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1
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R S A, R M, Sastri KT, G S M, A R A, V B. Precision medicine advances in cystic fibrosis: Exploring genetic pathways for targeted therapies. Life Sci 2024; 358:123186. [PMID: 39471902 DOI: 10.1016/j.lfs.2024.123186] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2024] [Revised: 10/14/2024] [Accepted: 10/24/2024] [Indexed: 11/01/2024]
Abstract
Personalized medicine has transformed the treatment of cystic fibrosis (CF), providing customized therapeutic approaches based on individual genetic profiles. This review explores the genetic foundations of CF, focusing on mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene and their implications for the development of the disease. The advent of genetic testing has enabled the association of specific mutations to disease severity, leading to the development of CFTR modulators like Ivacaftor, Lumacaftor, and Tezacaftor. Beyond CFTR mutations, genetic modifiers, including gene replacement therapy, genetic manipulation, lentivirus, and non-viral gene therapy formulations, along with environmental factors, play critical roles in influencing disease expression and outcomes. The identification of these modifiers is essential for optimizing therapeutic strategies. Emerging biomarkers, including inflammatory markers and pulmonary function indicators, aid in early disease detection and monitoring progression. Omics technologies are uncovering novel biomarkers, enabling more precise disease management. Pharmacogenomics has become integral to CF care, allowing for personalized approaches that consider genetic variations influencing drug metabolism, especially in antibiotics and anti-inflammatory therapies. The future of CF treatment lies in precision therapies, including CFTR modulators and cutting-edge techniques like gene therapy and CRISPR-Cas9 for mutation correction. As research evolves, these advances can improve patient outcomes while minimizing adverse effects. Ethical considerations and regulatory challenges remain critical as personalized medicine advances, ensuring equitable access and the long-term effectiveness of these innovative therapies.
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Affiliation(s)
- Abinesh R S
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Shivarathreeshwara Nagara, Mysuru, India
| | - Madhav R
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Shivarathreeshwara Nagara, Mysuru, India
| | - K Trideva Sastri
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Shivarathreeshwara Nagara, Mysuru, India.
| | - Meghana G S
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Shivarathreeshwara Nagara, Mysuru, India
| | - Akhila A R
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Shivarathreeshwara Nagara, Mysuru, India
| | - Balamuralidhara V
- Department of Pharmaceutics, JSS College of Pharmacy, JSS Academy of Higher Education & Research, Shivarathreeshwara Nagara, Mysuru, India
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2
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Plasschaert LW, MacDonald KD, Moffit JS. Current landscape of cystic fibrosis gene therapy. Front Pharmacol 2024; 15:1476331. [PMID: 39439894 PMCID: PMC11493704 DOI: 10.3389/fphar.2024.1476331] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2024] [Accepted: 09/26/2024] [Indexed: 10/25/2024] Open
Abstract
Cystic fibrosis is a life-threatening disease that is caused by mutations in CFTR, a gene which encodes an ion channel that supports proper function of several epithelial tissues, most critically the lung. Without CFTR, airway barrier mechanisms are impaired, allowing for chronic, recurrent infections that result in airway remodeling and deterioration of lung structure and function. Small molecule modulators can rescue existing, defective CFTR protein; however, they still leave a subset of people with CF with no current disease modifying treatments, aside from lung transplantation. Gene therapy directed to the lung is a promising strategy to modify CF disease in the organ most associated with morbidity and mortality. It is accomplished through delivery of a CFTR transgene with an airway permissive vector. Despite more than three decades of research in this area, a lung directed gene therapy has yet to be realized. There is hope that with improved delivery vectors, sufficient transduction of airway cells can achieve therapeutic levels of functional CFTR. In order to do this, preclinical programs need to meet a certain level of CFTR protein expression in vitro and in vivo through improved transduction, particularly in relevant airway cell types. Furthermore, clinical programs must be designed with sensitive methods to detect CFTR expression and function as well as methods to measure meaningful endpoints for lung structure, function and disease. Here, we discuss the current understanding of how much and where CFTR needs to be expressed, the most advanced vectors for CFTR delivery and clinical considerations for detecting CFTR protein and function in different patient subsets.
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Affiliation(s)
| | - Kelvin D. MacDonald
- Carbon Biosciences, Waltham, MA, United States
- Department of Pediatrics, Oregon Health and Science University, Portland, OR, United States
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3
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Oliver KE, Carlon MS, Pedemonte N, Lopes-Pacheco M. The revolution of personalized pharmacotherapies for cystic fibrosis: what does the future hold? Expert Opin Pharmacother 2023; 24:1545-1565. [PMID: 37379072 PMCID: PMC10528905 DOI: 10.1080/14656566.2023.2230129] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/17/2023] [Revised: 06/16/2023] [Accepted: 06/23/2023] [Indexed: 06/29/2023]
Abstract
INTRODUCTION Cystic fibrosis (CF), a potentially fatal genetic disease, is caused by loss-of-function mutations in the gene encoding for the CFTR chloride/bicarbonate channel. Modulator drugs rescuing mutant CFTR traffic and function are now in the clinic, providing unprecedented breakthrough therapies for people with CF (PwCF) carrying specific genotypes. However, several CFTR variants are unresponsive to these therapies. AREA COVERED We discussed several therapeutic approaches that are under development to tackle the fundamental cause of CF, including strategies targeting defective CFTR mRNA and/or protein expression and function. Alternatively, defective chloride secretion and dehydration in CF epithelia could be restored by exploiting pharmacological modulation of alternative targets, i.e., ion channels/transporters that concur with CFTR to maintain the airway surface liquid homeostasis (e.g., ENaC, TMEM16A, SLC26A4, SLC26A9, and ATP12A). Finally, we assessed progress and challenges in the development of gene-based therapies to replace or correct the mutant CFTR gene. EXPERT OPINION CFTR modulators are benefiting many PwCF responsive to these drugs, yielding substantial improvements in various clinical outcomes. Meanwhile, the CF therapy development pipeline continues to expand with the development of novel CFTR modulators and alternative therapeutic strategies with the ultimate goal of providing effective therapies for all PwCF in the foreseeable future.
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Affiliation(s)
- Kathryn E. Oliver
- Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia, USA
- Center for Cystic Fibrosis and Airways Disease Research, Emory University and Children’s Healthcare of Atlanta, Atlanta, Georgia, USA
| | - Marianne S. Carlon
- Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE), Department of Chronic Diseases and Metabolism, KU Leuven, Leuven, Belgium
- Center for Molecular Medicine, KU Leuven, Leuven, Belgium
| | | | - Miquéias Lopes-Pacheco
- Biosystems & Integrative Sciences Institute (BioISI), Faculty of Sciences, University of Lisbon, Lisbon, Portugal
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4
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Allen L, Allen L, Carr SB, Davies G, Downey D, Egan M, Forton JT, Gray R, Haworth C, Horsley A, Smyth AR, Southern KW, Davies JC. Future therapies for cystic fibrosis. Nat Commun 2023; 14:693. [PMID: 36755044 PMCID: PMC9907205 DOI: 10.1038/s41467-023-36244-2] [Citation(s) in RCA: 56] [Impact Index Per Article: 28.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2022] [Accepted: 01/20/2023] [Indexed: 02/10/2023] Open
Abstract
We are currently witnessing transformative change for people with cystic fibrosis with the introduction of small molecule, mutation-specific drugs capable of restoring function of the defective protein, cystic fibrosis transmembrane conductance regulator (CFTR). However, despite being a single gene disorder, there are multiple cystic fibrosis-causing genetic variants; mutation-specific drugs are not suitable for all genetic variants and also do not correct all the multisystem clinical manifestations of the disease. For many, there will remain a need for improved treatments. Those patients with gene variants responsive to CFTR modulators may have found these therapies to be transformational; research is now focusing on safely reducing the burden of symptom-directed treatment. However, modulators are not available in all parts of the globe, an issue which is further widening existing health inequalities. For patients who are not suitable for- or do not have access to- modulator drugs, alternative approaches are progressing through the trials pipeline. There will be challenges encountered in design and implementation of these trials, for which the established global CF infrastructure is a major advantage. Here, the Cystic Fibrosis National Research Strategy Group of the UK NIHR Respiratory Translational Research Collaboration looks to the future of cystic fibrosis therapies and consider priorities for future research and development.
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Affiliation(s)
| | | | - Siobhan B Carr
- Royal Brompton & Harefield Hospital, Guy's & St Thomas' Trust, London, UK
- National Heart & Lung Institute, Imperial College London, London, UK
| | - Gwyneth Davies
- UCL Great Ormond Street Institute of Child Health, University College London, London, UK
- Great Ormond Street Hospital for Children, London, UK
| | - Damian Downey
- Wellcome-Wolfson Institute for Experimental Medicine, Queen's University Belfast, Belfast, UK
| | | | - Julian T Forton
- Noah's Ark Children's Hospital for Wales, Cardiff, UK
- School of Medicine, Cardiff University, Cardiff, UK
| | - Robert Gray
- Centre for Inflammation Research, University of Edinburgh, Edinburgh, UK
- Western General Hospital, Edinburgh, UK
| | - Charles Haworth
- Royal Papworth Hospital and Department of Medicine, Cambridge, UK
- University of Cambridge, Cambridge, UK
| | - Alexander Horsley
- Division of Infection, Immunity and Respiratory Medicine, University of Manchester, Manchester, UK
- Manchester Adult CF Centre, Manchester University NHS Foundation Trust, Manchester, UK
| | - Alan R Smyth
- School of Medicine, University of Nottingham, Nottingham, UK
- NIHR Nottingham Biomedical Research Centre, Nottingham, UK
| | - Kevin W Southern
- Department of Women's and Children's Health, University of Liverpool, Liverpool, UK
- Institute in the Park, Alder Hey Children's Hospital, Liverpool, UK
| | - Jane C Davies
- Royal Brompton & Harefield Hospital, Guy's & St Thomas' Trust, London, UK.
- National Heart & Lung Institute, Imperial College London, London, UK.
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Ensinck MM, Carlon MS. One Size Does Not Fit All: The Past, Present and Future of Cystic Fibrosis Causal Therapies. Cells 2022; 11:cells11121868. [PMID: 35740997 PMCID: PMC9220995 DOI: 10.3390/cells11121868] [Citation(s) in RCA: 21] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2022] [Revised: 05/25/2022] [Accepted: 05/28/2022] [Indexed: 02/04/2023] Open
Abstract
Cystic fibrosis (CF) is the most common monogenic disorder, caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Over the last 30 years, tremendous progress has been made in understanding the molecular basis of CF and the development of treatments that target the underlying defects in CF. Currently, a highly effective CFTR modulator treatment (Kalydeco™/Trikafta™) is available for 90% of people with CF. In this review, we will give an extensive overview of past and ongoing efforts in the development of therapies targeting the molecular defects in CF. We will discuss strategies targeting the CFTR protein (i.e., CFTR modulators such as correctors and potentiators), its cellular environment (i.e., proteostasis modulation, stabilization at the plasma membrane), the CFTR mRNA (i.e., amplifiers, nonsense mediated mRNA decay suppressors, translational readthrough inducing drugs) or the CFTR gene (gene therapies). Finally, we will focus on how these efforts can be applied to the 15% of people with CF for whom no causal therapy is available yet.
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Affiliation(s)
- Marjolein M. Ensinck
- Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, 3000 Leuven, Flanders, Belgium;
| | - Marianne S. Carlon
- Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, 3000 Leuven, Flanders, Belgium;
- Laboratory of Respiratory Diseases and Thoracic Surgery (BREATHE), Department of Chronic Diseases and Metabolism, KU Leuven, 3000 Leuven, Flanders, Belgium
- Correspondence:
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6
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Effective viral-mediated lung gene therapy: is airway surface preparation necessary? Gene Ther 2022:10.1038/s41434-022-00332-7. [DOI: 10.1038/s41434-022-00332-7] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2021] [Revised: 03/01/2022] [Accepted: 03/04/2022] [Indexed: 12/20/2022]
Abstract
AbstractGene-based therapeutics are actively being pursued for the treatment of lung diseases. While promising advances have been made over the last decades, the absence of clinically available lung-directed genetic therapies highlights the difficulties associated with this effort. Largely, progress has been hindered by the presence of inherent physical and physiological airway barriers that significantly reduce the efficacy of gene transfer. These barriers include surface mucus, mucociliary action, cell-to-cell tight junctions, and the basolateral cell membrane location of viral receptors for many commonly used gene vectors. Accordingly, airway surface preparation methods have been developed to disrupt these barriers, creating a more conducive environment for gene uptake into the target airway cells. The two major approaches have been chemical and physical methods. Both have proven effective for increasing viral-mediated gene transfer pre-clinically, although with variable effect depending on the specific strategy employed. While such methods have been explored extensively in experimental settings, they have not been used clinically. This review covers the airway surface preparation strategies reported in the literature, the advantages and disadvantages of each method, as well as a discussion about applying this concept in the clinic.
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7
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Anticipating New Treatments for Cystic Fibrosis: A Global Survey of Researchers. J Clin Med 2022; 11:jcm11051283. [PMID: 35268374 PMCID: PMC8911007 DOI: 10.3390/jcm11051283] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2021] [Revised: 01/24/2022] [Accepted: 01/30/2022] [Indexed: 02/04/2023] Open
Abstract
Cystic fibrosis is a life-threatening disease that affects at least 100,000 people worldwide. It is caused by a defect in the cystic fibrosis transmembrane regulator (CFTR) gene and presently, 360 CFTR-causing mutations have been identified. Since the discovery of the CFTR gene, the expectation of developing treatments that can substantially increase the quality of life or even cure cystic fibrosis patients is growing. Yet, it is still uncertain today which developing treatments will be successful against cystic fibrosis. This study addresses this gap by assessing the opinions of over 524 cystic fibrosis researchers who participated in a global web-based survey. For most respondents, CFTR modulator therapies are the most likely to succeed in treating cystic fibrosis in the next 15 years, especially through the use of CFTR modulator combinations. Most respondents also believe that fixing or replacing the CFTR gene will lead to a cure for cystic fibrosis within 15 years, with CRISPR-Cas9 being the most likely genetic tool for this purpose.
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8
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Shteinberg M, Haq IJ, Polineni D, Davies JC. Cystic fibrosis. Lancet 2021; 397:2195-2211. [PMID: 34090606 DOI: 10.1016/s0140-6736(20)32542-3] [Citation(s) in RCA: 389] [Impact Index Per Article: 97.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/15/2020] [Revised: 10/03/2020] [Accepted: 11/20/2020] [Indexed: 12/11/2022]
Abstract
Cystic fibrosis is a monogenic disease considered to affect at least 100 000 people worldwide. Mutations in CFTR, the gene encoding the epithelial ion channel that normally transports chloride and bicarbonate, lead to impaired mucus hydration and clearance. Classical cystic fibrosis is thus characterised by chronic pulmonary infection and inflammation, pancreatic exocrine insufficiency, male infertility, and might include several comorbidities such as cystic fibrosis-related diabetes or cystic fibrosis liver disease. This autosomal recessive disease is diagnosed in many regions following newborn screening, whereas in other regions, diagnosis is based on a group of recognised multiorgan clinical manifestations, raised sweat chloride concentrations, or CFTR mutations. Disease that is less easily diagnosed, and in some cases affecting only one organ, can be seen in the context of gene variants leading to residual protein function. Management strategies, including augmenting mucociliary clearance and aggressively treating infections, have gradually improved life expectancy for people with cystic fibrosis. However, restoration of CFTR function via new small molecule modulator drugs is transforming the disease for many patients. Clinical trial pipelines are actively exploring many other approaches, which will be increasingly needed as survival improves and as the population of adults with cystic fibrosis increases. Here, we present the current understanding of CFTR mutations, protein function, and disease pathophysiology, consider strengths and limitations of current management strategies, and look to the future of multidisciplinary care for those with cystic fibrosis.
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Affiliation(s)
- Michal Shteinberg
- Pulmonology Institute and CF Center, Carmel Medical Center, Haifa, Israel; Rappaport Faculty of Medicine, The Technion-Israel Institute of Technology, Haifa, Israel
| | - Iram J Haq
- Translational and Clinical Research Institute, Newcastle University, Newcastle upon Tyne, UK; Great North Children's Hospital, Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK
| | | | - Jane C Davies
- National Heart and Lung Institute, Imperial College London, London, UK; Royal Brompton and Harefield, Guy's and St Thomas' NHS Foundation Trust, London, UK.
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9
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Maule G, Ensinck M, Bulcaen M, Carlon MS. Rewriting CFTR to cure cystic fibrosis. PROGRESS IN MOLECULAR BIOLOGY AND TRANSLATIONAL SCIENCE 2021; 182:185-224. [PMID: 34175042 DOI: 10.1016/bs.pmbts.2020.12.018] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Cystic fibrosis (CF) is an autosomal recessive monogenic disease caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. Although F508del is the most frequent mutation, there are in total 360 confirmed disease-causing CFTR mutations, impairing CFTR production, function and stability. Currently, the only causal treatments available are CFTR correctors and potentiators that directly target the mutant protein. While these pharmacological advances and better symptomatic care have improved life expectancy of people with CF, none of these treatments provides a cure. The discovery and development of programmable nucleases, in particular CRISPR nucleases and derived systems, rekindled the field of CF gene therapy, offering the possibility of a permanent correction of the CFTR gene. In this review we will discuss different strategies to restore CFTR function via gene editing correction of CFTR mutations or enhanced CFTR expression, and address how best to deliver these treatments to target cells.
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Affiliation(s)
- Giulia Maule
- Department CIBIO, University of Trento, Trento, Italy; Institute of Biophysics, National Research Council, Trento, Italy
| | - Marjolein Ensinck
- Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, Flanders, Belgium
| | - Mattijs Bulcaen
- Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, Flanders, Belgium
| | - Marianne S Carlon
- Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, Flanders, Belgium.
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10
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Parsons D, Donnelley M. Will Airway Gene Therapy for Cystic Fibrosis Improve Lung Function? New Imaging Technologies Can Help Us Find Out. Hum Gene Ther 2020; 31:973-984. [PMID: 32718206 DOI: 10.1089/hum.2020.153] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
The promise of genetic therapies has turned into reality in recent years, with new first-line treatments for fatal diseases now available to patients. The development and testing of genetic therapies for respiratory diseases such as cystic fibrosis (CF) has also progressed. The addition of gene editing to the genetic agent toolbox, and its early success in other organ systems, suggests we will see rapid expansion of gene correction options for CF in the future. Although substantial progress has been made in creating techniques and genetic agents that can be highly effective for CF correction in vitro, physiologically relevant functional in vivo changes have been largely prevented by poor delivery efficiency within the lungs. Somewhat hidden from view, however, is the absence of reliable, accurate, detailed, and noninvasive outcome measures that can detect subtle disease and treatment effects in the lungs of humans or animal models. The ability to measure the fundamental function of the lung-ventilation, the effective transport of air throughout the lung-has been constrained by the available measurement technologies. Without sensitive measurement methods, it is difficult to quantify the effectiveness of genetic therapies for CF. The mainstays of lung health assessment are spirometry, which cannot provide adequate disease localization and is not sensitive enough to detect small early changes in disease; and computed tomography, which provides structural rather than functional information. Magnetic resonance imaging using hyperpolarized gases is increasingly useful for lung ventilation assessment, and it removes the radiation risk that accompanies X-ray methods. A new lung imaging technique, X-ray velocimetry, can now offer highly detailed regional lung ventilation information well suited to the diagnosis, treatment, and monitoring needs of CF lung disease, particularly after the application of genetic therapies. In this review, we discuss the options now available for imaging-based lung function measurement in the generation and use of genetic and other therapies for treating CF lung disease.
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Affiliation(s)
- David Parsons
- Robinson Research Institute, University of Adelaide, Adelaide, Australia.,Adelaide Medical School, University of Adelaide, Adelaide, Australia.,Respiratory and Sleep Medicine, Women's and Children's Hospital, North Adelaide, Australia
| | - Martin Donnelley
- Robinson Research Institute, University of Adelaide, Adelaide, Australia.,Adelaide Medical School, University of Adelaide, Adelaide, Australia.,Respiratory and Sleep Medicine, Women's and Children's Hospital, North Adelaide, Australia
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11
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Erwood S, Laselva O, Bily TM, Brewer RA, Rutherford AH, Bear CE, Ivakine EA. Allele-Specific Prevention of Nonsense-Mediated Decay in Cystic Fibrosis Using Homology-Independent Genome Editing. Mol Ther Methods Clin Dev 2020; 17:1118-1128. [PMID: 32490033 PMCID: PMC7256445 DOI: 10.1016/j.omtm.2020.05.002] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2020] [Accepted: 05/07/2020] [Indexed: 02/07/2023]
Abstract
Nonsense-mediated decay (NMD) is a major pathogenic mechanism underlying a diversity of genetic disorders. Nonsense variants tend to lead to more severe disease phenotypes and are often difficult targets for small molecule therapeutic development as a result of insufficient protein production. The treatment of cystic fibrosis (CF), an autosomal recessive disease caused by mutations in the CFTR gene, exemplifies the challenge of therapeutically addressing nonsense mutations in human disease. Therapeutic development in CF has led to multiple, highly successful protein modulatory interventions, yet no targeted therapies have been approved for nonsense mutations. Here, we have designed a CRISPR-Cas9-based strategy for the targeted prevention of NMD of CFTR transcripts containing the second most common nonsense variant listed in CFTR2, W1282X. By introducing a deletion of the downstream genic region following the premature stop codon, we demonstrate significantly increased protein expression of this mutant variant. Notably, in combination with protein modulators, genome editing significantly increases the potentiated channel activity of W1282X-CFTR in human bronchial epithelial cells. Furthermore, we show how the outlined approach can be modified to permit allele-specific editing. The described approach can be extended to other late-occurring nonsense mutations in the CFTR gene or applied as a generalized approach for gene-specific prevention of NMD in disorders where a truncated protein product retains full or partial functionality.
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Affiliation(s)
- Steven Erwood
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada
| | - Onofrio Laselva
- Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Physiology, University of Toronto, Toronto, ON, Canada
| | - Teija M.I. Bily
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
| | - Reid A. Brewer
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
| | - Alexandra H. Rutherford
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
| | - Christine E. Bear
- Program in Molecular Medicine, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Physiology, University of Toronto, Toronto, ON, Canada
- Department of Biochemistry, University of Toronto, Toronto, ON, Canada
| | - Evgueni A. Ivakine
- Program in Genetics and Genome Biology, The Hospital for Sick Children Research Institute, Toronto, ON, Canada
- Department of Physiology, University of Toronto, Toronto, ON, Canada
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12
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Rang C, Keating D, Wilson J, Kotsimbos T. Re-imagining cystic fibrosis care: next generation thinking. Eur Respir J 2020; 55:13993003.02443-2019. [PMID: 32139465 DOI: 10.1183/13993003.02443-2019] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2019] [Accepted: 02/20/2020] [Indexed: 12/26/2022]
Abstract
Cystic fibrosis (CF) is a common multi-system genetically inherited condition, predominately found in individuals of Caucasian decent. Since the identification of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR) gene in 1989, and the subsequent improvement in understanding of CF pathophysiology, significant increases in life-expectancy have followed. Initially this was related to improvements in the management and systems of care for treating the various affected organ systems. These cornerstone treatments are still essential for CF patients born today. However, over the last decade, the major advance has been in therapies that target the resultant genetic defect: the dysfunctional CFTR protein. Small molecule agents that target this dysfunctional protein via a variety of mechanisms have led to lung function improvements, reductions in pulmonary exacerbation rates and increases in weight and quality-of-life indices. As more patients receive these agents earlier and earlier in life, it is likely that general CF care will increasingly pivot around these specific therapies, although it is also likely that effects other than those identified in the initial trials will be discovered and need to be managed. Despite great excitement for modulator therapies, they are unlikely to be suitable or available for all; whether this is due to a lack of availability for specific CFTR mutations, drug-reactions or the health economic set-up in certain countries. Nevertheless, the CF community must be applauded for its ongoing focus on research and development for this life-limiting disease. With time, personalised individualised therapy would ideally be the mainstay of CF care.
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Affiliation(s)
- Catherine Rang
- Cystic Fibrosis Service, Dept of Respiratory Medicine, Alfred Health, Melbourne, Australia
| | - Dominic Keating
- Cystic Fibrosis Service, Dept of Respiratory Medicine, Alfred Health, Melbourne, Australia.,Dept of Medicine, Monash University, Alfred Campus, Melbourne, Australia
| | - John Wilson
- Cystic Fibrosis Service, Dept of Respiratory Medicine, Alfred Health, Melbourne, Australia.,Dept of Medicine, Monash University, Alfred Campus, Melbourne, Australia
| | - Tom Kotsimbos
- Cystic Fibrosis Service, Dept of Respiratory Medicine, Alfred Health, Melbourne, Australia.,Dept of Medicine, Monash University, Alfred Campus, Melbourne, Australia
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13
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Drumm ML. Gene Editing for CF. Respir Med 2020. [DOI: 10.1007/978-3-030-42382-7_25] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
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Tosco A, Villella VR, Raia V, Kroemer G, Maiuri L. Cystic Fibrosis: New Insights into Therapeutic Approaches. CURRENT RESPIRATORY MEDICINE REVIEWS 2020. [DOI: 10.2174/1573398x15666190702151613] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Since the identification of Cystic Fibrosis (CF) as a disease in 1938 until 2012, only
therapies to treat symptoms rather than etiological therapies have been used to treat the disease. Over
the last few years, new technologies have been developed, and gene editing strategies are now
moving toward a one-time cure. This review will summarize recent advances in etiological therapies
that target the basic defect in the CF Transmembrane Receptor (CFTR), the protein that is mutated in
CF. We will discuss how newly identified compounds can directly target mutated CFTR to improve
its function. Moreover, we will discuss how proteostasis regulators can modify the environment in
which the mutant CFTR protein is synthesized and decayed, thus restoring CFTR function. The
future of CF therapies lies in combinatory therapies that may be personalized for each CF patient.
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Affiliation(s)
- Antonella Tosco
- Department of Translational Medical Sciences, Pediatric Unit, Regional Cystic Fibrosis Center, Federico II University, Naples 80131, Italy
| | - Valeria R. Villella
- Division of Genetics and Cell Biology, European Institute for Research in Cystic Fibrosis, San Raffaele Scientific Institute, Milan 20132, Italy
| | - Valeria Raia
- Department of Translational Medical Sciences, Pediatric Unit, Regional Cystic Fibrosis Center, Federico II University, Naples 80131, Italy
| | - Guido Kroemer
- Equipe11 labellisee Ligue Nationale Contrele Cancer, Centre de Recherche des Cordeliers, Paris, France
| | - Luigi Maiuri
- Division of Genetics and Cell Biology, European Institute for Research in Cystic Fibrosis, San Raffaele Scientific Institute, Milan 20132, Italy
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15
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Li H, Valkenier H, Thorne AG, Dias CM, Cooper JA, Kieffer M, Busschaert N, Gale PA, Sheppard DN, Davis AP. Anion carriers as potential treatments for cystic fibrosis: transport in cystic fibrosis cells, and additivity to channel-targeting drugs. Chem Sci 2019; 10:9663-9672. [PMID: 32055336 PMCID: PMC6984391 DOI: 10.1039/c9sc04242c] [Citation(s) in RCA: 67] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2019] [Accepted: 09/09/2019] [Indexed: 12/12/2022] Open
Abstract
Defective anion transport is a hallmark of the genetic disease cystic fibrosis (CF). One approach to restore anion transport to CF cells utilises alternative pathways for transmembrane anion transport, including artificial anion carriers (anionophores). Here, we screened 22 anionophores for biological activity using fluorescence emission from the halide-sensitive yellow fluorescent protein. Three compounds possessed anion transport activity similar to or greater than that of a bis-(p-nitrophenyl)ureidodecalin previously shown to have promising biological activity. Anion transport by these anionophores was concentration-dependent and persistent. All four anionophores mediated anion transport in CF cells, and their activity was additive to rescue of the predominant disease-causing variant F508del-CFTR using the clinically-licensed drugs lumacaftor and ivacaftor. Toxicity was variable but minimal at the lower end. The results provide further evidence that anionophores, by themselves or together with other treatments that restore anion transport, offer a potential therapeutic strategy for CF.
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Affiliation(s)
- Hongyu Li
- School of Physiology , Pharmacology and Neuroscience , University of Bristol , Biomedical Sciences Building, University Walk , Bristol BS8 1TD , UK .
| | - Hennie Valkenier
- School of Chemistry , University of Bristol , Cantock's Close , Bristol BS8 1TS , UK .
| | - Abigail G Thorne
- School of Chemistry , University of Bristol , Cantock's Close , Bristol BS8 1TS , UK .
| | - Christopher M Dias
- School of Chemistry , University of Bristol , Cantock's Close , Bristol BS8 1TS , UK .
| | - James A Cooper
- School of Chemistry , University of Bristol , Cantock's Close , Bristol BS8 1TS , UK .
| | - Marion Kieffer
- School of Chemistry , University of Bristol , Cantock's Close , Bristol BS8 1TS , UK .
| | | | - Philip A Gale
- Chemistry , University of Southampton , Southampton SO17 1BJ , UK .
| | - David N Sheppard
- School of Physiology , Pharmacology and Neuroscience , University of Bristol , Biomedical Sciences Building, University Walk , Bristol BS8 1TD , UK .
| | - Anthony P Davis
- School of Chemistry , University of Bristol , Cantock's Close , Bristol BS8 1TS , UK .
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16
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Maule G, Casini A, Montagna C, Ramalho AS, De Boeck K, Debyser Z, Carlon MS, Petris G, Cereseto A. Allele specific repair of splicing mutations in cystic fibrosis through AsCas12a genome editing. Nat Commun 2019; 10:3556. [PMID: 31391465 PMCID: PMC6685978 DOI: 10.1038/s41467-019-11454-9] [Citation(s) in RCA: 64] [Impact Index Per Article: 10.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/23/2018] [Accepted: 07/05/2019] [Indexed: 12/19/2022] Open
Abstract
Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the CFTR gene. The 3272-26A>G and 3849+10kbC>T CFTR mutations alter the correct splicing of the CFTR gene, generating new acceptor and donor splice sites respectively. Here we develop a genome editing approach to permanently correct these genetic defects, using a single crRNA and the Acidaminococcus sp. BV3L6, AsCas12a. This genetic repair strategy is highly precise, showing very strong discrimination between the wild-type and mutant sequence and a complete absence of detectable off-targets. The efficacy of this gene correction strategy is verified in intestinal organoids and airway epithelial cells derived from CF patients carrying the 3272-26A>G or 3849+10kbC>T mutations, showing efficient repair and complete functional recovery of the CFTR channel. These results demonstrate that allele-specific genome editing with AsCas12a can correct aberrant CFTR splicing mutations, paving the way for a permanent splicing correction in genetic diseases.
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Affiliation(s)
- Giulia Maule
- Centre for Integrative Biology (CIBIO), University of Trento, Via Sommarive 9, 38123, Trento, Italy
| | - Antonio Casini
- Centre for Integrative Biology (CIBIO), University of Trento, Via Sommarive 9, 38123, Trento, Italy
| | - Claudia Montagna
- Centre for Integrative Biology (CIBIO), University of Trento, Via Sommarive 9, 38123, Trento, Italy
| | - Anabela S Ramalho
- Department of Development and Regeneration, CF Centre, Woman and Child, KU Leuven, Herestraat 49, Leuven, 3000, Belgium
| | - Kris De Boeck
- Department of Development and Regeneration, CF Centre, Woman and Child, KU Leuven, Herestraat 49, Leuven, 3000, Belgium
- Pediatric Pulmonology, Department of Pediatrics, University Hospital Leuven, Herestraat 49, Leuven, 3000, Belgium
| | - Zeger Debyser
- Laboratory for Molecular Virology and Drug Discovery, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, Herestraat 49, Leuven, 3000, Belgium
| | - Marianne S Carlon
- Laboratory for Molecular Virology and Drug Discovery, Department of Pharmaceutical and Pharmacological Sciences, KU Leuven, Herestraat 49, Leuven, 3000, Belgium.
| | - Gianluca Petris
- Centre for Integrative Biology (CIBIO), University of Trento, Via Sommarive 9, 38123, Trento, Italy.
- Medical Research Council Laboratory of Molecular Biology, Cambridge Biomedical Campus, Francis Crick Avenue, Cambridge, CB2 0QH, UK.
| | - Anna Cereseto
- Centre for Integrative Biology (CIBIO), University of Trento, Via Sommarive 9, 38123, Trento, Italy.
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17
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Trandafir LM, Leon MM, Frasinariu O, Baciu G, Dodi G, Cojocaru E. Current Practices and Potential Nanotechnology Perspectives for Pain Related to Cystic Fibrosis. J Clin Med 2019; 8:1023. [PMID: 31336857 PMCID: PMC6678759 DOI: 10.3390/jcm8071023] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2019] [Revised: 07/08/2019] [Accepted: 07/10/2019] [Indexed: 12/31/2022] Open
Abstract
Pain is a complex, multidimensional process that negatively affects physical and mental functioning, clinical outcomes, quality of life, and productivity for cystic fibrosis (CF) patients. CF is an inherited multi-system disease that requires a complete approach in order to evaluate, monitor and treat patients. The landscape in CF care has changed significantly, with currently more adult patients than children worldwide. Despite the great advances in supportive care and in our understanding regarding its pathophysiology, there are still numerous aspects of CF pain that are not fully explained. This review aims to provide a critical overview of CF pain research that focuses on pain assessment, prevalence, characteristics, clinical association and the impact of pain in children and adults, along with innovative nanotechnology perspectives for CF management. Specifically, the paper evaluates the pain symptoms associated with CF and examines the relationship between pain symptoms and disease severity. The particularities of gastrointestinal, abdominal, musculoskeletal, pulmonary and chest pain, as well as pain associated with medical procedures are investigated in patients with CF. Disease-related pain is common for patients with CF, suggesting that pain assessment should be a routine part of their clinical care. A summary of the use of nanotechnology in CF and CF-related pain is also given. Further research is clearly needed to better understand the sources of pain and how to improve patients' quality of life.
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Affiliation(s)
- Laura M Trandafir
- Pediatric Department, Grigore T. Popa University of Medicine and Pharmacy of Iasi, 700115 Iasi, Romania
| | - Magdalena M Leon
- Medical I Department, Grigore T. Popa University of Medicine and Pharmacy of Iasi, 700115 Iasi, Romania
| | - Otilia Frasinariu
- Pediatric Department, Grigore T. Popa University of Medicine and Pharmacy of Iasi, 700115 Iasi, Romania
| | - Ginel Baciu
- Pediatric Department, "Dunărea de Jos" University of Galati, 800008 Galati, Romania
| | - Gianina Dodi
- Advanced Centre for Research-Development in Experimental Medicine, Grigore T. Popa University of Medicine and Pharmacy of Iasi, 700115 Iasi, Romania.
| | - Elena Cojocaru
- Morpho-Functional Sciences Department, Grigore T. Popa University of Medicine and Pharmacy of Iasi, 700115 Iasi, Romania
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18
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Mention K, Santos L, Harrison PT. Gene and Base Editing as a Therapeutic Option for Cystic Fibrosis-Learning from Other Diseases. Genes (Basel) 2019; 10:E387. [PMID: 31117296 PMCID: PMC6562706 DOI: 10.3390/genes10050387] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/15/2019] [Revised: 05/11/2019] [Accepted: 05/15/2019] [Indexed: 12/21/2022] Open
Abstract
Cystic fibrosis (CF) is a monogenic autosomal recessive disorder caused by mutations in the CFTR gene. There are at least 346 disease-causing variants in the CFTR gene, but effective small-molecule therapies exist for only ~10% of them. One option to treat all mutations is CFTR cDNA-based therapy, but clinical trials to date have only been able to stabilise rather than improve lung function disease in patients. While cDNA-based therapy is already a clinical reality for a number of diseases, some animal studies have clearly established that precision genome editing can be significantly more effective than cDNA addition. These observations have led to a number of gene-editing clinical trials for a small number of such genetic disorders. To date, gene-editing strategies to correct CFTR mutations have been conducted exclusively in cell models, with no in vivo gene-editing studies yet described. Here, we highlight some of the key breakthroughs in in vivo and ex vivo gene and base editing in animal models for other diseases and discuss what might be learned from these studies in the development of editing strategies that may be applied to cystic fibrosis as a potential therapeutic approach. There are many hurdles that need to be overcome, including the in vivo delivery of editing machinery or successful engraftment of ex vivo-edited cells, as well as minimising potential off-target effects. However, a successful proof-of-concept study for gene or base editing in one or more of the available CF animal models could pave the way towards a long-term therapeutic strategy for this disease.
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Affiliation(s)
- Karen Mention
- Department of Physiology, University College Cork, Cork T12 K8AF, Ireland.
| | - Lúcia Santos
- Department of Physiology, University College Cork, Cork T12 K8AF, Ireland.
- University of Lisboa Faculty of Sciences, BioISI-Biosystems & Integrative Sciences Institute, 1749-016 Lisboa, Portugal.
| | - Patrick T Harrison
- Department of Physiology, University College Cork, Cork T12 K8AF, Ireland.
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19
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Moses C, Kaur P. Applications of CRISPR systems in respiratory health: Entering a new 'red pen' era in genome editing. Respirology 2019; 24:628-637. [PMID: 30883991 DOI: 10.1111/resp.13527] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2018] [Revised: 02/20/2019] [Accepted: 02/25/2019] [Indexed: 12/16/2022]
Abstract
Respiratory diseases, such as influenza infection, acute tracheal bronchitis, pneumonia, tuberculosis, chronic obstructive pulmonary disease, asthma, lung cancer and nasopharyngeal carcinoma, continue to significantly impact human health. Diseases of the lung and respiratory tract are influenced by environmental conditions and socio-economic factors; however, many of these serious respiratory disorders are also rooted in genetic or epigenetic causes. Clustered regularly interspaced palindromic repeats (CRISPR) and CRISPR-associated (Cas) proteins, isolated from the immune system of prokaryotes, provide a tool to manipulate gene sequences and gene expression with significant implications for respiratory research. CRISPR/Cas systems allow preclinical modelling of causal factors involved in many respiratory diseases, providing new insights into their underlying mechanisms. CRISPR can also be used to screen for genes involved in respiratory processes, development and pathology, identifying novel disease drivers or drug targets. Finally, CRISPR/Cas systems can potentially correct genetic mutations and edit epigenetic marks that contribute to respiratory disorders, providing a form of personalized medicine that could be used in conjunction with other technologies such as stem cell reprogramming and transplantation. CRISPR gene editing is a young field of research, and concerns regarding its specificity, as well as the need for efficient and safe delivery methods, need to be addressed further. However, CRISPR/Cas systems represent a significant step forward for research and therapy in respiratory health, and it is likely we will see the breakthroughs generated from this technology continue.
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Affiliation(s)
| | - Parwinder Kaur
- Faculty of Science, School of Human Sciences and School of Agriculture and Environment, The University of Western Australia, Perth, WA, Australia
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20
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Marangi M, Pistritto G. Innovative Therapeutic Strategies for Cystic Fibrosis: Moving Forward to CRISPR Technique. Front Pharmacol 2018; 9:396. [PMID: 29731717 PMCID: PMC5920621 DOI: 10.3389/fphar.2018.00396] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/02/2018] [Accepted: 04/05/2018] [Indexed: 12/23/2022] Open
Abstract
One of the most revolutionary technologies in recent years in the field of molecular biology is CRISPR-Cas9. CRISPR technology is a promising tool for gene editing that provides researchers the opportunity to easily alter DNA sequences and modify gene function. Its many potential applications include correcting genetic defects, treating and preventing the spread of diseases. Cystic fibrosis (CF) is one of the most common lethal genetic diseases caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Although CF is an old acquaintance, there is still no effective/resolutive cure. Life expectancy has improved thanks to the combination of various treatments, but it is generally below average. Recently, a significant number of additional key medications have become licensed in Europe for the CF treatment including CFTR modulators. But innovative genomically-guided therapies have begun for CF and it is predictable that this will lead to rapid improvements in CF clinical disease and survival in the next decades. In this way, CRISPR-Cas9 approach may represent a valid tool to repair the CFTR mutation and hopeful results were obtained in tissue and animal models of CF disease.
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Affiliation(s)
- Michele Marangi
- Department of Economic Strategy of Pharmaceutical Products, Italian Medicines Agency, Rome, Italy
| | - Giuseppa Pistritto
- Department of Economic Strategy of Pharmaceutical Products, Italian Medicines Agency, Rome, Italy.,Department of Systems Medicine, University of Rome Tor Vergata, Rome, Italy
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21
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Assembly and Functional Analysis of an S/MAR Based Episome with the Cystic Fibrosis Transmembrane Conductance Regulator Gene. Int J Mol Sci 2018; 19:ijms19041220. [PMID: 29673202 PMCID: PMC5979583 DOI: 10.3390/ijms19041220] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2018] [Revised: 04/05/2018] [Accepted: 04/09/2018] [Indexed: 12/24/2022] Open
Abstract
Improving the efficacy of gene therapy vectors is still an important goal toward the development of safe and efficient gene therapy treatments. S/MAR (scaffold/matrix attached region)-based vectors are maintained extra-chromosomally in numerous cell types, which is similar to viral-based vectors. Additionally, when established as an episome, they show a very high mitotic stability. In the present study we tested the idea that addition of an S/MAR element to a CFTR (cystic fibrosis transmembrane conductance regulator) expression vector, may allow the establishment of a CFTR episome in bronchial epithelial cells. Starting from the observation that the S/MAR vector pEPI-EGFP (enhanced green fluorescence protein) is maintained as an episome in human bronchial epithelial cells, we assembled the CFTR vector pBQ-S/MAR. This vector, transfected in bronchial epithelial cells with mutated CFTR, supported long term wt CFTR expression and activity, which in turn positively impacted on the assembly of tight junctions in polarized epithelial cells. Additionally, the recovery of intact pBQ-S/MAR, but not the parental vector lacking the S/MAR element, from transfected cells after extensive proliferation, strongly suggested that pBQ-S/MAR was established as an episome. These results add a new element, the S/MAR, that can be considered to improve the persistence and safety of gene therapy vectors for cystic fibrosis pulmonary disease.
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22
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Harrison PT, Hart S. A beginner's guide to gene editing. Exp Physiol 2018; 103:439-448. [DOI: 10.1113/ep086047] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/09/2017] [Accepted: 12/19/2017] [Indexed: 12/11/2022]
Affiliation(s)
- Patrick T. Harrison
- Department of Physiology, BioSciences Institute; University College Cork; Cork Ireland
| | - Stephen Hart
- UCL Great Ormond Street Institute of Child Health; University College London; London UK
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23
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Nichols DP. Breathing Easier: A Well-tolerated Corrector for F508del. Am J Respir Crit Care Med 2018; 197:152-154. [DOI: 10.1164/rccm.201709-1957ed] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Affiliation(s)
- David P. Nichols
- Seattle Children’s HospitalUniversity of Washington School of MedicineSeattle, Washington
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24
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Ramsey BW, Welsh MJ. AJRCCM: 100-Year Anniversary. Progress along the Pathway of Discovery Leading to Treatment and Cure of Cystic Fibrosis. Am J Respir Crit Care Med 2017; 195:1092-1099. [PMID: 28459323 DOI: 10.1164/rccm.201702-0266ed] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Affiliation(s)
- Bonnie W Ramsey
- 1 Department of Pediatrics University of Washington School of Medicine Seattle, Washington.,2 Center for Clinical and Translational Research Seattle Children's Research Institute Seattle, Washington
| | - Michael J Welsh
- 3 Pappajohn Biomedical Institute.,4 Howard Hughes Medical Institute and.,5 Roy J. and Lucille A. Carver College of Medicine University of Iowa Iowa City, Iowa
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25
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Carlon MS, Vidović D, Birket S. Roadmap for an early gene therapy for cystic fibrosis airway disease. Prenat Diagn 2017; 37:1181-1190. [DOI: 10.1002/pd.5164] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2017] [Revised: 09/12/2017] [Accepted: 09/28/2017] [Indexed: 12/13/2022]
Affiliation(s)
- Marianne S. Carlon
- Molecular Virology and Gene Therapy; Department of Pharmaceutical and Pharmacological Sciences; KU Leuven Flanders Belgium
| | - Dragana Vidović
- Molecular Virology and Gene Therapy; Department of Pharmaceutical and Pharmacological Sciences; KU Leuven Flanders Belgium
- Current affiliation: Cellular Protein Chemistry, Faculty of Science; Utrecht University; The Netherlands
| | - Susan Birket
- Department of Medicine; University of Alabama at Birmingham; Birmingham AL USA
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26
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Abstract
Nucleic acid therapeutics are an established class of drugs that enable specific targeting of a gene of interest. This diverse family of drugs includes antisense oligonucleotides, siRNAs, and mRNA replacement therapies, which can elicit both gene repression and activation, primarily at the RNA level. Recent advances in medicinal chemistry have increased drug potency and enhanced delivery and distribution to a broad array of tissue and cell types. A key advantage of nucleic acid therapeutics is in their application to monogenic diseases. Cystic fibrosis (CF) is one such disease that affects ∼70,000 people globally. This severe disease is an excellent candidate for nucleic acid therapies, as it is due to a genetic defect in a single epithelial chloride channel. Although CF affects many tissues, the primary cause of patient mortality is lung disease. Here we review the various nucleic acid therapeutic modalities and their mechanisms of action, the opportunities and challenges associated with application of nucleic acid drugs to the lung pathology of CF, and the current state and prospects for nucleic acid drugs for the treatment of CF.
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Affiliation(s)
| | - Shuling Guo
- Ionis Pharmaceuticals, Inc. , Carlsbad, California
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27
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Hart SL, Harrison PT. Genetic therapies for cystic fibrosis lung disease. Curr Opin Pharmacol 2017; 34:119-124. [PMID: 29107808 DOI: 10.1016/j.coph.2017.10.006] [Citation(s) in RCA: 26] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2017] [Revised: 10/02/2017] [Accepted: 10/16/2017] [Indexed: 12/26/2022]
Abstract
Gene therapy for cystic fibrosis (CF) has been the subject of intense research over the last twenty-five years or more, using both viral and liposomal delivery methods, but so far without the emergence of a clinical therapy. New approaches to CF gene therapy involving recent improvements to vector systems, both viral and non-viral, as well as new nucleic acid technologies have led to renewed interest in the field. The field of therapeutic gene editing is rapidly developing with the emergence of CRISPR/Cas9 as well as chemically modified mRNA therapeutics. These new types of nucleic acid therapies are also a good fit with delivery by non-viral delivery approaches which has led to a renewed interest in lipid-based and other nanoformulations.
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Affiliation(s)
- Stephen L Hart
- Experimental and Personalised Medicines Section, Genetics and Genomic Medicine Programme, UCL Great Ormond Street Institute of Child Health, 30 Guilford St, London WC1N 1EH, UK.
| | - Patrick T Harrison
- Department of Physiology, BioSciences Institute, University College Cork, Cork, Ireland
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28
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Roesch EA, Drumm ML. Powerful tools for genetic modification: Advances in gene editing. Pediatr Pulmonol 2017; 52:S15-S20. [PMID: 28960896 DOI: 10.1002/ppul.23791] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/31/2017] [Accepted: 07/31/2017] [Indexed: 12/27/2022]
Abstract
Recent discoveries and technical advances in genetic engineering, methods called gene or genome editing, provide hope for repairing genes that cause diseases like cystic fibrosis (CF) or otherwise altering a gene for therapeutic benefit. There are both hopes and hurdles with these technologies, with new ideas emerging almost daily. Initial studies using intestinal organoid cultures carrying the common, F508del mutation have shown that gene editing by CRISPR/Cas9 can convert cells lacking CFTR function to cells with normal channel function, providing a precedent that this technology can be harnessed for CF. While this is an important precedent, the challenges that remain are not trivial. A logistical issue for this and many other genetic diseases is genetic heterogeneity. Approximately, 2000 mutations associated with CF have been found in CFTR, the gene responsible for CF, and thus a feasible strategy that would encompass all individuals affected by the disease is particularly difficult to envision. However, single strategies that would be applicable to all subjects affected by CF have been conceived and are being investigated. With all of these approaches, efficiency (the proportion of cells edited), accuracy (how often other sites in the genome are affected), and delivery of the gene editing components to the desired cells are perhaps the most significant, impending hurdles. Our understanding of each of these areas is increasing rapidly, and while it is impossible to predict when a successful strategy will reach the clinic, there is every reason to believe it is a question of "when" and not "if."
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Affiliation(s)
- Erica A Roesch
- Department of Pediatrics, Case Western Reserve University, Cleveland, Ohio
| | - Mitchell L Drumm
- Department of Pediatrics, Department of Genetics and Genome Sciences, Research Institute for Children's Health, Case Western Reserve University, Cleveland, Ohio
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29
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Guimbellot J, Sharma J, Rowe SM. Toward inclusive therapy with CFTR modulators: Progress and challenges. Pediatr Pulmonol 2017; 52:S4-S14. [PMID: 28881097 PMCID: PMC6208153 DOI: 10.1002/ppul.23773] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/04/2017] [Accepted: 06/29/2017] [Indexed: 12/29/2022]
Abstract
Cystic fibrosis is caused by gene mutations that result in an abnormal Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein on the surface of cells. CFTR modulators are a novel class of drugs that directly target the molecular defect. CFTR modulators include potentiators that result in improved activity of the channel; correctors that help the protein traffic to the cell surface properly; and readthrough agents that restore full-length CFTR by suppression of premature termination codons, among other novel classes more recently established. While some of these drugs, CFTR potentiators in particular, have provided remarkable improvements for CF patients, others have yet to achieve profoundly improved outcomes, and many CF patients are not yet impacted by CFTR modulators due to lack of knowledge regarding susceptibility of their mutations to treatment. One limitation to expanding these types of therapies to the maximum number of patients with CF is the lack of rigorously validated clinical biomarkers that can determine efficacy on an individual basis, as well as few pre-clinical tools that can predict whether an individual with a rare combination of mutant alleles will respond to a particular CFTR modulator regimen. In this review, we discuss the various groups of CFTR modulators and their status in clinical development, as well as address the current literature on biomarkers, pre-clinical cell-based tools, and the role of pharmacometrics in creating therapeutic strategies to improve the lives of all patients with cystic fibrosis, regardless of their specific mutation.
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Affiliation(s)
- Jennifer Guimbellot
- Departments of Pediatrics, The University of Alabama at Birmingham, Birmingham, Alabama
- Gregory Fleming James Cystic Fibrosis Research Center, The University of Alabama at Birmingham, Birmingham, Alabama
| | - Jyoti Sharma
- Gregory Fleming James Cystic Fibrosis Research Center, The University of Alabama at Birmingham, Birmingham, Alabama
- Cell Developmental and Integrative Biology, The University of Alabama at Birmingham, Birmingham, Alabama
| | - Steven M. Rowe
- Departments of Pediatrics, The University of Alabama at Birmingham, Birmingham, Alabama
- Gregory Fleming James Cystic Fibrosis Research Center, The University of Alabama at Birmingham, Birmingham, Alabama
- Cell Developmental and Integrative Biology, The University of Alabama at Birmingham, Birmingham, Alabama
- Departments of Medicine, The University of Alabama at Birmingham, Birmingham, Alabama
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30
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Sanz DJ, Hollywood JA, Scallan MF, Harrison PT. Cas9/gRNA targeted excision of cystic fibrosis-causing deep-intronic splicing mutations restores normal splicing of CFTR mRNA. PLoS One 2017; 12:e0184009. [PMID: 28863137 PMCID: PMC5581164 DOI: 10.1371/journal.pone.0184009] [Citation(s) in RCA: 65] [Impact Index Per Article: 8.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2017] [Accepted: 08/16/2017] [Indexed: 12/27/2022] Open
Abstract
Cystic Fibrosis is an autosomal recessive disorder caused by mutations in the CFTR gene. CRISPR mediated, template-dependent homology-directed gene editing has been used to correct the most common mutation, c.1521_1523delCTT / p.Phe508del (F508del) which affects ~70% of individuals, but the efficiency was relatively low. Here, we describe a high efficiency strategy for editing of three different rare CFTR mutations which together account for about 3% of individuals with Cystic Fibrosis. The mutations cause aberrant splicing of CFTR mRNA due to the creation of cryptic splice signals that result in the formation of pseudoexons containing premature stop codons c.1679+1634A>G (1811+1.6kbA>G) and c.3718-2477C>T (3849+10kbC>T), or an out-of-frame 5' extension to an existing exon c.3140-26A>G (3272-26A>G). We designed pairs of Cas9 guide RNAs to create targeted double-stranded breaks in CFTR either side of each mutation which resulted in high efficiency excision of the target genomic regions via non-homologous end-joining repair. When evaluated in a mini-gene splicing assay, we showed that targeted excision restored normal splicing for all three mutations. This approach could be used to correct aberrant splicing signals or remove disruptive transcription regulatory motifs caused by deep-intronic mutations in a range of other genetic disorders.
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Affiliation(s)
- David J. Sanz
- Department of Physiology, BioSciences Institute, University College Cork, Cork, Ireland
| | - Jennifer A. Hollywood
- Department of Physiology, BioSciences Institute, University College Cork, Cork, Ireland
- School of Microbiology, University College Cork, Cork, Ireland
| | | | - Patrick T. Harrison
- Department of Physiology, BioSciences Institute, University College Cork, Cork, Ireland
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von Dossow V, Costa J, D'Ovidio F, Marczin N. Worldwide trends in heart and lung transplantation: Guarding the most precious gift ever. Best Pract Res Clin Anaesthesiol 2017; 31:141-152. [PMID: 29110788 DOI: 10.1016/j.bpa.2017.08.001] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2017] [Accepted: 08/03/2017] [Indexed: 01/17/2023]
Abstract
Transplantation is sadly a therapy to die for. The survival of a recipient with end-stage heart or lung disease requires the demise of a human being through brain death or cessation of circulation, with the noblest final act of offering one's organs to another. However, transplantation is constrained by severe hemodynamic, regulatory, inflammatory, and metabolic stresses in the donor, rendering the majority of offered organs unsuitable for transplantation. Coupled with our inability to acquire exact molecular and cellular information and missed opportunities for effectively modulating deteriorations of donors and allografts, anesthesia and critical care contributes to ongoing organ shortages. Progress is made with improving waiting lists by bridging patients for transplantation using mechanical support. However, this represents more complex recipients, higher risk transplant operations, and increased resource utilization. The advent of ex vivo perfusion allows implementing novel diagnostic and therapeutic strategies with real potential of reconditioning less ideal organs. This review advocates a paradigm change in critical care management of the potential donor for improving retrieval practices and for more intellectual involvement of our specialties in organ preservation, ex vivo evaluation and reconditioning, and the need for great advancement in our efficiency in converting unacceptable allografts to suitable donor organs.
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Affiliation(s)
- Vera von Dossow
- Department of Anesthesiology, Ludwig-Maximilians-University of Munich, Germany
| | - Joseph Costa
- Department of Surgery, Division of Cardiothoracic Surgery and Transplantation, Columbia University Medical Center, New York, NY, USA
| | - Frank D'Ovidio
- Department of Surgery, Division of Cardiothoracic Surgery and Transplantation, Columbia University Medical Center, New York, NY, USA
| | - Nandor Marczin
- Section of Anaesthetics, Pain Medicine and Intensive Care, Department of Surgery and Cancer, Faculty of Medicine, Imperial College London, London, UK; Department of Anaesthesia, Royal Brompton and Harefield NHS Foundation Trust, Harefield Hospital, Harefield, Middlesex, UK; Centre of Anaesthesia and Intensive Care, Semmelweis University, Budapest, Hungary.
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Marson FAL, Bertuzzo CS, Ribeiro JD. Personalized or Precision Medicine? The Example of Cystic Fibrosis. Front Pharmacol 2017; 8:390. [PMID: 28676762 PMCID: PMC5476708 DOI: 10.3389/fphar.2017.00390] [Citation(s) in RCA: 44] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2017] [Accepted: 06/02/2017] [Indexed: 01/01/2023] Open
Abstract
The advent of the knowledge on human genetics, by the identification of disease-associated variants, culminated in the understanding of human variability. With the genetic knowledge, the specificity of the clinical phenotype and the drug response of each individual were understood. Using the cystic fibrosis (CF) as an example, the new terms that emerged such as personalized medicine and precision medicine can be characterized. The genetic knowledge in CF is broad and the presence of a monogenic disease caused by mutations in the CFTR gene enables the phenotype–genotype association studies (including the response to drugs), considering the wide clinical and laboratory spectrum dependent on the mutual action of genotype, environment, and lifestyle. Regarding the CF disease, personalized medicine is the treatment directed at the symptoms, and this treatment is adjusted depending on the patient’s phenotype. However, more recently, the term precision medicine began to be widely used, although its correct application and understanding are still vague and poorly characterized. In precision medicine, we understand the individual as a response to the interrelation between environment, lifestyle, and genetic factors, which enabled the advent of new therapeutic models, such as conventional drugs adjustment by individual patient dosage and drug type and response, development of new drugs (read through, broker, enhancer, stabilizer, and amplifier compounds), genome editing by homologous recombination, zinc finger nucleases, TALEN (transcription activator-like effector nuclease), CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR-associated endonuclease 9), and gene therapy. Thus, we introduced the terms personalized medicine and precision medicine based on the CF.
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Affiliation(s)
- Fernando A L Marson
- Department of Medical Genetics, Faculty of Medical Sciences, State University of CampinasCampinas, Brazil.,Department of Pediatrics, Faculty of Medical Sciences, State University of CampinasCampinas, Brazil
| | - Carmen S Bertuzzo
- Department of Medical Genetics, Faculty of Medical Sciences, State University of CampinasCampinas, Brazil
| | - José D Ribeiro
- Department of Pediatrics, Faculty of Medical Sciences, State University of CampinasCampinas, Brazil
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Rowbotham NJ, Smyth AR. The patient voice in research - Supporting actor or starring role? J Cyst Fibros 2017; 16:313-314. [PMID: 28285933 DOI: 10.1016/j.jcf.2017.03.001] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Affiliation(s)
| | - Alan R Smyth
- School of Medicine, University of Nottingham, United Kingdom.
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Lucarelli M. New era of cystic fibrosis: Full mutational analysis and personalized therapy. World J Med Genet 2017; 7:1-9. [DOI: 10.5496/wjmg.v7.i1.1] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/06/2016] [Revised: 12/19/2016] [Accepted: 02/21/2017] [Indexed: 02/07/2023] Open
Abstract
Despite its apparently simple genetics, cystic fibrosis (CF) is a rather complex genetic disease. A lot of variability in the steps of the path from the cystic fibrosis transmembrane conductance regulator (CFTR) gene to the clinical manifestations originates an uncertain genotype - phenotype relationship. A major determinant of this uncertainty is the incomplete knowledge of the CFTR mutated genotypes, due to the high number of CFTR mutations and to the higher number of their combinations in trans and in cis. Also the very limited knowledge of functional effects of CFTR mutated alleles severely impairs our diagnostic and prognostic ability. The final phenotypic modulation exerted by CFTR modifier genes and interactome further complicates the framework. The next generation sequencing approach is a rapid, low-cost and high-throughput tool that allows a near complete structural characterization of CFTR mutated genotypes, as well as of genotypes of several other genes cooperating to the final CF clinical manifestations. This powerful method perfectly complements the new personalized therapeutic approach for CF. Drugs active on specific CFTR mutational classes are already available for CF patients or are in phase 3 trials. A complete genetic characterization has been becoming crucial for a correct personalized therapy. However, the need of a functional classification of each CFTR mutation potently arises. Future big efforts towards an ever more detailed knowledge of both structural and functional CFTR defects, coupled to parallel personalized therapeutic interventions decisive for CF cure can be foreseen.
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Affiliation(s)
- Marco Lucarelli
- Department of Cellular Biotechnologies and Hematology, Sapienza University of Rome, Laboratory affiliated to Istituto Pasteur Italia - Fondazione Cenci Bolognetti, 00161 Rome, Italy
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Abstract
Cystic fibrosis is an inherited multi-system disease associated with chronic lung infection, malabsorption, salt loss syndromes, male infertility and leading to numerous comorbidities. The landscape in cystic fibrosis care has changed markedly with currently more adult patients than children in many countries. Over 2000 different mutations in the CFTR gene have been reported and the majority are extremely rare. Understanding how CFTR mutations translate to disturbed synthesis or function of the CFTR protein has opened the way to 'personalized' treatments to correct the basic defect. The first 2 drugs have reached the clinic: a CFTR potentiator to augment CFTR channel function, and the combination of this potentiator with a corrector to increase CFTR expression at the cell membrane. To obtain robust correction of CFTR expression at the cell membrane, combinations of correctors with additive efficacy are under investigation. Other mutation type-specific treatments under clinical investigation are premature stop codon-read through drugs and antisense oligonucleotides that correct the basic defect at the mRNA level. Restoring the defective gene by gene editing can already be achieved ex vivo. Mutation agnostic treatments are explored as well: stabilizing CFTR expression at the cell membrane, circumventing the CFTR channel by blocking or activating other ion channels, and gene therapy. Combinations of these therapies can be anticipated. The pipeline of corrective strategies under clinical investigation is increasing continuously and a rising number of pharmaceutical companies are entering the field.
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Affiliation(s)
- Isabelle Fajac
- Université Paris Descartes, Sorbonne Paris Cité, Paris, France; AP-HP, Hôpital Cochin, Service de Physiologie et Explorations Fonctionnelles, Paris, France.
| | - Kris De Boeck
- University Hospital Gasthuisberg, Department of Pediatric Pulmonology, University of Leuven, Belgium.
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