this message in the cell is RNA, which transcribes the gene instructions and leaves the nucleus to be read and translated into protein. This protein then performs functions within the cell (see above image).

How can RNAi be used in ataxia?

In specific forms of ataxia, a gene mutation may provide the instructions for a protein that acts improperly and leads to disease. RNAi is a method of silencing RNA that interferes with the reading of this message, keeping a protein from being made. It works by generating a small interfering RNA in the laboratory that matches the gene of interest. When this small interfering RNA enters the cell, it binds the matching messenger RNA copied from a gene. When these two RNAs bind, the cell is triggered to cut up the message and destroy it. This means the disease-causing protein is never made. (see below image)

RNAi works by binding the mRNA, preventing it from being transcribed by the ribosome. This stops protein from being made.
How does RNAi work? It binds matching messenger RNA. This stops it from being translated by the ribosome into protein. Graphic designed by Colleen Stoyas and illustrated by Celeste Suart.

While RNAi is straightforward in the lab, getting it to work in humans can be tricky. The small interfering RNA cannot be taken in a pill, because it will not survive digestion. Additionally, the small interfering RNA is degraded along with the target messenger RNA, and so it must be continually administered. Using a viral payload, or encapsulating the interfering RNA in the coat proteins of a virus, has successfully delivered RNAi therapies in mouse models of SCA1, SCA3, and SCA7. In this method the virus integrates into your cells, which can then continue to produce the small interfering RNA. This means a single dose could potentially be all that is needed. Viral delivery to the brain is complicated, but not impossible. More work remains to be done clinically in order to determine if RNAi therapy is viable in a viral payload to treat multiple forms of spinocerebellar ataxia.

If you would like to learn more about RNAi, take a look at this video by TED-ED or entry in the Encyclopedia Britannica.

Snapshot written by Dr. Colleen Stoyas and edited by Frida Niss.

Continue reading “Snapshot: What is RNAi?”

Snapshot: What is RAN translation?

In many diseases caused by repeat expansion mutations in the DNA, harmful proteins containing repetitive stretches are found to build up in the brain. The repeat expansion mutation, when translated into a protein, results in an abnormally expanded repeat tract that can affect the function of the protein and have harmful consequences for the cells. Following a study published in 2011, we know that repeat expansion mutations can make additional harmful repeat-containing proteins by a process called Repeat Associated Non-AUG translation or RAN translation.

How are proteins made?

To get from DNA to protein, there are two main steps. The first step involves the conversion of a gene in the DNA into an instructional file called messenger RNA (mRNA). The second step is translation, this is where the cellular machinery responsible for making proteins uses mRNA as a template to make the protein encoded by the gene.

During translation mRNA is “read” in sets of three bases. Each set of three bases is called a codon and each codon codes for one amino acid. There is a specific codon that signals where to start making the protein, this codon is AUG. From the point where the cellular machinery “reads” the start codon, the mRNA is “read” one codon at a time and the matching amino acid is added onto the growing protein.

What happens when there is a repeat expansion mutation?

As the name suggests, Repeat Associated Non-AUG (RAN) translation is a protein translation mechanism that happens without a start codon. RAN translation occurs when the mRNA contains a repeat expansion that causes the mRNA to fold into RAN-promoting secondary structures. Because RAN translation starts without an AUG start codon, the mRNA can be “read” in different ways.

Let’s consider a CAG repeat expansion to illustrate this process. In the CAG “reading frame” a polyglutamine containing protein would be made because the codon CAG leads to incorporation of the amino acid glutamine. But a CAG repeat expansion could also be “read” as an AGC or a GCA repeat expansion if you don’t know where in the sequence to start “reading”. When “read” as AGC, the cellular machinery would incorporate the amino acid serine, making a polyserine repeat protein. In the GCA frame a polyalanine repeat protein would be made. This has been shown to happen in Huntington’s disease (HD). In HD, RAN-translated polyserine and polyalanine proteins accumulate in HD patients’ brains, along with the AUG-initiated mutant huntingtin protein containing a polyglutamine expansion.

Diagram show how different DNA sequences can be "read" and translated as different proteins
Overview of repeat proteins that can be produced by RAN-translation from a CAG expansion transcript. Designed by Mónica Bañez-Coronel.

To complicate matters more, RAN translation can happen from different repeat expansions, including those in regions of the DNA that aren’t normally made into proteins at all. Through the process of RAN translation, repeat expansion mutations in the DNA can give rise to multiple different proteins that aren’t made in healthy individuals. RAN proteins have now been identified in several neurodegenerative diseases where they have been shown to be toxic to cells, including in HD, spinocerebellar ataxia type 8, myotonic dystrophy type 1 and 2, and C9orf72 amyotrophic lateral sclerosis (ALS).

To learn more about the implications of RAN proteins for repeat expansion diseases see this article by Stanford Medicine News Center.

To learn more about the process of translation see this article by Nature.

https://www.nature.com/scitable/topicpage/translation-dna-to-mrna-to-protein-393/

For the original article describing RAN translation see this article by PNAS, and this article by Neuron about RAN translated proteins in Huntington’s disease.

Snapshot written by Dr. Hannah Shorrock and edited by Dr. Mónica Bañez-Coronel.