May 2016 Funded Microgrants

May 2016 Funded Microgrants

A prospective, randomized, double-blinded, cross-over trial of deep brain stimulation for essential voice tremor: A combined surgical and imaging study

Essential Voice Tremor (EVT) is a rare disease with an estimated prevalence of 3 in 100,000. Patients with EVT lose their ability to speak due to involuntary rhythmic contractions in the vocal musculature. Current treatments such as medications, Botox, and voice therapy only provide temporary relief and thus, we are exploring new strategies to provide EVT patients with long-term improvement. Deep Brain Stimulation (DBS) is a neurosurgical procedure that involves the implantation of electrodes to deliver electrical stimuli to specific brain regions. It is the gold-standard treatment for limb tremor and can eliminate tremor instantaneously with a resultant improvement in quality of life. For the first time ever, we wish to study a pure cohort of six voice tremor patients who will undergo DBS surgery and functional brain imaging. Our team has anecdotal evidence that the DBS therapy used for combined limb and voice tremor also had an unexpected benefit in vocal dysfunction. This pilot trial has been designed to test the hypothesis that DBS can improve the vocal dysfunction in EVT.

Results - We found that the motor circuit controlled by the cerebellum (not basal ganglia) is preferentially involved in modulating speech motor control. We also found that that this effect, like language, is a subcortically lateralised to the dominant thalamus despite the vocal cords being bilaterally innervated. Along with our publications, these findings were presented at several prestigious international neurosurgery conferences in which the Rare Disease Foundation was acknowledged.

Thanks to your initial support, we have secured nearly $100k of peer-reviewed funding for a Phase 1 surgical trial of international significance which will undoubtedly bring UBC recognition but most importantly, improve the lives of our medically-refractory patients. 

Avecillas-Chasin JM, Poologaindran A, Morrison MD, Rammage LA, Honey CR. Unilateral Thalamic Deep Brain Stimulation for Voice Tremor. Stereotact Funct Neurosurg. 2019 Jan 9:1-8.

Use of bedside home videos to impact patient and provider satisfaction with the inpatient care of children with medical complexity

Children with medical complexity (CMC) have multiple chronic diseases, high health care needs and functional impairments. Many of them have significant neurodevelopmental differences and rare or undiagnosed diseases. As a result, care from unfamiliar providers can be very challenging, particularly during acute illnesses in the inpatient setting. To improve the inpatient experience for CMC, their families and providers there is a need to develop and evaluate clinical interventions that could minimize communication barriers, such as information sharing and a lack of prior knowledge of the child’s condition. We propose to use bedside home videos to provide families with an opportunity to share information about their child that they feel is important for unfamiliar care providers to know. We hypothesize that this may give families more confidence in their child’s inpatient team and also may give unfamiliar providers important information regarding the child’s typical status.

Translation read-through-inducing drugs as a potential cure for children with microvillus inclusion disease

Microvillus inclusion disease is a very rare and severe genetic bowel disorder that affects infants and young children. Patients suffer from unstoppable diarrhea, do not tolerate normal food,  and require life-long intravenous feeding and bowel transplantation as a last resort for survival. Most patients die at young age and a cure is desperately needed. We will determine the potential of novel compounds that can suppress specific gene mutations to better treat this disease and make a difference for these patients.

Interrogation of the CRISPR/Cas9 system for duplication removal in DMD

Incorrect instructions can lead to a wrong message. For example, a recipe that reads: “Add 1 diced avocado and 1 teaspoon of salt and 1 teaspoon of salt.” will give us guacamole that is too salty. This situation is analogous to Duchenne Muscular Dystrophy (DMD) patients with duplication in dystrophin gene, which accounts for about 10% of the patient population. The CRISPR/Cas9 gene editing technology can virtually edit any location in the gene using two components, namely guide RNA and Cas9 protein, which resemble a computer mouse and “delete” key in a keyboard, respectively. The guide RNA will target the duplicated part of the sentence, e.g. “and 1 teaspoon of salt”, which will subsequently be deleted by the Cas9, resulting in a correct recipe to follow. We have previously shown that using this novel gene editing technology we are able to precisely cut out the duplication in the cells of a DMD patient and restore the missing dystrophin protein. We next plan to use a second generation version of the CRISPR/technology that is more clinically relevant and test to see if we can do the same for a wider range of DMD patients with duplications of various sizes. Successful completion of this project will open up entirely new treatment strategies for individuals affected by duplications in the DMD gene.

Growth failure and craniosynostosis potentially caused by activating mutations in EZH2

In 2011, my lab discovered that mutations in EZH2 cause Weaver syndrome, a rare disease that causes wholebody overgrowth and large head size.1 In early 2015, we also found that mutations in EED cause a very similar overgrowth syndrome, which we have tentatively named Cohen-Gibson syndrome. Interestingly, the EED protein pairs up directly with EZH2 and is necessary for EZH2 to do its work. This explains why the two syndromes have such similar features – the proteins work together to do the same thing as part of a “protein complex” called PRC2. In both Weaver syndrome and Cohen-Gibson syndrome, the mutations are brand new mutations that are not usually inherited from either parent – they are “genetic accidents” that occur in the sperm or egg just before or just after conception. Earlier this year, our work showed that mutations that cause Weaver syndrome reduce the activity of the EZH2-EED enzyme complex.4 Because of our expertise in this area, diagnostic labs across the world have started to ask us about mutations they find in these genes in their patients. One lab in Bergamo, Italy asked us to look at a brand new EZH2 mutation they had identified in a 3½ year-old girl who has growth failure and a small head. Because her medical features more or less “opposite” to Weaver syndrome (which features overgrowth and large heads), we believe that her mutation should have the opposite effects on EZH2 – making the enzyme hyperactive rather than less active. If we can show that her EZH2 enzyme is hyperactive, she would be eligible for treatment with EZH2 inhibitors (drugs that reduce the activity of the EZH2 enzyme). We can’t use these drugs in Weaver syndrome, because we think they would make these patients worse by reducing their EZH2 activity even more. However, these drugs might make this little girl with the “anti-Weaver” syndrome better. We are asking for this microgrant to fund enzyme activity studies to look at this extremely rare EZH2 mutation that seems to be causing a unique growth failure syndrome. No other patients in the world have yet been reported with this syndrome.

Results - Using a similar in vitro assay, mutant EZH2 (p.Ala738Thr or A738T) pre-assembled into PRC2 showed increased EZH2-mediated H3K27 methylation activity.These results support the hypothesis that gain-of-function mutations in EZH2 cause growth failure, in a reciprocal manner to loss-of-function mutations in EZH2 causing WS. Impact: These data support consideration of an N-of-1 clinical trial of low-dose EZH2 inhibitors to help stimulate this patient’s physical growth.

Primary Immunodeficiencies in Critically Ill Children

The risk of a severe infection that leads to near-death for otherwise healthy children is very small. Other scientists have shown a strong link between this risk and problems with genes related to how the immune system works. We are hoping to investigate this further by looking closely at the immune system and genes in the small number of children who suffer from severe infections, and seeing if there are any problems. This will help us better understand the rare child who gets sick, why they get sick, and how to prevent them from getting sick in the future. 

Use of bortezomib (Velcade®) as a radiosensitizer in Ewing’s sarcoma

Ewing’s sarcoma (EWS) is an aggressive bone tumour affecting approximately 1 case per one million people. It most often develops in the long bones, ribs, pelvis and spine, but can also appear in soft tissues surrounding the bone. Treatment for EWS involves shrinking the tumour with chemotherapy, then removing it with surgery or killing it with radiation therapy. Although 5-year survival rates with these approaches are 60-75%, they have not changed much in the last 20 years. Radiation therapy is effective as a sole treatment or in combination with surgery, but it also damages the growing skeleton; around 50% of EWS patients are children or adolescents, limiting the applicability of radiation therapy in these patients. 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, and we now propose to test whether bortezomib can selectively increase the susceptibility of EWS to radiation therapy. Positive results from this short study will support a grant application for testing in an animal model of EWS, followed by rapid translation in human clinical trials. We anticipate that the use of bortezomib will represent a safe and highly effective way of increasing the selective killing of EWS cells, maximising tumour destruction while minimising the potential detrimental effects of radiation on the growing skeleton of patients with Ewing’s sarcoma.

Results - Primary results from this study show bortezomib is an effective cytotoxic agent against a panel of Ewing’s sarcoma cell lines. We have demonstrated that bortezomib is also effective in reducing the number of viable Ewing’s sarcoma cells over time in an in vitro system (Figure 1). We have demonstrated that Ewing’s sarcoma cell lines, derived from patients, are sensitive to bortezomib, with reductions in overall cell number and in colony formation (Figure 2a,2b), suggesting that the drug would likely inhibit growth of the tumour in vivo. This result, although preliminary in nature, is the first clear evidence that a proteasome inhibitor may have potential efficacy against Ewing’s sarcoma. We have also shown that a combination of bortezomib plus irradiation has a synergistic cytocidal effect on Ewing’s Sarcoma cell lines (Figure 4). Exposure to a relatively low dose of bortezomib increases tumour kill more than either bortezomib alone or radiation alone. These results support our research hypothesis, but additional preclinical work needs to be undertaken to determine whether the drug also functions as a radiosensitizer in vivo. To this end, we plan to submit a request to the Rare Disease Trust later this year to fund a mouse study to address this question. Positive results from the mouse study would provide strong justification for clinical trials in humans, and these could be initiated relatively quickly since bortezomib has already been approved for use in human cancer.

Ongoing Work : The results are pilot studies to be repeated on a bigger scale to confirm reproducibility and clinical relevance. As a result of the trends we have seen in in vitro, we plan to submit a second proposal to the Rare Disease Foundation to extend this model into an in vivo system. This will enable us to model the effects of bortezomib as a drug and the effects of combined drug-radiation therapy in a more biologically relevant model as the one we have published6

In addition, we have applied for funding from the CCLGC to utilise frozen Ewing’s Sarcoma patient samples in order to expend the in vitro model to a larger subset of heterogenic patient tissue. Taken as a whole, these ongoing projects will allow our laboratory to develop a broad set of data to support a larger programmatic grant application on paediatric cancers to the MRC or Wellcome Trust.

Comprehensively preventing and treating bone fragility in patients with cystic fibrosis

Cystic fibrosis (CF) is a life-limiting genetic disorder that affects 1 in 10,000 people world-wide (more common in Europeans). One of the well-known features of CF is unusually thick mucus secretions that plug airways causing tissue damage and destruction due to inflammation and infection. This problem with abnormal secretions affect other organ systems as well – including the pancreas (that makes enzymes that help digest food), which contributes to malnutrition, as well as the reproductive organs (especially of men) resulting in infertility. Advances in treatment over the last two decades have greatly improved life expectancy of CF patients, but this has uncovered the fact that patients with CF have a very high risk of developing bone fragility (osteoporosis). There is an important urgent need for better and safer treatment options for bone fragility in CF patients as current anti-osteoporosis medications may do more harm than good, particularly in CF. Our plan is to prevent and treat bone loss in CF patients by both ensuring key bone-building nutrients are properly absorbed while at the same time strategically limiting inflammation that promotes bone destruction. With this microgrant, we aim to show this holistic approach has the potential to significantly improve the bone health and overall well-being of patients with CF without resorting to the use of potentially harmful bone drugs that may further negatively impact their immune systems.

Results - CF-related bone disease is an issue that grows as CF patient care improves. The goal of this study was to find the relationships between clinical factors, blood markers, and bone mineral density (BMD) and to identify predictors for bone loss. This study was conducted at the St. Paul’s Hospital Adult CF Clinic (Vancouver, Canada) and looked at 56 participants with at least two bone scans (DXA scans) separated by at least three years, and a blood sample collected within 12 months of the first bone scan. The blood markers that were looked at included osteoprotegerin (OPG), interleukin (IL)-1ß, IL-6, IL-8, and tumor necrosis factor (TNF)-alpha. The clinical factors, including age, lung function (FEV1% predicted), and CF-related diabetes, were collected within 6 months of the first bone scan. After the analysis of the relationships between blood markers, clinical factors, and bone scans, we found that older age and lower lung function had a strong relationship with lower BMD of the hip and femoral neck. CF-related diabetes was related with the BMD in the femoral neck only. Blood marker IL-1ß had a strong relationship with all the bone scans except for the spine. This relationship remained strong even after adjustments for clinical factors. The relationship between blood markers and the changes of BMD over time is still being analyzed. In conclusion, IL-1ß has a relationship with the hip and femoral neck BMD, showing that the outer surface of the bone may be more sensitive to high amounts of IL-1ß   in people with CF. Il-1ß not only signals bone cells to absorb bone tissue but also stops bone cells from creating bone tissues. Since other blood markers did not have a relationship with these bone scans, this suggests that IL-1ß could play an important role in bone health in CF and be of interest for future research.

Establishing an intranet-based referral network for Spina Bifida patients transitioning to adult care

Advances in Surgical care over several decades have allowed children with neural tube defects to survive into adulthood.  Annually, there are small but regular numbers of patients with spina bifida, and its attendant sequelae of neurosurgical, orthopedic, urological and general surgical concern that require ongoing care from adult health care providers. Projected estimates for the BCCH spinal cord clinic are an average of 9 graduates per year over the next 5 years. Our goal is to establish a patient accessible, intranet-based referral network that will greatly improve transition care, specifically for patients graduating from the BCCH spinal cord clinic.

Defining TBR1 syndrome: So we know the gene, but what is the disease?

In the last 5 years, human geneticists have identified an astounding number of novel disease genes. These discoveries are often reported in the form of an aggregate summary of findings in large cohort studies, which means that the provided phenotypic details can be extremely brief. In order to counsel families on what to expect or to improve medical and supportive care, we need first to understand what the newly discovered disorder is exactly.  Clues from a phenotype can sometimes inform the underlying biology in ways that stimulate the development of therapeutic strategies. De novo variants in TBR1 have been reported in 8 individuals with phenotypes of “autism,” “intellectual disability,” “microcephaly,” and “growth restriction.” That is all we know about the phenotype associated with TBR1 mutations. This project will capture the medical and developmental stories of many more individuals with TBR1 syndrome and synthesize this material into a helpful medical guide for clinicians and lay information for families.

RYR3 in epilepsy

We are following a family with two affected siblings, with seizures, psychiatric problems, short stature, severe asthma and lung disease. The family was enrolled in the TIDEX whole exome sequencing study. The genetic study results came back suggesting that a gene that senses calcium inside the cells, called RYR3 is affected, having two previously not seen changes in its genetic code. There are a handful of other patients in the world who have been described to have changes in this gene who have presented with seizures and epilepsy, but nobody has been able to demonstrate the connection between the gene and the epilepsy. We know that calcium has major role in the function of a brain cell, and also has significant effect on release of different chemical substances that can cause seizures. If we could show in laboratory experiments that if we change the genetic code in this RYR3 receptor, than we will have changes in the concentration of calcium inside the brain cell, and measurement of the chemical release is also altered, then we could make the link between the gene and the epilepsy. Validating this finding would mean not only that the family finally will have an answer after many years of search for a diagnosis, but could eventually lead to a novel treatment of their epilepsy. It would also bring an answer to other families whose children have changes in this gene. Better understanding of RYR3 pathophysiology also could have relevance to RYR1 and 2 defects (causes of malignant hyperthermia, seizures, cardiac disease) as well as epilepsy in general.

High-throughput screening for small molecule therapeutics for hypertrophic cardiomyopathy in Leopard syndrome

Leopard Syndrome (LS) is a rare genetic disease that belongs to a family of disorders termed “RASopathies”. Most of the LS patients (>75%) exhibit hypertrophic cardiomyopathy (HCM) which is characterized by an enlargement of the cardiac cells. HCM is a life threatening condition that usually manifests in childhood and can lead to early death. Although clinical features of LS patients with a mutation in the RAF1 gene are typical of LS (pulmonary stenosis, deafness, retardation of growth), remarkably, >95% of LS patients with a S257L RAF1 mutation display HCM, which is usually already present at birth. Despite some progress in understanding the disease, there is still no specific treatment for HCM in LS patients. Therefore, we propose to identify new therapies to treat HCM in LS by performing a high-throughput drug screen on human cardiac cells derived from LS RAF1 induced-pluripotent stem cells (iPSCs).

 The above research grant was generously supported by Global Genes (

Testing the autoimmune hypothesis for ROHHAD

The hypothalamus is a region of the brain that controls many of the automatic functions of the body, such as appetite, breathing, heart rate and temperature control. In ROHHAD (which stands for Rapid-Onset Obesity with Hypothalamic Dysfunction, Hypoventilation and Autonomic Dysregulation), the hypothalamus seems to be malfunctioning, though we do not know why. This leads first to a massively increased appetite and rapid weight gain in very young children (2-4 years old) who had previously been very healthy. It also causes other malfunctions of the body’s automatic processes. Most seriously, these kids will not breathe at the rate they should, and in some cases will actually stop breathing (especially while sleeping) and can die. This is not caused by their obesity, but rather by a failure of their brain to signal their body to breathe. This disorder is extremely rare, with only about 80 cases reported in the literature so far. Some doctors believe that these malfunctions of the hypothalamus might be caused by an autoimmune attack of that brain region – meaning that for some reason, the body starts to produce antibodies that recognize some marker (an antigen) on neurons in the hypothalamus and signal the immune system to attack and kill those neurons (the way it would normally attack and kill invading virus particles or bacterial cells). There have been a few hints that this might be happening, observed in a few cases of ROHHAD, but a comprehensive study on a large group of ROHHAD patients has never been carried out. We propose to undertake such a study, using a large group of ROHHAD patients available to us to look for evidence that the patients are producing antibodies that can recognize some marker on neurons of the hypothalamus. Determining the underlying basis for this disease is of utmost importance in the quest for treatment and prevention. The results of this study will tell us whether the autoimmune hypothesis is one that warrants further study, or whether it is an unlikely mechanism. It will also tell us whether it will be beneficial to patients to continue to experiment with immune therapy as a possible treatment for ROHHAD.

Illuminating the experience of caring for a patient with scleroderma through focus groups

Informal caregivers provide support for a family member or friend with a health condition. Informal caregivers are unpaid and typically do not receive formal training. Patient’s diagnosed with the rare disease scleroderma (systemic sclerosis, SSc) experience debilitating symptoms, including functional disability, pain, fatigue, disfigurement and feelings of helplessness. Most research on SSc focuses on people with SSc, but not the experiences of caregivers, even though many patients with SSc are supported by an informal caregiver. Helping to care for a loved one with an incurable disease, such as SSc, can result in significant disruptions and challenges to daily living. Informal caregivers in SSc report decreased quality of life and are often forced to reduce time spent in paid employment. The proposed project will help to discover the challenges that informal caregivers face and explore services that may be beneficial to lessen the burden associated with providing care to a patient with SSc.

Results - Systemic sclerosis (SSc), also known as scleroderma, is a rare autoimmune disease with debilitating consequences. Many people diagnosed with SSc receive care and support from an informal caregiver to help manage the disease. Little research has considered the experiences of informal caregivers helping to support a loved one diagnosed with SSc. For this reason, we conducted a total of three focus groups, two in New Orleans and one in Montreal, to gain an understanding of the specific challenges faced by informal caregivers to persons with SSc. A total of 13 informal caregivers participated in these focus groups. As a result of the focus groups, a list of 65 unique challenges were identified as being associated with caregiving for a loved one with SSc. Participants also generated a list of 18 potential support services that could be implemented to decrease the burden associated with caregiving. Participants rated the importance of each challenge, and the likelihood of using each of the support services identified.

The highest rated caregiver challenges reported were: not having access to a caregiver support group, others’ lack of awareness of SSc, understanding the emotional needs of the care recipient, and managing emotions as a caregiver. The highest rated support services were internet-based chat groups or forums with knowledgeable health care provider, internet resource sites designed for caregivers, internet based psychological and emotional support tools (e.g. guidance on cognitive tools, use of journaling), specific sessions or workshops focused on caregiver needs, and online informational resources explaining physiological symptoms of SSc. The results gathered from the focus groups will allow for the generation of a survey for dissemination to a larger sample of caregivers for persons with SSc. The final goal of this project is to implement an intervention to help caregivers manage the burden and challenges associated with caring for a loved one with SSc.


1. Rice, D.B., & Thombs, B.D. (2016). Focus Group Discussions to Inform the Development of Services for Informal Caregivers of Patients with Scleroderma.Presented at Annual National Scleroderma Conference, Montreal, Quebec, September 16-17, 2016.

2. Rice, D.B., Cañedo-Ayala, M., Gumuchian, S.T., Thombs, B.D. Informal Caregivers for Scleroderma Focus Groups. Submit to Laurentian Conference of Rheumatology, Estérel, Quebec, May 11-13, 2017.

3. Rice, D.B., Cañedo-Ayala, M., Gumuchian, S.T., & Hagedoorn, M., & Thombs, B.D Nominal Focus Groups to Identify Stakeholder Priorities and Inform Survey Development: An Example with Informal Caregivers of People with Scleroderma. Working Document 

Publication: BMJ Publication March 2018

A hypothesized link between seizures and two syndromes: GLUT1, DYRK1A, and a potential new target for glucose control

Seizures affect about 1 in 100 children, and the number of possible causes is vast. It is the task of the pediatric neurologist to control the seizures, find the cause, and refine the treatment plan according to the cause. Whenever there may be an underlying cause that can be directly treatable, such as a brain tumor, infection, or biochemical imbalance, investigations must be prioritized to find these problems. One such underlying disorder is a genetic problem with transporting glucose to the brain, known as GLUT1 deficiency, which can be treated by feeding the brain with ketones rather than glucose via a special diet. This diagnosis is strongly suspected when measurement of the fluid around the brain shows a low glucose level. At our centre, a child with low glucose in this fluid, and a good response to dietary therapy, surprisingly did not have a genetic mutation in the gene for GLUT1 deficiency, but instead had a mutation in the DYRK1A gene, a recently described cause for a syndrome of intellectual disability and seizures. DYRK1A plays an important role in cells by changing the structure of other proteins to activate them. We hypothesize that DYRK1A may be important for activating GLUT1. If we prove this hypothesis, through the study of the proteins in the cells of individuals with dysfunctional DYRK1A, then we will have discovered a new target for modulating the control of glucose levels, particularly in the brain, but possibly throughout the body as well.

In vivo drug screening of a zebrafish model of hereditary spastic paraplegia (HSP)

Hereditary spastic paraplegia (HSP) is a group of rare inherited disorders, characterized by the loss of upper motor neurons, leading to lower extremities weakness and spasticity and for which no cure is available. In this project, we have developed zebrafish models of HSP recapitulating some features of the human disease. We will use these models for an in vivo drug screen of small molecules that can attenuate or even rescue their condition. Candidate compounds will subsequently be tested in pre-clinical and clinical studies. Our findings may assist in accelerating the development of drugs for the treatment of HSP e.g. as for a drug we discovered in ALS models and moved to clinical trial

Improving Access to Care for Ugandan Children with Rare Surgical Diseases

Each year, our interdisciplinary pediatric surgical team collaborates with Ugandan counterparts in the delivery of surgical care of children from socioeconomically-challenged families in Uganda, where in 2014, around 10% of infants died during or shortly after birth. Many of the children undergoing surgery suffer from rare surgical diseases. We seek to more fully understand the early identification, diagnosis, and treatment of these babies who live in rural settings, where pediatric surgical specialists are not close at hand. We aim to strengthen referral networks, awareness, advocacy and timely access to care. To this end, we will develop a survey that will document the referral, diagnosis, and management of neonates in the first 30 days of life. The survey will be sent to pediatricians, midwives, surgeons, family doctors, nurses, and hospital managers in Uganda to fully understand their current practice of care. Secondly, we will hold a focus group at the annual pediatric surgical camp to discuss best practices around referral and care of these. Most importantly, our study team will interview the parents of patients who present at the pediatric surgical camp with rare surgical diagnoses to understand their care journey. All the information obtained will then be synthesized into a network report.

GBA2 validation

We are following a young adult patient who had problems since her childhood with developmental delay, mild intellectual disability, who presented in her late teens with problems walking. She now needs to walk with a cane or holding onto somebody’s arm. She has also balance problems, and had many falls. Recent genetic studies showed that she has two changes in the gene associated with rare forms of Hereditary Spastic Paraparesis, called GBA2. This gene encodes an enzyme that helps in the cells linking sugar and fat metabolism. We know well the enzyme GBA1, which causes a well known metabolic disease, Gaucher disease, but have very few patients described to have GBA2 enzyme deficiency. The genetic result could not characterize the variants found in this gene in the patient, only as variant of uncertain significance. We could validate the genetic finding and prove that the mutations are disease causing if we could perform enzyme testing in the patient’s blood, and perform some more testing of this protein in patient’s skin fibroblast culture, including Western blots to test the protein quantity, to see if it is decreased. If we could prove that this patient’s genetic findings are causing her symptoms, then she would be a candidate for potential enzyme replacement therapy that is developed for now only for Gaucher disease, maybe with some modifications. This might halt the progression of her symptoms of worsening abilities to walk. She could also be candidate for substrate reduction therapy that would help also with her symptoms.

Harnessing protein acetylation as an alternative treatment strategy for Diamond-Blackfan Anemia

Diamond-Blackfan anemia (DBA) is a collection of diseases resulting from defects in protein synthesis machines called ribosomes. Defects in ribosomes cause problems with the inner part of the bone called the marrow. Patients experience fatigue and weakness, have developmental defects and are cancer prone. Blood transfusions or bone marrow transplants are common treatment options for DBA. We will test whether modulating special chemical groups called acetylations on defective ribosomes will serve as a less invasive treatment option. Our project will use human cells and baker’s yeast, wherein ribosomes are identical to those in humans. This combined approach will speed delivery of our findings to the clinic.

Results - Blackfan Diamond anemia is a rare disease resulting from defects in ribosomal proteins, which form part of a machine in the cell called a ribosome that is responsible for protein synthesis. Ribosome function is highly conserved across species and even kingdoms, i.e. ribosomes work in a very similar way in humans, animals, plants and fungi such as yeast. This makes the budding yeast a model system ideal to tease out the function and regulation of these proteins. We originally identified acetylation sites on yeast versions of the proteins that are affected in Blackfan anemia (Rpl15 and Rps17) using mass spectrometry. We found that nicotinamide, an inhibitor of proteins that remove acetylations, increased acetylation on the proteins we studied. Mutation of the Rtt109 acetyltransferase, which adds acetylation, led to decreased acetylation on these proteins. We hypothesized that manipulation of acetylation would impact protein levels/function and that this could form the basis of a treatment for Blackfan Diamond anemia in the future. However, using different molecular biology approaches we identified no changes in protein levels or mobility (possibly indicative of phosphorylation or ubiquitylation) in the presence or absence of RTT109. We now hypothesize that acetylation of ribosomal proteins may be a rare event indicative of ribosomal dysfunction. Indeed, there are previous reports of Rtt109 interacting physically with proteins being involved in ribosomal quality control. We continue to examine the links between ribosome function and protein acetylation using additional mass spectrometry experiments in both yeast and human cells. In addition, we have carried out a screen for yeast homologs of rare disease genes that cause hypersensitivity to the sirtuin inhibitor nicotinamide when overexpressed. This additional work, which was also supported in part by this microgrant, has spurred an interest in rare disease biology and will ensure that we have additional pathways to examine in the future.

Internet-based accredited education for health care providers of youth with Duchenne Muscular Dystrophy transitioning to adult care

Duchenne Muscular Dystrophy (DMD) is an inherited, progressive, life-limiting neuromuscular disorder affecting ~1 in 4,500 boys.  Most boys with DMD are diagnosed by 3 years of age with delayed gross motor skills; the majority loses the ability to walk by age 13. Until very recently, most DMD patients died during childhood. With the introduction of new treatments, the mean age of death is now 30, and with exciting genetic and molecular discoveries and new potential treatments these young adults may survive longer still.  All of these young patients now need to be transitioned to adult care, but family physicians and adult specialists do not have the knowledge and skill competencies required to manage these patients. In addition, youth leave a system of care and providers they have known since early childhood and lose their eligibility for many benefits and support services.  Difficult transitions can lead to decreased clinic attendance and to the patients not taking their medications appropriately. The consequences are often a worsening of the condition, decreased quality of life or even death. The goal of this project is to improve transitions to adult care by providing education on care of DMD patients to adult health professionals in a self-paced, interactive, accredited online module.

Delineating the genetic basis of severe intestinal dysmotility syndromes

Abnormal movement (motility) of the intestine can be the result of abnormal development of the bowel’s nervous system or the bowel’s muscle.  It is impossible to tell which problem exists except by taking samples of the intestine to examine under the microscope.  No one understands exactly how or why these problems occur.  Because they sometimes occur in many family members, it was suspected that genes may play a role. In some families, mutations in genes affecting muscle or nerves have been found.  It is unknown whether children who have abnormal motility without a family history have similar genetic changes.  This study will look for mutations in genes to better understand the reason for the dysmotility.  Most of the treatments that we currently have available do not help with the dysmotility, but help minimize the consequences such as malnutrition.  A better understanding of how and why this happens will help to “group” similar children together to understand how best to provide better medical and nutritional care, based on the reason for the problem.  It will also be helpful to understand what to expect in each condition, to better plan management for children and families. All in all, the current study strives for personalized medicine to improve outcomes for a group of children suffering life threatening ultra rare disease.

Identification of biomarkers in the TDP-43 pathology of ALS

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that attacks the neurons which control the voluntary muscle movement called motor neurons. The loss of these neurons leads to a progressive paralysis of skeletal muscles. There is no cure for this disease and the lack of answers about the pathologic mechanisms underlying this disease is reflected by the limited availability of effective treatments or biomarkers. The goal of this project is to evaluate the potential of a key protein linked to how neurons manage exposure to environmental stress as a biomarker for ALS.

Elucidating neurobehavioral phenotype in Sotos syndrome

Individuals with SS exhibit characteristic facial findings, large body size, intellectual disability, and behavioral differences. Through the proposed research, we hope to gain a better understanding of their cognitive functioning and hone in on the exact behavioral characteristics common to individuals with SS. This will better define strengths and weaknesses in individuals with SS and may help them to better function in school and in life. A better understanding of the intellectual disability and behavioral problems in SS may direct us to therapies that could benefit these individuals. Furthermore, objective data from the neuropsychological testing will also serve as a baseline outcome measure for any potential future drug trials to treat the neurobehavioral aspects of SS.

The above research grant was generously supported by Global Genes (

Determining the genetic cause of CAMRQ syndrome using the zebrafish model

Individuals with SS exhibit characteristic facial findings, large body size, intellectual disability, and behavioral differences. Through the proposed research, we hope to gain a better understanding of their cognitive functioning and hone in on the exact behavioral characteristics common to individuals with SS. This will better define strengths and weaknesses in individuals with SS and may help them to better function in school and in life. A better understanding of the intellectual disability and behavioral problems in SS may direct us to therapies that could benefit these individuals. Furthermore, objective data from the neuropsychological testing will also serve as a baseline outcome measure for any potential future drug trials to treat the neurobehavioral aspects of SS.

CRISPR/Cas9-mediated editing for deletion mutations in DMD via AAV-DJ delivery

Duchenne muscular dystrophy (DMD) is an X-linked recessive neuromuscular disease, affecting approximately 1 in 5000 boys. The disease is caused by the lack of dystrophin protein expression due to mutations in the DMD gene. The lack of dystrophin leads to progressive muscle deterioration, loss of ambulation, cardiac and respiratory complications, and limited life expectancy due to the destabilization of the dystrophin glycoprotein complex in skeletal muscle Currently, there is no cure with only symptomatic management available. To correct the underlying deletion mutations in the DMD gene, which account for 65% of all DMD cases5, I will explore the potential of a genome engineering technology called clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPRassociated protein 9 (Cas9) for exon skipping and knock-in approaches in vitro via adeno-associated virus (AAV)-mediated delivery.

Unravelling the genetic basis of a novel, rare autoinflammatory disease: the potential role of a Clec12A genetic variant

We have identified a protein that may contribute to the pathogenesis of a very rare autoinflammatory disease (only 1 case in the world) of unknown cause. We propose to determine if this protein renders immune cells more susceptible to activation which would explain the recurrent inflammatory episodes suffered by the patient. A better understanding of the disease process will help in tailoring a more effective therapy for this patient.

Exploring the genetic basis for Vitamin A deficiency in Midline Brain Anomalies

Vitamin A plays a key role in the development of the brain and spinal cord. The cases of several infants with brain anomalies and insufficient levels of vitamin A were recently described. The infants’ vitamin A intake was adequate, and their mothers had no vitamin A deficiency, thus the low levels of vitamin A that may have led to the brain anomalies must have been caused by defects in vitamin A absorption or transfer. None of the children had other clinical signs related to systemic Vitamin A deficiency. This project will conduct genetic analyses to find out whether there are gene defects in vitamin A related genes in these infants that are related to brain development. This will ultimately allow us to provide counseling and better management to families affected by these rare but devastating brain disorders.

The effect of positioning error on the Hilgenreiner epiphyseal angle as compared to the femoral neck-shaft angle in slipped capital femoral epiphysis: A geometrical modeling study

Slipped capital femoral epiphysis (SCFE) is a painful condition in adolescent hips where the upper end (ball) of the thighbone (femur) slips backward out of the correct position. In severe cases, patients need surgery to correct the position of the bone. Surgeons use a particular measurement from X-rays to determine how severe the slip is. However, many patients are not able to put their hips in the ideal position for X-rays due to pain. It is not clear how much measurement error these non-ideal X-rays cause. With computer models, we can find out how much error there is in non-ideal positions and compare it to the error in an alternative measurement. This will tell us if the current measurement works well, or if the alternative measurement will be more reliable in the surgical planning and post-surgical treatments for adolescents with SCFE.

Academic performance of children with non-syndromic single-suture craniosynostosis

Craniosynostosis is a birth defect in which one or more of the growth seams between the bones of a baby’s skull close too early, before the brain is fully developed. This can lead to the brain and head not growing in their natural shapes. In the majority of craniosynostosis cases only one seam is affected, and the condition is non-syndromic in nature. Children with NSC may be at risk for difficulties in cognitive, language and motor skills. Surgical treatment of NSC is recommended to decrease this risk. The potential relationship between surgical treatment and decreased neurological risk drives the need to investigate the academic outcomes in these children. This study aims to evaluate the individual academic achievement levels of school-aged children with NSC using Ontario’s standardized education testing of reading, writing and mathematics at Grade 3 and 6, and to compare these achievements with those of other children. This study provides important direction to risk factors (age of surgery, type of surgery, diagnosis, length of surgery) related to academic delay in children with NSC. This will help identify those children in need and direct educational resources appropriately.

Results - Forty-two children with Single Suture Craniosynostosis (SSC) were evaluated: 36 underwent operative management and 6 were managed non-operatively. The EQAO scores of children with SSC revealed average performance across academic subjects compared with their peers. However, their reading scores were significantly lower over time. Change in writing and math scores were not statistical significant. Twenty-five patients had special education data available for both Grade 3 and 6. One child (4%) required special education in Grade 3, while the need for special education significantly increased (n = 4, 20%) in Grade 6. Conclusions: Children with SSC may experience increased cognitive difficulties and special education needs in grade school. Cognitive delays observed in SSC patients may become more evident as these children progress through grade school. 

Publications: Steen K, Roy A-A, Hong B, Ho ES, Stephens D, Phillips JH, Forrest CR (Submitted) Academic Achievement in School-Aged Single Suture Craniosynostosis Patients Over Time, Cleft Palate-Craniofacial Journal. 

Presentations: Steen K, Roy A-A, Ho ES, Stephens D, Phillips J, Forrest CR. Academic achievement in school-aged single-suture craniosynostosis patients over time, Canadian Society of Plastic Surgeons, 72th Annual Meeting, June 19-23, 2018, Jasper, Canada.

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