PI/Investigator: Robert Wilson, MD, PhD – Children's Hospital of Philadelphia

Award type: General Research Grant

Grant Title: Drug and drug target validation for Friedreich ataxia

Lay summary: There are no approved therapies for Friedreich ataxia (FA). Frataxin functions in the mitochondria, the primary energy-producing part of cells, specifically in the formation of iron-sulfur-clusters (ISCs), which are important co-factors for the function of many enzymes, both inside and outside the mitochondria. This group found that a stress-response pathway that includes a protein called p38 is hyperactivated in FA cells, likely as a result of ongoing oxidative stress and DNA damage. Their working hypothesis is that chronic hyperactivation of the p38 stress-response pathway, which regulates a key protein involved in ISC formation, represents a maladaptive feedback loop that further suppresses ISC formation in FA cells, hence inhibition of the p38 pathway allows FA cells to partially bypass the need for frataxin. Preliminary studies have also implicated a cell-death pathway called ferroptosis, as well as DNA damage, both of which cause p38 activation and both of which are consequences of ISC deficiencies. These investigators hypothesize that ferroptosis inhibitors could facilitate cellular repair processes and decrease p38 activation, with positive functional consequences for FA cells, and that deficiencies in ISC-containing DNA repair enzymes worsen DNA damage and contribute to p38 activation and FA pathogenesis. The overall objectives for this application are (i) to validate the p38 MAP kinase pathway and ferroptosis as targets for FA therapeutics, and (ii) to identify and prioritize lead compounds for clinical development. To this end, drugs and drug targets for these pathways will be tested by using known inhibitors, as well as genetic approaches, and by quantifying FA-associated phenotypes in new FA models. The Specific Aims of these proposal are: Aim 1. Validate the p38 stress-response pathway as a therapeutic target for FA and evaluate drugs targeting this pathway. They will test the hypothesis that chronic hyperactivation of the p38 pathway in FA cells represents a maladaptive feedback loop, and that inhibiting this pathway allows FA cells to partially bypass the need for frataxin. Aim 2. Validate the ferroptosis pathway as a therapeutic target for FA and evaluate drugs targeting this pathway. They will test the hypothesis that ferroptosis contributes to the pathophysiology of FA, and that inhibitors of ferroptosis will have positive functional consequences for FA cells. The proposed research is innovative because novel pathways for the treatment of FA will be tested, and these pathways will be tested in novel models of the disorder. The expectation is that the results will have an important positive impact, identifying promising treatment options and thereby reducing the burden of this serious degenerative disorder.

PI/Investigator: Benoit D'Autréaux, PhD, Mixed Research Unit (UMR 9198), Centre National de la Recherche Scientifique (CNRS), Commissariat à l'Energie Atomique et aux énergies alternatives (CEA), Paris-Saclay University

Award type: General Research Grant

Grant Title: Cell-free high throughput screening assays for the discovery of compounds replacing frataxin in FA

Lay summary: Friedreich's ataxia (FA) is a neurodegenerative and cardiac disease caused by genetic mutations in the gene encoding the protein frataxin (FXN). FXN is an important protein in the synthesis of cofactors of enzymes called iron-sulfur clusters, which are involved in a multitude of essential biological functions such as energy production and defense against oxidative stress. Although promising gene therapy and pharmacologic approaches are currently under development for FA, there is still no treatment to cure or even slow disease progression.

A novel strategy to develop FA therapeutics would be to either replace FXN function or enhance the activity of residual FXN in FA patients. This group has developed a biochemical assay that can measure the results of FXN activity by faithfully reproducing the physiological conditions of Fe-S cluster biosynthesis using a reconstituted machinery with isolated proteins. They found that FXN stimulates Fe-S cluster biosynthesis by enhancing the mobilization of sulfur. Their goal is to use this reconstituted system to develop a cell-free high throughput assay to randomly screen large chemical libraries (in the range of 100,000 compounds) and identify molecules that could produce the same effect as FXN does on Fe-S cluster synthesis formation. They will also test candidate molecules: sulfur donors and molecules that were previously identified by their ability to mitigate the cardiac phenotype of FA. The molecules selected in these assays will then be evaluated in vivo, using a new fly model of FA. The advantage of this animal model is that it allows rapid evaluation of drug candidates, which will facilitate future drug-design procedures to improve the efficiency of the molecules selected in the screens.

This novel high throughput cell-free assay will allow the researchers to select a large number of molecules directly targeting the primary defect in FA that could lead to the discovery of the first drugs specifically active in the replacement of FXN, which would result in a new class of potential FA therapeutics.

PI/Investigator: Hélène Puccio, PhD - INSERM and IGBMC, France

Award type: Kyle Bryant Translational Research Award

Grant Title: High-throughput drug screen on primary sensory neurons depleted for frataxin

Lay summary: Many small molecule-based therapeutic strategies are currently in development for Friedreich's Ataxia (FA), including compounds identified for their ability to increase levels of the frataxin protein mostly by opening the DNA compaction induced by the GAA repeats or by protecting the frataxin protein from degradation. Some compounds aim at ameliorating the health of mitochondria (the power plant of the cells) or at reducing oxidative stress. These potential therapeutics are in clinical development but to date many do not seem to significantly rescue the symptoms or stop the disease progression. Screenings of pharmaceutical compounds are often performed on models (biological systems recapitulating the disease) mildly affected, requiring an additional treatment to mimic the disease, or simply lacking the tissue specificity: FA patient fibroblasts, yeast models, immortalized cell lines or more recently drosophila models. To date there has not been any high-throughput screening (HTS) of drugs on sensory neurons deficient for frataxin, a cell type primarily affected in FA. Establishing such a strategy in this cell type is rather challenging: DRG neurons primary cultures (i.e. neurons collected directly from the animal) are not available in unlimited quantities and often show great variability from culture to culture; moreover, an HTS demands to successfully grow and maintain the desired cells in plates with a high number of very small wells. This group has successfully developed optimal culture conditions for primary DRG neurons in 96 well plates and established a robust and reproducible simple assay to test drug efficiency. They propose to perform a high throughput screening of therapeutic compounds capable of bypassing frataxin deficiency in DRG sensory neurons with the aim to identify potential therapeutics for FA with increased specificity.

PI/Investigator: Javier Santos, PhD - Departamento de Fisiología y Biología Molecular y Celular. Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Consejo Nacional de Investigaciones Científicas y Técnicas, Argentina

Award type: General Research Grant

Grant Title: Structural Dynamics and the Consolidation of Protein Function in Protein Complexes Involved in the Biosynthesis of Iron-Sulfur Clusters: Quaternary Addition of Small Trojan Tutor Proteins

Lay summary: Iron-sulfur (Fe-S) clusters are essential cofactors present in all known forms of life. To exert their functions, hundreds of proteins require such cofactors. In eukaryotic organisms, the biogenesis of most Fe-S clusters takes place in the mitochondria. The process involves the interactions and activities of several key proteins, which form a supercomplex. Namely these proteins are: Frataxin, NFS1, ISCU, and ACP and ISD11. Mutations in these building blocks lead to severe human diseases. For instance, mutations in Frataxin, or a deficiency in its expression, results in Friedreich's Ataxia (FRDA). In particular, Frataxin upregulates the activity of the supercomplex. Our laboratory studies the stability, internal motions, and functionality of Frataxin mutants to better understand the root cause of loss of function in the pathogenic variants. We propose delivering a small “Trojan” tutor protein with high affinity for Frataxin to the mitochondrial matrix, inside the cell. Modulation of Frataxin stability and activity may exert control on the supercomplex function. Furthermore, we will search for “Trojan” tutors against other proteins of the supercomplex. In this way, we will increase our chances of successfully improving Fe-S cluster biosynthesis.

Publications:

• Rescuing the Rescuer: On the Protein Complex between Human Mitochondrial Acyl Carrier Protein and ISD11. Herrera MG, Pignataro MF, Noguera ME, Cruz KM and Santos J. ACS Chemical Biology ACS Chemical Biology 2018 Jun 15;13(6):1455-1462.
• Biophysical Characterization of the Recombinant Human Frataxin Precursor. Castro IH, Ferrari A, Herrera MG, Noguera ME, Maso L, Benini M, Rufini A, Testi R, Costantini P and Santos J*. 2018. FEBS Open Bio. doi:10.1002/2211-5463.12376.
• 3- Insights on the conformational dynamics of human frataxin through modifications of loop-1. Noguera ME, Aran M, Smal C, Vazquez DS, Herrera MG, Roman EA, Alaimo N, Gallo M, Santos J. Arch Biochem Biophys. 2017 Dec 15;636:123-137.
• Human Frataxin Folds Via an Intermediate State. Role of the C-Terminal Region. Faraj SE, González-Lebrero RM, Roman EA, Javier Santos*. Scientific Reports, Nature. 2016 Feb 9;6:20782
• Structural characterization of metal binding to a cold-adapted frataxin. Noguera ME, Roman EA, Rigal RB, Cousido-Siah A, Mitschler A, Podjarny A, and Santos J*. J Biol Inorg Chem. 2015 Jun;20(4):653-64.
• A helix-coil transition induced by the metal ion interaction with a grafted iron-binding site of the CyaY protein family. Vazquez DS, Agudelo WA, Yone A, Vizioli N, Arán M, González Flecha FL, González Lebrero MC, Santos J*. Dalton Trans. 2015 Feb 7;44(5):2370-9
• The Alteration of the C-terminal Region of Human Frataxin Distorts its Structural Dynamics and Function. 7- Faraj SE, Roman EA, Aran M, Gallo M, Santos J*. FEBS J. 2014 Aug;281(15):3397-3419.
• The role of the N-terminal tail for the oligomerization, folding and stability of human frataxin. Faraj SE, Venturutti L, Roman EA, Marino-Buslje CB, Mignone A, Tosatto SCE, Delfino JM*, and Santos J*. FEBS OPEN BIO, 2013 Jul 24;3:310-20.
• Frataxin from Psychromonas ingrahamii as a model to study stability modulation within the CyaY protein family. Roman EA, Faraj SE, Cousido-Siah A, Mitschler A, Podjarny A, Santos J*. Biochim Biophys Acta. 2013. Jun;1834(6):1168-80.
• Protein Stability and Dynamics Modulation: The Case of Human Frataxin. Roman EA, Faraj SE, Gallo M, Salvay AG, Ferreiro DU, Santos J*. PLoS ONE 2012. 7(9): e45743.

PI/Investigator: David Corey, PhD - UT Southwestern, Dallas, Texas
PI/Investigator: Marek Napierala, PhD - University of Alabama at Birmingham

Award type: General Research Grant

Grant Title: Development of oligonucleotide activators of FXN expression

Lay summary: Friedreich's ataxia (FRDA) is an incurable genetic disorder caused by reduced expression of the mitochondrial protein frataxin (FXN). Agents that increase expression of FXN would correct the disease-causing defect and are a promising approach to therapy. FRDA patients have an expanded GAA repeat region within intron one of FXN and this expanded repeat causes transcriptional silencing by a mechanism that has yet to be definitively described. We designed duplex RNAs and single stranded locked nucleic acid (LNA) oligonucleotides (short man-made strands of DNA) to recognize the repeat region and interfere with contacts that contribute to decreased transcription. We found that both duplex RNAs and LNA oligonucleotides caused increased expression of FXN protein in cells derived from FRDA patients. The increase in FXN expression was similar to the level found in normal cells. Both duplex RNAs and LNA oligonucleotides belong to classes of molecule that are being developed clinical. The combination of our promising initial results with experience using similar models in clinical trials suggests a path towards clinical translation towards treatment of FRDA. We now report extending this research to testing more compounds in more cells. Our findings extend the generality of our approach and provide a broader database for evaluating how to move forward with drug discovery.

Publications

• Liu J, Hu J, Ludlow AT, Pham JT, Shay JW, Rothstein JD, Corey DR. (2017) "c9orf72 Disease-Related Foci Are Each Composed of One Mutant Expanded Repeat RNA." Cell Chem Biol. 24: 141-148.
• Matsui M and Corey DR. (2016) "Noncoding RNAs as drug targets." Nature Rev. Drug. Discov. 16: 167-179.

PI/Investigator: Stephanie Cherqui, PhD - University of California, San Diego

Award type: General research grant

Grant Title: Mechanism of neuronal rescue by microglia expressing frataxin and characterization of neuronal defects in Friedreich's ataxia

Lay summary: Dr. Cherqui and her group showed that, in a mouse model of FRDA, bone marrow stem cells, especially the hematopoietic stem cells (HSCs) that give rise to blood cells, hold great promise to treat FRDA. Indeed, transplantation of healthy HSCs led to the prevention of the neurological, and muscular complications in the mouse model. The mechanism by which frataxin-expressing HSCs protected the tissues is mediated by the differentiation of the transplanted HSCs into microglia/macrophages (type of immune cells) within tissues, that will transfer the functional frataxin to the brain cells (neurons) and muscle and heart cells (cardiomyocytes and myocytes). The exact mechanism of frataxin transfer is still unknown and determining the mechanism of action is crucial for designing adequate safety and efficacy studies to meet FDA requirements for future clinical application of HSC transplantation for FRDA. Because these questions cannot be answered in vivo, this group will utilize induced pluripotent stem cells (iPSCs) derived from FRDA patients and healthy donor fibroblasts to generate microglia, neurons, and cerebral organoids (mini-brains) to study the microglial-neuronal interaction and, potentially, the mitochondria transfer. We will also utilize these new models of FRDA to investigate the pathogenesis of the neurodegeneration in this disease, and the potential impact of the frataxin-deficient microglia in this degenerative process. Indeed, using their 2D and 3D FA models, they already observed significant phenotype in the FA-neurons and FA-microglia compared to controls. This work will lead to a better understanding of the mechanism by which HSC transplantation leads to improvement of the neurological symptoms, will further the understanding of FRDA neuropathogenesis, and potentially identify new therapeutic targets to prevent or slow neuronal death in FRDA patients.

Co-sponsor: fara Australia

PI/Investigator: Hélène Puccio, PhD - Institute NeuroMyoGène, Lyon, France

Award type: General Research Grant

Grant Title: Defining the therapeutic window and threshold for neuronal gene therapy in Friedreich Ataxia

Lay summary: Gene replacement therapy is an experimental technique that uses genetic material to treat or prevent a disease. One promising gene replacement therapeutic approach for Friedreich ataxia (FA) is to deliver the frataxin gene to affected cells, using viral vectors. Several groups have recently shown that this type of approach could prevent and treat the disease in relevant mouse models. However, there are a number of questions that need to be addressed to optimize the development of a safe therapeutic protocol. In particular, it is now accepted in the field that too much frataxin expression can be detrimental to the normal function of the cell. It is therefore essential to develop a therapeutic vector that will mimic the normal endogenous expression of frataxin. Furthermore, it is important to estimate the number of neurons that need to be corrected to produce a clinical benefit. Recent studies have shown that the sensory ataxia in FA might be partly related to problems during the development, and it is therefore important to determine if gene replacement can correct this very early damage to the tissue. Dr. Puccio and her team will address these questions with a particular focus on the neurological aspects, using two relevant mouse models of the disease, and a novel viral vector that expresses frataxin at near physiological levels.

PI/Investigator: Jill Napierala, PhD - University of Alabama at Birmingham

Award type: General Research Grant

Grant Title: Regulation of frataxin expression - implications for Friedreich's ataxia therapy

Lay summary: Friedreich's ataxia (FRDA), a severe progressive neurodegenerative disorder, is caused by an increasing number of specific DNA sequences, termed GAA repeats, that are present in the Friedreich's ataxia gene (FXN). This error in DNA causes a block in the flow of information from DNA to RNA, and ultimately leads to a deficiency of the final FXN product, a protein called frataxin. One of the major types of therapeutic approaches for FRDA currently being developed tries to counteract this frataxin deficiency. In order for the therapy to be successful, we need to determine what is the minimum amount of frataxin increase that will be beneficial for patients as well as what is the maximum possible increase of frataxin that will not cause any negative consequences. This is called a therapeutic window and is an essential parameter for therapy development for FRDA. Also, to better understand dosing of potential drugs that could increase frataxin levels, results of the proposed work will determine the ways that frataxin production, maintenance and removal are controlled. In summary, this work is contributing to the development of critical therapeutic guidelines for treatment of frataxin deficiency in Friedreich's ataxia.

PI/Investigator: Natalia Gomez-Ospina, MD, PhD - Stanford University

Award type: General Research Grant

Grant Title: Development of autologous transplantation of genetically corrected hematopoietic stem cells for Friedreich Ataxia

Lay summary: Previous studies have suggested that bone marrow transplantation can improve symptoms in a mouse model of Friedreich's ataxia. A conclusive demonstration of the efficacy and feasibility of such an approach for this disease would enable a one-time treatment for this otherwise devastating disease. If such benefit were to be firmly established, as it has in other neurological diseases, donor-blood stem cell transplantation could quickly become a treatment alternative for individuals with Friedreich's ataxia, while launching the development of patient-derived stem cell transplantation approaches in which the patient's own stem cells have been genetically corrected. Dr. Gomez-Ospina proposes to investigate the application of blood stem cell transplantation to treat Friedreich's ataxia by: 1) performing stem cell transplantation experiments into a new model of Friedreich's ataxia and assessing the molecular and functional outcomes, 2) using genetic engineering to investigate whether expression of the protein Frataxin is required in specific blood cell types for the therapeutic benefit of stem cell transplant, and 3) establishing an in vitro model to assess if cellular bodies called mitochondria undergo inter-cellular transfer to confer function to neighboring cells. These studies will provide support for further development of a blood stem cell-based therapy for Friedreich's ataxia.
Co-funding: FARA Ireland

PI/Investigator: Christina Cortez-Jugo, PhD - University of Melbourne, Australia

Award type: Award for Innovative Mindset

Grant Title: Targeted Delivery of Frataxin DNA to Proprioceptors of the Dorsal Root Ganglia

Lay summary: This application proposes to use nanoparticles instead of viruses to deliver the frataxin gene in gene therapy approaches. Nanoparticles are small aggregates made of polymers, lipids and other molecules that can deliver a specific cargo (in this case the frataxin gene) to cells. This investigator proposes to improve the delivery of the cargo to the specific cell types that are mostly affected in FA, by coating the nanoparticle surface with antibodies and ligands that will promote their binding and uptake by the target cells. The ability to target specific cells affected in FA allows to increase the concentration of the therapeutic where it is needed, reduce dosage, cost of treatment and side effects.

Co-sponsor: CureFA Foundation and fara Australia

PI/Investigator: Shannon Boye, PhD - University of Florida

Award type: General Research Grant

Grant Title: AAV-mediated therapy for visual impairment associated with Friedreich's ataxia

Lay summary: Friedreich's Ataxia (FA) patients typically present before the second decade with loss of muscular function, speech impediments, and cardiomyopathy. In addition to these peripheral complications, defects of the central nervous system including hearing and vision loss ultimately manifest. Progress is being made to develop a whole body (systemic) gene therapy to treat FA. While systemically delivered Adeno associated virus (AAV) can efficiently target the muscle and heart, and injections into the spinal cord (intrathecal) can efficiently reach the brain, neither are likely to mediate sufficient levels of therapeutic transgene in the retina to prevent vision loss. A more directed approach is needed to treat the ocular phenotype of this disease which includes loss of retinal ganglion cells (RGCs), thinning of the nerve fiber layer, optic nerve atrophy, nystagmus and loss of visual field. The purpose of this study is twofold; 1) to characterize the natural history of a retina specific Fxn knock out mouse model, and 2) to develop a retina-targeted, AAV-based therapy for preserving vision in FA patients. The latter could be administered alone in advanced stage patients, or in combination with systemic/intrathecal treatments in those patients with less advanced disease.

PI/Investigator: Ronald Crystal, MD - Weill Cornell Medical College, NY, USA

Award type: General Research Grant

Grant Title: Gene Therapy for Cardiac Manifestations of Friedreich's Ataxia

Lay summary: Friedrich's ataxia (FA) is a common, fatal hereditary movement disorder. While the neurologic disease limits mobility, 60% of individuals with FA die from progressive cardiomyopathy. This group plans to initiate a clinical study of AAVrh.10hFXN (a serotype rh.10 adeno-associated virus coding for human frataxin) to reverse the cardiac manifestations of FA. They have completed: (1) demonstration of efficacy of AAVrh.10hFXN in 2 different mouse models of the cardiac disease associated with FA; (2) development of methods to manufacture AAVrh.10hFXN for use in humans in our clinical (GMP) manufacturing facility; (3) carried out short-term toxicology studies of AAVrh.10hFXN in the heart of nonhuman primates; and (4) have carried out extensive toxicology studies in experimental animals and humans of the AAVrh.10 vectors. The goal of this proposal is two-fold: (1) to carry out a murine dose-ranging toxicology study with intravenous administration of AAVrh.10hFXN to insure safety (scaled to humans), of the proposed doses in the clinical trial; and (2) to leverage the clinical evaluation of individuals with FARA in a parallel proposal (Pagovich, O., PI, "Corneal Confocal Microscopy Quantitative Imaging of Corneal Nerves as a Biomarker of Neurologic Disease in Friedreich's Ataxia"), to carry out the cardiac-related screening studies to identify potential candidates for the clinical trial.

PI/Investigator: Mirella Dottori, PhD - University of Wollongong, Australia

Award type: General Research Grant

Grant Title: Nanoparticle-Mediated Gene Delivery of Frataxin to Neurons

Lay summary: A major component towards treating Friedreich's Ataxia (FRDA) is to identify the most optimal approach for delivering therapeutic molecules into the human nervous system. Advances in bioengineering have developed assembled chemical compounds, called ‘nanoparticles', that have the capability of encapsulating protein or DNA molecules and penetrate into cells. Once inside the cell, nanoparticles release their contents, thereby essentially serving as specialized carrier system for delivering therapeutic agents. There are many different nanoparticle types that differ in their physical and chemical properties that influence which cells they can or cannot penetrate into. The major aim of this application is to determine the optimal nanoparticle type that can penetrate human neurons and deliver DNA that encodes for Frataxin protein. With stem cell technologies, human neurons can now be grown in 3D aggregates, such that they resemble neural-like tissue. This group proposes to use this system to test different nanoparticle types in a high-throughput manner. The outcome of these studies will fast-track the development of nanoparticle materials for their use in treating FRDA.

PI/Investigator: Daniele Lettieri-Barbato, PhD – University of Rome Tor Vergata, Italy

Award type: General Research Grant

Grant Title: Testing the efficacy of dietary butyrate in ameliorating ataxic symptoms in Friedreich's ataxia mouse models

Lay summary: Together with degeneration in the brain and spinal cord, FRDA patients frequently develop metabolic complications that culminate in heart disease and type 2 diabetes, which significantly aggravate FRDA progression. A direct link has been discovered in recent years between the gut and the brain and alteration of the composition of the microbial gut population is associated with several neurodegenerative diseases. In this project, by using mouse models of FRDA, Dr. Lettieri-Barbato and his team will investigate whether the gut microbiome is altered in FRDA with a particular focus on bacteria producing short chain fatty acids, including butyrate, as these molecules possess both a neuroprotective and anti-diabetic function. FRDA mice will be treated with a diet rich in butyrate and the effect on motor function and molecular hallmarks of the disease will be analyzed in the cerebellum at single cell level. If butyrate will demonstrate efficacy in mitigating the neuromotor symptoms and molecular deregulation identified in the cerebellum of FRDA mouse models, translation from bench to bedside could be highly feasible as this physiologically produced molecule has been shown to be safe in humans.

Co-sponsor: AFAF

PI/Investigator: Elena Dedkova, PhD - University of California, Davis

Award type: General Research Grant

Grant Title: Cardiac benefit of drugs that stimulate Nrf2 and HCA2 pathways in Friedreich's ataxia

Lay summary: The most common cause of death in Friedriech's ataxia (FA) is cardiomyopathy, thus a therapeutic drug that increases frataxin, iron-sulfur and mitochondrial functions has the potential to ameliorate the most lethal consequence of FA. Ixchel Pharma, a UC Davis spinout company, has developed a novel chemical entity, IMF, that increases frataxin, iron-sulfur and mitochondrial functions in the FXNKD mouse model of FA, that closely resembles the human FA condition. IMF dosing in these FXNKD mice rescues their cardiac pathology. Furthermore, IMF dosing increases survival in the frataxin MCK-Cre mouse, which only lacks frataxin in the heart, and dies because of this loss. Thus, IMF is potentially a novel therapeutic with greater efficacy than the related DMF, and operating through other mechanisms. This group proposes that IMF works by increasing frataxin expression and mitochondrial gene expression, which appears to be upstream from the target of RTA408, Omaveloxolone, which does not increase frataxin and works to benefit oxidative status downstream of frataxin deficiency. Thus, this group proposes to compare efficacy of IMF and RTA408 in mice, and if IMF has higher efficacy, then to carry out pharmacokinetics, metabolism and toxicity studies of IMF. If successful this would take IMF to the next level of preclinical development.

PI/Investigator: Rucha Sarwade, PhD - Monash University, Australia

Award type: Postdoctoral fellowship

Grant Title: Investigating epigenetic silencing in Friedreich's Ataxia

Lay summary: Several models have been proposed to explain FXN gene silencing. Two eminent amongst them are; 1. Formation of unusual triplex DNA structures and R-loops that interferes with the RNA pol II processivity leading to transcriptional blockage, 2. Formation of heterochromatin. While research findings are consistent with both possibilities, neither of them adequately explains transcriptional silencing of FXN gene. This project aims to fill this critical knowledge gap and uncover sequential epigenetic events that are crucial to design effective treatment strategies for Friedreich's Ataxia (FRDA). In a parallel universe, studies on plants that have a peculiar growth defect due to an intronic triplet repeat expansion led to interesting observations. This plant model shares striking parallels at the molecular level with FRDA, suggestive of potential common underlying biology. In the plant model, repeat expansion causes accumulation of specific non-coding RNA species called siRNAs. These siRNAs lead to gene silencing by RdDM (RNA dependent DNA methylation) -dependent epigenetic modifications. Interestingly, repeat expansion-associated plant phenotype was rescued by mutations in enzymes that can cause post-translational modification of proteins. Excitingly, HETEROCHROMATIN PROTEIN 1 (HP1) that has been shown to be associated with epigenetic silencing in FRDA is known to be affected by such post-translational modifications. Dr. Sarwade hypothesizes that RNA -mediated epigenetic changes occurring at the FXN locus are maintained by protein modifications of chromatin modifiers such as, HP1. Through this fellowship, Dr. Sarwade intends to test whether the learnings from the plant research also translate to FRDA, using cell lines derived from patients.

Co-sponsor: fara Australia

PI/Investigator: Chen Liang, PhD - University of Rochester

Award type: Postdoctoral Fellowship

Grant Title: Pathophysiology of Muscle Dysfunction in Friedreich's Ataxia

Lay summary: FRDA is a deadly neurodegenerative disorder that is associated with significant muscle wasting and weakness. However, the reasons for this have not been extensively examined. Given that skeletal muscle plays a critical role in movement and daily activity, there is a pressing need to enhance our understanding of how altered skeletal muscle function contributes to the development and progression of FRDA. A major barrier to achieving this goal has been the lack of an animal model that accurately reflect the clinical features of FRDA in skeletal muscle. To overcome this barrier, Dr. Liang will generate two new pre-clinical mouse models, constitutive and inducible muscle-specific Fxn knockout mice. These mouse models will enable detailed analyses of the role of Fxn in muscle development and function, as well as the acute effect on muscle function of Fxn deficiency in adulthood. These mouse models will also be used to identify and test the efficacy of new interventions designed to counteract the deficits that result from Fxn deficiency during development and adulthood. This investigator and her mentors hypothesize that reduced Fxn expression in skeletal muscle results in muscle dysfunction and that restoration of Fxn expression ameliorates muscle dysfunction and prolongs survival. They further hypothesize that Fxn deficiency in skeletal muscle arises from mitochondrial damage due to mitochondrial Ca2+ overload and that muscle function is improved by blocking the activity of mitochondrial permeability transition pore (mPTP) through ablation of cyclophilin D. These hypotheses will be comprehensively evaluated using a multidisciplinary approach that includes muscle physiology, molecular biology, biochemistry, and Ca2+ imaging. The outcome of this study will provide new insights into the pathogenic mechanisms and treatment of the debilitating muscle dysfunction experienced by FRDA patients.

PI/Investigator: Sara Anjomani-Virmouni, PhD - Brunel University, London

Award type: General research grant

Grant Title: Sphingolipid rheostat as a potential target for Friedreich's Ataxia

Lay summary: Evidence indicates that defective sphingolipid metabolism may contribute to different neurodegenerative conditions, including Alzheimer’s disease, Huntington’s disease and Parkinson’s disease. Sphingolipid is a group of lipids important for the activity of the brain and therefore, disturbances in their metabolism can have a huge impact on brain function. Dr Anjomani Virmouni and her group have recently found that the sphingolipid levels and their related genes are altered in FRDA mouse and human samples, which may play a critical role in the disease progression. The aim of this project is to have a clear picture of the sphingolipid changes in FRDA and to identify potential and novel targets for the development of therapeutic strategies in the disease. This may involve the design of novel drugs or repurposing of currently licensed drugs.

PI/Investigator: Marco Carpenter, PhD - Children’s Hospital of Philadelphia

Award type: Award for Innovative Mindset

Grant Title: Nuclear frataxin and the regulation of macrophage activation

Lay summary: Little is known about the role of frataxin in nuclei or about the mechanisms that control its appearance within the nuclear volume. Key evidence suggest Frataxin has novel DNA repair functions and that this function is important in the pathogenesis of FRDA: 1) Iron accumulation is an inconsistent and late event in FRDA cells and animal models 2) Increased sensitivity to reactive oxygen species and impaired DNA repair appears to be an early event in the pathogenesis of FRDA 3) frataxin isoform expression and proteolytic processing target frataxin protein to the nucleus in some cell-types. These observations encourage a new perspective on frataxin beyond mitochondrial mechanisms, such as a role in the regulation of DNA protection related to oxidative damage and transcriptional regulation in macrophages. Dr. Carpenter proposes to optimize methodologies to profile and quantify frataxin interactions in the nucleus. This approach is the first to investigate nuclear frataxin-mediated gene regulation in response to increased oxidative stress using a combination of classic and novel methodologies. Findings from this proposal will inform the design of novel therapies targeting nuclear frataxin.

Co-sponsor: fara Australia

PI/Investigators: Ricardo Mouro Pinto, PhD – Massachusetts General Hospital, Harvard Medical School & Caroline Benn, PhD – LoQus23 Therapeutics

Award type: General research grant

Grant Title: Testing whether somatic GAA expansions are a therapeutic target for FRDA

Lay summary: The inherited GAA repeat expansion in FA undergoes even further expansion in some parts of the body, in a process called “somatic instability”. These are the tissues that are most affected by the disease, such as the heart and the dorsal root ganglia in the spinal cord. In other neurodegenerative diseases that are also caused by repeat expansions, there is increasing evidence that somatic instability may contribute to disease. For this reason, researchers and companies are designing drugs that target somatic instability. Before these drugs can be considered potential therapies for FA, the contribution of somatic instability to disease progression needs to be established. The inherited GAA expansion causes a decrease in the production of the frataxin protein, which causes problems with energy generation in the cell. The question is whether somatic instability reduces frataxin production even further, making matters much worse in tissues such as the heart and dorsal root ganglia. To test this, Dr. Mouro Pinto and his collaborators will use an FA mouse model of somatic instability, in combination with patient-derived cell systems. The latter lack the multi-cellular environment of an animal, but are simpler and can give faster answers, while the former takes some time, but will give important clues on what is going on in a whole organism. The information obtained from this project is crucial to understanding whether drugs targeting somatic instability will make a viable therapeutic strategy to treat FA.

Supported by the Crisp Family Fund

PI/Investigator: Jordi Magrane, PhD - Weill Cornell Medicine

Award type: General research grant

Grant Title: Assessment of early somatosensory impairment in the KIKO mouse model of Friedreich's ataxia

Lay summary: Alterations in the normal development of the nervous system may explain some of the earliest pathology in Friedreich's ataxia (FRDA). Clinical and histological evidence suggest that FRDA patients may suffer from an impaired neurodevelopmental process that may occur during embryonic development or during early years after birth. No research studies have directly examined the consequences of frataxin reduction on the maturation of the nervous system during perinatal and early postnatal stages in mouse models of the disease. Dr. Magrane and his group have obtained experimental evidence using a well-characterized FRDA mouse model that frataxin deficiency causes a delay in the appearance of sensory and motor functions in neonates. Based on these novel observations, they aim to further investigate pathology in FRDA mouse pups affecting the nervous system structure and function, and to study the molecular mechanisms involved, with a focus on mitochondria. Successful completion of these studies will not only be important for our understanding of the pathogenesis of FRDA but will also validate the use of this model to evaluate the impact of early therapeutic approaches targeting the sensorimotor system.

PI/Investigator: Nadia D'Ambrosi, PhD - University of Rome Tor Vergata, Italy

Award type: General research grant

Grant Title: Role of iron-dependent dysfunctions in microglia toxicity

Lay summary: It is widely recognized that the nervous system is a primary target in Friedreich's ataxia (FRDA), with specific brain and spinal cord regions displaying significant atrophy and degeneration. Neuronal loss is usually a permanent event, since nerve cells are poorly replaced by adult stem cells, leading therefore to derangement of brain areas, with dramatic consequences. In most diseases characterized by nerve cells injury, neuronal fate relies not only on intrinsic pathological mechanisms, rather neuronal loss is often preceded and accompanied by activation of neighboring non-neuronal cells, that contribute to establish the neuronal outcome of these diseases. Microglia, in particular, are considered the immune cells of the central nervous system, where they represent 10% of the total cell population. They constantly surveil the extracellular environment and respond to multiple stimuli, in order to kill and remove possible harmful agents. Malfunctioning neurons are a source of signals that activate adverse responses by microglia, culminating into further damage to neurons, recognized by microglia as undesirable elements that have to be removed. Nevertheless, under certain circumstances, microglia can also exert trophic and pro-survival actions that can support neuron integrity. Therefore, the possibility to switch microglia towards this beneficial side can be of advantage. The multifaceted role of microglia is well defined in neurodegenerative conditions such as Alzheimer’s and Lou Gehrig’s, as well as fronto-temporal dementia, where the replacing of harmful microglia with a beneficial type has provided many encouraging results in slowing-down the progression of the diseases. In FRDA, there is a limited knowledge about the toxic functions assumed by microglia as a consequence of frataxin-dependent iron accumulation. However, important papers published in very recent years, demonstrated that reactive microglia populate affected brain areas and that the transplantation of healthy microglia protects to a certain extent tissue damage in FRDA mice models. These data provide a robust indication that microglia can contribute to neuronal demise also in FRDA. With this project, Dr. D’Ambrosi aims to obtain a direct demonstration that FRDA-related pathological mechanisms encompass the neurotoxic conversion of microglia. To this purpose, her group will employ microglia derived from healthy and FRDA affected mice to dissect how iron accumulation, derived by frataxin loss-of-function, drives incorrect mechanisms in microglia that alter their mitochondria, their production of damaging free oxygen radicals and their release of pro-inflammatory molecules, transforming them into possible neurons’ foes. Finally, they will directly demonstrate how microglia from FRDA mice impair neuron survival and thus how these cells contribute to neuronal demise in the disease. The clear identification of microglia as main players in the pathology will open new venues for a possible treatment strategy.

Co-sponsor: National Ataxia Foundation

PI/Investigator: Elisabetta Indelicato, MD, PhD - Medical University of Innsbruck, Austria

Award type: Postdoctoral Research Award

Grant Title: Hepcidin-Ferroportin axis in Friedreich's ataxia

Lay summary: One of the open questions in Friedreich´s ataxia (FRDA) is elucidating the role of iron in the development of the disease. In heart and brain of FRDA abnormal iron deposits are found in the mitochondria, the cell organelles mostly affected by the disease. Abnormal iron deposition is believed to trigger damage of mitochondria and contribute to disease manifestations. Over the past years, research in other fields identified the protein hepdicin (HAMP) as the main regulator of iron metabolism in the human body. HAMP is produced by the liver and in conditions of iron excess binds to ferroportin (FPN), an iron-exporting protein located in the cell membrane. This binding triggers FPN destruction and thus prevent the outflow of the ingested iron from the intestinal cells in the blood. Few studies in autopsies and in mouse models suggested that HAMP pathway may be altered in FRDA. To explore HAMP involvement in FRDA, Dr. Indelicato designed a pilot study in which she will measure levels of HAMP, iron and copper parameters, erythropoietin and erythroferrone (another human hormones with iron-regulating properties), as well as frataxin and FPN expression in blood cells of FRDA patients, carriers and control subjects. Moreover, Dr. Indelicato will study if iron accumulates in organs other than the nervous system and the heart by means of an MRI examination of the abdomen, which can visualize iron content in liver, pancreas and spleen. The aim of this proposal is to elucidate if the whole-body regulation of iron through the key factors HAMP and FNP is impaired in FRDA. These findings will help to guide therapeutic interventions on iron metabolism in FRDA and will help to understand if levels of HAMP may provide indirect information on disease state in FRDA.

Co-sponsors: fara Australia and FARA Ireland

PI/Investigator: Erin Seifert, PhD - Thomas Jefferson University

Award type: General research grant

Grant Title: Metabolic (mal)adaptation of heart and skeletal muscle to frataxin depletion

Lay summary: Dr. Seifert's lab has previously shown that in an animal model of FA (“UCLA” mice), heart function can be normal despite >98% loss of Frataxin (Fxn) and evidence of iron overload; normal function likely relies on preservation of fat oxidation and activation of processes that protect the heart. Interestingly, the hearts of Fxn-depleted mice show perturbations in 3 major signaling pathways, indicating that the Fxn-depleted heart is not normal. Some of these pathways may help to preserve heart function in the face of Fxn depletion, while others may lead to poor heart function if left unchecked. To understand how each pathway impacts the Fxn-depleted heart, the team will take advantage of existing small molecules: rapamycin (inhibits mTORC1 pathway), AICAR (activates AMPK pathway), and ISRIB (blocks the Integrated Stress Response pathway). The goal of this project is to test these small molecules in the UCLA mice, which have normal/compensated heart function, and in “MCK” mice that have complete Fxn loss in muscle soon after birth and show dramatic cardiac pathology. This group of investigators will determine if each of the 3 small molecules has a beneficial or harmful effect on heart function, size and metabolism in both mouse models. They will also determine if these molecules can protect skeletal muscle in these animals from declining in mass and strength. This study will fundamentally address whether the observed changes in signaling pathways are beneficial or maladaptive, and whether these pathways can be useful therapeutic targets.

PI/Investigator: Arnulf H. Koeppen, MD - VA Medical Center, Albany

Award type: General research grant

Grant Title: The pathogenesis of the major neural lesions in Friedreich ataxia: dorsal root ganglion and dentate nucleus

Lay summary: The mutation in Friedreich ataxia (FA) was established in 1996, but the mechanisms by which brain, spinal cord, dorsal root ganglia (DRG), and sensory peripheral nerves are damaged have remained elusive. The principal investigator hypothesizes that each one of the vulnerable tissues contains a unique set of proteins that undergo changes in the level of expression in response to frataxin deficiency in FA. The goal of this research is to identify those proteins-of-interest in FA that are up-regulated or down-regulated relative to levels in non-FA subjects, and to determine the functional consequences of these differences. Discovery and identification of FA-relevant proteins is accomplished by a combination of antibody microarrays with tissue lysates; slide technology (immunohistochemistry and immunofluorescence), gel electrophoresis and Western blotting. Antibody microarrays offer enough sensitivity to detect structural and signaling proteins relative to total protein. The success of proteomic methods depends on validation of antibodies and their reactivity with identifiable proteins. The investigator uses tissue samples obtained at the time of autopsy that were generously donated by the families of patients who succumbed to FA. The project has already yielded several important conclusions among which are: (1) Frataxin deficiency causes developmental hypoplasia of DRG; and (2) further destruction is due to proliferation of satellite cells that normally surround and nourish the nerve cells of DRG.

PI/Investigator: Yutaka Yoshida, PhD - Burke Neurological Institute & Joriene de Nooij, PhD – Columbia University

Award type: General research grant

Grant Title: Constancy of FRDA phenotypes across neuronal types and development

Lay summary: Despite remarkable progress, there remain considerable gaps in our knowledge of FRDA pathology in neuronal tissues. This lack of knowledge limits our ability to predict which new types of medication may be more effective in treating the disease and deserve a higher priority. This proposal is focused on three such areas that remain understudied. First, is the mechanism of disease the same in all neuronal tissues? Second, is the effect of the loss of FXN the same in a developing neuron as in a mature neuron? Lastly, does the loss of FXN in the developing neuron lead to permanent (genomic) alterations that cause or exacerbate disease in later life? Based on expertise in the development of two FRDA-affected neuronal tissues: proprioceptive sensory neurons (PSNs) and corticospinal neurons (CSNs), these investigators propose to perform a detailed comparative phenotypic and molecular analysis to examine the similarities or dissimilarities between the FRDA pathology in these two different tissues, and during their development. The result of these studies will inform design strategies for new therapeutics, or guide the evaluation of existing experimental treatments, particularly for those that pertain to neurological FRDA phenotypes.

Co-funding: CureFA Foundation

PI/Investigator: Stephen Chan, MD, PhD - University of Pittsburgh School of Medicine

Award type: General research grant

Grant Title: Frataxin deficiency as a cause of endothelial senescence and vasculature remodeling in Friedreich's ataxia

Lay summary: Friedreich's ataxia (FRDA) and deficiency of a factor called frataxin (FXN) results in a nerve disease affecting coordination and a condition called hypertrophic cardiomyopathy (HCM), marked by an abnormal thickening of the heart. While most of the research in FRDA has focused on nerves and heart muscle, alterations in blood vessels of the heart may worsen the disease in FRDA. But, the role of FXN in these blood vessels has never been defined. Dr. Chan and his team propose that FXN deficiency in FRDA causes DNA damage in lung and heart blood vessel cells to promote heart disease. First, they aim at determining whether FXN deficiency controls damage to genetic material called DNA in blood vessel cells. By studying human blood vessel cells cultured in a dish, this group will artificially manipulate the levels of FXN to determine the effects on DNA damage as well as cell growth and survival. To show the relevance of these findings to FRDA, they will study blood vessel cells originating from stem cells of patients with FRDA, coupled with analyses of heart tissue in FRDA patients. This group also will determine whether FXN deficiency in blood vessels causes heart disease. They have custom generated mice that carry an FXN mutation in blood vessel cells, heart muscle cells, both cell types, or neither. Using these mice, they plan to define the unique roles of FXN in these specific cell types and how they contribute uniquely to heart disease in FRDA. Finally, the team will determine if the drug ABT-263, known to be effective in reversing effects of DNA damage, can be effective in improving heart disease in mice relevant to FRDA.

PI/Investigator: Jennifer Phillips-Cremins, PhD – University of Pennsylvania

Award type: General research grant

Grant Title: Elucidating the link between genome topology and repeat instability in Friedreich's Ataxia

Lay summary: DNA from a single human cell is more than 6 feet long when stretched out end to end. Over the last two decades, scientists have focused on elucidating the sequence of the linear DNA. It is now well established that information encoded in the DNA sequence shapes normal development of human traits, however the mechanisms underlying most diseases remain poorly understood. Recent technological advances have revealed that the 6 feet long DNA sequence is folded into sophisticated 3D configurations that enable it to fit into a nucleus the size of the head of a pin. This lab and others have recently discovered that the DNA's 3D structure is a critical regulator of gene expression patterns essential for normal human development. Cremins and her team have uncovered a striking new link between 3D genome folding and trinucleotide repeat (TNR) expansion disorders, like Friedreich's ataxia (FRDA). They have discovered that nearly all genes that cause TNR disorders are folded into the same unique 3D structure. This result is important because it provides new insight into the locations in the genome that are particularly vulnerable to mutation by incorrect sequence expansion. This group proposes to investigate the role of the 3D Epigenome in FRDA. They will (1) create maps of 3D genome misfolding and linear epigenetic mark alterations in induced pluripotent stem cell-derived neurons and cardiomyocytes from FRDA patients, (2) quantify the FRATAXIN GAA repeat tract length as a function of somatic cell state, and (3) computationally integrate 3D genome and linear epigenetic marks to assess the link to GAA tract length and FRATAXIN expression. Knowledge gained by this work will empower the long-term goal to engineer the 3D genome to reverse gene expression defects in human disease.

PI/Investigator: Seth Masters, PhD - Walter and Eliza Hall Institute, Melbourne Australia

Award type: Award for Innovative Mindset

Grant Title: Targeting neuroinflammation in Friedreich's Ataxia

Lay summary: This grant proposal addresses the role of inflammation in FA. There is evidence of inflammation due to low levels of frataxin in cell lines, in mouse models, and in patients with FA, however mechanistic insights into the trigger for inflammation in FA are missing. This investigator hypothesizes that a specific innate immune pathway drives inflammation in FA. The cellular pathway normally detects virus/bacteria, but is also known to respond to damaged mitochondria. In FA, the damage to mitochondria due to frataxin loss would cause activation of this specific innate immune pathway. The aim of this proposal is to demonstrate mitochondrial damage leading to activation of this innate immune pathway with associated inflammatory biomarkers which may help stratify and direct therapy for FA.

Co-sponsor: CureFA Foundation and fara Australia

PI/Investigator: Giovanni Manfredi, MD, PhD - Weill Cornell Medicine & Helene Puccio, PhD - Institute NeuroMyoGène, Lyon, France

Award type: Award for Innovative Mindset

Grant Title: Mitochondrial integrated stress response in FA cardiomyopathy

Lay summary: The hypothesis of this proposal stems from the observation that death from cardiomyopathy in FA occurs in the third or fourth decade of life, but surprisingly the FA heart often maintains adequate function until shortly before death. This suggests that the FA heart is able to adapt, at least initially, to the defects caused by the loss of frataxin. This adaptation likely involves metabolic rewiring to allow the utilization of alternative energy sources and other adaptive events. The investigators propose that these events recapitulate the “mitochondrial integrated stress response” (well characterized in other mitochondrial diseases) which is an evolutionarily conserved response designed to help the organisms face periods of acute stress, but if chronic and unresolved, becomes “maladaptive”. This proposal aims at determining whether there is evidence of mitochondrial integrated stress response in the heart of an FA mouse model.

Co-sponsor: CureFA Foundation and fara Australia

PI/Investigator: Sanjay Bidichandani, PhD - University of Oklahoma

Award type: Award for Innovative Mindset

Grant Title: Single Cell Gene Expression Analysis in Friedreich Ataxia

Lay summary: This application proposes a new model to explain the (apparently) high residual level of frataxin seen in FA patients. The current understanding is that FA develops when cells express levels of frataxin protein that are 10‐20% of normal levels. Thus, when compared to other loss-of-function conditions, where disease is typically triggered at residual protein levels/activity of <5%, FA seems to manifest at relatively higher residual protein levels. The hypothesis put forward in this grant proposal is that most FA cells actually express very low levels of frataxin (<5%, as in other recessive disorders), and that a substantial minority of cells show “escape from gene silencing” and thus express near normal levels of transcript due to FXN genes that, despite containing an expanded GAA repeat, are not fully silenced. This group plans to test this hypothesis by analyzing FXN expression and silencing in individual cells. This model has important implications for the understanding of FA pathogenesis, and for the development of treatments.

Co-sponsor: CureFA Foundation and fara Australia

PI/Investigator: Katia Aquilano, PhD – University of Rome Tor Vergata, Italy

Award type: General Research Grant

Grant Title: Studying the role of white adipose tissue dysfunction in the development of metabolic complications in Friedreich's ataxia

Lay summary: Patients with Friedreich's ataxia (FRDA) manifest an increased risk of developing type 2 diabetes (T2D). Recent studies have indicated that in addition to showing increased visceral adiposity, patients undergo a low-grade inflammatory state. The expansion of white adipose tissue (WAT) plays a fundamental role in the onset of T2D as it becomes insulin-resistant and a source of inflammatory molecules (adipokines). In this project, this group proposes to characterize WAT at metabolic and immunological level in a murine disease model (KIKO) in order to test whether dysfunction of WAT could be operative. In parallel they will study human primary adipocytes in which FXN deficiency has been induced. A strict interaction exists between gut microbiota and WAT, as some members of the microbial community are able to release molecules (e.g. short chain fatty acids) that exert beneficial effects on WAT physiology. Therefore, in KIKO mice and FRDA patients these investigators will also evaluate gut microbiota composition. Moreover, in plasma samples from FRDA patients and KIKO mice they will analyze key adipokines that have been found altered in T2D patients. Finally, in KIKO mice this group will test the effectiveness of the short-chain fatty acid butyrate in reverting WAT dysfunction and improving T2D-related markers. Performing this research, the hope is to identify WAT as an important player in the setting of metabolic complications typical of FRDA and suggest butyrate as a natural and safe therapeutic tool.

Co-sponsor: fara Australia

PI/Investigator: Henry Houlden, MD, PhD - University College London, UK

Award type: Bronya J. Keats International Research Collaboration Award

Grant Title: Identification of genetic modifiers of Friedreich's ataxia

Lay summary: The GAA expansion length associated with Friedreich ataxia (FRDA) correlates with severity of symptoms and inversely with age of onset, particularly for the shorter allele (GAA1), with a prediction of 2.3 years earlier onset for every 100 GAA repeats added to GAA1. However, the GAA repeat size only accounts for between 36% to 56% of the variation in age of onset and the GAA expansion content has only been investigated for large scale interruptions. This suggests that other contributory factors such as non-coding or coding modifying genetic variation, environmental factors or small repeat sequence interruptions, may influence age of onset and severity. Identifying these factors in FRDA will be important in a number of ways, such as: 1. To define the individual genetic profile of each patient to help determine disease progression, predict clinical problems and understand allele lengths and age at onset, 2. To stratify patients more effectively for treatment trials and 3. The modifying molecular pathways identified will certainly improve our understanding of FRDA and may in themselves be potential therapeutic targets.

This international group collected a large series of FRDA from Europe/South America with DNA and core clinical features. To identify genetic modifying factors in FRDA, the following aims will be carried out:

A. A genome wide association study (GWAS) to identify genetic variants associated with (a) FRDA age at onset (AAO), (b) disease progression in FRDA and (c) investigate the overlap of modifiers associated with other repeat disorders (such as HD and SCAs). This part of project will be funded by Vertex Pharmaceuticals in a joint project with FARA. Vertex will fund genotyping of FRDA in each FRDA patient at University College London, to enable the collaboration with colleagues in the United States (Prof David Lynch and colleagues), where Vertex is currently funding SNP genotyping of their FRDA cases (to be completed in 2020) and enable comparison of data.

B. Identify biological pathways and gene networks influencing FRDA age at onset and severity.

C. Long-read sequencing of the FRDA GAA expansion to assess variation/and repeat changes in the FRDA expansion tract and the 5' and 3' flanking regions of the expansion, we will also carry out PacBio long-read sequencing of 400 FRDA patients.

The group is keen to be in contact with other research groups and clinical teams that are interested to be part of this study: email h.houlden@ucl.ac.uk

PI/Investigator: Richard Possemato, PhD, Assistant Professor - NYU Langone Medical Center

Award type: General Research Grant

Grant Title: Contribution of Ferroptosis to Friedreich's Ataxia

Lay summary: Iron-sulfur clusters (ISCs) are essential protein cofactors, whose dysregulation is linked to a wide range of diseases including Friedreich's Ataxia (FA) [1, 2]. This is because frataxin is required to make ISCs, and because ISCs are an important part of proteins that respond to iron (iron-responsive proteins or IRPs) [3]. Upon loss of the ISC from IRP1, the protein changes shape and enables binding to iron-responsive elements (IREs) in specific genes, resulting in changes in the amounts of protein made. This response is known as the iron-starvation response [4]. This group recently found that cells in which several core components of the ISC machinery, including frataxin, are suppressed are sensitized to a form of oxidative cell death termed ferroptosis [5]. The group's overarching hypothesis is that much of the pathology of FA occurs as a consequence of a chronic hyperactive iron-starvation response. They will test this hypothesis by evaluating the impact of a novel ferroptosis inhibitor in an established mouse model of FA, and by developing novel tools to block the iron starvation response downstream of frataxin inhibition.

Publications:

• Netz DJ, Mascarenhas J, Stehling O, Pierik AJ, Lill R. Maturation of cytosolic and nuclear iron-sulfur proteins. Trends in cell biology. 2014;24(5):303-12.
• Stehling O, Wilbrecht C, Lill R. Mitochondrial iron-sulfur protein biogenesis and human disease. Biochimie. 2014;100:61-77.
• Fleming RE, Ponka P. Iron overload in human disease. The New England journal of medicine. 2012;366(4):348-59.
• Casey JL, Hentze MW, Koeller DM, Caughman SW, Rouault TA, Klausner RD, Harford JB. Iron responsive elements: regulatory RNA sequences that control mRNA levels and translation. Science. 1988;240(4854):924-8.
• Alvarez SW, Sviderskiy VO, Terzi EM, Papagiannakopoulos T, Moreira AL, Adams S, Sabatini DM, Birsoy K, Possemato R. NFS1 undergoes positive selection in lung tumours and protects cells from ferroptosis. Nature. 2017;551(7682):639-43.

PI/Investigator: Jill Napierala, PhD - University of Alabama at Birmingham

Award type: General Research Grant

Grant Title: Defining the pathogenic mechanism of the Frataxin G130V mutation

Lay summary: Friedreich's ataxia (FRDA) is a progressive neurodegenerative disease caused by reduced expression of the mitochondrial protein Frataxin (FXN). Most individuals with FRDA have large expansions of repetitive DNA sequences in both copies of the FXN gene, while some have an expansion in one copy and a missense or nonsense mutation in the other. Both genotypes (homozygous and compound heterozygous) result in reduced levels of mature FXN protein when compared with healthy controls. The most prevalent missense mutation changes one amino acid at position 130 (G130V) in the FXN protein. Individuals with the FXN G130V mutation exhibit clinical features distinct from FRDA individuals with repeat expansions in both copies of FXN, including retained reflexes, preserved sensory responses, spared speech, and slower disease progression. Unexpectedly, a lower amount of FXN protein is detected in samples from FRDA G130V patients than in samples from FRDA patients with two FXN repeat expansions, who often endure a more severe and faster progressing disease. Therefore, the clinical presentation of FRDA G130V patients does not appear to agree with the extremely low levels of mature FXN protein detected in patient samples. This clinical distinction suggests a unique G130V-associated pathogenesis that has not yet been investigated. The FXN protein is shortened from a precursor form (FXN-P) to an intermediate (FXN-I) form, and finally a mature (FXN-M) form. We discovered that the ratio of FXN-G130V-I to FXN-G130V-M is higher than that observed for the FXN-WT protein, which is almost all completely shortened to the mature form (FXN-WT-M). We hypothesize that the G130V mutation impairs this processing of FXN and/or destabilizes the mature form. The unprocessed FXN-G130V-I form is functional and compensates for the decreased amount of FXN-M, thus slowing disease progression in FRDA G130V patients. Little is known regarding the levels, processing, and function of the FXN-G130V protein in cells due to lack of reagents and models that can distinguish the mutant G130V protein from the non-mutated protein produced from the FXN copy with the repeat expansion. We have designed and generated unique patient-derived cell line and mouse models to define the levels and function of FXN-G130V protein in living systems. The data collected from these studies will address whether the G130V mutation provides a protective effect to FRDA G130V patient cells by increasing the ratio of a functional FXN-I form that compensates for loss of the FXN-M form. This discovery could be developed into a therapeutic strategy to benefit all individuals living with FRDA.

PI/Investigator: Massimo Pandolfo, MD - Hôpital Erasme, Belgium

PI/Investigator: Hélène Puccio, PhD - IGBMC, France

Award type: General Research Grant

Grant Title: Inflammation and metabolic changes in the nervous system in Friedreich ataxia: relevance for pathogenesis and identification of biomarkers

Lay summary: The overall objective of this project is to investigate the pathogenic role of metabolic changes and neuroinflammation in Friedreich ataxia (FRDA) neuropathology, and to identify and validate related biomarkers to be used as candidate surrogate outcomes in clinical trials. The neuropathology of FRDA is characterized by marked differences in the vulnerability of neuronal systems. The reason(s) for such specific vulnerabilities are still unknown. Exploring changes in RNA and protein levels, metabolites, and inflammatory markers in different nervous system structures and biofluids from FRDA models may provide clues about pathogenesis and specific vulnerability. Furthermore, data from models can guide the search and allow cross-validation of biomarkers of disease status and/or progression in human patients. We plan to perform unbiased proteomic analysis and focused analysis of metabolic and inflammation markers in plasma and CSF of FRDA patients. Analysis of both plasma and CSF will be performed to dissect contribution from central nervous system (CNS) and peripheral tissues, both affected in FRDA but with different time courses. Data will be cross-validated with findings in two mouse models, , as well as in human induced pluripotent cell (hiPSC)-derived neurons, including proprioceptive neurons.

PI/Investigator: Paola Costantini, PhD – University of Padova, Italy

Award type: General Research Grant

Grant Title: Three-dimensional mature cardiac microtissues from human induced pluripotent stem cells to explore mitochondrial dynamics, cardiac function and therapeutic options in Friedreich Ataxia

Lay summary: Myocardial energy production in significantly impaired in Friedreich ataxia patients and this reduction may even precede the development of tissue damage in the heart. These investigators have recently found that the shape of mitochondria, which is crucial for their function, is altered in isolated Friedreich ataxia patient cells and can be restored by genetically increasing the levels of the mitochondria shaping protein OPA1. In this project they will use an innovative tool, cardiac microtissues or “mini-hearts”, obtained from patient stem cells to investigate 1) if remodeling of the mitochondrial shape can correct cardiac dysfunction and 2) if increasing the levels of frataxin can correct the mitochondrial and cardiac defects. Modeling cardiac dysfunctions in Friedreich ataxia is a crucial and challenging task and cardiac microtissue is an advanced cellular model that will allow to mimic more closely the human myocardium. Defining the molecular features of the disease, in an appropriate cellular context, will facilitate the development of novel therapeutic strategies for Friedreich ataxia.

PI/Investigator: Vania Broccoli, PhD - San Raffaele Scientific Institute and CNR Institute of Neuroscience, Milan, Italy

Award type: General Research Grant

Grant Title: Advancing stem cell-based modeling of the proprioceptive neuronal circuit with dorsal root ganglia organoids and its impairment in Friedreich's ataxia

Lay summary: A major limitation in research on Friedreich's ataxia (FRDA) is the lack of experimental models to study dorsal root ganglia (DRG) sensory neurons. To fill this gap, the Broccoli group generated dorsal root ganglia organoids (DRGOs) by 3D in vitro differentiation of human reprogrammed stem cells (iPSCs). Organoids are cellular structures obtained in vitro where the different cell populations organize in the 3D space, closely recapitulating the organization and geometry of the tissue in vivo. Remarkably, the DRGOs generated by in vitro differentiation of iPSCs shared with native DRGs the expression of a large set of peripheral markers and robust electrophysiological activity. Furthermore, when co-cultured with human intrafusal muscle fibers, DRGO sensory neurons contacted their peripheral targets. Thus, for the first time an ordered circuit between the sensory neurons and their natural peripheral targets was established. Importantly, these investigators generated DRGOs from FRDA patient specific iPSCs and were able to recapitulate several aspects of the pathology including FXN silencing, diminished survival and defects in morphology and impaired formation of muscle spindles. Remarkably, these pathological features were extensively rescued when the entire FXN intron-1 was removed in iPSCs by using the CRISPR/Cas9 technology. This proposal will determine the exact neural cell diversity in DRGOs by single-cell RNA sequencing and establish an assay to evaluate the functional properties of the DRGO neuronal-muscle spindle connection in custom-made microfluidic devices. FRDA DRGOs will then be analyzed for pathophysiological dysfunctions in calcium handling, iron metabolism, inflammatory response and dynamics of mitochondria along axons by advanced imaging in novel microfluidic platforms. Finally, CRISPR/Cas9 corrected FRDA DRGOs will be analyzed to determine the extent of the recovery obtained after gene correction. This program will allow to determine the FRDA pathophysiological roots in a new biological system consisting of DRGO-derived human sensory neurons contacting the muscle cells, thus, modeling one of the neuronal circuits mostly affected in FRDA patients and responsible for gait and motor coordination dysfunctions.

Co-sponsor: FARA Ireland

PI/Investigator: Vijayendran Chandran, PhD - Departments of Pediatrics and Neuroscience, University of Florida, USA

Award type: General Research Grant

Grant Title: Understanding tissue-specific reversibility in Friedreich's ataxia

Lay summary: Friedreich's ataxia (FRDA), the most common inherited ataxia, is caused by recessive mutations that reduce the levels of frataxin (FXN), a mitochondrial iron binding protein. There is no effective treatment for FRDA. The causative basis of FRDA is under-expression of the FXN gene. Testing effective therapies for FRDA has been hindered by a paucity of animal models that faithfully recapitulate human symptoms. The Chandran group recently developed a comprehensive and mechanistically tractable FRDA mouse model (FRDAkd) with inducible and reversible FXN knockdown to study temporal disease progression and/or recovery. Temporal knockdown of FXN in FRDAkd mice causes multiple phenotypic deficits paralleling those observed in humans with FRDA. Strong tissue specificity is observed in both our FRDAkd mouse and in FRDA patients. This proposal is designed to understand which FRDA associated deficits are reversed due to tissue-specific FXN restoration. Genetic approaches will be used to determine if the tissue-specific rescue of FXN knockdown in FRDAkd mice attenuates behavioral and pathological deficits. The results will significantly improve our understanding of the disease mechanism and lay the groundwork for targeted therapies in FRDA.

Publications

• Chandran V, Gao K, Swarup V, Versano R, Dong H, Jordan MC, and Geschwind DH. (2017) "Inducible and reversible phenotypes in a novel mouse model of Friedreich's Ataxia." Elife. 6:e30054.

PI/Investigator: Hélène Puccio PhD, Permanent research director (DR1) at the Institut National de la Santé et de la Recherche Médicale (INSERM). Group leader of the team "Fundamental and pathophysiological mechanisms implicated in ataxia" in the department of Translational Medicine and Neurogenetics at the Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC).

Award type: General Research Grant

Grant Title: Characterization of new humanized mouse model (TG(FXN)YG8Pook/800J) carrying 650-800 GAA repeats and stem cell therapies

Lay summary: Although several mouse models of FA have already been generated, no model to date faithfully reproduces the genetics and the phenotype associated with partial loss of frataxin. A new humanized mouse model carrying 800 GAA repeats within the human frataxin locus and knockout for the endogenous mouse frataxin gene (Tg(FXN)YG8Pook/800J) was recently derived at the Jackson laboratory (from the original Tg(FXN)YG8Pook/200 model from Dr. Pook's laboratory). While not yet characterized, this newly generated Tg(FXN)YG8Pook/800J model lab presents with very low levels of frataxin ubiquitously, based on data shared by Dr. Cat Lutz at the Jackson laboratory. It is genetically the most faithful model to human disease and should allow us to not only better characterize the epigenetic effects of the GAA expansion mutation, but also could be a more faithful model to uncover the pathophysiological mechanism of the disease. The main objective of this proposal is to fully characterized this new model for the FA community.

PI/Investigator: Joriene De Nooij, PhD - Columbia University, USA

Award type: General Research Grant

Grant Title: Modeling Friedreich ataxia in human iPSC-derived sensory neuron subtypes.

Lay summary: Friedreich ataxia (FA) patients present a complex set of clinical features, including ataxia, cardiomyopathy, diabetes mellitus, dysarthria, hearing loss, scoliosis, and visual loss (1-3). Distinctive characteristics of the FA disease phenotype are the progressive limb and gait ataxia and the absence of tendon reflexes (areflexia), symptoms that are consistently observed at early stages of the disease (4,5). The gait ataxia and areflexia correlates with a progressive loss of sensory neurons (SNs) in dorsal root ganglia (DRG) (6-7). Interestingly, while the frataxin gene (Fxn) appears expressed in most DRG neurons (8-10), individual SN subclasses (e.g., mechanoreceptors, proprioceptors or nociceptors) appear differently affected by a loss of Fxn. For example, most if not all FA patients experience a loss in the sense of touch or limb position – senses that are mediated by skin mechanoreceptors and proprioceptors, respectively (8). In contrast, few patients exhibit a reduced sensitivity to pain or temperature - senses associated with nociceptors (8, but see 12). However, exactly which or how each of the specific SNs in DRG are affected by a Fxn deficiency remains poorly understood. This proposal aims to define the molecular pathways that underlie the vulnerability of DRG SNs subtypes to the loss of Fxn. We seek to do this by modeling FA using patient-derived induced pluripotent stem cells (iPSCs) that we differentiate into these distinct SN subtypes. We will use CRISPR/Cas9 gene-editing strategies to generate selective fluorescent labels for these SN subtypes, allowing us to create reporter lines to correctly identify and characterize each subtype in both FA and control SNs. The underlying therapeutic goal of this work is to identify the sensory neuronal differences in FA, and to exploit that knowledge in the development of better treatment strategies.

Lay abstract references:

• Pandolfo M. (2009) "Friedreich ataxia: the clinical picture." J Neurol. 256:Suppl 1:3-8.
• Parkinson MH, Boesch S, Nachbauer W, Mariotti C, Giunti P. (2013) "Clinical features of Friedreich's ataxia: classical and atypical phenotypes." J Neurochem. 126:Suppl 1:103-17.
• Abrahao A, Pedroso JL, Braga-Neto P, Bor-Seng-Shu E, de Carvalho Aguiar P, Barsottini OG. (2015) "Milestones in Friedreich ataxia: more than a century and still learning." Neurogenetics. 16:151-60.
• Harding AE. (1981) "Friedreich's ataxia: a clinical and genetic study of 90 families with an analysis of early diagnostic criteria and intrafamilial clustering of clinical features." Brain. 104:589-620.
• Stephenson J, Zesiewicz T, Gooch C, Wecker L, Sullivan K, Jahan I, Kim SH. (2015) "Gait and balance in adults with Friedreich's ataxia." Gait Posture. 41:603-7.
• Caruso G, Santoro L, Perretti A, Massini R, Pelosi L, Crisci C, Ragno M, Campanella G, Filla A. (1987) "Friedreich's ataxia: electrophysiologic and histologic findings in patients and relatives." Muscle Nerve. 10:503-15.
• Koeppen AH, Morral JA, Davis AN, Qian J, Petrocine SV, Knutson MD, Gibson WM, Cusack MJ, Li D. (2009) "The dorsal root ganglion in Friedreich's ataxia." Acta Neuropathol. 118:763-76
• Jiralerspong S, Liu Y, Montermini L, Stifani S, Pandolfo M. (1997) "Frataxin shows developmentally regulated tissue-specific expression in the mouse embryo." Neurobiol Dis. 4:103-13.
• Koutnikova H, Campuzano V, Foury F, Dollé P, Cazzalini O, Koenig M. (1997) "Studies of human, mouse and yeast homologues indicate a mitochondrial function for frataxin." Nat Genet. 16:345-51.
• Lein ES, Hawrylycz MJ, Ao N, Ayres M, Bensinger A, Bernard A, Boe AF, Boguski MS, Brockway KS, Byrnes EJ, Chen L, Chen L, Chen TM, Chin MC, Chong J, Crook BE, Czaplinska A, Dang CN, Datta S, Dee NR, Desaki AL, Desta T, Diep E, Dolbeare TA, Donelan MJ, Dong HW, Dougherty JG, Duncan BJ, Ebbert AJ, Eichele G, Estin LK, Faber C, Facer BA, Fields R, Fischer SR, Fliss TP, Frensley C, Gates SN, Glattfelder KJ, Halverson KR, Hart MR, Hohmann JG, Howell MP, Jeung DP, Johnson RA, Karr PT, Kawal R, Kidney JM, Knapik RH, Kuan CL, Lake JH, Laramee AR, Larsen KD, Lau C, Lemon TA, Liang AJ, Liu Y, Luong LT, Michaels J, Morgan JJ, Morgan RJ, Mortrud MT, Mosqueda NF, Ng LL, Ng R, Orta GJ, Overly CC, Pak TH, Parry SE, Pathak SD, Pearson OC, Puchalski RB, Riley ZL, Rockett HR, Rowland SA, Royall JJ, Ruiz MJ, Sarno NR, Schaffnit K, Shapovalova NV, Sivisay T, Slaughterbeck CR, Smith SC, Smith KA, Smith BI, Sodt AJ, Stewart NN, Stumpf KR, Sunkin SM, Sutram M, Tam A, Teemer CD, Thaller C, Thompson CL, Varnam LR, Visel A, Whitlock RM, Wohnoutka PE, Wolkey CK, Wong VY, Wood M, Yaylaoglu MB, Young RC, Youngstrom BL, Yuan XF, Zhang B, Zwingman TA, Jones AR. 2007) "Genome-wide atlas of gene expression in the adult mouse brain." Nature 445:168- 176.
• Saunders, P.W. (1913) "Sensory changes in Friedreich's disease." Brain. 36:166.
• Nolano M, Provitera V, Crisci C, Saltalamacchia AM, Wendelschafer-Crabb G, Kennedy WR, Filla A, Santoro L, Caruso G. (2001) "Small fibers involvement in Friedreich's ataxia." Ann Neurol. 50:17-25.
Publications:
• de Nooij JC, Simon CM, Simon A, Doobar S, Steel KP, Banks RW, Mentis GZ, Bewick GS, Jessell TM. (2015) "The PDZ-domain protein Whirlin facilitates mechanosensory signaling in mammalian proprioceptors." J Neurosci. 35:3073-84.
• de Nooij JC, Doobar S, Jessell TM. (2013) "Etv1 inactivation reveals proprioceptor subclasses that reflect the level of NT3 expression in muscle targets." Neuron 77:1055-68.
• Kramer I, Sigrist M, de Nooij JC, Taniuchi I, Jessell TM, Arber S. (2006) "A role for Runx transcription factor signaling in dorsal root ganglion sensory neuron diversification." Neuron. 49:379-93.

PI/Investigator: Jaclyn Tamaroff, MD – Children's Hospital of Philadelphia, USA

Award type: Postdoctoral Fellowship

Grant Title: Mechanisms of diabetes mellitus related to Friedreich's Ataxia

Lay summary: Friedreich's Ataxia (FA) related diabetes affects 5% to 40% of individuals with FA. Diabetes occurs more frequently in individuals with FA than in the general population and does not behave entirely like either “typical” type 1 or type 2 diabetes. FA-related diabetes appears to be associated with worse clinical outcomes, including decreased ability to perform activities of daily living. Even though we have long known about the association between FA and diabetes, there are currently no evidence-supported screening or management guidelines. In order to develop these guidelines, we need to better understand how diabetes develops in the context of FA.

Some studies assessing how the body uses sugar (glucose) have been done in adults with FA. However, similar studies have not yet been done in children. This is important because children with FA-related diabetes present differently than adults. For example, children with FA-related diabetes may present suddenly with severe illness requiring hospitalization. In contrast, some adults with FA-related diabetes may have years of “pre-diabetes” prior to developing overt diabetes.

Dr. Tamaroff plans to enroll twenty children, ages 7 to 17 years, who have FA but are not known to have diabetes in order to better understand how their bodies process glucose. To do this, oral glucose tolerance tests (OGTTs) will be performed and continuous glucose monitors (CGMs) will be placed on patients' upper arms. OGTTs are used in endocrinology clinics to diagnose pre-diabetes or diabetes by evaluating the body's response to drinking a standardized glucose beverage. In a research setting, additional hormones that affect glucose, including insulin, and downstream products of glucose breakdown can be measured. A labeled (stable isotope) glucose in the beverage can also be used to understand how the body uses glucose in even more detail. CGMs add to this information by giving a “real-world” snapshot of what the blood sugar levels are throughout the day and night over two weeks.

Additionally, using registry data, Dr. Tamaroff will study how FA-related diabetes is managed currently. By putting together all of the FA-related diabetes cases, she hopes to learn from what worked and what did not to generate insights for clinicians and advice for individuals with FA-related diabetes. Taken together, her studies will help to develop better strategies for screening, prevention, and treatment of FA-related diabetes.

PI/Investigator: Odelya Pagovich, MD - Weill Cornell Medical College, NY, USA

Award type: General Research Grant

Grant Title: Corneal Confocal Microscopy Quantitative Imaging of Corneal Nerves as a Biomarker of Neurologic Disease in Friedreich's Ataxia

Lay summary: Friedreich's ataxia (FA) is an inherited disease in which neurons lose function over time leading to loss of control of muscles and affecting other organs of the body. The onset and severity of FA varies among affected individuals, so the progression of the disease, and someday, the effectiveness of a cure, must be judged by physicians in individual patients based on semi-quantitative, functional rating scales. While those scales have been highly informative of disease progression, physicians and patients would benefit from an objective, quantitative biomarker that correlated with the progression of the disease. In other neurological diseases, non-invasive examination of the number and structure of nerves in the cornea has been assessed as an indication of disease progression. Recent studies by our group demonstrated that corneal confocal microscopy (CCM) may also provide a biomarker for FA. Abnormal nerve morphology and a reduction in the nerve fiber length and density in the cornea correlated with the severity of the genetic abnormality and the neurologic manifestations of FA. In order to determine whether CCM has the potential to be a biomarker for FA, this proposal will examine whether CCM correlates with the progression of disease over time (aim 1) and whether changes in corneal nerve morphology occur before the clinical manifestations (aim 2). To address these questions, participants enrolled in our earlier study will be invited to return for a second analysis to determine whether the corneal nerves have continued to degrade, and new, young study subjects with genetic confirmation of the disease who have not yet experienced significant disease manifestations will be recruited for an initial assessment and a one year follow-up assessment of their corneal nerves. In summary, this study seeks to evaluate CCM as a tool for physicians and study subjects to follow the onset, progression and ultimately, a potential therapy for FA.

PI/Investigator: Chad Heatwole, MD - University of Rochester Medical Center, NY

Award type: General Research Grant

Grant Title: Developing a Clinically Relevant Disease Specific Patient Reported Outcome Measures for use in Friedreich's Ataxia Therapeutic Trials and FDA Drug Labeling Claims

Lay summary: This research will utilize existing methods and infrastructure to develop and validate disease-specific, patient-reported outcome measures for clinical trials of patients with Friedreich's ataxia. This proposal will shift and refine current research paradigms by producing instruments that will efficiently identify relevant changes in several areas of Friedreich's ataxia patient health. All instruments will be developed and validated in accordance with FDA guidelines for use in drug labeling claims. In addition, input will be obtained from the Friedreich's ataxia research community to optimize the acceptance and use of each of these instruments. These measures will provide researchers with valuable tools to use in clinical trials of pediatric and adult Friedreich's ataxia patients. Although the validation techniques proposed in this study are considered industry standard by many, they have never been implemented on this scale for pediatric and adult Friedreich's ataxia. At the completion of our work, the Friedreich's ataxia research community will have valid and highly responsive outcome measures to aid in therapeutic assessment and therapeutic development in Friedreich's ataxia.

Publications

• Chad Heatwole, Rita Bode, Nicholas Johnson, Jeanne Dekdebrun, Nuran Dilek, Katy Eichinger, James E. Hilbert, Eric Logigian, Elizabeth Luebbe, William Martens, Michael P. McDermott, Shree Pandya, Araya Puwanant, Nan Rothrock, Charles Thornton, Barbara G. Vickrey, David Victorson, Richard T. Moxley, III (2017) The Myotonic Dystrophy Health Index: Correlations with Clinical Tests and Patient Function. Muscle Nerve. Author manuscript; available in PMC 2017 Feb 1
• Heatwole C, Bode R, Johnson N, et al. Patient-reported impact of symptoms in myotonic dystrophy type 1 (PRISM-1) (2012) . Neurology. 79(4):348-357
• Heatwole C, Bode R, Johnson N, et al. (2013) The myotonic dystrophy health index: Initial evaluation of a new outcome measure. Muscle Nerve.

PI/Investigator: David Herrmann, MBBCh – University of Rochester Medical Center, NY

PI/Investigator: Peter Creigh, MD – University of Rochester Medical Center, NY

Award type: General Research Grant

Grant Title: In-Vivo Confocal Imaging of Meissner's Corpuscles as a Biomarker in Friedreich's Ataxia (FA)

Lay summary: This is an extension of an observational study designed to determine the potential role of Meissner Corpuscle (MC) Imaging as a biomarker in FA. In the first phase, 16 FA patients and 16 healthy controls were recruited and studied over 12 months with a series of potential biomarkers. Several of these biomarkers looked promising, and this study is being extended to include an additional 11 participants with FA, and the study will continue for up to 24 months. This is a two-part study. An initial cross-sectional phase assessed the utility of MC imaging, quantitative sensory testing (QST) (touch-pressure, vibration (timed and quantitative) and cold detection thresholds), as biomarkers in FA. These candidate biomarkers will now be assessed longitudinally in a larger cohort of patients and for a longer period in this extension phase. The study will include a screening and baseline visit on the same day and 3 subsequent visits, conducted 6, 12 and 24 months after the initial study visit. The original study included only screening, baseline 6 and 12 month visits, and in this extension a 24 month measurement will be added.

Co-Sponsor: Voyager Therapeutics

PI/Investigator: Jarmon Lees, PhD - St Vincent's Institute of Medical Research, Melbourne, Australia

Award type: Postdoctoral Research Award

Grant Title: Investigating sympathetic nervous dysregulation in the pathogenesis of cardiomyopathy in Friedreich's ataxia

Lay summary: Major hurdles in the development of effective therapies to combat Friedreich's ataxia (FRDA)-associated heart disease include the difficulty of obtaining human heart tissue for study, and animal models of FRDA that do not sufficiently recapitulate the human disease. To this end, disease modelling using human induced pluripotent stem cells (iPSCs) may provide a solution by delivering models that faithfully represent human diseases. iPSCs can multiply indefinitely and be differentiated into any cell type in the body. Hence, FRDA heart tissue which can be made from FRDA-iPSCs, is a promising tool for the study of FRDA heart disease and for the development of new therapies.

Clinical studies of FRDA have reported dysfunction of the heart's autonomic nervous system, the electrical system that controls how fast or slow the heart beats. In FRDA this manifests as a faster resting heart rate, increased noradrenaline production, and a higher incidence of cardiac arrhythmias. It is plausible that both the heart disease and arrhythmias, commonly manifested in FRDA patients, are linked to this dysfunction of the autonomic nervous system. Previous research has focused mainly on the heart muscle (i.e. cardiomyocytes), but has not examined the heart's autonomic nervous system.

In this proposal, we aim to investigate the causes of the heart disease in FRDA by generating cardiomyocytes and autonomic neurons from FRDA-iPSCs to study the development of FRDA-associated heart disease. We will examine the cardiomyocytes and neurons in isolation, and then assess the interaction between the cardiomyocytes and the neurons by growing the cells using our cutting-edge 3D multicellular beating cardiac organoids composed of cardiomyocytes, autonomic neurons and blood vessels. This new model of human heart tissue is a marked improvement over older simplistic models that lack blood vessels and neurons which are essential cellular components in the heart for disease development and the discovery of new effective treatments. This organoid model is an advanced pre-clinical human platform that can faithfully recapitulate heart disease. This proposal aims to unravel the causes of heart disease in FRDA, with a particular focus on the contribution of dysfunctional autonomic neuronal activity. The outcomes of this study will potentially provide a novel target for therapeutic interventions to limit or prevent morbidity and mortality in FRDA.

Co-sponsor: fara Australia

PI/Investigator: Marcondes França Jr, MD, PhD and Thiago de Rezende, MSc – University of Campinas, Brazil

Award type: Postdoctoral Research Award

Grant Title: Cardiac Imaging Biomarkers in Friedreich's Ataxia

Lay summary: Biomarkers are urgently needed to assist in the clinical care and the design of clinical trials for Friedreich´s ataxia (FRDA). Neuroimaging-based parameters have emerged as potential candidates, although further studies are necessary to validate them. In particular, cardiac magnetic resonance imaging (cMRI) has emerged as a promising diagnostic technique. In FRDA, cardiac studies have not received as much research interest as the neurological manifestations, despite heart failure being the main cause of death in such patients. For this reason, further studies in this area are needed to provide relevant information on the natural history and pathophysiology of the disease, in order to identify useful and sensitive biomarkers for clinical trial and follow-up. Therefore, this research proposal centers on the following objectives: 1. To characterize and quantify cardiac damage in pediatric patients with FRDA, patients with long standing FRDA and patients with late-onset FRDA (LOFA); 2. To characterize longitudinal progression of such damage in each of these groups; 3. To investigate whether image parameters correlate with clinical measures, mainly ataxia severity.

PI/Investigator: Gilles Naeije, MD, PhD - Hôpital Erasme, Université libre de Bruxelles, Belgium

Award type: Award for Innovative Mindset

Grant Title: Dentato-Thalamo-Cortical tracts in Friedreich Ataxia: impact of its modulation on Friedreich Ataxia symptoms and brain functional architecture

Lay summary: The cerebellum modulates a wide range of motor and cognitive behaviors thanks to reciprocal connections between the cerebellum and the brain cortex. The main cerebellar output structure is the dentate nuclei that target the brain cortex through the dentato-thalamo-cortical tracts (DTC). Dentate nuclei progressive atrophy and associated DTC impairment are core to the development and progression of Friedreich Ataxia motor and non-motor symptoms. Cerebellar transcranial direct current stimulation (ctDCS) is a non-invasive and clinically friendly technique that may improve DTC functioning. ctDCS has shown efficacy in improving motor and cognitive performances in degenerative ataxia of mixed origins but its mechanisms of action are poorly characterized. The aim of this project is first to assess the potential efficacy of ctDCS to alleviate Friedreich symptoms and second to understand the relationship between DTC and brain functional architecture in Friedreich Ataxia. To do so, ctDCS and its potential clinical benefits evaluation will be combined to non-invasive brain functional imaging investigations of cerebral resting state (i.e., in the absence of any explicit task) activity pre and post ctDCS stimulation, using functional magnetic resonance imaging and magnetoencephalography. Ultimately, this study may provide evidence for an electrophysiological alternative to drug treatment in Friedreich Ataxia and a better understanding of DTC role in motor and cognitive behaviors.

Co-sponsor: fara Australia