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Biology subjects

AMENDOLA, M.

Publications and source records attributed to AMENDOLA, M..

2 recordsLinked to original sources

Base editing of β0 thalassemia mutations as a therapeutic strategy for β-hemoglobinopathies: efficacy and genotoxicity studies

Gene therapy has emerged as a promising curative treatment for {beta}-hemoglobinopathies, the most common genetic disorders worldwide. However, current approved approaches still have some limitations in terms of safety and efficacy. Here, we used highly processive adenine base editors (ABE) variants to precisely correct some of the most prevalent and severe {beta}-thalassemia-causing mutations in the {beta}-globin gene. Efficient editing of hematopoietic stem/progenitor cells (HSPCs) led to potent {beta}-globin expression in their erythroid progeny and persistent correction of both {beta}-thalassemia and sickle cell-{beta}-thalassemia phenotypes. Safety of this strategy was confirmed in HSPCs in vitro and in vivo by the absence of gene dysregulation or any meaningful impact on the DNA mutational burden, the RNA deamination level, the {beta}-globin gene locus integrity and the clonality of the HSPC graft. Overall, base editing-mediated gene correction is a safe and effective strategy for treating {beta}-hemoglobinopathies. One sentence summaryPreclinical safety and efficacy studies of a new gene therapy approach for patients with severe {beta}-hemoglobinopathies.

genetics↗

Novel cytometry-based characterization of lysosomal storage disease affected patient's cells

Wolman disease (WD) is a severe lysosomal storage disorder characterized by fatal lipid accumulation caused by the deficiency of a lipid metabolic enzyme, Lysosomal Acid Lipase (LAL), involved in the lysosomal hydrolysis of cholesterols and triglycerides. Due to the imbalance of lipids homeostasis, WD patients suffer from severe hepatosplenomegaly, hepatic failure and adrenal calcification resulting in a premature infant death within the first year of age. In this work, we explored multiple imaging analyses to fully characterize the phenotype of LAL deficient cells. In particular, we stained WD patients fibroblasts for intracellular lipid droplets (LD) and lysosomes and we analysed staining intensity and granularity as well as an increased number of LD and lysosomes using fluorescence wide field microscopy, confocal microscopy, conventional and image flow cytometry. Noteworthy, we showed that lipid homeostasis was restored upon delivery of a functional LAL transgene. Finally, since fibroblasts cannot be used as routine clinical test as they are difficult to collect from WD patients, we confirmed our observations in LAL deficient human blood cell lines and in peripheral blood mononuclear cells (PBMC) from LAL deficient (LAL-D) mouse model, as a proxy for easily accessible WD PBMC. Overall, we expect that this novel imaging analysis pipeline will help to diagnose WD, follow its progression and evaluate the success of enzyme replacement therapy or gene correction strategies for WD as well as other lysosomal storage disorders.

pathology↗