Advancing the Battle against Cystic Fibrosis: Stem Cell and Gene Therapy Insights

Abstract

Cystic fibrosis (CF) is a hereditary disorder characterized by mutations in the CFTR gene, leading to impaired chloride ion transport and subsequent thickening of mucus in various organs, particularly the lungs. Despite significant progress in CF management, current treatments focus mainly on symptom relief and do not address the underlying genetic defects. Stem cell and gene therapies present promising avenues for tackling CF at its root cause. Stem cells, including embryonic, induced pluripotent, mesenchymal, hematopoietic, and lung progenitor cells, offer regenerative potential by differentiating into specialized cells and modulating immune responses. Similarly, gene therapy aims to correct CFTR gene mutations by delivering functional copies of the gene into affected cells. Various approaches, such as viral and nonviral vectors, gene editing with CRISPR-Cas9, small interfering RNA (siRNA) therapy, and mRNA therapy, are being explored to achieve gene correction. Despite their potential, challenges such as safety concerns, ethical considerations, delivery system optimization, and long-term efficacy remain. This review provides a comprehensive overview of the current understanding of CF pathophysiology, the rationale for exploring stem cell and gene therapies, the types of therapies available, their mechanisms of action, and the challenges and future directions in the field. By addressing these challenges, stem cell and gene therapies hold promise for transforming CF management and improving the quality of life of affected individuals.

References

[1] De BoeckK. Cystic fibrosis in the year 2020: A disease with a new face. Acta Paediatr, 2020, 109(5): 893-899

[2] ChenQ, ShenY, ZhengJ. A review of cystic fibrosis: Basic and clinical aspects. Animal Model Exp Med, 2021, 4(3): 220-232

[3] ElbornJS. Cystic fibrosis. Lancet, 2016, 388(10059): 2519-2531

[4] SomogyiV, ChaudhuriN, TorrisiSE, et al. . The therapy of idiopathic pulmonary fibrosis: what is next?. Eur Respir Rev, 2019, 28: 190021

[5] BarbenJ, CastellaniC, MunckA, et al. . Updated guidance on the management of children with cystic fibrosis transmembrane conductance regulator-related metabolic syndrome/cystic fibrosis screen positive, inconclusive diagnosis (CRMS/CFSPID). J Cyst Fibros, 2021, 20(5): 810-819

[6] KingCS, BrownAW, AryalS, et al. . Critical Care of the Adult Patient With Cystic Fibrosis. Chest, 2019, 155(1): 202-214

[7] D’AbroscaF, GarabelliB, SavioG, et al. . Comparing airways clearance techniques in chronic obstructive pulmonary disease and bronchiectasis: positive expiratory pressure or temporary positive expiratory pressure? A retrospective study. Braz J Phys Ther, 2017, 21(1): 15-23

[8] Dwyer T. Huff and puff of exercise for airway clearance in cystic fibrosis: how clear is the evidence? Thorax, 2021,Apr 29:thoraxjnl-2020-216622

[9] SchofieldLM, DuffA, BrennanC. Airway Clearance Techniques for Primary Ciliary Dyskinesia; is the Cystic Fibrosis literature portable?. Paediatr Respir Rev, 2018, 25: 73-77

[10] SmithS, RowbothamNJ, CharbekE. Inhaled antibiotics for pulmonary exacerbations in cystic fibrosis. Cochrane Database Syst Rev, 2022, 8(8): CD008319

[11] Calvo-LermaJ, Martínez-BaronaS, MasipE, et al. . Pancreatic enzyme replacement therapy in cystic fibrosis: dose, variability and coefficient of fat absorption. Rev Esp Enferm Dig, 2017, 109(10): 684-689

[12] SatheM, HouwenR. Meconium ileus in Cystic Fibrosis. J Cyst Fibros, 2017, 16(Suppl2): S32-S39

[13] AmaralMD. Novel personalized therapies for cystic fibrosis: treating the basic defect in all patients. J Intern Med, 2015, 277(2): 155-166

[14] SmithS, BorchardtM. Analyzing the use and impact of elexacaftor/tezacaftor/ivacaftor on total cost of care and other health care resource utilization in a commercially insured population. J Manag Care Spec Pharm, 2022, 28(7): 721-731

[15] GramegnaA, ContariniM, BindoF, AlibertiS, BlasiF. Elexacaftor-tezacaftor-ivacaftor: The new paradigm to treat people with cystic fibrosis with at least one p.Phe508del mutation. Curr Opin Pharmacol, 2021, 57: 81-88

[16] UlrichM, WorlitzschD, ViglioS, et al. . Alveolar inflammation in cystic fibrosis. J Cyst Fibros, 2010, 9(3): 217-227

[17] ChenJY, QiaoK, LiuF, et al. . Lung transplantation as therapeutic option in acute respiratory distress syndrome for coronavirus disease 2019-related pulmonary fibrosis. Chin Med J (Engl), 2020, 133(12): 1390-1396

[18] SnellG, ReedA, SternM, et al. . The evolution of lung transplantation for cystic fibrosis: A 2017 update. J Cyst Fibros, 2017, 16(5): 553-564

[19] GrasemannH, RatjenF. Emerging therapies for cystic fibrosis lung disease. Expert Opin Emerg Drugs, 2010, 15(4): 653-659

[20] LuQ, El-HashashAHK. Cell-based therapy for idiopathic pulmonary fibrosis. Stem Cell Investig, 2019, 6: 22

[21] KottonDN, MorriseyEE. Lung regeneration: mechanisms, applications and emerging stem cell populations. Nat Med, 2014, 20(8): 822-832

[22] Villate-BeitiaI, ZarateJ, PurasG, et al. . Gene delivery to the lungs: pulmonary gene therapy for cystic fibrosis. Drug Dev Ind Pharm, 2017, 43(7): 1071-1081

[23] CholonDM, GentzschM. Recent progress in translational cystic fibrosis research using precision medicine strategies. J Cyst Fibros, 2018, 17(2S): S52-S60

[24] QuonBS, RoweSM. New and emerging targeted therapies for cystic fibrosis. BMJ, 2016, 352: i859

[25] WangG, BunnellBA, PainterRG, et al. . Adult stem cells from bone marrow stroma differentiate into airway epithelial cells: potential therapy for cystic fibrosis. Proc Natl Acad Sci U S A, 2005, 102(1): 186-191

[26] LeeJA, ChoA, HuangEN, et al. . Gene therapy for cystic fibrosis: new tools for precision medicine. J Transl Med, 2021, 19(1): 452

[27] PrankeI, GolecA, HinzpeterA, et al. . Emerging Therapeutic Approaches for Cystic Fibrosis. From Gene Editing to Personalized Medicine. Front Pharmacol, 2019, 10: 121

[28] Marquez LozaLI, CooneyAL, DongQ, et al. . Increased CFTR expression and function from an optimized lentiviral vector for cystic fibrosis gene therapy. Mol Ther Methods Clin Dev, 2021, 21: 94-106

[29] IkonomouL, MagnussonM, DriesR, et al. . Stem cells, cell therapies, and bioengineering in lung biology and disease 2021. Am J Physiol Lung Cell Mol Physiol, 2022, 323(3): L341-L354

[30] EbrahimiA, AhmadiH, Pourfraidon GhasrodashtiZ, et al. . Therapeutic effects of stem cells in different body systems, a novel method that is yet to gain trust: A comprehensive review. Bosn J Basic Med Sci, 2021, 21(6): 672-701

[31] QinL, LiuN, BaoCL, et al. . Mesenchymal stem cells in fibrotic diseases-the two sides of the same coin. Acta Pharmacol Sin, 2023, 44(2): 268-287

[32] BonfieldTL, LennonD, GhoshSK, DiMarinoAM, WeinbergA, CaplanAI. Cell based therapy aides in infection and inflammation resolution in the murine model of cystic fibrosis lung disease. Stem Cell Discovery, 2013, 3(2): 139-153

[33] BericalA, LeeRE, RandellSH, et al. . Challenges Facing Airway Epithelial Cell-Based Therapy for Cystic Fibrosis. Front Pharmacol, 2019, 10: 74

[34] MahmoudiM, HosseinkhaniH, HosseinkhaniM, et al. . Magnetic resonance imaging tracking of stem cells in vivo using iron oxide nanoparticles as a tool for the advancement of clinical regenerative medicine. Chem Rev, 2011, 111(2): 253-280

[35] IltisAS, KosterG, ReevesE, et al. . Ethical, legal, regulatory, and policy issues concerning embryoids: a systematic review of the literature. Stem Cell Res Ther, 2023, 14(1): 209

[36] HaworthR, SharpeM. Accept or Reject: The Role of Immune Tolerance in the Development of Stem Cell Therapies and Possible Future Approaches. Toxicol Pathol, 2021, 49(7): 1308-1316

[37] MouH, ZhaoR, SherwoodR, et al. . Generation of multipotent lung and airway progenitors from mouse ESCs and patient-specific cystic fibrosis iPSCs. Cell Stem Cell, 2012, 10(4): 385-397

[38] HawkinsFJ, SuzukiS, BeermannML, et al. . Derivation of Airway Basal Stem Cells from Human Pluripotent Stem Cells. Cell Stem Cell, 2021, 28(1): 79-95.e8

[39] Ajilore PO, Yang HY, Kerasidis A, et al. Curative Measures for Cystic Fibrosis: A Perspective on Current Stem Cell-Based, Gene, and Small Molecule Therapies. Georgetown Medical Review, 2022,6(1)

[40] HardingJ, Vintersten-NagyK, YangH, et al. . Immune-privileged tissues formed from immunologically cloaked mouse embryonic stem cells survive long term in allogeneic hosts. Nat Biomed Eng, 2024, 8(4): 427-442

[41] PessôaLVF, BressanFF, FreudeKK. Induced pluripotent stem cells throughout the animal kingdom: Availability and applications. World J Stem Cells, 2019, 11(8): 491-505

[42] BragançaJ, LopesJA, Mendes-SilvaL, et al. . Induced pluripotent stem cells, a giant leap for mankind therapeutic applications. World J Stem Cells, 2019, 11(7): 421-430

[43] FilaretoA, ParkerS, DarabiR, et al. . An ex vivo gene therapy approach to treat muscular dystrophy using inducible pluripotent stem cells. Nat Commun, 2013, 4: 1549

[44] DingDC, ShyuWC, LinSZ. Mesenchymal stem cells. Cell Transplant, 2011, 20(1): 5-14

[45] NykänenAI, LiuM, KeshavjeeS. Mesenchymal Stromal Cell Therapy in Lung Transplantation. Bioengineering (Basel), 2023, 10(6): 728

[46] CruzFF, RoccoPRM. The potential of mesenchymal stem cell therapy for chronic lung disease. Expert Rev Respir Med, 2020, 14(1): 31-39

[47] KhaddourK, HanaCK, MewawallaP. Hematopoietic Stem Cell Transplantation. StatPearls, 2024 Treasure Island (FL) StatPearls Publishing [Updated 2023 May 6]

[48] RaoI, CrisafulliL, PaulisM, et al. . Hematopoietic Cells from Pluripotent Stem Cells: Hope and Promise for the Treatment of Inherited Blood Disorders. Cells, 2022, 11(3): 557

[49] BushLM, HealyCP, JavdanSB, et al. . Biological Cells as Therapeutic Delivery Vehicles. Trends Pharmacol Sci, 2021, 42(2): 106-118

[50] KingNE, SuzukiS, BarillàC, et al. . Correction of Airway Stem Cells: Genome Editing Approaches for the Treatment of Cystic Fibrosis. Hum Gene Ther, 2020, 31(17–18): 956-972

[51] Conrad C, Magnen M, Tsui J, et al. Decoding functional hematopoietic progenitor cells in the adult human lung. Preprint. Res Sq, 2024,rs.3.rs-3576483

[52] ChengW, ZengY, WangD. Stem cell-based therapy for pulmonary fibrosis. Stem Cell Res Ther, 2022, 13(1): 492

[53] PopowskiK, LutzH, HuS, et al. . Exosome therapeutics for lung regenerative medicine. J Extracell Vesicles, 2020, 9(1): 1785161

[54] MasuiA, HiraiT, GotohS. Perspectives of future lung toxicology studies using human pluripotent stem cells. Arch Toxicol, 2022, 96(2): 389-402

[55] LiuP, MaoY, XieY, et al. . Stem cells for treatment of liver fibrosis/cirrhosis: clinical progress and therapeutic potential. Stem Cell Res Ther, 2022, 13(1): 356

[56] HongKU, ReynoldsSD, WatkinsS, et al. . Basal cells are a multipotent progenitor capable of renewing the bronchial epithelium. Am J Pathol, 2004, 164(2): 577-588

[57] XieT, LynnH, ParksWC, et al. . Abnormal respiratory progenitors in fibrotic lung injury. Stem Cell Res Ther, 2022, 13(1): 64

[58] RockJR, RandellSH, HoganBL. Airway basal stem cells: a perspective on their roles in epithelial homeostasis and remodeling. Dis Model Mech, 2010, 3(9–10): 545-556

[59] KleinD. Lung Multipotent Stem Cells of Mesenchymal Nature: Cellular Basis, Clinical Relevance, and Implications for Stem Cell Therapy. Antioxid Redox Signal, 2021, 35(3): 204-216

[60] BuschSM, LorenzanaZ, RyanAL. Implications for Extracellular Matrix Interactions With Human Lung Basal Stem Cells in Lung Development, Disease, and Airway Modeling. Front Pharmacol, 2021, 12: 645858

[61] YangS, YuM. Role of Goblet Cells in Intestinal Barrier and Mucosal Immunity. J Inflamm Res, 2021, 14: 3171-3183

[62] ScudieriP, MusanteI, VenturiniA, et al. . Ionocytes and CFTR Chloride Channel Expression in Normal and Cystic Fibrosis Nasal and Bronchial Epithelial Cells. Cells, 2020, 9(9): 2090

[63] YuanF, GasserGN, LemireE, et al. . Transgenic ferret models define pulmonary ionocyte diversity and function. Nature, 2023, 621(7980): 857-867

[64] ForbesSJ, RosenthalN. Preparing the ground for tissue regeneration: from mechanism to therapy. Nat Med, 2014, 20(8): 857-869

[65] MurphySV, AtalaA. Cell therapy for cystic fibrosis. J Tissue Eng Regen Med, 2015, 9(3): 210-223

[66] LiX, YueS, LuoZ. Mesenchymal stem cells in idiopathic pulmonary fibrosis. Oncotarget, 2017, 8(60): 102600-102616

[67] MoodleyY, VaghjianiV, ChanJ, et al. . Anti-inflammatory effects of adult stem cells in sustained lung injury: a comparative study. PLoS One, 2013, 8(8): e69299

[68] van GeffenC, DeißlerA, QuanteM, et al. . Regulatory Immune Cells in Idiopathic Pulmonary Fibrosis: Friends or Foes?. Front Immunol, 2021, 12: 663203

[69] TangXD, JiTT, DongJR, et al. . Pathogenesis and Treatment of Cytokine Storm Induced by Infectious Diseases. Int J Mol Sci, 2021, 22(23): 13009

[70] TakaoS, NakashimaT, MasudaT, et al. . Human bone marrow-derived mesenchymal stromal cells cultured in serumfree media demonstrate enhanced antifibrotic abilities via prolonged survival and robust regulatory T cell induction in murine bleomycin-induced pulmonary fibrosis. Stem Cell Res Ther, 2021, 12(1): 506

[71] ChowL, JohnsonV, ImpastatoR, et al. . Antibacterial activity of human mesenchymal stem cells mediated directly by constitutively secreted factors and indirectly by activation of innate immune effector cells. Stem Cells Transl Med, 2020, 9(2): 235-249

[72] Santos-FernandezE, Martin-SoutoL, AntoranA, et al. . Microbiota and fungal-bacterial interactions in the cystic fibrosis lung. FEMS Microbiol Rev, 2023, 47(3): fuad029

[73] MeiSH, HaitsmaJJ, Dos SantosCC, et al. . Mesenchymal stem cells reduce inflammation while enhancing bacterial clearance and improving survival in sepsis. Am J Respir Crit Care Med, 2010, 182(8): 1047-1057

[74] XiaJ, MinaminoS, KuwabaraK, et al. . Stem cell secretome as a new booster for regenerative medicine. Biosci Trends, 2019, 13(4): 299-307

[75] ShiY, ShiH, NomiA, et al. . Mesenchymal stem cell-derived extracellular vesicles: a new impetus of promoting angiogenesis in tissue regeneration. Cytotherapy, 2019, 21(5): 497-508

[76] LiewLC, HoBX, SohBS. Mending a broken heart: current strategies and limitations of cell-based therapy. Stem Cell Res Ther, 2020, 11(1): 138

[77] WiśniewskaJ, SadowskaA, WójtowiczA, et al. . Perspective on Stem Cell Therapy in Organ Fibrosis: Animal Models and Human Studies. Life (Basel), 2021, 11(10): 1068

[78] ZinterMS, HumeJR. Effects of Hematopoietic Cell Transplantation on the Pulmonary Immune Response to Infection. Front Pediatr, 2021, 9: 634566

[79] CharitosIA, BalliniA, CantoreS, et al. . Stem Cells: A Historical Review about Biological, Religious, and Ethical Issues. Stem Cells Int, 2021, 2021: 9978837

[80] van RietS, NinaberDK, MikkersHMM, et al. . In vitro modelling of alveolar repair at the air-liquid interface using alveolar epithelial cells derived from human induced pluripotent stem cells. Sci Rep, 2020, 10(1): 5499

[81] OveiliE, VafaeiS, BazavarH, et al. . The potential use of mesenchymal stem cells-derived exosomes as microRNAs delivery systems in different diseases. Cell Commun Signal, 2023, 21(1): 20

[82] DalyS, O’SullivanA, MacLoughlinR. Cellular Immunotherapy and the Lung. Vaccines (Basel), 2021, 9(9): 1018

[83] ParanjapyeA, RuffinM, HarrisA, et al. . Genetic variation in CFTR and modifier loci may modulate cystic fibrosis disease severity. J Cyst Fibros, 2020, 19(Suppl1): S10-S14

[84] JeongMH, HanH, LagaresD, et al. . Recent Advances in Molecular Diagnosis of Pulmonary Fibrosis for Precision Medicine. ACS Pharmacol Transl Sci, 2022, 5(8): 520-538

[85] LiangJ, HuangW, JiangL, et al. . Concise Review: Reduction of Adverse Cardiac Scarring Facilitates Pluripotent Stem Cell-Based Therapy for Myocardial Infarction. Stem Cells, 2019, 37(7): 844-854

[86] DriscollD, FarniaS, KefalasP, et al. . Concise Review: The High Cost of High Tech Medicine: Planning Ahead for Market Access. Stem Cells Transl Med, 2017, 6(8): 1723-1729

[87] DyeBR, HillDR, FergusonMA, et al. . In vitro generation of human pluripotent stem cell derived lung organoids. Elife, 2015, 4: e05098

[88] MondrinosMJ, KoutzakiS, JiwanmallE, et al. . Engineering three-dimensional pulmonary tissue constructs. Tissue Eng, 2006, 12(4): 717-728

[89] WongAP, BearCE, ChinS, et al. . Directed differentiation of human pluripotent stem cells into mature airway epithelia expressing functional CFTR protein. Nat Biotechnol, 2012, 30(9): 876-882

[90] MouH, ZhaoR, SherwoodR, et al. . Generation of multipotent lung and airway progenitors from mouse ESCs and patient-specific cystic fibrosis iPSCs. Cell Stem Cell, 2012, 10(4): 385-397

[91] FirthAL, MenonT, ParkerGS, et al. . Functional Gene Correction for Cystic Fibrosis in Lung Epithelial Cells Generated from Patient iPSCs. Cell Rep, 2015, 12(9): 1385-1390

[92] SampaziotisF, de BritoMC, MadrigalP, et al. . Cholangiocytes derived from human induced pluripotent stem cells for disease modeling and drug validation. Nat Biotechnol, 2015, 33(8): 845-852

[93] LiouTG. The Clinical Biology of Cystic Fibrosis Transmembrane Regulator Protein: Its Role and Function in Extrapulmonary Disease. Chest, 2019, 155(3): 605-616

[94] FajacI, WainwrightCE. New treatments targeting the basic defects in cystic fibrosis. Presse Med, 2017, 46(6): e165-e175 Pt 2

[95] BandaraRA, ChenZR, HuJ. Potential of helper-dependent Adenoviral vectors in CRISPR-cas9-mediated lung gene therapy. Cell Biosci, 2021, 11(1): 145

[96] Christopher BoydA, GuoS, HuangL, et al. . New approaches to genetic therapies for cystic fibrosis. J Cyst Fibros, 2020, 19(Suppl1): S54-S59

[97] HackettPB, LargaespadaDA, SwitzerKC, et al. . Evaluating risks of insertional mutagenesis by DNA transposons in gene therapy. Transl Res, 2013, 161(4): 265-283

[98] HanssensLS, DuchateauJ, CasimirGJ. CFTR Protein: Not Just a Chloride Channel?. Cells, 2021, 10(11): 2844

[99] FohnerAE, McDonaghEM, ClancyJP, et al. . PharmGKB: ivacaftor pathway, pharmacokinetics/pharmacodynamics. Pharmacogenet Genomics, 2017, 27(1): 39-42

[100] QuonBS, RoweSM. New and emerging targeted therapies for cystic fibrosis. BMJ, 2016, 352: i859

[101] FarinhaCM, CanatoS. From the endoplasmic reticulum to the plasma membrane: mechanisms of CFTR folding and trafficking. Cell Mol Life Sci, 2017, 74(1): 39-55

[102] Lopes-PachecoM. CFTR Modulators: The Changing Face of Cystic Fibrosis in the Era of Precision Medicine. Front Pharmacol, 2020, 10: 1662

[103] Sermet-GaudelusI, GirodonE, VermeulenF, et al. . ECFS standards of care on CFTR-related disorders: Diagnostic criteria of CFTR dysfunction. J Cyst Fibros, 2022, 21(6): 922-936

[104] De PalmaFDE, RaiaV, KroemerG, et al. . The Multifaceted Roles of MicroRNAs in Cystic Fibrosis. Diagnostics (Basel), 2020, 10(12): 1102

[105] McGarryME, GibbER, OatesGR, et al. . Left behind: The potential impact of CFTR modulators on racial and ethnic disparities in cystic fibrosis. Paediatr Respir Rev, 2022, 42: 3542

[106] ScotetV, L’HostisC, FérecC. The Changing Epidemiology of Cystic Fibrosis: Incidence, Survival and Impact of the CFTR Gene Discovery. Genes (Basel), 2020, 11(6): 589

[107] TangY, YanZ, EngelhardtJF. Viral Vectors, Animal Models, and Cellular Targets for Gene Therapy of Cystic Fibrosis Lung Disease. Hum Gene Ther, 2020, 31(9–10): 524-537

[108] BulchaJT, WangY, MaH, et al. . Viral vector platforms within the gene therapy landscape. Signal Transduct Target Ther, 2021, 6(1): 53

[109] Sainz-RamosM, Villate-BeitiaI, GallegoI, et al. . Nonviral mediated gene therapy in human cystic fibrosis airway epithelial cells recovers chloride channel functionality. Int J Pharm, 2020, 588: 119757

[110] WangH, QinL, ZhangX, et al. . Mechanisms and challenges of nanocarriers as non-viral vectors of therapeutic genes for enhanced pulmonary delivery. J Control Release, 2022, 352: 970-993

[111] AsmamawM, ZawdieB. Mechanism and Applications of CRISPR/Cas-9-Mediated Genome Editing. Biologics, 2021, 15: 353-361

[112] AbdelnourSA, XieL, HassaninAA, et al. . The Potential of CRISPR/Cas9 Gene Editing as a Treatment Strategy for Inherited Diseases. Front Cell Dev Biol, 2021, 9: 699597

[113] BrothersKB, DevereauxM, SadeRM. Bespoke Babies: Genome Editing in Cystic Fibrosis Embryos. Ann Thorac Surg, 2019, 108(4): 995-999

[114] MollaKA, ShihJ, WheatleyMS, et al. . Predictable NHEJ Insertion and Assessment of HDR Editing Strategies in Plants. Front Genome Ed, 2022, 4: 825236

[115] ShamshirgaranY, LiuJ, SumerH, et al. . Tools for Efficient Genome Editing; ZFN, TALEN, and CRISPR. Methods Mol Biol, 2022, 2495: 29-46

[116] FleischerA, Vallejo-DíezS, Martín-FernándezJM, et al. . iPSC-Derived Intestinal Organoids from Cystic Fibrosis Patients Acquire CFTR Activity upon TALEN-Mediated Repair of the p.F508del Mutation. Mol Ther Methods Clin Dev, 2020, 17: 858870

[117] SchwankG, KooBK, SasselliV, et al. . Functional repair of CFTR by CRISPR/Cas9 in intestinal stem cell organoids of cystic fibrosis patients. Cell Stem Cell, 2013, 13(6): 653-658

[118] WangW, LiW, MaN, et al. . Non-viral gene delivery methods. Curr Pharm Biotechnol, 2013, 14(1): 46-60

[119] ZhangXH, TeeLY, WangXG, et al. . Off-target Effects in CRISPR/Cas9-mediated Genome Engineering. Mol Ther Nucleic Acids, 2015, 4(11): e264

[120] YeeJK. Off-target effects of engineered nucleases. FEBS J, 2016, 283(17): 3239-3248

[121] NewmanA, StarrsL, BurgioG. Cas9 Cuts and Consequences; Detecting, Predicting, and Mitigating CRISPR/Cas9 On- and Off-Target Damage: Techniques for Detecting, Predicting, and Mitigating the On- and off-target Effects of Cas9 Editing. Bioessays, 2020, 42(9): e2000047

[122] ManghwarH, LiB, DingX, et al. . CRISPR/Cas Systems in Genome Editing: Methodologies and Tools for sgRNA Design, Off-Target Evaluation, and Strategies to Mitigate Off-Target Effects. Adv Sci (Weinh), 2020, 7(6): 1902312

[123] HanHA, PangJKS, SohBS. Mitigating off-target effects in CRISPR/Cas9-mediated in vivo gene editing. J Mol Med (Berl), 2020, 98(5): 615-632

[124] SavulescuJ, PughJ, DouglasT, et al. . The moral imperative to continue gene editing research on human embryos. Protein Cell, 2015, 6(7): 476-479

[125] TurrizianiJV. Designer Babies: The Need for Regulation on the Quest For Perfection, 2014 595 Law School Student Scholarship

[126] Da Silva SanchezA, PaunovskaK, CristianA, et al. . Treating Cystic Fibrosis with mRNA and CRISPR. Hum Gene Ther, 2020, 31(17–18): 940-955

[127] ViartV, BergougnouxA, BoniniJ, et al. . Transcription factors and miRNAs that regulate fetal to adult CFTR expression change are new targets for cystic fibrosis. Eur Respir J, 2015, 45(1): 116-128

[128] ConteG, CostabileG, BaldassiD, et al. . Hybrid Lipid/Polymer Nanoparticles to Tackle the Cystic Fibrosis Mucus Barrier in siRNA Delivery to the Lungs: Does PEGylation Make the Difference?. ACS Appl Mater Interfaces, 2022, 14(6): 7565-7578

[129] d’AngeloI, ConteC, La RotondaMI, et al. . Improving the efficacy of inhaled drugs in cystic fibrosis: challenges and emerging drug delivery strategies. Adv Drug Deliv Rev, 2014, 75: 92-111

[130] SteinleH, BehringA, SchlensakC, et al. . Concise Review: Application of In Vitro Transcribed Messenger RNA for Cellular Engineering and Reprogramming: Progress and Challenges. Stem Cells, 2017, 35(1): 68-79

[131] SahinU, KarikóK, TüreciÖ. mRNA-based therapeutics—developing a new class of drugs. Nat Rev Drug Discov, 2014, 13(10): 759-780

[132] KatzMG, FargnoliAS, WilliamsRD, et al. . Gene therapy delivery systems for enhancing viral and nonviral vectors for cardiac diseases: current concepts and future applications. Hum Gene Ther, 2013, 24(11): 914-927

[133] GhoshS, BrownAM, JenkinsC, et al. . Viral Vector Systems for Gene Therapy: A Comprehensive Literature Review of Progress and Biosafety Challenges. Appl Biosaf, 2020, 25(1): 7-18

[134] LiW, SzokaFC Jr. Lipid-based nanoparticles for nucleic acid delivery. Pharm Res, 2007, 24(3): 438-449

[135] FilipczakN, PanJ, YalamartySSK, et al. . Recent advancements in liposome technology. Adv Drug Deliv Rev, 2020, 156: 4-22

[136] PariharA, PrajapatiBG, PaliwalH, et al. . Advanced pulmonary drug delivery formulations for the treatment of cystic fibrosis. Drug Discov Today, 2023, 28(10): 103729

[137] BalantičK, MiklavčičD, KrižajI, et al. . The good and the bad of cell membrane electroporation. Acta Chim Slov, 2021, 68(4): 753-764

[138] Mehier-HumbertS, GuyRH. Physical methods for gene transfer: improving the kinetics of gene delivery into cells. Adv Drug Deliv Rev, 2005, 57(5): 733-753

[139] Llargués-SistacG, BonjochL, Castellvi-BelS. HAP1, a new revolutionary cell model for gene editing using CRISPR-Cas9. Front Cell Dev Biol, 2023, 11: 1111488

[140] Villate-BeitiaI, ZarateJ, PurasG, et al. . Gene delivery to the lungs: pulmonary gene therapy for cystic fibrosis. Drug Dev Ind Pharm, 2017, 43(7): 1071-1081

[141] AltonEWFW, ArmstrongDK, AshbyD, et al. . Repeated nebulisation of non-viral CFTR gene therapy in patients with cystic fibrosis: a randomised, double-blind, placebo-controlled, phase 2b trial. Lancet Respir Med, 2015, 3(9): 684-691

[142] RamamoorthM, NarvekarA. Non viral vectors in gene therapy- an overview. J Clin Diagn Res, 2015, 9(1): GE01-GE6

[143] ThomasCE, EhrhardtA, KayMA. Progress and problems with the use of viral vectors for gene therapy. Nat Rev Genet, 2003, 4(5): 346-358

[144] ZaissAK, MuruveDA. Immune responses to adeno-associated virus vectors. Curr Gene Ther, 2005, 5(3): 323-331

[145] FajacI, De BoeckK. New horizons for cystic fibrosis treatment. Pharmacol Ther, 2017, 170: 205-211

[146] ButtMH, ZamanM, AhmadA, et al. . Appraisal for the Potential of Viral and Nonviral Vectors in Gene Therapy: A Review. Genes (Basel), 2022, 13(8): 1370

[147] NaldiniL. Gene therapy returns to centre stage. Nature, 2015, 526(7573): 351-360

[148] WangD, TaiPWL, GaoG. Adeno-associated virus vector as a platform for gene therapy delivery. Nat Rev Drug Discov, 2019, 18(5): 358-378

[149] LiH, YangY, HongW, et al. . Applications of genome editing technology in the targeted therapy of human diseases: mechanisms, advances and prospects. Signal Transduct Target Ther, 2020, 5(1): 1

[150] GuimbellotJ, SharmaJ, RoweSM. Toward inclusive therapy with CFTR modulators: Progress and challenges. Pediatr Pulmonol, 2017, 52(S48): S4-S14

[151] ShalabyK, AouidaM, El-AgnafO. Tissue-Specific Delivery of CRISPR Therapeutics: Strategies and Mechanisms of Non-Viral Vectors. Int J Mol Sci, 2020, 21(19): 7353

[152] FoldvariM, ChenDW, NafissiN, et al. . Non-viral gene therapy: Gains and challenges of non-invasive administration methods. J Control Release, 2016, 240: 165-190

[153] LiaoL, ShiB, ChangH, et al. . Heparin improves BMSC cell therapy: Anticoagulant treatment by heparin improves the safety and therapeutic effect of bone marrow-derived mesenchymal stem cell cytotherapy. Theranostics, 2017, 7(1): 106-116

[154] DandawateP, PadhyeS, AhmadA, et al. . Novel strategies targeting cancer stem cells through phytochemicals and their analogs. Drug Deliv Transl Res, 2013, 3(2): 165-182

[155] ZhouHS, LiuDP, LiangCC. Challenges and strategies: the immune responses in gene therapy. Med Res Rev, 2004, 24(6): 748-761

[156] GonçalvesGAR, PaivaRMA. Gene therapy: advances, challenges and perspectives. Einstein (Sao Paulo), 2017, 15(3): 369-375

[157] CooneyAL, McCrayPB Jr, SinnPL. Cystic Fibrosis Gene Therapy: Looking Back, Looking Forward. Genes (Basel), 2018, 9(11): 538

[158] HayesD Jr, KoppBT, HillCL, et al. . Cell Therapy for Cystic Fibrosis Lung Disease: Regenerative Basal Cell Amplification. Stem Cells Transl Med, 2019, 8(3): 225-235

[159] ConeseM, AscenzioniF, BoydAC, et al. . Gene and cell therapy for cystic fibrosis: from bench to bedside. J Cyst Fibros, 2011, 10(Suppl2): S114-S128

[160] ConradC, GuptaR, MohanH, et al. . Genetically engineered stem cells for therapeutic gene delivery. Curr Gene Ther, 2007, 7(4): 249-260

[161] MyersTJ, Granero-MoltoF, LongobardiL, et al. . Mesenchymal stem cells at the intersection of cell and gene therapy. Expert Opin Biol Ther, 2010, 10(12): 1663-1679

[162] CraneAM, KramerP, BuiJH, et al. . Targeted correction and restored function of the CFTR gene in cystic fibrosis induced pluripotent stem cells. Stem Cell Reports, 2015, 4(4): 569-577

[163] MorganRA, GrayD, LomovaA, et al. . Hematopoietic Stem Cell Gene Therapy: Progress and Lessons Learned. Cell Stem Cell, 2017, 21(5): 574-590

[164] McCarronA, CmielewskiP, DrysdaleV, et al. . Effective viral-mediated lung gene therapy: is airway surface preparation necessary?. Gene Ther, 2023, 30(6): 469-477

[165] LanYW, ChooKB, ChenCM, et al. . Hypoxia-preconditioned mesenchymal stem cells attenuate bleomycin-induced pulmonary fibrosis. Stem Cell Res Ther, 2015, 6(1): 97

[166] GauglitzGG, JeschkeMG. Combined gene and stem cell therapy for cutaneous wound healing. Mol Pharm, 2011, 8(5): 1471-1479

[167] CaoB, BruderJ, KovesdiI, et al. . Muscle stem cells can act as antigen-presenting cells: implication for gene therapy. Gene Ther, 2004, 11(17): 1321-1330

[168] AhmedAU, AlexiadesNG, LesniakMS. The use of neural stem cells in cancer gene therapy: predicting the path to the clinic. Curr Opin Mol Ther, 2010, 12(5): 546-552

[169] ZhangJ, HuangX, WangH, et al. . The challenges and promises of allogeneic mesenchymal stem cells for use as a cell-based therapy. Stem Cell Res Ther, 2015, 6: 234

[170] BrownN, SongL, KolluNR, et al. . Adeno-Associated Virus Vectors and Stem Cells: Friends or Foes?. Hum Gene Ther, 2017, 28(6): 450-463

[171] NakanishiM, OtsuM. Development of Sendai virus vectors and their potential applications in gene therapy and regenerative medicine. Curr Gene Ther, 2012, 12(5): 410-416

[172] Martínez-GonzálezI, RocaO, MasclansJR, et al. . Human mesenchymal stem cells overexpressing the IL-33 antagonist soluble IL-1 receptor-like-1 attenuate endotoxin-induced acute lung injury. Am J Respir Cell Mol Biol, 2013, 49(4): 552-562

[173] WanT, PingY. Delivery of genome-editing biomacromolecules for treatment of lung genetic disorders. Adv Drug Deliv Rev, 2021, 168: 196-216

[174] DeuseT, HuX, GravinaA, et al. . Hypoimmunogenic derivatives of induced pluripotent stem cells evade immune rejection in fully immunocompetent allogeneic recipients. Nat Biotechnol, 2019, 37(3): 252-258

[175] GoldringCE, DuffyPA, BenvenistyN, et al. . Assessing the safety of stem cell therapeutics. Cell Stem Cell, 2011, 8(6): 618-628

[176] VolarevicV, MarkovicBS, GazdicM, et al. . Ethical and Safety Issues of Stem Cell-Based Therapy. Int J Med Sci, 2018, 15(1): 36-45

[177] DawsonL, Bateman-HouseAS, Mueller AgnewD, et al. . Safety issues in cell-based intervention trials. Fertil Steril, 2003, 80(5): 1077-1085

[178] AfifySM, SenoM. Conversion of Stem Cells to Cancer Stem Cells: Undercurrent of Cancer Initiation. Cancers (Basel), 2019, 11(3): 345

[179] FoxIJ, DaleyGQ, GoldmanSA, et al. . Stem cell therapy. Use of differentiated pluripotent stem cells as replacement therapy for treating disease. Science, 2014, 345(6199): 1247391

[180] YipBH. Recent Advances in CRISPR/Cas9 Delivery Strategies. Biomolecules, 2020, 10(6): 839

[181] XuX, WanT, XinH, et al. . Delivery of CRISPR/Cas9 for therapeutic genome editing. J Gene Med, 2019, 21(7): e3107

[182] ShouY, MaZ, LuT, et al. . Unique risk factors for insertional mutagenesis in a mouse model of XSCID gene therapy. Proc Natl Acad Sci U S A, 2006, 103(31): 11730-11735

[183] Iglesias-LópezC, AgustíA, ObachM, et al. . Regulatory Framework for Advanced Therapy Medicinal Products in Europe and United States. Front Pharmacol, 2019, 10: 921

[184] HolmS. Going to the roots of the stem cell controversy. Bioethics, 2002, 16(6): 493-507

[185] RivaL, PetriniC. A few ethical issues in translational research for gene and cell therapy. J Transl Med, 2019, 17(1): 395

[186] LoB, ParhamL. Ethical issues in stem cell research. Endocr Rev, 2009, 30(3): 204-213

[187] CollerBS. Ethics of Human Genome Editing. Annu Rev Med, 2019, 70: 289-305

[188] RothenbergKH. Genetic information and health insurance: state legislative approaches. J Law Med Ethics, 1995, 23(4): 312-319

[189] Piotrowski-DaspitAS, BaroneC, LinCY, et al. . In vivo correction of cystic fibrosis mediated by PNA nanoparticles. Sci Adv, 2022, 8(40): eabo0522

[190] SunY, ChatterjeeS, LianX, et al. . In vivo editing of lung stem cells for durable gene correction in mice. Science, 2024, 384(6701): 1196-1202

[191] KulhankovaK, TraoreS, ChengX, et al. . Shuttle peptide delivers base editor RNPs to rhesus monkey airway epithelial cells in vivo. Nat Commun, 2023, 14(1): 8051

[192] WangG. Genome Editing for Cystic Fibrosis. Cells, 2023, 12(12): 1555

[193] AllenL, AllenL, CarrSB, et al. . Future therapies for cystic fibrosis. Nat Commun, 2023, 14(1): 693

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