Multiple sclerosis
Multiple sclerosis (MS) is the most common disabling neurological disease among young adults.1 MS is an autoimmune disease in which the insulating layer (or myeline sheath) surrounding nerves in the central nervous system (CNS) is damaged (Figure 1). The exact mode of action is still unknown.1 These attacks cause nerve damage resulting in loss of motor control and impaired sensory function.
The most common form of MS is Relapsing-Remitting MS (RRMS), where the attacks occur in localised flares lasting approximately one day. The resulting nerve damage may lead to various symptoms such as spontaneous blindness, or the loss of limb functioning.1 Since the attacks are typically abrupt and deeply disturbing, many people seek medical care fast. However, the path to diagnosis can be long and challenging.2
Diagnosis
The current leading diagnostic method is based on the McDonald criteria, which heavily rely on MRI findings. Since RRMS occurs in flares, it is primarily determined when two flares correspond with two new appearances of damage on the MRI, separated in both time and space. However, these McDonald criteria are not always satisfied after the first scan.3 Often the damage observed with MRI could be attributed to several other conditions, making the result inconclusive.3 When only a single flare has occurred neurologists speak of a clinically isolated syndrome (CIS), which cannot be diagnosed as MS with just MRI. Waiting for a second flare to occur can take years.
Once MRI confirms a potential case of MS (Figure 2), it is often necessary to perform a lumbar puncture. This involves taking a sample of the cerebrospinal fluid (CSF), that can contain antibodies, protein fragments and white blood cells associated with MS or exclude the presence of other diseases. This is a highly invasive procedure, and it is not uncommon that patients refuse.5 Even when all the options mentioned above have been exhausted, around 10% of the patients cannot be diagnosed for months or even years.2
The lack of conclusive evidence for MS can cause patients to not receive a proper diagnosis for MS, resulting in a physical and mental burden on them.6 Waiting for a diagnosis without treatment is problematic, because the attacks cause lasting damage. Timely treatment can avoid this damage, but people can only access medication that prevents flares and accumulated damage when they are diagnosed (Figure 3). This highlights the need to develop new tests that contribute to improved diagnoses for complex cases of MS, with minimal patient discomfort.
Accessible
To learn more about the diagnosis of MS we conducted interviews with doctors, researchers and patients, as can be seen in our integrated HP page. Based on the interviews with our stakeholders, we established a set of requirements for an ideal diagnostic test. In one word, we need the test to be accessible. This is because improving the diagnosis does not only happen in the examination room, but also in laboratories and accountants’ offices. For diagnostic tools to be useful in a clinical setting, according to our stakeholders, they must qualify for several criteria (Figure 4).
The first key requirement of accessibility is being cost-efficient. In Europe, access to neurologists can be constrained due to long waiting lists. In non-western countries, the health care system may have even more restricted options, since specialised equipment is not, or scarcely available.7 These facts must be considered; the cost of a medical procedure must be minimised for it to be accessible to a wide audience.
Secondly it should be easy to perform. A lot of costs are hidden in the manhours of the expert work of lab specialists and neurologists. This labour should be minimised, both in sampling, processing and analysing. This would allow them to focus their time and expertise on helping more patients.
The diagnostic test must also have a long shelf-life. Many biotechnological techniques work well in the lab, but all components required are prone to degradation. Replacing the test components regularly can be an excessive economic burden for small hospitals. Therefore, for a test to be accessible, it must be able to be stored for a long time in the fridge, or ideally at room temperature.
Last but certainly not least, the test should be as minimally invasive as possible. Medical professionals typically try to avoid invasive methods unless they are strictly necessary, but even when they are crucial to obtain a diagnosis, patients always have the right to refuse. A minimally invasive procedure, such as taking a blood sample, maximises the chance patients will accept the test, and improves patient comfort.
miRNAs as biomarkers for MS diagnosis
New diagnostic tools for MS are currently in development, and the identification of new biomarkers, small molecules, antibodies or proteins that can conclusively confirm the presence of the disease is ongoing.6 There are several molecular indicators of MS including neurofilament proteins, antibodies, inflammation and invasion of immune cell into the CNS.3 However, since all of these biomarkers also correlate with other diseases, a biomarker that is specific to MS has not yet been identified.
A new group of biomarkers that is gaining interest in the field of diagnostics are microRNAs (miRNA).8 These small RNA molecules, about 19 to 25 nucleotides in length, are always present in bodily fluids, including blood. They influence cellular processes by regulating gene expression. Based on the type of miRNA, it will target different genes and stimulate or inhibit basic cellular functions like inflammation and replication. This effect will vary depending on the concentration of the miRNA. Each miRNA is typically ubiquitous, playing a role in many parts of the body. As a result, the concentration of a single miRNA does not give a lot of information about an individual’s health. However, recent studies have shown that diseases can cause a specific pattern of up- or downregulation of miRNA levels (Figure 5).8 This means that by looking at the change in concentrations of several miRNA’s, you can correlate the pattern to a specific disease.
Besides the miRNA-based test that miRADAR is proposing, there are several other techniques to quantify the concentration of miRNA, such as RT-qPCR and microarrays. However, these have not been widely used in clinical settings due to the specific drawbacks of each technique.9,10
For RT-qPCR this is because of the lack of consistency. The different commercial RT-qPCR kits vary in effectiveness. Between studies this already causes discrepancies9 in final concentration based on the same sample. Notably, small changes in protocols between departments, and changes due to handling and storage have a significant impact on the results in clinical settings. Therefore, it is lacking in terms of accessibility.
Microarrays on the other hand lack specificity and sensitivity, requiring very high concentrations of miRNA with no way to differentiate between the healthy levels, and elevated concentrations. Other techniques suffer similarly from the problems mentioned above: excessive complexity and cost, low sensitivity or a lack of specificity.10 Those problems disqualify them for accessible large scale clinical testing of the dilute and complex miRNA samples from blood.
The benefit of a single standardised and integrated miRNA quantification and detection kit is based on reliability. When you perform the test, you know that due to standardised assembly of the test, it will function the exact same way each time. If you want to know more, click here to visit our design page where we delve into the specifics of the design of our test.
miRADAR
Based on these recent studies on miRNA patterns in MS, we see the opportunity to develop a tool for neurologists to improve the diagnosis for patients. When all modern techniques such as an MRI with McDonald criteria are available, neurologists can typically diagnose 70% of patients within four weeks. However, due to inconclusive MRI images 30% of patients must wait for over two months, and for 10% of these patients it can take years.
We target our diagnostic test at this latter patient group, as well as at those patients opposed against more invasive diagnostic methods. Overall, this project is focused on improving the diagnosis of complex cases of MS, based on the integrated detection of multiple miRNAs.
Our diagnostic tool would be an addition to the existing procedures to help neurologists expedite the diagnostic process, ensuring that patients can receive the appropriate medication in a timelier manner.