+ Robert Wilson, MD, PhD | Funding period: Feb 1, 2020 - Jan 31, 2022
Drug and drug target validation for Friedreich ataxia
PI/Investigator: Robert Wilson, MD, PhD
– Children's Hospital of PhiladelphiaAward type:
General Research Grant Grant Title:
Drug and drug target validation for Friedreich ataxiaLay 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.
+ Benoit D'Autréaux, PhD | Funding period: Dec 1, 2019 – Nov 30, 2021
Cell-free high throughput screening assays for the discovery of compounds replacing frataxin in FA
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 UniversityAward type:
General Research GrantGrant Title:
Cell-free high throughput screening assays for the discovery of compounds replacing frataxin in FALay 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.
+ Javier Santos, PhD | Funding period: Mar 15, 2019 - Mar 14, 2021
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
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, ArgentinaAward type:
General Research GrantGrant 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 ProteinsLay 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.
+ Joseph Baur, PhD & Shana McCormack, MD | Funding period: Jul 1, 2018 – Jun 30, 2020
NAD+ Precursor Supplementation in Friedreich's Ataxia
PI/Investigator: Joseph Baur, PhD
- Perelman School of Medicine, University of PennsylvaniaPI/Investigator: Shana McCormack, MD
- Children's Hospital of Philadelphia Award type:
Keith Michael Andrus Cardiac Research AwardGrant Title:
NAD+ Precursor Supplementation in Friedreich's AtaxiaLay summary:
Nicotinamide adenine dinucleotide (NAD+) is important for bioenergetic and metabolic processes. NAD+ deficiency has been implicated in heart failure and there is increasing interest restoring its concentration as a therapeutic strategy. Intriguingly, hyperacetylation of mitochondrial proteins is emerging as a consistent feature of failing hearts, and is dramatic in a severe model of FA cardiomyopathy (the cardiac-specific frataxin knockout). The sirtuin SIRT3 is responsible for removing acetylation from many mitochondrial proteins. Thus, Martin et al. (JCI Insight, 2017) hypothesized that supplementing cardiac NAD+ (via the precursor nicotinamide mononucleotide, NMN) might enhance SIRT3 activity, reduce acetylation, and delay heart failure. NMN did indeed improve cardiac function in the heart specific frataxin KO in a SIRT3-dependent manner, but paradoxically did not have a major effect on acetylation. These observations support NAD+ metabolism as a therapeutic target in FA, but raise many important questions about the underlying mechanisms. We propose to look at the effects of NAD+ precursor supplementation in two additional mouse models of FA.
We will also test short-term NMN for tolerability and effects on cardiac bioenergetics in adults with FA without overt heart failure. We will assess the feasibility and safety of short-term (1 week) NAD+ precursor supplementation (NMN, dosing based on studies including NCT03151239) in individuals with FA (n=6) via an open-label study. Although we will primarily focus on feasibility and safety, we will also look for normalization of bioenergetics. Together, these experiments will provide insight and, if promising, guide the design of a longer phase 2/3 interventional study of NAD+ supplementation in FA. Publications
- Martin, A. S. et al. (2017) Nicotinamide mononucleotide requires SIRT3 to improve cardiac function and bioenergetics in a Friedreich's ataxia cardiomyopathy model. JCI Insight 2(14). doi: 10.1172/jci.insight.93885. PubMed PMID: 28724806; PMCID: PMC5518566
+ Diane Ward, PhD | Funding period: Jan 1, 2018 - Jun 30, 2020
Identification of the mechanism of oxidant-mediated Yfh1/frataxin turnover and screen for compounds that suppress the effects of reduced Yfh1/frataxin levels
PI/Investigator: Diane Ward, PhD
- University of Utah, USA Award type:
General Research GrantGrant Title:
Identification of the mechanism of oxidant-mediated Yfh1/frataxin turnover and screen for compounds that suppress the effects of reduced Yfh1/frataxin levels Lay summary:
Friedreich ataxia (FRDA) is a lethal human disorder resulting from mutations in the FXN gene, which give rise to decreased levels of the frataxin protein. The frataxin protein is targeted to the mitochondrial matrix where it plays a critical role in the synthesis of mitochondrial iron-sulfur (Fe-S) clusters. These clusters are essential prosthetic groups involved in oxygen binding and enzymatic activity. Frataxin, like many of the proteins involved in mitochondrial Fe-S cluster synthesis is highly conserved among eukaryotes. The yeast gene YFH1, encodes for the homologue of mammalian frataxin. Loss of YFH1 in yeast results in increased mitochondrial oxidants, increased mitochondrial iron accumulation and decreased Fe-S cluster synthesis. Importantly, overexpression of the human frataxin in yeast complements the loss of Yfh1 and corrects these phenotypes. As such, we utilize yeast as a model system to study FRDA and iron metabolism because of the conserved cellular function and relative ease of genetic manipulation. Using yeast, we discovered that mutation of a yeast gene ERG29 results in defective sterol synthesis, leading to increased levels of sterol intermediates. We found that these intermediates interact with iron generating mitochondrial oxidants, and that the increased mitochondrial oxidants resulted in the loss of Yfh1. Mutant ERG29-induced loss of Yfh1 led to decreased Fe-S cluster synthesis, and ultimately cell death due to decreased mitochondrial respiratory activity. The finding that mitochondrial oxidants affect Fe-S cluster activity through decreased frataxin is not restricted to yeast or to sterol intermediates (1). Together, these results support the hypothesis that conditions that increase mitochondrial oxidants will result in frataxin/Yfh1 degradation. Based on these findings, we propose to determine the mechanism of oxidant-induced loss of frataxin/Yfh1. We will also take advantage of the strong phenotype of Yfh1 and Erg29 knockdown in yeast to screen for compounds that can suppress the effects of loss of Yfh1. Determining the mechanism of how increased mitochondrial oxidants and iron affect frataxin/Yfh1 stability and identifying compounds that suppress the effects of reduced frataxin/Yfh1 levels will guide in identifying efficacious treatments for FRDA and may possibly provide novel treatments for human diseases in which mitochondrial function is compromised.Lay abstract references
- Mouli, S., Nanayakkara, G., AlAlasmari, A., Eldoumani, H., Fu, X., Berlin, A., Lohani, M., Nie, B., Arnold, R. D., Kavazis, A., Smith, F., Beyers, R., Denney, T., Dhanasekaran, M., Zhong, J., Quindry, J. and Amin, R. (2015) "The role of frataxin in doxorubicin-mediated cardiac hypertrophy." Am J Physiol Heart Circ Physiol. 309:H844-859.
- Li, L., Kaplan, J. and Ward, D. M. (2017) "The glucose sensor Snf1 and the transcription factors Msn2 and Msn4 regulate transcription of the vacuolar iron importer gene CCC1 and iron resistance in yeast." J Biol Chem. 292:15577-15586.
- Seguin, A, Takahashi-Makise, N, Yien, YY, Huston, NC, Musso, G, Wallace, JA, Bradley, T, Bergonia, H, Matsumoto, M, Igarashi, K, Phillips, JD, Paw, BH, Kaplan, J and Ward, DM. (2017)
- "Reductions in Abcb10 affect the heme biosynthesis transcriptional profile". J. Biol. Chem. 292:16284-16299.
- Yaish, HM, Farrell, CP, Christensen, RD, MacQueen, BC, Jackson, LK, Trochez-Enciso, J, Kaplan, J, Ward, DM, Salah WK and Phillips, JD. (2017) "Two novel mutations in TMPRSS6 associated with iron-refractory iron deficiency anemia in a mother and child." Blood Cells Mol. Dis. 65:38-40. PMID: 28460265.
+ Jon Watts, PhD | Funding period: Feb 1, 2018 - Jan 31, 2020
Activating frataxin expression in animals using chemically modified oligonucleotides
PI/Investigator: Jon Watts, PhD
- RNA Therapeutics Institute, UMass Medical School, USA Award type:
General Research GrantGrant Title:
Activating frataxin expression in animals using chemically modified oligonucleotidesLay summary:
Friedrich's Ataxia (FA) is an incurable genetic disease caused by insufficient expression of the mitochondrial protein frataxin. This reduced expression is primarily driven by a GAA triplet repeat expansion within the first intron of the frataxin gene, and this mutant expansion leads to formation of an "R-loop" or heterochromatin formation and transcriptional gene silencing . A number of strategies are being developed for treating FA, including approaches to improve mitochondrial function, release epigenetic silencing, or modulate pathways downstream of frataxin . Nevertheless, specific activation of frataxin expression is the ideal therapeutic strategy, addressing the disease at its root cause. Pioneering work by David Corey's laboratory at the University of Texas Southwestern Medical Center showed that frataxin expression could be activated by treating FA patient-derived fibroblasts with oligonucleotides complementary to the expanded repeat . My group recently worked with Dr. Corey's group and others to further explore the chemistry of these repeat-targeted oligonucleotides . The next key step in the development of oligonucleotide therapeutics for FA is their testing in animal models. Gene expression and regulation can differ in one tissue relative to another, and in cultured cells relative to organisms. While animal models of FA are imperfect, it is essential to test whether the frataxin activation observed in cultured cells is robust and reproducible in the more complex context of in vivo use. My laboratory has made progress on optimizing the medicinal chemistry of oligonucleotides for the central nervous system, and we are actively working on methods to improve the potency of oligonucleotides in peripheral tissues including heart. Together with our colleagues at the RNA Therapeutics Institute, we have been working on strategies including conjugation of small molecules and hydrophobic groups, use of advanced sugar-modified nucleotides, and development of multimeric oligonucleotide structures. We aim to deepen the preclinical data on lead oligonucleotide compounds, as well as lay the groundwork for clinical development of these compounds.Grant made possible with support from the Crisp Family FundLay abstract references
- Groh M, Lufino MMP, Wade-Martins R and Gromak N. (2014) "R-loops Associated with Triplet Repeat Expansions Promote Gene Silencing in Friedreich Ataxia and Fragile X Syndrome." PLOS Genetics. 10: e1004318.
- Strawser CJ, Schadt KA and Lynch DR. (2014) "Therapeutic approaches for the treatment of Friedreich's ataxia." Expert Rev. Neurotherapeutics. 14: 949-957.
- Li L, Matsui M and Corey DR. (2016). "Activating frataxin expression by repeat-targeted nucleic acids." Nat Commun. 7: 10606.
- Li L, Shen X, Liu Z, Norrbom M, Prakash TP, O'Reilly D, Sharma VK, Damha MJ, Watts JK, Rigo F and Corey DR. (2018) "Activation of Frataxin Protein Expression by Antisense Oligonucleotides Targeting the Mutant Expanded Repeat." Nucleic Acid Ther., in press.
- Pendergraff HM, Krishnamurthy PM, Debacker AJ, Moazami MP, Sharma VK, Niitsoo L, Yu Y, Tan YN, Haitchi HM, and Watts JK. (2017) "Locked Nucleic Acid Gapmers and Conjugates Potently Silence ADAM33, an Asthma-Associated Metalloprotease with Nuclear-Localized mRNA." Mol Ther Nucleic Acids. 8158-168.
- Khvorova A., Watts J.K., (2017) "The chemical evolution of oligonucleotide therapies of clinical utility." Nat. Biotechnol. 35: 238-248.
+ David Corey, PhD | Funding period: Jul 1, 2016 - Jan 14, 2021
Development of Oligonucleotide Activators of FXN Expression
PI/Investigator: David Corey, PhD
- UT Southwestern, Dallas, Texas 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.
+ Mirella Dottori, PhD | Funding period: Sep 1, 2019 - Aug 31, 2021
Nanoparticle-Mediated Gene Delivery of Frataxin to Neurons
PI/Investigator: Mirella Dottori, PhD
- University of Wollongong, AustraliaAward type:
General Research GrantGrant Title:
Nanoparticle-Mediated Gene Delivery of Frataxin to NeuronsLay 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.
+ Ronald Crystal, MD | Funding period: Oct 1, 2019 - Sep 30, 2020
Gene Therapy for Cardiac Manifestations of Friedreich's Ataxia
PI/Investigator: Ronald Crystal, MD
- Weill Cornell Medical College, NY, USAAward type:
General Research GrantGrant Title:
Gene Therapy for Cardiac Manifestations of Friedreich's AtaxiaLay 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.
+ CRISPR Therapeutics & Marek Napierala, PhD | Funding period: Oct 1, 2017 – Mar 31, 2020
CRISPR / Cas9 mediated deletion of the human FXN intronic trinucleotide repeat as a therapeutic approach for Friedreich's Ataxia
PI/Investigator: Tony Ho, MD
of Crispr Therapeutics PI/Investigator: Marek Napierala, PhD
- University of Alabama at Birmingham Award type: Kyle Bryant Translational Research Award Grant Title: CRISPR / Cas9
mediated deletion of the human FXN intronic trinucleotide repeat as a therapeutic approach for Friedreich's ataxiaLay summary:
Friedreich's ataxia (FA) is an incurable neurodegenerative disease caused most commonly by a pathologic expansion of a naturally occurring short DNA sequence repeat in both copies of the FXN gene. The FXN gene codes for the protein frataxin, which is needed for the proper functioning of mitochondria, the powerhouse of the cell. Having two copies of the FXN gene with expanded repeats significantly reduces the amount of frataxin protein produced and is associated with a number of serious cellular abnormalities. While frataxin protein is produced by most cells in the body, some cells are more sensitive to the degree of frataxin deficiency than others. In particular, neurons in the brain, eye, and spinal cord as well as muscle cells in the heart appear disproportionally vulnerable to damage and degeneration in the setting of reduced FXN protein. Dysfunction in these cells leads to progressive imbalance and discoordination, loss of visual acuity, and life-threatening cardiomyopathy that develops in patients diagnosed with FA. It has been shown that removing the expanded repeat sequences from FXN gene, using different DNA editing platforms (i.e. molecular scissors that can specifically excise the repeat sequences), can consistently restore the production of frataxin protein and prevent degeneration of affected cells (1-2). The CRISPR / Cas9 system, a recently discovered DNA editing strategy, is technically simpler compared to other forms of molecular scissors. In this grant, we propose to use the CRISPR / Cas9 system to remove the expanded repeat sequence from the FXN gene in FA patient-derived stem cells and in a FA mouse model. We will screen for and identify CRISPR reagents that efficiently remove the expanded repeat sequences and will optimize the delivery of these reagents to patient-derived cells and to the FA-model mouse. We will examine the restoration of frataxin protein function after repeat expansion removal. This proof of concept study will enable the generation of CRISPR-based therapeutic reagents and strategies to deliver these reagents to patient cells thereby potentially supporting the development of an effective therapy for FA. Co-sponsor: The Cure FA FoundationLay abstract references
- Li Y, Polak U, Bhalla AD, Rozwadowska N, Butler JS, Lynch DR, Dent SYR, Napierala M. (2015) "Excision of Expanded GAA Repeats Alleviates the Molecular Phenotype of Friedreich's Ataxia." Mol Ther. 23:1055-1065.
- Oullet DL, Cherif K, Rousseau J, and Temblay JP. (2017) "Deletion of the GAA repeats from the human frataxin gene using CRISPR-Cas9 system in YG8R-derived cells and mouse models of Friedreich ataxia." Gene Therapy. 24: 265-274.
+ Stephanie Cherqui, PhD | Funding period: Feb 1, 2018 - Jan 31, 2020
Stem Cell Gene Therapy for Friedreich's Ataxia
PI/Investigator: Stephanie Cherqui, PhD
- University of California, San Diego, USAAward type:
General Research GrantGrant Title:
Stem cell gene therapy for Friedreich's ataxiaLay summary:
Friedreich's ataxia (FRDA) is a multi-systemic autosomal recessive genetic disorder caused by a mutation in the frataxin gene, which reduces the amount of frataxin made by the cell. Frataxin is a mitochondrial protein involved in iron metabolism and is critical for proper mitochondrial function and health. Consistent with the cellular role of frataxin, FRDA is characterized by ataxia, neurodegeneration, muscle weakness, and cardiomyopathy – there is no treatment for this lethal disease. Our lab is interested in the role of inflammation in disease progression, and in the ability to modulate the immune system for therapeutic gain (1-3). Specifically, we study the capacity of hematopoietic stem and progenitor cells (HSPCs) to differentiate into immune cells that can migrate to sites of inflammation in the body and halt the progression of disease. Recently, we tested this therapeutic strategy in the Y8GR mouse model of FRDA (4). This mouse model expresses exclusively the mutated human FXN transgene, thus mimicking the transcriptional deficiency and clinical phenotype seen in FRDA patients. We transplanted normal (i.e., no frataxin mutation), control HSPCs from wild-type mice into 2-month old FRDA YG8R mice and found that this therapy worked quite beyond our expectation. Control HSPCs transplanted in the YG8R mouse differentiated into macrophages that migrated to all sites of inflammation injury, such as the brain, spinal cord, dorsal root ganglia, skeletal muscle and heart of the YG8R mouse. The neurologic and muscular complications in the YG8R mouse were completely prevented as observed 7-months post-transplantation (latest time point tested) after only a single infusion of control HSPCs. As well, it appears that part of the therapeutic gain from this treatment occurs from the transfer of functional frataxin protein from the control HPSC-derived macrophages to the affected tissues in the YG8R mouse. Taken together, we hypothesize that this strategy could also treat FRDA patients. Given the high risk of morbidity and mortality associated with HSPC transplantation from a donor (i.e., allogeneic transplantation), our objective is to develop "an autologous" HSPC gene therapy approach for FRDA, whereby patient cells are extracted, corrected and returned to the patient. To accomplish our objective, we will first develop a gene-correction therapy strategy for FRDA HSPCs and determine their ability of halting disease progression in YG8R mice. Furthermore, we will treat YG8R mice in late-stage disease and determine the ability of HSPCs to reverse preexisting complications. This work represents the first autologous gene-corrected HSPC transplantation treatment strategy for FRDA and builds the foundation for a clinical application of this strategy.Grant made possible with support from the Crisp Family FundLay abstract references
- Harrison F, Yeagy BA, Rocca CJ, Kohn DB, Salomon DR, Cherqui S. (2013) "Hematopoietic stem cell gene therapy in the mouse model of cystinosis." Mol Ther. 21:433-444.
- Naphade S, Sharma J, Gaide Chevronnay HP, Shook MA, Yeagy BA, Rocca CJ, Ur SN, Lau AJ, Courtoy PJ, Cherqui S. (2015) "Lysosomal cross-correction by hematopoietic stem cell-derived macrophages via tunneling nanotubes." Stem Cells. 33:301-309.
- Gaide Chevronnay HP, Jansen V, Van Der Smissen P, Rocca CJ, Liao XH, Refetoff S, Pierreux CE, Cherqui S*, and Courtoy P*. (2016) "Hematopoietic stem cell transplantation can normalize thyroid function in a cystinosis mouse model." Endocrinology. 57:1363-1371 *co-senior author
- Rocca CJ, Goodman SM, Dulin JN, Haquang JH, Gertsman I, Blondelle J, Smith JLM, Heyser CJ, Cherqui S. (2017) "Hematopoietic stem cell transplantation prevents development of Friedreich's Ataxia in a humanized mouse model." Sci Transl Med. 9:eeaj2347.
+ Henry Houlden, MD, PhD | Funding period: May 1, 2020 - Apr 30, 2022
Identification of genetic modifiers of Friedreich's ataxia
Henry Houlden, MD, PhD - University College London, UKAward type:
Bronya J. Keats International Research Collaboration AwardGrant Title:
Identification of genetic modifiers of Friedreich's ataxiaLay 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 email@example.com
+ Richard Possemato, PhD | Funding period: Jul 1, 2019 – Jun 31, 2021
Contribution of Ferroptosis to Friedreich's Ataxia
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) . 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 . 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 . 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.
+ Jill Napierala, PhD | Funding period: Mar 1, 2019 - Feb 28, 2021
Defining the pathogenic mechanism of the Frataxin G130V mutation
PI/Investigator: Jill Napierala, PhD
- University of Alabama at BirminghamAward type:
General Research GrantGrant Title:
Defining the pathogenic mechanism of the Frataxin G130V mutationLay 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.
+ Massimo Pandolfo, MD & Hélène Puccio, PhD | Funding period: Oct 1, 2018 – Sep 30, 2020
Inflammation and metabolic changes in the nervous system in Friedreich ataxia: relevance for pathogenesis and identification of biomarkers
PI/Investigator: Massimo Pandolfo, MD
- Hôpital Erasme, BelgiumPI/Investigator: Hélène Puccio, PhD
- IGBMC, FranceAward type:
General Research GrantGrant Title:
Inflammation and metabolic changes in the nervous system in Friedreich ataxia: relevance for pathogenesis and identification of biomarkersLay 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.
+ Sara Anjomani Virmouni, PhD | Funding period: Apr 1, 2018 – Jun 30, 2020
Elucidation of the metabolic signature of Friedreich's ataxia
PI/Investigator: Sara Anjomani Virmouni, PhD
- Brunel University London & Insitute of Cancer research, UK. Award type:
General Research Grant Grant Title:
Elucidation of the metabolic signature of Friedreich's ataxia Lay summary:
In recent years, there has been a growing interest in the use of metabolomics in neurodegenerative diseases such as Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis. However, insufficient resources have been dedicated to studying alterations in the levels of small molecules, metabolites, and lipids in FRDA. Therefore, we aim to identify unique metabolic signatures of FRDA human and mouse model samples using mass spectrometry–based metabolomics approaches. To our knowledge, the current, novel YG8LR mouse model represents the largest GAA repeat expansion-containing model of all available FRDA mouse models. Due to the presence of the very large hyperexpansions in these mice, similar to those detected in FRDA patients, and also due to the inverse correlation between the GAA repeat size and the disease severity, we propose to employ the YG8LR mice for this project. Our approach to studying the role of metabolic dysfunction in FA disease pathogenesis is threefold: 1) We aim to evaluate the intracellular oxidative stress and mitochondrial health and function in the novel YG8LR transgenic mouse model, and to assess the bioenergetics profiles of FRDA human and mouse cell lines using tools to analyze oxidative metabolism; 2) We aim to investigate the effect of frataxin reduction on metabolic pathways involved in mitochondrial function and energy metabolism in FRDA human and mouse model samples using several highly-sensitive mass spectrometric-based methods; and lastly 3) We aim to delineate the impact of targeting relevant metabolic enzymes to rescue the deficits of central metabolism in FRDA both in vitro and in vivo. We expect that the findings of this proposal will provide a unique opportunity to devise novel therapeutic strategies for FRDA patient diagnosis and treatment through targeting their unique metabolism. Efforts to use metabolic phenotyping as a novel therapeutic approach may not only be limited to FRDA therapeutics, but could be also useful for pharmaceutical companies which are active in developing drugs for different metabolic diseases and neurological disorders. Publications:
- Santoro A, Anjomani Virmouni S, Paradies E, Villalobos V, Al-Mahdawi S, Khoo M, Porcelli V, Vozza A, Perrone M, Denora N, Taroni F, Merla G, Palmieri L, Pook MA, Marobbio CMT: Diazoxide as a novel frataxin-increasing therapy for Friedreich Ataxia. Human Molecular Genetics 2018, ddy016, https://doi.org/10.1093/hmg/ddy016.
- Gupta A, Anjomani-Virmouni S, Koundouros N, Dimitriadi M, Choo-Wing R, Valle A, Zheng Y, Chiu YH, Agnihotri S, Zadeh G, Asara JM, Anastasiou D, Arends MJ, Cantley LC, Poulogiannis G. (2017) "PARK2 Depletion Connects Energy and Oxidative Stress to PI3K/Akt Activation via PTEN S-Nitrosylation." Mol Cell. 65:999-1013.e7.
- Mardakheh FK, Sailem HZ, Kümper S, Tape CJ, McCully RR, Paul A, Anjomani-Virmouni S, Jørgensen C, Poulogiannis G, Marshall CJ, Bakal C. (2016) "Proteomics profiling of interactome dynamics by colocalisation analysis (COLA)." Mol Biosyst. 13:92-105.
- Anjomani-Virmouni S, Al-Mahdawi S, Sandi C, Yasaei H, Giunti P, Slijepcevic P, Pook MA: Identification of telomere dysfunction in Friedreich ataxia. Molecular Neurodegeneration 2015, 10:22.
- Anjomani Virmouni S, Ezzatizadeh V, Sandi C, Sandi M, Al-Mahdawi S, Chutake Y, Pook MA: A novel GAA-repeat-expansion-based mouse model of Friedreich's ataxia. Disease models & mechanisms 2015, 8:225-235.
- Anjomani Virmouni S, Sandi C, Al-Mahdawi S, Pook MA: Cellular, molecular and functional characterisation of YAC transgenic mouse models of Friedreich ataxia. PLoS One 2014, 9:e107416.
+ Jordi Magrane, PhD | Funding period: Sep 1, 2017 - Apr 30, 2020
Functional analysis of primary sensory neurons and (proprio) sensory pathology in Friedreich's ataxia
PI/Investigator: Jordi Magrane, PhD
- Cornell University, NY, USA Award type:
General Research Grant Grant Title:
Functional analysis of primary sensory neurons and (proprio) sensory pathology in Friedreich's ataxia Lay summary:
Friedreich's ataxia (FA) is a physiologically complex disorder that affects several tissues over the course of the disease. Early clinical symptoms in FA patients include areflexia, sensory loss, progressive limb and gait ataxia, and weakness. Impairments in patient's sensory systems consist of a loss of large dorsal root ganglion (DRG) neurons, atrophy of sensory neuron (SN) axons in the dorsal columns, atrophy in the dorsal nucleus of Clarke. Additionally, the patient sensory system exhibits degeneration of the dorsal spinocerebellar tract, corticospinal tract, primary sensory nuclei (cochlear nucleus, lateral geniculate nucleus), and dentate nucleus of the cerebellum. Functionally, defects in conduction velocity along sensory fibers of FA patients have also described. These observations raise the question whether the neuronal pathology in FA results from abnormalities in sensory circuitry. We initially focused on several aspects of mitochondria morphology and dynamics in in vitro and in vivo models of FA, and studied the peripheral nervous system and spinal cords of adult FA mouse models. This proposal aims to study whether reduced frataxin (Fxn) levels have an early impact on DRG SNs, with ambitions to shed light into the progression of sensory loss in FA. Our objectives are: 1) To explore the mechanisms of mitochondrial and neuronal dysfunction in embryonic SNs derived from a FA mouse model; and 2) to investigate early abnormalities in the sensory circuitry by using in vivo and ex vivo imaging.Publications
- Bolea I, Gan WB, Manfredi G, Magrane J (2014) "Imaging of mitochondrial dynamics in motor and sensory axons of living mice." Methods Enzymol, 547:97-110.
- Lin H, Magrane J, Rattelle A, Stepanova A, Galkin A, Clark EM, Dong Y, Halawani SM, and Lynch DR. (2017) "Early cerebellar deficits in mitochondrial biogenesis and respiratory chain complexes in the KIKO mouse model of Friedreich ataxia." Disease Models & Mechanisms. 10:1343-1352.
+ Vania Broccoli, PhD | Funding period: May 1, 2020 - April 30, 2022
Advancing stem cell-based modeling of the proprioceptive neuronal circuit with dorsal root ganglia organoids and its impairment in Friedreich's ataxia
Vania Broccoli, PhD - San Raffaele Scientific Institute and CNR Institute of Neuroscience, Milan, ItalyAward 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 ataxiaLay 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
+ Vijayendran Chandran, PhD | Funding period: Dec 1, 2019 - Nov 30, 2021
Understanding tissue-specific reversibility in Friedreich's ataxia
PI/Investigator: Vijayendran Chandran, PhD
- Departments of Pediatrics and Neuroscience, University of Florida, USAAward type:
General Research Grant Grant Title:
Understanding tissue-specific reversibility in Friedreich's ataxiaLay 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.
+ Hélène Puccio, PhD | Funding period: Sep 1, 2019 – Aug 31, 2021
Characterization of new humanized mouse model (TG(FXN)YG8Pook/800J) carrying 650-800 GAA repeats
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 GrantGrant Title:
Characterization of new humanized mouse model (TG(FXN)YG8Pook/800J) carrying 650-800 GAA repeats and stem cell therapiesLay 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.
+ Joriene De Nooij, PhD | Funding period: Aug 1, 2018 – Jul 31, 2020
Modeling Friedreich Ataxia in human iPSC-derived sensory neuron subtypes.
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.
- 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.
+ Jaclyn Tamaroff, MD | Funding period: Jul 1, 2020 – Jun 30, 2021
Mechanisms of diabetes mellitus related to Friedreich's Ataxia
+ Odelya Pagovich, MD | Funding period: Oct 1, 2019 - Sep 30, 2021
Corneal Confocal Microscopy Quantitative Imaging of Corneal Nerves as a Biomarker of Neurologic Disease in Friedreich's Ataxia
Odelya Pagovich, MD - Weill Cornell Medical College, NY, USAAward type:
General Research GrantGrant Title:
Corneal Confocal Microscopy Quantitative Imaging of Corneal Nerves as a Biomarker of Neurologic Disease in Friedreich's AtaxiaLay 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.
+ David Herrmann, MD & Peter Creigh, MD | Funding period: Sep 1, 2018 – Feb 28, 2021
In-Vivo Confocal Imaging of Meissner's Corpuscles as a Biomarker in Friedreich's Ataxia (FA)
David Herrmann, MBBCh – University of Rochester Medical Center, NYPI/Investigator:
Peter Creigh, MD – University of Rochester Medical Center, NYAward type:
General Research GrantGrant 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
+ Ian Harding, PhD | Funding period: Oct 1, 2018 – Sep 30, 2020
Neuroinflammation in Friedreich Ataxia: Mechanism, Biomarker, and Therapeutic Target
PI/Investigator: Ian Harding, PhD
- Monash University, AustraliaAward type:
General Research GrantGrant Title:
Neuroinflammation in Friedreich Ataxia: Mechanism, Biomarker, and Therapeutic TargetLay summary:
This study aims to establish if in vivo biomarkers of inflammation exist in the brain and spinal cord of individuals with Friedreich ataxia (FRDA). Although clinical presentation and progression are variable in individuals with FRDA, a universal feature is ataxia and loss of motor control secondary to the significant neuropathology that typifies FRDA. Sustained activation of immune-responsive cells in the brain – termed neuroinflammation – may represent one mechanism contributing to this progressive neuropathology. Recent preclinical and post-mortem studies in FRDA report increased inflammatory metabolites and gliosis in the nervous system. Importantly, cell line and animal studies indicate that blocking the inflammatory response in FRDA may ameliorate cell death. The link between chronic neuroinflammation and progressive neurodegeneration has also become increasingly well-establish in other degenerative disorders, including Alzheimer's and Parkinson's diseases. This project will be the first to evaluate in vivo neuroinflammation and its link with measures of neurodegeneration in individuals with FRDA using a novel combination of magnetic resonance imaging (MRI) and positron emission tomography (PET) brain imaging approaches. Given there are currently no treatments that are proven to alter the devastating natural history of FRDA, identifying markers of neuroinflammation and uncovering its role in driving or exacerbating neuropathology in FRDA will be key to improving the understanding of disease mechanisms, tracking disease progression, and identifying and monitoring novel treatment approaches.Co-sponsor: fara Australia and FARA Ireland Publications
- Selvadurai LP, Harding IH, Corben LA, Georgiou-Karistianis N. (2018). Cerebral abnormalities in Friedreich Ataxia: A review. Neuroscience & Biobehavioral Reviews 84: 394-406.
- Harding IH, Corben LA, Delatycki MB, Stagnitti MR, Storey E, Egan GF, Georgiou-Karistianis N. (2017). Cerebral compensation during motor function in Friedreich ataxia: The IMAGE-FRDA study. Movement Disorders 32(8): 1221-1229.
- Selvadurai LP*, Harding IH*, Corben LA, Stagnitti MR, Storey E, Egan GF, Delatycki MB, Georgiou- Karistianis N. (2016). Cerebral and cerebellar grey matter atrophy in Friedreich ataxia: The IMAGE-FRDA study. J Neurology 263(11): 2215-2223. *Equal Contribution
- Harding IH, Raniga P, Delatycki MB, Stagnitti MR, Corben LA, Storey E, Georgiou-Karistianis N, Egan GF. (2016). Tissue atrophy and elevated iron concentration in the extrapyramidal motor system in Friedreich ataxia: The IMAGE-FRDA study. J Neurology Neurosurgery and Psychiatry 87: 1261-1263.
- Harding IH, Corben LA, Storey E, Egan GF, Stagnitti MR, Poudel GR, Delatycki MB, Georgiou- Karistianis N. (2016). Fronto-cerebellar dysfunction and dysconnectivity underlying cognition in Friedreich ataxia: The IMAGE-FRDA study. Human Brain Mapping 37: 338-350.
+ Chad Heatwole, MD | Funding period: Oct 1, 2018 – Sep 30, 2020
Developing a Clinically Relevant Disease Specific Patient Reported Outcome Measures for use in Friedreich's Ataxia Therapeutic Trials and FDA Drug Labeling Claims
PI/Investigator: Chad Heatwole, MD
- University of Rochester Medical Center, NY Award type:
General Research GrantGrant Title:
Developing a Clinically Relevant Disease Specific Patient Reported Outcome Measures for use in Friedreich's Ataxia Therapeutic Trials and FDA Drug Labeling ClaimsLay 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.
For more informational about grants awarded for Outcome Measures & Biomarkers,
please visit our page on the Center of Excellence in FA
+ Marcondes França Jr, MD, PhD and Thiago de Rezende, MSc | Funding period: Jun 1, 2020 - May 31, 2022
Cardiac Imaging Biomarkers in Friedreich's Ataxia
Marcondes França Jr, MD, PhD and Thiago de Rezende, MSc – University of Campinas, BrazilAward type:
Postdoctoral Research AwardGrant Title:
Cardiac Imaging Biomarkers in Friedreich's AtaxiaLay 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.
+ Arnulf H. Koeppen, MD | Funding period: Jan 15, 2019 - Jan 14, 2021
Cytoskeletal, heat shock, and blood vessel proteins in Friedreich cardiomyopathy
PI/Investigator: Arnulf H. Koeppen, MD
- VA Medical Center, AlbanyAward type: Keith Michael Andrus Cardiac Research Award Grant Title:
Cytoskeletal, heat shock, and blood vessel proteins in Friedreich cardiomyopathyLay summary:
Heart failure with or without arrhythmia is the most common cause of death in patients with Friedreich ataxia (FA). However, in patients with modest expansions of guanine-adenine-adenine (GAA) trinucleotide repeats in the shorter allele, heart disease may be entirely absent (Koeppen, et al., 2016). Several different mechanisms may contribute to heart failure in FA, directly and indirectly associated with reduced frataxin. Inflammation may be the principal mechanism underlying fiber necrosis and myocardial scarring (Koeppen, et al., 2015), but it cannot account for cardiomyocyte hypertrophy or the remarkable disorganization of intercalated discs (ICD) (Koeppen, et al., 2016). In the human heart, ICD are fully developed at the age of 6 years (Peters, et al., 1994), and it is unclear how ICD become so chaotic in most cases of FA. Hearts in FA may also be subject to ischemia, but the literature contains little attention to this mechanism. In most cases of FA, cardiomyocytes contain small perinuclear iron-positive granules (Lamarche, et al., 1980).
This study set out to look at the mechanisms of progressive cardiomyopathy in FA patients. Based on previous data collected by the investigator, he hypothesized (a) that frataxin deficiency in mitochondria disables the function of the desmin interactome in the maintenance of normal heart fiber cytoskeleton, including ICD; (b) that frataxin deficiency leads to the addition of extra fragmentary ICD along the course of heart fibers; and (c) that αB-crystallin and desmin aggregation impair iron export from cardiomyocytes. He also proposed that capillary hyperplasia in FA hearts causes ischemia due to "luxury perfusion" and that FA hearts do not respond to oxidative stress in FA as reflected by the unexpected downregulation of HSP72 and HSP27. The objectives of this study are therefore (1) to establish the destruction of the desmin interactome and its effect on ICD by immunohistochemistry and single- and double-label immunofluorescence of multiple heart proteins, including those of the ICD; (2) to provide semiquantitative analysis of these proteins by Western blots with validated antibodies; and (3) to explore the microvascularity of FA hearts by a quantitative morphological method. Publications:
- The significance of intercalated discs in the pathogenesis of Friedreich cardiomyopathy. Koeppen AH, Becker AB, Feustel PJ, Gelman BB, Mazurkiewicz JE. J Neurol Sci. 2016 Aug 15;367:171-6. doi: 10.1016/j.jns.2016.06.006. Epub 2016 Jun 4. PMID: 27423584
- Pathology of Intercalated Discs in Friedreich Cardiomyopathy. Ramirez RL, Becker AB, Mazurkiewicz JE, Feustel PJ, Gelman BB, Koeppen AH. J Am Coll Cardiol. 2015 Oct 13;66(15):1739-40. doi: 10.1016/j.jacc.2015.06.1355.. PMID: 26449146
- The pathogenesis of cardiomyopathy in Friedreich ataxia. Koeppen AH, Ramirez RL, Becker AB, Bjork ST, Levi S, Santambrogio P, Parsons PJ, Kruger PC, Yang KX, Feustel PJ, Mazurkiewicz JE. PLoS One. 2015 Mar 4;10(3):e0116396. doi: 10.1371/journal.pone.0116396. eCollection 2015. .PMID: 25738292