Recent research has focused on tackling the issue of the unmet medical need of an objective and quantitative biomarker for Huntington’s disease (HD). The discovery of biomarkers specific to the different stages of HD could help physicians monitor the physical changes of an individual’s HD during asymptomatic stages, and help improve the quality of treatment at all stages by allowing a better measurable assessment of an individual’s HD.
What are biomarkers?
Molecular, histologic, radiographic, or physiologic characteristics of a disease are all types of biomarkers. The FDA defines a biomarker as a “characteristic that is measured as an indicator of normal biological processes, pathogenic processes, or responses to an intervention”1. Biomarkers come in difference forms, and can serve various purposes in disease research. Clinically, for example, a biomarker can be used as a measurable indicator of the severity or presence of a disease state, or the effects of a treatment.
If you have ever gone to the doctor, it is likely that you have been asked to take an x-ray, or receive a blood or saliva test. Each one of these tests and x-rays is looking for a specific biomarker, an indicator of the state of the body that helps doctors predict an individual’s risk of a specific disease. For example, if you come into a doctor’s office with a sore throat, they might take a swab sample from your throat in order to test for the presence of Streptococcus pyogene, the bacteria that causes strep throat, in order to diagnose you with strep throat. In this scenario, the bacteria is the biomarker 2. In the future, you could be tested again for the presence of this biomarker to evaluate if the strep throat is gone, or is still present.
Scientists have taken a huge interest in biomarkers because of this powerful ability to give insight into the state of the human body. One such scientist is Dr. Joshua Chiappelli at the University of Maryland. In a research study, Dr. Chiappelli collected hundreds of saliva samples, and tested them in hopes of finding a biomarker specific to schizophrenia. Through his research, he found that elevated levels of a type of molecule called “Kynurenic acid” in saliva at any age could be an indicator of an individual’s risk to a specific type of schizophrenia 3 . Similar studies like this are currently underway in hopes of finding biomarkers specific to the progressive stages of HD.
Biomarker research and HD
The goal of many HD biomarker studies is to find ideal biomarkers that are closely linked and specific to the state of someone’s HD. Because of its progressive nature, there are no defined stages for HD; however, researchers and physicians often use this categorization of symptoms to better describe the progression of HD. For example, the period in which no motor signs are present is commonly referred to as the “premanifest phase”. This means that at this stage, an individual’s risks for the disease cannot be detected using existing neurological assessments. Currently, some of the clinical diagnosis and progress evaluations of HD rely on a neurological assessment of motor and cognitive ability, such as chorea or psychological symptoms, and blood tests for the presence of CAG repeats 4. Although these current methods open the possibility for preparedness and planning ahead with disease-modifying treatments, there is a pertinent need for more reliable biomarkers that measure “disease activity”, as well as biomarkers that help in evaluating the effectiveness of therapeutic interventions. Recent research studies have pursued this need through investigating the ongoing cellular processes of HD, with hopes of better understanding the state and conditions of the body at the microscopic level before and after the onset of HD symptoms. Ultimately, scientists hope to discover a “premanifest phase biomarker” which would ideally help in monitoring changes in asymptomatic individuals. Biomarkers such as this could also help in providing an objective measurement when assessing the progression and severity of a patient’s HD, which could contribute to the improvement and appropriateness of clinical care.
Biomarker research is extensive, and requires multiple steps after a biomarker is discovered to reach the point of clinical adoption and approved use in patients. Figure 1 displays the step by step phases of biomarker development. Thus far, many of the biomarker discoveries made for HD have been unable to surpass the second step (highlighted in red in Figure 1). This is due to variability between research studies that aim to replicate the results of the discovered biomarker, which cause conflicting results.
Figure 1 . Phases of biomarker development
“Wet Biomarker” research for HD
To address this issue, Professors Edina Silajdzi and Maria Bjorkqvist published an article in the Journal of Huntington’s Disease, in which they evaluate almost all studies done on “wet biomarkers” 5. They defined a wet biomarker as a potential biomarker that is objectively measured in a bodily fluid, such as blood, urine, saliva, or cerebrospinal fluid (CSF). In addition to reviewing wet biomarker research studies, the authors make detailed recommendations for improving biomarker research for HD. Specifically, they pinpoint biological variation, study design, patient selection, and even time at which an experimental sample was taken as potential considerations for improvement. The studies they reviewed, along with their findings, are described below.
Immune markers 6
The immune system has been previously suggested to play a role in HD progression 7. Specifically, a number of studies have highlighted the existence of an immune response in HD patients. In one study, scientists at King’s College London performed an extensive screen of HD patient plasma and identified elevated levels of a specific immune system protein in HD patients 8.
The authors hypothesised that a specific immune protein called “proinflammatory cytokine IL-6,” or IL-6, induces the release of proteins that increase or decrease in concentration in response to inflammation. These proteins further trigger a cascade by activating other immune proteins and factors responsible for cleaning up any dead cells in the body. A follow-up study demonstrated that IL-6 was elevated in HD subjects, even during the premanifest phase of HD. Thus, the study proposed the possibility of immune proteins, specifically inflammatory proteins such as IL-6, in plasma being potential biomarkers for HD progression tracking. However, no other study has been successful in replicating these results, which has prevented scientists from arriving at a consensus that this is a biomarker for HD
Another group of studies focused on investigating a liver protein called C-reactive protein (CRP), which normally rises when there is inflammation in your body, as a potential biomarker in HD. One study found decreased levels of CRP in one HD subject group 9, while several other studies reported increased CRP levels in HD subjects 10. The scientists of this study explained that one potential explanation for increased CRP levels is an inflammatory response in HD patients caused by mutant huntingtin expression. However, antipsychotics, a type of medication to control psychotic symptoms often taken by HD patients, were also described to instigate an inflammatory response; thus, this study expressed the possibility of this CRP increase being reflective of the antipsychotic use by the HD subjects they studied 11, further adding to discrepancies in CRP biomarker research.
Research shows that there is a “negative energy balance” in HD patients, which means that their energy expenditure typically exceeds their caloric intake 12. This metabolic factor contributes to the peripheral manifestations of HD, such as weight loss and muscle wasting. Because of HD’s impact on the body’s metabolism, many researchers have focused on testing “metabolites”— proteins or molecules involved in metabolism — as potential biomarkers for HD. One study analysed the serum, the fluid that remains when proteins involved in blood clotting are removed, from control subjects and premanifest and early stage HD subjects 13. The study found that changes in amino acid metabolism occur even before the onset of symptoms. Other studies that have focused on the levels of amino acids in HD patients consistently reported these changes to be correlated with weight loss, disease progression and abnormal triplet repeat expansion; however, conflicting results were received 5.
Further metabolic research has aimed at investigating different protein markers of metabolism in HD, such as those involved in cholesterol metabolism 14. Cholesterol is a waxy, fat-like substance that’s found in all the cells of the body, and is needed to make hormones, vitamin D, and important substances involved in food digestion 15. Although no significant changes were demonstrated in peripheral levels of total cholesterol, a specific brain cholesterol metabolite called “ – 24(S) hydroxycholesterol,”- or 24OHC, was consistently observed to be reduced in the plasma of individuals with HD 16. Reduced levels of cholesterol precursors (the inactive form of a protein that can be turned into an active form) called lanosterol and lathosterol and a bile acid precursor (a type of cholesterol involved in fat digestion) called “27-hydroxycholesterol” were also demonstrated 14. However, one of the suggestions made for further validation of this is to follow subjects longitudinally, or long-term, to examine the rate of change of metabolic markers as the disease progresses 5.
Novel biomarker discoveries
An Oculomotor Biomarker
Figure 2. Experiment outline
A 2020 study published in the Journal of Neurology took a different approach at identifying potential HD biomarkers 17. In hopes of addressing the lack of a quantitative and reliable way to clinically diagnose HD, a group of scientists focused on the potential for the altered eye movements in HD patients to be physical motor biomarkers of HD. Thus, they performed an eye-movement analysis to develop a novel eye-movement based model to diagnose and monitor the progression of (HD).
The study evaluated the eye movement in 25 participants, 10 of whom were early symptomatic for HD, 10 were in the premanifest phase for HD, and 5 were healthy controls. The research group developed machine learning algorithms for disease state classification to create an algorithm able to predict and differentiate HD patients from healthy individuals with high accuracy. These algorithms were able to differentiate HD patients from healthy individuals with high accuracy. Overall, the study demonstrated the feasibility of considering quantitative eye-movement assessments as objective biomarkers for HD that could serve as a support tool in medical decisions and therapeutic development.
Neurofilament Light protein as a Biomarker
In 2017, a study investigated whether neurofilament light protein NfL (also known as NF-L) in blood is a potential prognostic marker of neurodegeneration in patients with Huntington’s disease 18. NF-L is the smallest of three subunits that make up neurofilaments, which are major components of the neuronal cytoskeleton; NF-L is released from neurons when they become damages 19. In individuals with HD, four small-scale cross-sectional studies found raised concentrations of NF-L in their cerebrospinal fluid (CSF), the clear body fluid that surrounds the brain and spinal cord 20. In the 2017 study, plasma samples from 97 control patients and 201 individuals at risk for HD were analyzed. In 104 individuals with premanifest HD, NF-L concentration in their plasma was associated with subsequent clinical onset during a 3-year follow-up period. Thus, this study proposed that concentrations of NF-L in CSF and plasma were correlated in HD mutation carriers. NF- L in plasma thus shows promise as another potential prognostic blood biomarker of disease onset and progression in HD.
Although HD biofluid marker literature shows no clinically validated biomarkers for HD, much progress has been made in better understanding what improvements need to be made to promote wet biomarker discovery. Every research study reveals a lesson learned for the next, as they provide guidelines that benefit future biomarker studies in HD. The development of a reliable biomarker would provide a more objective and quantitative measuring tool for HD compared to clinically based methods currently used in HD trials. Additionally, further pursuing biomarker research will aid in the development of therapies for HD by providing a more accurate and specific means of evaluation. Taken together, this biomarker research analysis offers new potential avenues of biomarker research for HD, and serves as a useful tool source of insight for future biomarker researchers.
- FDA: About Biomarkers and Qualification
- AAMC. Strep Throat Test information
- Chiapelli, et.al. “Salivary kynurenic acid response to psychological stress: inverse relationship to cortical glutamate in schizophrenia” Neuropsychopharmacology. This article explains the research study done that found Kynurenic acid to be a biomarker in Schizophrenia patients
- Reilmann, et.al. “Diagnostic criteria for Huntington’s disease based on natural history” Journal of Movement Disorders. This article explains a history of diagnostic criteria for Huntington’s disease.
- Silajdziˇ c´, E. Bjorkqvist, M. “A Critical Evaluation of Wet Biomarkers for Huntington’s Disease: Current Status and Ways Forward”. Journal of Huntington’s Disease. This article describes a meta-analysis of biomarker research for HD up to 2018
- Ellrichmann, et.al. “The Role of the Immune System in Huntington’s Disease”. Journal of Immunology Research. This article gives an overview of the role of the immune system in HD
- Dalrymple, A. “Proteomic Profiling of Plasma in Huntington’s Disease Reveals Neuroinflammatory Activation and Biomarker Candidates”. Journal of proteome research. This article describes the research involved in studying proteins in neuroinflammation in HD as biomarkers
- Silajdžić, et. al. “F06 A critical evaluation of inflammatory markers in Huntington’s disease plasma”. Journal of Neurology, Neurosurgery, & Psychiatry. This article explains the study involved in F06
- Sánchez-López, F. “Oxidative stress and inflammation biomarkers in the blood of patients with Huntington’s disease”. Journal of Progress in Neurosurgery, Neurology and Neurosciences. Research study that analyzes oxidative stress and inflammation biomarkers in blood of patients with HD
- Bouwens, et.al. “Acute-phase proteins in relation to neuropsychiatric symptoms and use of psychotropic medication in Huntington׳s disease”. Journal of European Neuropsychopharmacology. Research study that provides evidence that HD mutation carriers who use antipsychotics are prone to develop an acute-phase response.
- Goodman, et.al. “The metabolic profile of early Huntington’s disease- a combined human and transgenic mouse study”. Journal of Experimental Neurology. This article explains research study focused on loss of weight loss in HD
- Underwood, et.al. “Huntington disease patients and transgenic mice have similar pro-catabolic serum metabolite profiles”. Journal of Neurology. Research article focused on serum metabolites
- MedlinePlus. Cholesterol.
- Leoni, et.al. “Plasma 24S-hydroxycholesterol and caudate MRI in pre-manifest and early Huntington’s disease”. The Journal of Neurology. Research study on the potential plasma biomarker 24OHC
- Frank, et.al. “Quantitative Oculomotor Biomarkers for Huntington’s Disease (1948)”. Journal of Neurology. Research article for study investigating eye movement as a potential biomarker
- Byrne, et.al. “Neurofilament light protein in blood as a potential biomarker of neurodegeneration in Huntington’s disease: a retrospective cohort analysis” (2017). The Lancet. Neurology. Research article for study investigating NF-L as a potential biomarker
- Wild, et.al. “Quantification of mutant huntingtin protein in cerebrospinal fluid from Huntington’s disease patients” (2015). The Journal of Clinical Investigation. Research article for study investigating NF-L in the CSF of HD patients