Studying in-situ, in-vivo, and in-silico the pleiotropic action of collagen-VI to identify potential late-onset risks in COL6-CMD patients

Awardee: Jeanette Erdmann

Institution: Universität zu Lübeck

Grant Amount: $48,876

Funding Period: February 1, 2022 - January 31, 2023

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Summary:

Due to better healthcare, COL6-CMD patients have a significantly longer life expectancy today than a few decades ago. For future health management of these patients early recognition of potential late-onset disease risks such as aneurysms, cardiovascular, and intestinal diseases can be vital. We will make use of col6a2 KO zebrafish (by morpholino antisense oligonucleotides) to comprehensively study the pleiotropic action of collagen-VI. Moreover, we will leverage human genetic data from UK biobank to identify by phenome-wide association study associations between genetic variants in COL6A2 gene and potential disease risks. Both strategies may help us to identify potential late-onset risks in COL6-CMD patients.

Rescuing oligodendrogenesis and myelination as possible treatment for Pitt-Hopkins patients

Awardee: Simone Mesman

Institution: Swammerdam Institute of Life Sciences, University of Amsterdam

Grant Amount: $78,530

Funding Period: February 1, 2022 - January 31, 2023

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Summary:

In the brain of Pitt-Hopkins patients myelination, fatty sheets surrounding axons of neurons, is affected. The correct construction of these myelin sheets is crucial for proper brain functioning and neuronal communication. Incorrect myelination generally results in affected brain functioning and could be an underlying cause for defects in brain functioning as detected in PTHS. We propose to study myelination under inflence of PTHS-related Tcf4 mutations, by investigating myelination profiles in PTHS patients and by studying the effects of these mutations on oligodendrogenesis, the generation of oligodendrocytes the cell-type that produces myelin.

Multiplex analysis of variant effects for SLC2A1 gene

Awardee: Christina Gurnett

Institution: Washington University, St Louis

Grant Amount: $64,465

Funding Period: February 1, 2022 - January 31, 2023

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Summary:

The goal of this research is to quantitatively determine the functional impact of all possible genetic variants in SLC2A1 and construct algorithms to accurately predict disease onset and severity correlating through calibration with known pathogenic and benign variants. Toward this goal, we propose to employ a high-throughput framework to assess the functional impact of genetic variants in the SLC2A1. We will introduce hundreds of SLC2A1 variants individually into the haploid cell line (HAP1) via multiplex homology-directed-repair (HDR) using CRISPR and a donor library, so that each cell obtains a single variant knocked into the endogenous SLC2A1 gene. Damaging variants that completely disrupt SLC2A1 function (and glucose uptake) will result in impaired cell growth and will drop out of a population of cells sequenced at different time points. We have preliminary data demonstrating the utility of this approach to quantify the functional effects of 15 SLC2A1 variants. We now propose to scale this assay to generate comprehensive, quantitative functional data for the entire SLC2A1 coding region.

Discovery of glycogen synthase inhibitors for validation as a novel therapeutic target for adult polyglucosan body disease (APBD)

Awardee: Wyatt Yue

Institution: Newcastle University, UK

Grant Amount: $99,025

Funding Period: February 1, 2022 - January 31, 2023

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Summary:

Adult polyglucosan body disease is caused by a defective gene encoding glycogen branching enzyme GBE1, resulting in its low activity. The GBE1 enzyme is essential for making glycogen very compact inside the cell, otherwise the glycogen that is being synthesised by another enzyme glycogen synthase GYS1 will form clogging clumps. Drug development programmes for APBD and related diseases have largely sought to deliver an artificial version of the GBE1 gene, or turn down the native GYS1 gene, both emerging gene therapy approaches that remain experimental and costly for the long run. Our vision is to develop a daily pill for APBD patients as a transformative oral therapy. In the first step towards this goal, we aim to develop small molecules that act on the GYS1 enzyme as a drug starting point. To achieve this, we will take advantage of our unique knowledge about the shape of GYS1 enzyme, as well as cutting-edge computational and screening methods to find small molecules.

Discovery of new therapies for hypertrophic cardiomyopathy in RASopathies using an innovative hiPSC-based high-throughput drug screening platform

Awardee: Fabrice Jaffré

Institution: Weill Cornell Medical College

Grant Amount: $75,431

Funding Period: February 1, 2022 - January 31, 2023

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Summary:

Children with RASopathies often present with severe cardiomyopathies and have a 22% mortality rate by the end of the first year of life. Currently, no specific treatment exists for RASopathy children with hypertrophic cardiomyopathy, therefore there is an urgent need to identify novel therapeutic strategies. The overall goal of this proposal is to uncover innovative therapeutic approaches using human induced pluripotent stem cell-derived cardiomyocytes as a RASopathy disease model and as innovative human 2D and 3D high-throughput drug screening platforms. Completion of this proposal will identify therapeutic molecules for RASopathy children with hypertrophic cardiomyopathy at an unparalleled speed.

EXPLOITING NEURONS- AND BRAIN-DERIVED EXOSOMES AS BIOMARKER FOR CDKL5 DEFICIENCY DISORDER

Awardee: Maurizio Giustetto

Institution: Univ. of Torino - Dept. of Neuroscience

Grant Amount: $75,000

Funding Period: February 1, 2022 - January 31, 2023

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Summary:

The identification of objectively measurable parameters is urgently needed to speed up the diagnosis of CDKL5 deficiency disorder (CDD) and to evaluate the outcomes of both pre-clinical and clinical trials. Although exosomes, nanovescicles seceted by all cell types, can be exploited as unbiased, quantitative and non-invasive biomarker for clinical diagnosis, so far, no information is available on the molecular profile of exosomes in CDD. The goal of our proposal is to fill this gap and by establishing and validating a novel molecular, reliable biomarker for CDD patients.

ASO therapy in CDKL5 deficiency disorder

Awardee: Michela Fagiolini, PhD

Institution: Boston Children's Hospital

Award Amount: $150,000

Funding Period: May 1, 2021 - April 31, 2022

Comparison of Brain Biodistribution of Two CDKL5 Gene Therapy Vector Leads

Awardee: Dolan Sondhi, PhD

Institution: Weill Cornell Medicine

Award Amount: $150,000

Funding Period: May 1, 2021 - April 31, 2022

Advancing a mosaic CDKL5 zebrafish model for high-throughput screening approaches

Awardee: Christopher McGraw, MD, PhD

Institution: Boston Children's Hospital

Award Amount: $150,000

Funding Period: May 1, 2021 - April 31, 2022

Defining ciliary signaling and targets in CDKL5 Deficiency Disorder

Awardee: Peter K. Jackson, PhD

Institution: Stanford University School of Medicine

Award Amount: $150,000

Funding Period: May 1, 2021 - April 31, 2022

Impact of Cdkl5 deficiency on cortical dynamics and network architecture

Awardee: Michael J. Higley, MD, PhD

Institution: Yale University

Award Amount: $150,000

Funding Period: May 1, 2021 - April 31, 2022

Systems Analysis of the CDKL5-Deficiency Disorder (CDKL5-DD) Cerebrospinal Fluid Proteome

Awardee: Victor Faundez, MD, PhD

Institution: Emory University

Award Amount: $150,000

Funding Period: May 1, 2021 - April 31, 2022

Spatial profiling of scRNAseq signatures in human parathyroid glands

Awardee: Julie Ann Sosa

Institution: University of California at San Francisco

Grant Amount: $50,000

Funding Period: April 1, 2021 - March 31, 2022

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The objectives of this project were to: (1) utilize transcriptomic methods to define individual cell types within the human parathyroid, and (2) employ digital spatial profiling to visualize the localization of these cell types within the native parathyroid gland architecture. The developmental pilot phase work supported by the grant enabled us to establish a solid foundation of procedural optimization and proof of concept data for scaling our single cell sequencing efforts to a larger, more broadly representative cohort of donor parathyroid glands. 

The scientific objectives completed during the one-year project period are essential for comprehensive mapping of the human parathyroid gland.  The specific landmarks achieved include: demonstration that our live organ procurement work flow preserves tissue viability and maintains intact biochemical function; validation of recovery efficiency, parathyroid marker expression and cellular integrity in suspension; comparative assessment of whole cell vs nuclear isolation for downstream molecular analysis; validation of a novel split-pool sequencing approach that greatly improves capture efficiency, reduces selective recovery bias, and eliminates library construction batch effect concerns; digital spatial profiling of archived normal parathyroid gland sections to demonstrate the capture and whole transcriptome interrogation of specific cellular subsets demarcated by marker gene expression; and the molecular data from these studies showing that the cellular composition and transcriptional profiles of parathyroid gland tissue are dynamic rather than static.  This last finding reveals that the cellular content and biochemical activity of the parathyroid gland may be physiologically conditional, suggesting that functional reconstitution of the parathyroid gland is not a fixed target, but instead requires complementation of adaptive capacity in addition to terminally differentiated cellular phenotypes.  These key data will inform future and ongoing studies to reconstitute native parathyroid gland function.

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Publication:

Digital spatial profiling of human parathyroid tumors reveals cellular and molecular alterations linked to vitamin D deficiency

Chia-Ling Tu, Wenhan Chang, Julie A Sosa, James Koh

PNAS Nexus, Volume 2, Issue 3, March 2023, pgad073

Understanding RASopathies to better inform diagnosis and treatment.

Awardee: Vanessa Fear

Institution: The University of Western Australia

Award Amount: $69,885

Funding Period: February 1, 2021 - January 31, 2022

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Summary:

The RASopathies are a set of syndromes that include cardiofaciocutaneous (CFC) Syndrome, Noonan Syndrome, Noonan Syndrome with lentigines, and Costello Syndrome.  The syndromes are characterised by overlapping disease phenotype and there is a need to distinguish the different RASopathies in order to facilitate accurate patient diagnosis and identify better treatments. In this study we compare changes in patient DNA (genetic variants) that are causative of CFC and Noonan Syndrome. Further, we investigate a potential disease causing patient genetic variant to determine if they have CFC or Noonan Syndrome. The study harnesses gene editing technology to introduce genetic variants into stem cells, which are then matured into nerve cells. The nerve cell maturation process is monitored to identify syndrome-specific changes to inform syndrome classification in the patient, and to provide a better understanding of both CFC and Noonan Syndrome.

Targeting muscle stem cells for the treatment of Collagen VI muscular dystrophies

Awardee: Nicholas Dumont

Institution: CHU Sainte-Justine research center (University of Montreal)

Award Amount: $42,406

Funding Period: February 1, 2021 - January 31, 2022

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Summary:

Mutations in one of the genes encoding for Collagen VI cause Ullrich muscular dystrophy (severe form) or Bethlem myopathy (milder form). These rare genetic diseases are characterized by progressive muscle weakness and degeneration, which can lead to functional incapacities such as impaired or delayed walking. The effect of collagen VI deficiency on muscle degeneration has been characterized; however, its impact on muscle stem cells, the engine of muscle repair, is unknown. Therefore, the overall goal of this project is to investigate if the myogenesis capacity (formation of new muscle tissue) of muscle stem cells is affected by the lack of collagen VI. We will collect samples from patients affected by collagen-VI muscular dystrophies to study muscle stem cell defects in vitro. Moreover, we will use a 3D muscle-in-a-dish system to screen for therapeutic drugs that enhance the myogenesis capacity of muscle stem cells. Overall, this project will provide a better comprehension of this rare muscular disease, and it will open the way to new therapeutic avenues.

Novel tools for drug repurposing in COL6 myopathies: accelerating the route towards therapy.

Awardee: Paolo Bonaldo

Institution: University of Padova, Department of Molecular Medicine

Award Amount: $42,406

Funding Period: February 1, 2021 - January 31, 2022

Global Registry for Inherited Neuropathies (GRIN) - Natural History Study Data Assessment of Charcot-Marie-Tooth (CMT) Patients (age 3-25) to Support Virtual Clinical Studies

Awardee: Florian Thomas

Institution: Hackensack University Medical Center

Award Amount: $55,090

Funding Period: February 1, 2021 - January 31, 2022

Role of CTHRC1 in the Pathogenesis and Therapy of LAM

Awardee: Elizabeth Henske

Institution: BWH

Award Amount: $70,769

Funding Period: February 1, 2021 - January 31, 2022

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Summary:

This project is focused on a protein, CTHRC1 (collagen triple-helix repeat containing 1), that has never before been studied in LAM.  CTHRC1 is a protein that is usually secreted by cells and can be detected in the blood.  In other diseases, CTHRC1 is linked to the rate of cellular growth, and in several types of cancer, a high level of CTHRC1 in the blood is associated with a poor clinical prognosis.  

 In a new line of investigation in our lab, Dr. Nico Alesi has discovered that levels of CTHRC1 are elevated in cellular models of LAM.  CTHRC1 is also increased in human angiomyolipomas and in LAM cells.   Interestingly, levels of CTHRC1 are not suppressed by Rapamycin.  In TSC2-deficient cells, inhibition of CTHRC1 decreases cell growth. 

 These data suggest that CTHRC1 is a newly recognized driver of LAM cell growth.  Because levels of CTHRC1 are not affected by the mTOR inhibitor Rapamycin, CTHRC1 could help to explain why LAM cells are not eliminated during therapy with mTOR inhibitors.  Identifying therapeutic strategies to eliminate LAM cells is a key goal of this work. 

REACT: a reversible knock-out mouse model to explore treatment strategies for the SETBP1 haploinsufficiency disease

Awardee: Rocco Piazza

Institution: University of Milano - Bicocca

Award Amount: $40,373

Funding Period: February 1, 2021 - January 31, 2022

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Summary:

The SETBP1 gene is located on chromosome 18q21.1; it encodes for a protein of 1596 residues with a predicted molecular weight of 170 kD and a predominantly nuclear localization. Genetic abnormalities occurring in the SETBP1 gene are responsible for the onset of two different disorders: 1) SETBP1 haploinsufficiency (SH), a disorder characterized by varying degrees of intellectual disability, developmental as well as speech delays and caused by sub-megabase deletions occurring in SETBP1 locus. 2) Schinzel-Giedion Syndrome (SGS), a rare disease with multiple severe congenital malformations and fatal outcome, caused by de novo, single nucleotide SETBP1 mutations. The pathogenic mechanisms responsible for the onset of SH and SGS are probably tightly connected albeit opposite, as SH is caused by a decrease in SETBP1 protein while SGS is caused by its accumulation. The involvement of the central nervous system in both disorders suggests that SETBP1 itself plays a critical role in this context. Here, we propose to generate and to functionally validate a reversible knock-out mouse model for the SH syndrome. In this model, a blocking cassette flanked with loxP recombination sites would be inserted at intron level in the normal Setbp1 locus by homologous recombination, resulting in a mouse that is unable to express Setbp1 at normal level, therefore mimicking the human SH condition. Then, the usage of specific Cre mouse lines, where the recombinase is either expressed starting from the embryo, only in the adult, or is tamoxifen-inducible, would allow the removal of the blocking cassette and reactivation of the Setbp1 expression at normal levels.

The project herein presented will provide insightful information on the molecular consequences of the reactivation of SETBP1 protein in a knock-out/haploinsufficient model that mimic the SH syndrome. Our new in vivo model will constitute a valuable platform to dissect the molecular mechanisms at the basis of the brain damage following SETBP1 haploinsufficiency and, even more importantly, to study the effect of SETBP1 reactivation at different time-points during the life of the mouse model.

Neuromodulation of prefrontal circuits in a mouse model of SETBP1 disorder

Awardee: Audrey Brumback

Institution: The University of Texas at Austin

Award Amount: $40,373

Funding Period: February 1, 2021 - January 31, 2022