Edina Poletto
Research Assistant, Pediatrics - Genetics
Contact
https://orcid.org/0000-0003-0554-0295All Publications
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Improved engraftment and therapeutic efficacy by human genome-edited hematopoietic stem cells with Busulfan-based myeloablation.
Molecular therapy. Methods & clinical development
2022; 25: 392-409
Abstract
Autologous hematopoietic stem cell transplantation using genome-edited cells can become a definitive therapy for hematological and non-hematological disorders with neurological involvement. Proof-of-concept studies using human genome-edited hematopoietic stem cells have been hindered by the low efficiency of engraftment of the edited cells in the bone marrow and their modest efficacy in the CNS. To address these challenges, we tested a myeloablative conditioning regimen based on Busulfan in an immunocompromised model of mucopolysaccharidosis type 1. Compared with sub-lethal irradiation, Busulfan conditioning enhanced the engraftment of edited CD34+ cells in the bone marrow, as well the long-term homing and survival of bone-marrow-derived cells in viscera, and in the CNS, resulting in higher transgene expression and biochemical correction in these organs. Edited cell selection using a clinically compatible marker resulted in a population withlow engraftment potential. We conclude that conditioning can impact the engraftment of edited hematopoietic stem cells.Furthermore, Busulfan-conditioned recipients have a higher expression of therapeutic proteins in target organs, particularly in the CNS, constituting a better conditioning approach for non-hematological diseases with neurological involvement.
View details for DOI 10.1016/j.omtm.2022.04.009
View details for PubMedID 35573043
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Genome Editing for Mucopolysaccharidoses.
International journal of molecular sciences
2020; 21 (2)
Abstract
Genome editing holds the promise of one-off and potentially curative therapies for many patients with genetic diseases. This is especially true for patients affected by mucopolysaccharidoses as the disease pathophysiology is amenable to correction using multiple approaches. Ex vivo and in vivo genome editing platforms have been tested primarily on MSPI and MPSII, with in vivo approaches having reached clinical testing in both diseases. Though we still await proof of efficacy in humans, the therapeutic tools established for these two diseases should pave the way for other mucopolysaccharidoses. Herein, we review the current preclinical and clinical development studies, using genome editing as a therapeutic approach for these diseases. The development of new genome editing platforms and the variety of genetic modifications possible with each tool provide potential applications of genome editing for mucopolysaccharidoses, which vastly exceed the potential of current approaches. We expect that in a not-so-distant future, more genome editing-based strategies will be established, and individual diseases will be treated through multiple approaches.
View details for DOI 10.3390/ijms21020500
View details for PubMedID 31941077
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Engineering monocyte/macrophage−specific glucocerebrosidase expression in human hematopoietic stem cells using genome editing
Nature Communications
2020; 11: 1-14
View details for DOI 10.1038/s41467-020-17148-x
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Liposomal CRISPR/Cas9-Mediated Local Genome Editing for Joint Disease in Mucopolysaccharidosis Type I
PHARMACEUTICS
2026; 18 (3)
Abstract
Background/Objectives: Mucopolysaccharidosis type I (MPS I) is a lysosomal storage disorder caused by α-L-iduronidase (IDUA) deficiency, leading to progressive glycosaminoglycan (GAG) accumulation and severe joint involvement. Gene editing represents a promising alternative to restore localized enzyme production. Therefore, this study aimed to evaluate the feasibility, efficacy, and safety of in situ genome editing through intra-articular administration of a nonviral CRISPR/Cas9 system to increase localized IDUA expression in an MPS I mouse model. Methods: Cationic liposomes were formulated to deliver plasmids encoding the CRISPR/Cas9 system targeted to the ROSA26 locus along with an IDUA donor sequence. In vitro assays were performed in fibroblast-like synoviocytes (FLSs) isolated from MPS I mice to assess cytotoxicity, gene editing efficiency, and IDUA activity. In vivo, MPS I mice received intra-articular injections in the knee joints, either as a single dose (short-term study) or monthly for three months (long-term study). IDUA activity, GAG levels, and genome editing efficiency were evaluated in joint tissues, synovial fluid, serum, and major organs. Results: Gene-edited FLS showed sustained IDUA activity for up to 30 days with low cytotoxicity. In vivo, intra-articular administration resulted in a significant increase in IDUA activity in joint tissue and synovial fluid without detectable systemic IDUA. Long-term treatment led to persistent joint-localized IDUA activity, significant reductions (>50%) in GAG levels, and detectable genome editing in joint DNA. Conclusions: Intra-articular delivery of CRISPR/Cas9 via cationic liposomes enables safe and effective localized genome editing, representing a promising strategy for treating joint manifestations of MPS I.
View details for DOI 10.3390/pharmaceutics18030281
View details for Web of Science ID 001726943100001
View details for PubMedID 41900767
View details for PubMedCentralID PMC13029379
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Molecular Profile of Mucopolysaccharidosis Type I Patients in Brazil.
Journal of inherited metabolic disease
2026; 49 (1): e70144
Abstract
Mucopolysaccharidosis type I (MPS I) is an autosomal recessive disease caused by mutations in the IDUA gene, resulting in decreased activity of the lysosomal enzyme alpha-l-iduronidase (IDUA) and consequent accumulation of glycosaminoglycans in the lysosomes. There are more than 300 disease-causing variants reported in the IDUA gene, and the mutational profile varies considerably worldwide. In this study, we performed molecular analysis on 119 MPS I patients, representing the largest Brazilian cohort studied so far. Forty-seven different mutations were identified in our sample, and 13 of them are newly described: c.48delG, c.78delC, c.159-23_159-1del23, p.Gln125Ter, p.Trp175Ter, c.590-6ins4G, c.763delC, c.973-1G>A, p.Asp349Glu, p.Asn350Lys, p.Lys384Asn, c.1403-12_1403-4del9, and p.Lys546Ter. In silico analysis of novel variants suggests they are possibly pathogenic, supporting previous biochemical and clinical diagnoses. Among recurrent mutations, p.Trp402Ter and p.Pro533Arg are the most frequent in Brazil, found in 42.4% and 16% of the alleles, respectively. These results reveal the great allelic heterogeneity of IDUA variants in Brazilian patients. Unraveling the genetic profile of MPS I patients may improve the diagnosis and management of this rare disease.
View details for DOI 10.1002/jimd.70144
View details for PubMedID 41582445
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Newborn Intravenous Injection of Liposomal CRISPR/Cas9 Complex Has No Incidence of Off-Targets or Tumors in Mice.
Pharmaceutics
2025; 17 (5)
Abstract
Background: Genome editing at specific loci is an innovative therapeutic approach; however, it faces many challenges, so optimizing delivery vectors is essential to enhance the safety and efficacy of the CRISPR/Cas9 system. This study investigated whether the hydrodynamic administration of liposomal CRISPR/Cas9 complexes (LCs) in newborn mice induces off-target events or tumors. Methods: Liposomes were obtained through microfluidization. The CRISPR/Cas9 plasmid and a donor plasmid containing the Idua cDNA (alpha-L-iduronidase enzyme) were incorporated by adsorption, and complexes (LCs) were characterized regarding physicochemical properties. C57BL/6 newborn mice were divided in two groups, one received the complexes through hydrodynamic intravenous injection (n = 15) and the other was used as control (n = 15). After 21 months, mice were euthanized and organs were analyzed regarding histological characteristics. Lungs and liver were analyzed by qPCR searching for potential off-target sites in chromosomes 2, 5, 11, and 17 and on-target site in chromosome 6, identified by COSMID. Sequences were analyzed using an ICE tool for indels detection. Results: LCs exhibited 136 nm mean vesicle diameter with PDI below 0.15 and a zeta potential around +43 mV. Immediate biodistribution was predominant in the lungs and liver. There was no significant increase in tumor induction (20% in LCs vs. 33% in control). Molecular analyses indicated 0% off-target effects and around 3% on-target events. Conclusions: We conclude that this set of experiments demonstrates the potential of the chosen gRNA sequence to perform safe gene editing at the murine ROSA26 locus, corroborating the safety of the CRISPR/Cas9 gene editing platform.
View details for DOI 10.3390/pharmaceutics17050656
View details for PubMedID 40430946
View details for PubMedCentralID PMC12114706
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Genome-edited autologous stem cell transplantation with enhanced brain conditioning to correct progranulin deficiency
ACADEMIC PRESS INC ELSEVIER SCIENCE. 2025
View details for DOI 10.1016/j.ymgme.2024.108687
View details for Web of Science ID 001428859700075
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Hematopoietic stem cell transplantation for lysosomal disorders: Unraveling the mechanisms for CNS repopulation
ACADEMIC PRESS INC ELSEVIER SCIENCE. 2025: 5-6
View details for DOI 10.1016/j.ymgme.2024.108641
View details for Web of Science ID 001428859700034
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Basic and translational research in rare diseases in low- and middle-income countries: challenges and solutions.
Journal of community genetics
2024
Abstract
In this document, we discuss the main difficulties faced by investigators in low- and middle-income countries (LMICs) and propose potential solutions. Challenges include the scarcity of experts in rare disorders, higher costs of supplies, underfunding, and limited patient advocacy groups. Establishing collaborations among patient advocacy groups, governments, investigators, and other stakeholders to create action plans can address many of these issues and promote research into rare diseases.
View details for DOI 10.1007/s12687-024-00759-y
View details for PubMedID 39671061
View details for PubMedCentralID 10562568
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Laronidase-loaded liposomes reach the brain and other hard-to-treat organs after noninvasive nasal administration.
International journal of pharmaceutics
2024: 124355
Abstract
Mucopolysaccharidosis type I (MPS I) is caused by lack of the lysosomal enzyme α-L-iduronidase (IDUA), responsible for the degradation of the glycosaminoglycans (GAGs) dermatan and heparan sulfate, leading to multisystemic signs and symptoms. Enzyme replacement therapy (ERT) is a treatment that consists of weekly intravenous administrations of laronidase, a recombinant version of IDUA. However, ERT have limited access to certain tissues, such as bone, cartilage, and brain, and laronidase fails to trespass the BBB. In this sense, this study reports the development and characterization of laronidase-loaded liposomes for the treatment of MPS I mice. Liposomal complexes were obtained by the thin film formation method followed by microfluidization. The main characterization results showed vesicle size of 103.0 ± 3.3 nm, monodisperse populations of vesicles, zeta potential around + 30.0 ± 2.1 mV, and mucoadhesion strength of 5.69 ± 0.14 mN. Treatment of MPS I mouse fibroblasts showed significant increase in enzyme activity. Nasal administration of complexes to MPS I mice resulted in significant increase in laronidase activity in the brain cortex, heart, lungs, kidneys, eyes, and serum. The overall results demonstrate the feasibility of nasal administration of laronidase-loaded liposomes to deliver enzyme in difficult-to-reach tissues, circumventing ERT issues and bringing hope as a potential treatment for MPS I.
View details for DOI 10.1016/j.ijpharm.2024.124355
View details for PubMedID 38897489
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Clinical development of autologous genome-edited hematopoietic stem cells to treat mucopolysaccharidosis type I
ACADEMIC PRESS INC ELSEVIER SCIENCE. 2024
View details for DOI 10.1016/j.ymgme.2023.108000
View details for Web of Science ID 001192169300262
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Genome-edited hematopoietic stem cells as a curative approach for Gaucher disease type 1
ACADEMIC PRESS INC ELSEVIER SCIENCE. 2024
View details for DOI 10.1016/j.ymgme.2023.107998
View details for Web of Science ID 001192169300260
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Editorial: Genome editing in stem cells.
Frontiers in genome editing
2024; 6: 1357369
View details for DOI 10.3389/fgeed.2024.1357369
View details for PubMedID 38304247
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Pilot study of newborn screening for six lysosomal diseases in Brazil
MOLECULAR GENETICS AND METABOLISM
2023; 140 (1-2): 107654
Abstract
Lysosomal diseases (LDs) are progressive life-threatening disorders that are usually asymptomatic at birth. Specific treatments are available for several LDs, and early intervention improves patient's outcomes. Thus, these diseases benefit from newborn screening (NBS). We have performed a pilot study for six LDs in Brazil by tandem mass spectrometry.Dried blood spot (DBS) samples of unselected newborns were analyzed by the Neo-LSD™ kit (Perkin-Elmer) by MS/MS. Samples with low enzyme activity were submitted to the evaluation of specific biomarkers by ultra-performance liquid chromatography tandem-mass spectrometry as the second-tier, and were analyzed by a next-generation sequencing (NGS) multi-gene panel as the third-tier. All tests were performed in the same DBS sample.In 20,066 newborns analyzed, 15 samples showed activity of one enzyme below the cutoff. Two newborns had biochemical and molecular results compatible with Fabry disease, and five newborns had biochemical results and pathogenic variants or variants of unknown significance (VUS) in GAA.This study indicates that the use of enzyme assay as the first-tier test gives an acceptably low number of positive results that requires second/third tier testing. The possibility to run all tests in a DBS sample makes this protocol applicable to large-scale NBS programs.
View details for DOI 10.1016/j.ymgme.2023.107654
View details for Web of Science ID 001123690900001
View details for PubMedID 37507255
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Microglia Replacement with Bone Marrow-Derived Cells after Transient Inhibition of the Colony-Stimulating Factor 1 Receptor (CSF1R) is Superior to Standard Myeloablative Conditioning in Neuropathic Lysosomal Storage Diseases
CELL PRESS. 2023: 160-161
View details for Web of Science ID 001045144200296
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Engineering Human Hematopoietic Stem and Progenitor Cells for Lineage-Specific Expression of Galactocerebrosidase Using Genome Editing
CELL PRESS. 2023: 782
View details for Web of Science ID 001045144203407
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Ex vivo gene therapy for lysosomal storage disorders: future perspectives
EXPERT OPINION ON BIOLOGICAL THERAPY
2023: 1-12
Abstract
Lysosomal storage disorders (LSD) are a group of monogenic rare diseases caused by pathogenic variants in genes that encode proteins related to lysosomal function. These disorders are good candidates for gene therapy for different reasons: they are monogenic, most of lysosomal proteins are enzymes that can be secreted and cross-correct neighboring cells, and small quantities of these proteins are able to produce clinical benefits in many cases. Ex vivo gene therapy allows for autologous transplant of modified cells from different sources, including stem cells and hematopoietic precursors.Here, we summarize the main gene therapy and genome editing strategies that are currently being used as ex vivo gene therapy approaches for lysosomal disorders, highlighting important characteristics, such as vectors used, strategies, types of cells that are modified and main results in different disorders.Clinical trials are already ongoing, and soon approved therapies for LSD based on ex vivo gene therapy approaches should reach the market.
View details for DOI 10.1080/14712598.2023.2192348
View details for Web of Science ID 000954082500001
View details for PubMedID 36920351
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Diagnosis and Emerging Treatment Strategies for Mucopolysaccharidosis VII (Sly Syndrome).
Therapeutics and clinical risk management
2022; 18: 1143-1155
Abstract
Mucopolysaccharidosis VII (MPS VII, Sly syndrome) is an ultra-rare lysosomal disease caused by a deficiency of the enzyme β-glucuronidase (GUS). The diagnosis is suspected based on a range of symptoms that are common to many other MPS types, and it is confirmed through biochemical and molecular studies. Besides supportive treatment, current and emerging treatments include enzyme replacement therapy, hematopoietic stem cell transplantation, and gene therapy. This review summarizes the clinical manifestations, diagnosis, and emerging treatments for MPS VII.
View details for DOI 10.2147/TCRM.S351300
View details for PubMedID 36578769
View details for PubMedCentralID PMC9791935
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Experience of the NPC Brazil Network with a Comprehensive Program for the Screening and Diagnosis of Niemann-Pick Disease Type C
INTERNATIONAL JOURNAL OF NEONATAL SCREENING
2022; 8 (3)
Abstract
Niemann-Pick disease type C (NPC) is a lysosomal disorder caused by impaired cholesterol metabolism. Levels of lysosphingomyelin 509 (LysoSM509) have been shown elevated in dried blood spots (DBS) of NPC and acid sphingomyelinase deficiency patients. In this study, we report our experience using a two-tier approach (1st tier is the quantification of lysoSM509 by ultra-performance liquid chromatography tandem mass spectrometry followed by the 2nd tier with next-generation sequencing of the NPC1 and NPC2 genes). DBS samples from 450 suspected patients were received by the NPC Brazil network. Of these, 33 samples had elevated levels of lysoSM509, and in 25 of them, variants classified as pathogenic, likely pathogenic, or of unknown significance were identified in the NPC1 or NPC2 genes by next-generation sequencing. The quantification of lysoSM509 in DBS as a first-tier test for the diagnosis of NPC followed by molecular analysis of the NPC1 and NPC2 genes almost doubled the detection rate when compared to the performance of chitotriosidase activity as a first-tier biomarker, and it could likely be increased with the addition of a third tier with MLPA of the two genes involved. This strategy seems suitable for the neonatal screening (NBS) of NPC if this disease is eventually adopted by NBS programs.
View details for DOI 10.3390/ijns8030039
View details for Web of Science ID 000858300700001
View details for PubMedID 35892469
View details for PubMedCentralID PMC9326630
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Biochemical diagnosis of aromatic-L-amino acid decarboxylase deficiency (AADCD) by assay of AADC activity in plasma using liquid chromatography/tandem mass spectrometry
MOLECULAR GENETICS AND METABOLISM REPORTS
2022; 32: 100888
Abstract
Aromatic l-amino acid decarboxylase (AADC, EC 4.1.1.28) deficiency is a rare genetic disorder characterized by developmental delay, oculogyric crises, autonomic dysfunction and other problems, caused by biallelic mutations in the DDC gene leading to deficient activity of aromatic l-amino acid decarboxylase, an enzyme involved in the formation of important neurotransmitters, such as dopamine and serotonin. A clinical development program of gene therapy for AADC deficiency is ongoing. An important step for the success of this therapy is the early and precise identification of the affected individuals, but it has been estimated that around 90% of the cases remain undiagnosed. The availability measurement of the AADC activity is mandatory for an accurate biochemical diagnosis. Based on these statements, our objectives were to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) method suitable for the determination of the AADC activity, and to evaluate its capacity to confirm the deficiency of AADC in potential patients in Brazil. The AADC activities were measured in plasma samples of seven AADC deficient patients and 35 healthy controls, after enzymatic reaction and LC-MS/MS analysis of dopamine, the main reaction product. The results obtained showed clear discrimination between confirmed AADC deficient patients and healthy controls. The method presented here could be incorporated in the IEM laboratories for confirmation of the diagnosis of when a suspicion of AADC deficiency is present due to clinical signs and/or abnormal biomarkers, including when an increased level of 3-O-methyldopa (3-OMD) is found in dried blood spots (DBS) samples from high-risk patients or from newborn screening programs.
View details for DOI 10.1016/j.ymgmr.2022.100888
View details for Web of Science ID 000836463600003
View details for PubMedID 35769135
View details for PubMedCentralID PMC9234702