2715 – CK2 activity changes in the Okur-Chung Neurodevelopmental syndrome
Okur-Chung Neurodevelopmental Disorder Syndrome (OCNDS) is a newly described condition characterized by motor, brain (e.g. intellectual disability, epilepsy) and a range of multisystem symptoms. It is linked to newly occurring changes (mutations) in the gene that encodes for a ubiquitous, important protein kinase, termed CK2. Thus far, the patients are mainly children and adolescents, but as genome-wide sequencing becomes more available, OCNDS will be most probably be detected more often and extend to the adult population. To date, beyond the description of clinical cases, there is no knowledge about the impact of the different mutations on CK2, nor are there any other mechanistic studies published. Our project is aimed at unraveling how CK2 mutations affect its activity and physiological function, to link specific mutations to specific roles of CK2 in our bodies and ultimately to disease symptoms. This work is much needed since it will lay the basis for a better understanding of the condition and for the rational pursuit of future avenues of treatment.
2753 - Modern illness or a thing of the past? Surveillance study of childhood/adolescent Sydenham's chorea in the UK and the Republic of Ireland
Sydenham’s chorea (SC) is a disease affecting the brains of some children and young people. SC may follow after a throat infection with a type of bacteria (Streptococcus). Symptoms are abnormal body movements, which range from mild to severe and may disrupt a child’s ability to carry out activities of daily living like writing and walking. SC is currently considered a ‘rare disease’; but little is known about how many children nowadays are affected, what happens to them after diagnosis, or about their needs. We plan to study children with SC aged between 0 and 16 years in the United Kingdom. We will work with the British Paediatric Surveillance Unit and ask paediatric doctors to report when they first see a child with symptoms of SC. We will then send a follow-up questionnaire to the doctor asking about symptoms, treatment, recovery and the impact on the child’s daily life, for example, educational difficulties occurring as a result of SC. Our study will have a direct impact on patient care. It will raise awareness and contribute to improved training for health professionals, which we hope will lead to quicker diagnosis for children with SC and similar rare conditions. By finding out how many children are currently affected and about their needs, the study will help in planning services. Working with the Sydenham’s Chorea Association, a family charity, we will also use the findings to produce better information for children and families about what to expect from a diagnosis of SC, for example, by developing an animated video.
2764 - An unusual SMARCB1 gene variant causing schwannomatosis disease
A rare disorder, called schwannomatosis, causes people to develop tumours on the nerves anywhere in their body. These tumours can cause severe pain. There are two genes, called SMARCB1 and LZTR1, which, when damaged, are known to cause schwannomatosis. The damaged version of either of these genes can be passed on through families. Damage to one
of these two genes is known to be disease-causing in about half of people with schwannomatosis. In the other half, the cause is unknown. We are investigating a large family with schwannomatosis, in which no change was found in SMARCB1 or LZTR1 in the clinic. Recently, through our research, we have found an unusual type of change in SMARCB1 in this family. This type of change is not detected by normal clinical tests, but we think it is causing their disease. The current study will help us to confirm the link between the novel gene change and schwannomatosis disease. Confirmation of this link would allow specific testing of other family members who may be at risk of having the same disease. It will also help doctors and genetic counsellors to provide the most appropriate clinical care and inform decision making such as prenatal diagnosis for family planning. This type of gene change has not been seen before in schwannomatosis.
2771 - Exploring biomarkers of inflammation in CF patients with eosinophilia
Cystic fibrosis(CF), is a genetic disease with persistent infection and inflammation seen in the lungs. CF patients suffer from pulmonary exacerbations(PEx) which consist of increased symptoms and worsening lung disease. These episodes are usually treated with antibiotics that target bacteria colonizing the lungs, however, up to a quarter of patients experience permanent loss of lung function even after treatment which suggests the current course of treatment for PEx is not ideal for everyone. We hypothesize that this may be due in part to eosinophils which are a specific type of white blood cell that are present during TH2, or “allergic” inflammation. Because recently we found that almost 50% of our hospitalized CF patients had an elevated number of eosinophils (eosinophilia) in the blood, it is important to characterize what is driving this response as eosinophils are known to cause airway destruction. When eosinophils are activated by our immune systems, they release toxic products that can damage lung cells and contribute to the inflammatory response already present in CF due to the primary lung infection. Additionally, previous studies have shown that antibiotics do not decrease markers of eosinophilic inflammation therefore this is going untreated in CF patients with eosinophilia and may help explain why we found our patients had longer hospital stays and remained symptomatic longer. By measuring eosinophilic/TH2 inflammatory markers on admission and discharge, we can compare how those with eosinophilia differ from non-eosinophilic patients and potentially find new biomarkers that more accurately subtype patients and predict treatment response. We believe this would allow CF doctors to offer more tailored therapy (corticosteroids or anti-inflammatory agents) at the onset of symptoms, potentially avoiding unnecessary long courses of harsh antibiotics as well as improve quality of life by salvaging lung function and shortening hospital stays.
2774 - Determining the cause of cell loss in inherited blinding diseases
Inherited conditions that cause death of the light-sensitive cells within the eye, photoreceptors, are rare, irreversible causes of vision loss and blindness. Cumulatively, inherited photoreceptor diseases affect ~1/3000 individuals. Blindness has a dramatic effect on quality of life, but currently there are few effective treatments to combat photoreceptor-associated blinding diseases. This project focuses on a gene that has genetic variants known to cause two different, rare photoreceptor diseases. These diseases are retinitis pigmentosa (RP), a condition wherein rods (dim-light photoreceptors) degenerate, leading to night blindness and peripheral vision loss, and occult macular dystrophy (OMD), a condition involving cone (daytime/colour photoreceptors) loss, resulting in reduced visual acuity and central vision loss. How and why variants in one gene can lead to two differing diseases that affect different photoreceptor types is unclear, which makes the development of therapies to combat vision loss in these conditions very difficult. This work will investigate how mutations in this gene cause disease in order to determine why rods and cones become diseased differentially, and how photoreceptor diseases can be prevented/treated. We will accomplish this by creating animal and cell models of RP and OMD. Zebrafish are ideal for studying photoreceptors, as they have a diversity of photoreceptors similar to humans. Disease-causing variants in this gene will be introduced into zebrafish and how the diseases progress will be characterized. We have already introduced RP-causing mutations in the zebrafish. Similarly, disease-causing variants will be introduced into cultured human cells to observe how the changes impact cell function. These models will allow for characterization of RP and OMD progression, including early disease indicators, and elucidation of disease mechanism(s), which will allow for development of targeted therapies to prevent vision loss.
2779 - Rapid identification of children with rare immune-mediated inflammatory disorders that could benefit from anti-interferon treatment
Inflammation is a natural part of the immune response to infection or injury and under normal circumstances, should resolve naturally. There is mounting evidence that persistent (chronic) inflammation may cause and/or advance many of the most common human diseases. The most common anti-inflammatory treatments suppress the entire immune system and make patients prone to infection and other complications. Scientists now know that inflammation is not the same in every person or in every situation and there are dozens of molecules that can turn the process on and off. This opens up new possibilities for treatments, called “biologics” that target single molecules that are responsible for different types of inflammation. Due to the specificity, these drugs work better and have reduced side effects compared to drugs that suppress the entire immune system. ‘Type 1 interferons (IFN)’ drive a particular type of inflammation that is associated with lupus, type I diabetes, and a host of rare diseases in children. Several “anti-IFN” treatments are already available or are in the final stage of clinical trials. Doctors at our hospital have access to some of these treatments and already used them to successfully treat a family with a rare immune disorder; prior to this, the family had many visits to different specialists and months of rigorous and expensive workup to identify the problem. We anticipate as many as 100 more children at BCCH followed in rheumatology, neurology and infectious diseases may have a disease that could benefit from this type of therapy. The aim of this study is to put in place a quick (<3 days turnaround time) and relatively inexpensive (<$45 per test) test to identify children with persistent type I IFN inflammation that may benefit from targeted therapy.
2781 - Promoting nutrient absorption in children with microvillus inclusion disease
Microvillus inclusion disease is a very rare and severe genetic bowel disorder that affects infants and young children. Microvillus inclusion disease is caused by mutations in the MYO5B gene. Degeneration of their small bowel causes the patients’ inability to absorb nutrients from the diet and makes them life-long dependent on intravenous feeding. Unfortunately, most patients also die from the severe complications that accompany long-term intravenous feeding. Recently, we identified a cellular defect in MVID that may be accountable for the degeneration of the small bowel and, importantly, can be treated with FDA-approved medicines. In this study, we will employ a mouse model of MVID to investigate the potential of FDA-approved medicines to remedy this cellular defect, improve nutrient absorption and decrease the need for intravenous feeding, and in this way increase survival chances. The results of this study have potential impact for other rare diseases associated with mutations in the MYO5B gene and for other rare malabsorption diseases associated with the degeneration of the small bowel. Further, the results of this study will improve our understanding of the biology of the small bowel and nutrient absorption in health and disease.
2782 - Molecular chaperones to restore the function of mutated myosin Vb in microvillus inclusion disease
Microvillus inclusion disease is a very rare and severe inherited bowel disorder that affects infants and young children. There is no cure and most patients die during infancy. Microvillus inclusion disease is caused by mutations in a gene called MYO5B. The MYO5B gene contains the information for the production of a protein called myosin Vb. The myosin Vb protein needs to be folded in order to work properly. As a result of the mutations in the gene, the myosin Vb protein cannot be folded correctly and this results in its loss of function and onset of the disease. The normal folding of proteins in cells is facilitated by so-called chaperones. The stimulation of the activity of these chaperones have been shown to correct other mutant proteins that are misfolded and give rise to other rare diseases. However, whether this also can work for misfolded myosin proteins is not known. In this study we will investigate whether stimulation of the activity of these chaperones can also improve the folding of the mutant myosin Vb protein or even correct misfolded myosin Vb proteins. If successful, this study opens the door to a novel treatment strategy that targets the root of this devastating disease. Further, if successful this study may have therapeutic potential for other rare diseases that are caused by the misfolding of other myosin proteins.
2784 - Identifying Novel Targets for Antisense Oligonucleotide Therapy to Treat Dysferlinopathy
Dysferlinopathies are rare muscular dystrophies caused by mutations in the dysferlin gene, and include limb-girdle muscular dystrophy IIB (LGMDIIB), Miyoshi myopathy, and distal myopathy with anterior tibial onset (DMAT)(1). The prevalence is approximately 1/100,000-200,000(2), depending on the populations. Currently, no specific treatment is available for dysferlinopathy. Dysferlin (DYSF) is required for efficient cell membrane repair (1). We aim to develop a new therapy for dysferlinopathy using exon skipping strategy. Exon skipping typically uses an artificial DNA-like molecule (antisense oligonucleotide) that targets one of the potential target sites on an exon, a part of DNA that carries the information necessary to produce a protein. Because several shorter forms of DYSF are associated with milder symptoms and can ameliorate the symptoms of the model mice (3, 4), we will rescue dysferlinopathy by removing the mutated part and creating shorter but functional forms of DYSF using antisense oligonucleotides. We demonstrated that multi-exon skipping of DYSF exons 28-29 rescues plasma membrane resealing in dysferlinopathy patient cells (funded by Rare disease foundation last year) (5).
To explore other target sites for exon skipping therapy, we will make three shorter forms of DYSF (micro-Dysferlins) and test the function in dysferlinopathy patient cells. To evaluate the membrane repair ability, the cells introduced with the micro-Dysferlins will be damaged by a laser, and the repair process will be monitored by microscopy in real-time. We will identify novel exon skipping targets in DYSF by characterizing the relationship between exon deletion pattern and plasma membrane resealing ability. Successful completion of this study will identify the parts required for the membrane repair function of DYSF, and provide a proof of principle for applying exon-skipping therapy for dysferlinopathy.
2787 - Use of Bortezomib (Velcade®) to Enhance Radiation Sensitivity in Ewing’s Sarcoma
Ewing’s sarcoma (EWS) is an aggressive bone tumour affecting accounting for 2% of childhood cancers. It develops in the long bones, ribs, pelvis and spine, but can also appear in soft tissues surrounding the bone. The therapeutic approach to these patients involves surgery, chemotherapy and radiation therapy to target the tumour. With this current treatment approach only 60% of patient survive longer than 5 years. High dose Radiation therapy, although effective causes damage to the growing skeleton and reduces bone density, leaving patients vulnerable to fracture. Bortezomib (Velcade®), a chemotherapy that is already approved for use in multiple myeloma, has been shown to increase the effectiveness of radiation therapy in several laboratory models of human cancer. We have shown that Velcade is effective in killing EWS cells and reduces the radiation therapy dose required to be effective. We are now validating these effects in a clinically relevant animal model of EWS, which, if successful we plan to rapidly translate into human clinical trials. We anticipate that the use of bortezomib will represent a safe and highly effective way of maximising tumour destruction while minimising the potential detrimental effects of radiation on the growing skeleton of patients with Ewing’s sarcoma.
2788 - Limb-Girdle Muscular Dystrophy Type 2b as a new type of primary genetic dyslipidemia
Using patient and animal models our lab has demonstrated that plasma lipids, particularly cholesterol, may play a critical role in the progression of muscular dystrophy (MD). There are currently no therapeutic options available for sufferers of a rare, non-lethal form of MD known as Limb-girdle MD type 2b. This pilot study aims to assess the degree to which plasma lipids contribute to muscle wasting in this unique population, in order to determine whether widely available medications used to treat dyslipidemia (or elevated plasma cholesterol) can be repurposed as an effective pharmacotherapy.
2791 - Simplified Home and Ambulatory Monitoring in Patients Evaluated for Inherited Arrhythmia (SHAPE-IA)
Inherited heart rhythm disorders (IHRD) are rare conditions that are often familial and can present with devastating complications. For example, various IHRDs including Long QT syndrome, Brugada syndrome, Catecholaminergic Polymorphic Ventricular Tachycardia, and Arrhythmogenic Right Ventricular Cardiomyopathy have a lifetime prevalence between 1 in 2000 to 1 in 5000 individuals. However, a patient with an IHRD can present suddenly with fainting or sudden death of the affected individual.
The Inherited Arrrhythmia Program routinely evaluates patients for IHRD. Patients will undertake a series of tests ranging from 24-hour heart rate monitoring (“Holter monitor”) to advanced cardiac tests including ultrasound and MRI scans. Performing all these investigations can be challenging, particularly for patients living in remote areas with limited health care access.
Cardiac patch monitors are a new form of monitoring technology that is being increasingly used in the detection of arrhythmias, including atrial fibrillation. Compared to conventional monitoring technology, cardiac patch monitors provide the opportunity for patients to initiate their own monitoring without needing to go to the local hospital or travel to the tertiary center. Patch monitors have all the capabilities of conventional “Holter monitoring”, but have the added benefit of longer monitoring and convenience (i.e. less bulky, no wires, and waterproof).
2794 - Determining the functional impact of missense variants in RYR1 gene
Next-generation DNA-sequencing has largely contributed to gene discoveries in rare diseases. However, many rare diseases are still without a molecular cause due to heterogeneity, limitations of the approach and the difficulty in variants interpretation. In the case of myopathies, it is estimated that at least 25% still lack a molecular diagnosis after exhaustive clinical and genetic evaluation. More specifically, muscle disorders caused by mutations in the gene RYR1 are highly heterogeneous contributing the challenge of identifying disease causing variants. In this context, functional validation of variants of unknown significance is necessary to confirm the pathogenic status of a variant. Our project will have a direct impact on patients affected with RYR1 disorders and their families. Obtaining a molecular diagnosis will allow a proper genetic counseling, better risk assessment and clinical management as well as reducing the psychological hardship associated with the long diagnostic odyssey. More importantly, RYR1 mutations can be associated with malignant hyperthermia, which consist in an adverse reaction to an anesthetic medication. Most individuals are only aware of this disease once exposed to the medication. The confirmation of the pathogenic status of RYR1 variants will contribute to the clinical management of these individuals. Finally, our project will contribute towards a better understanding of biological processes underlying these RYR1-related disorders.
2796 - McMaster Pediatric ITP Registry
Immune thrombocytopenia (ITP) is a rare bleeding disorder characterized by the loss of platelets, which are cells that help prevent bleeding and bruising. When the platelet count is low, patients will experience increased bleeding in the form of nosebleeds, bruises, or even serious brain bleeds. The risk of a life-threatening bleeding event in pediatric patients can be devastating and causes extreme anxiety for patients and their families. The cause of childhood ITP is largely understudied, which is problematic for doctors trying to treat these patients. It is known that some adult patients with ITP develop a factor (antibody) that attacks platelets, and we are going to explore this idea in a pediatric population. We will collect blood from patients under 18 years of age who have ITP to study the cells that make antibodies, and the nature of the antibodies produced. In our study, we will develop a database of pediatric ITP patients so we can better track common patterns of disease. We will investigate whether anti-platelet antibodies are present and determine the sites that these antibodies bind to. In addition, we will study the patients’ platelets. This will help explain what is happening in childhood ITP, which will lead to more effective patient management moving forward. Overall, our study will help doctors, patients, and their families better understand an understudied, rare, pediatric bleeding disease.
2803 - Evaluation of the molecular effects of two novel variants of the CYP26B1 gene found in siblings with skeletogenesis defects
In this proposal, we aim to investigate the mechanism of action of two variants of the CYP26B1 gene found in siblings with defects in their skeletal development and learning disabilities. Through Whole Exome Sequencing, we recently found both siblings harbour two rare variants in the CYP26B1 gene. There are only a few reports of patients harbouring CYP26B1 variants, but they have similar physical findings as our patients. The CYP26B1 gene has the instructions to make cytochrome P450RAI-2, an enzyme responsible for retinoic acid (RA) breakdown. The control of RA concentration is critical during early embryo development for the regulation of many genes driving growth. Breakdown of RA concentration during this period is regulated by the P450RAI-2 enzyme. Although we believe the CYP26B1 variants in our patients impairs the normal function of P450RAI-2, it does not disturb the enzyme’s active site, and thus should not affect RA concentration. We hypothesize that the mechanism by which these two variants cause dysregulation of RA levels is through disruption of the interactions of this enzyme with other proteins or regulatory factors. We aim to prove our hypothesis by first measuring and confirming abnormal RA levels in our patients, and then identifying possible changes in interactions of the enzyme with other proteins using a technology called BioID. This knowledge will enable us to re-classify the variants as “disease causing” and allow us to effectively provide accurate genetic counselling for this family. Furthermore, our data may also reveal new biological information that is important in the understanding of the rare condition caused by CYP26B1 genetic variants.
2807 - Development of an oligodendrocyte-specific genome editing strategy for the treatment of Pelizaeus-Merzbacher Disease
Pelizaeus-Merzbacher Disease (PMD) is a rare pediatric disorder that affects around 1:100,000 boys at birth. The disease is caused by mutations of the PLP1 gene, encoding Proteolipid Protein 1, a key structural protein of the myelin, which is an insulating layer that wraps around nerve fibers to facilitate the flow of information in the nervous system. PMD belongs to a group of disorders called leukodystrophies, characterized by defective myelination of the central nervous system (CNS). PLP1 is mainly expressed in oligodendrocytes that are the cells responsible for myelination in the CNS. Thus, mutations in PLP1 result in abnormal myelination, which ultimately leads to severe locomotor and cognitive impairment, and limited life expectancy. Only symptom management, but no curative treatment options are available for PMD and the other leukodystrophies.
This project aims at establishing a novel delivery system to implement CRISPR/Cas9 genome editing therapies for individuals affected by PMD. Our goal is to develop a methodology that will allow robust and efficient targeting of oligodendrocyte cells, which are the main cell type affected in the disease. To achieve this goal, we will test and identify the best promoter and clinically relevant viral vector to enhance CRISPR/Cas9 expression in human oligodendrocyte-like cell lines.
Successful completion of this project will support the development and translation of novel genome editing strategies for individual affected by PMD. In addition, it will have a significant impact on the advancement of new therapeutic options for the broad group of oligodendrocytes-related genetic disorders, such as the leukodystrophies.
2808 - Characterizing a patient mutation in a novel gene underlying Joubert Syndrome
Joubert Syndrome (JS) is a genetically diverse, recessive disorder characterized by defective brain development that affects approximately 1 in 100000 live births (1). In addition to the neurological symptoms, patients with JS may suffer from an array of symptoms in other organs such as kidney dysfunction, liver fibrosis and retinal defects (1). Presently there is no cure for JS and, while symptom management strategies exist, it remains difficult to implement uniform standard care as disease symptoms and severity vary among patients. Despite having greater than 30 associated genes identified thus far, the exact genetic basis of disease for many JS patients remain undetermined (2).
A novel de novo heterozygous mutation in Tubulin Beta Class 1 (TUBB) has been identified in a JS patient with unknown genetic etiology that results in an amino-acid change in the protein sequence. This candidate disease-causing mutation represents a novel gene (TUBB) and inheritance pattern (autosomal dominant) for JS.
Using patient-derived fibroblasts and other established human cell lines I will assess the pathogenicity of the TUBB mutation. Through biomolecular techniques I will elucidate the mechanism by which the mutated TUBB protein causes or contributes to disease, shedding light on the biological processes underlying JS.
This study will give JS patients without a genetic diagnosis one more opportunity to receive one. Positive results will support the addition of TUBB to the panel of genes to test in JS cases going forward. Also, showing that JS can be inherited as a dominant condition will change the criteria used by bioinformaticians when they analyze patients’ genomes searching for mutations, enabling them to pick up variants that have been so far disregarded. Moreover, understanding the pathogenic mechanism of the mutated TUBB protein will reveal points of intervention relevant to the restoration of proper neurological functions and provide possible therapeutic avenues for JS.
2809 - Body composition of normally growing children with intestinal failure: a pilot study
Children with intestinal failure (IF) are unable to absorb enough nutrients and fluids through their intestine to grow properly. They need to be fed by vein with parenteral nutrition. To know if the nutrition provided is appropriate, height and weights are monitored on a regular basis and plotted on growth charts. However, heights and weights cannot tell if children are gaining enough muscle or too much fat. This can only be assessed by measuring their body composition. Body composition measurements tell us the amount of fat and lean mass in the body. Measuring body composition is important because too much fat is linked to poor health in children and adults. Increased body fat in children can lead to diabetes, hypertension and heart disease in adults. And low lean mass can lead to osteoporosis. Yet, body composition is not usually measured in children in the clinical setting. Recent studies showed that children with IF have lower lean body mass (LBM) and higher fat mass than healthy children. This information shows that it is important to measure body composition of children with IF so that their nutrition can be adjusted to achieve a better outcome. Bioelectrical impedance analysis (BIA) is an easy, noninvasive method to measure body composition in children. It is safer than dual X-ray absorptiometry (DXA) because the subjects are not exposed to radiation as when DXA is done. It can therefore be used as a routine test in the clinical setting. The goal of this pilot study is to measure lean and fat mass in children with IF at our hospital using BIA and to compare it to children with same age, gender, and race. The results of this study will provide information to help design a future study of body composition in children with IF and better care for children with IF. It will allow clinicians to adjust the nutrition being provided to achieve a better lean and fat mass instead of only relying on weight and height.
2810 - Facilitating the introduction of genomics medicine and definitive diagnoses to African children and adolescents with undiagnosed neuromuscular diseases
Inherited neuromuscular disease (NMD) affects many children and adolescents globally. These include hereditary diseases of the nerves (neuropathies), muscles (myopathies / dystrophies) and motor control units (motor neurone disease). In South Africa we have genetic tests to diagnose the commonest NMD conditions viz. Duchenne’s and Becker’s muscle dystrophy, myotonic dystrophy, RYR1 related centronuclear myopathy, Charcot-Marie-Tooth-1 and SMN1-spinal muscular atrophy. However, individuals with less common/rare conditions, are unable to access definitive molecular diagnoses which impacts on management of their condition and on accessing new therapies as they become available for clinical trials.
Our NMD team was recently awarded a UK-MRC strategic award to establish an International Centre for Genomic Research in Neuromuscular Diseases. The investigators applying for this microgrant, are all key collaborators in the project at three South African sites. The UK-MRC award, which starts in mid-2019, will focus on the genetic analysis of undiagnosed patients in a tiered approach, as well as supporting the training of two local NMD neurology fellows. The first-tier diagnostic approach will use gene panels of genes variants known to cause NMD in studied populations viz. Europeans and Asians. As our African populations are unstudied, we anticipate that many/most of our patients will remain without a diagnosis and therefore will move forward to next generation sequencing to discover pathogenic genes. This strategy has the potential to identify new NMD risk genes which may be prevalent and unique in Africans. The program insists on global data sharing and the discovery of new treatment targets to the benefit of all is a prominent goal of the project. Although the grant will cover the sequencing and analysis pipeline, we require funds to develop a long-term DNA extraction and storage base