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Fanconi Anemia (FA) is a debilitating genetic disorder with a wide range of severe symptoms including bone marrow failure and predisposition to cancer. CRISPR-Cas genome editing manipulates genotypes by harnessing DNA repair and has been proposed as a potential cure for FA. But FA is caused by deficiencies in DNA repair itself, preventing the use of editing strategies such as homology directed repair. Recently developed base editing (BE) systems do not rely on double stranded DNA breaks and might be used to target mutations in FA genes, but this remains to be tested. Here we develop a proof of concept therapeutic base editing strategy to address two of the most prevalent FANCA mutations in patient hematopoietic stem and progenitor cells. We find that optimizing adenine base editor construct, vector type, guide RNA format, and delivery conditions leads to very effective genetic modification in multiple FA patient backgrounds. Optimized base editing restored FANCA expression, molecular function of the FA pathway, and phenotypic resistance to crosslinking agents. ABE8e mediated editing in primary hematopoietic stem and progenitor cells from FA patients was both genotypically effective and restored FA pathway function, indicating the potential of base editing strategies for future clinical application in FA., Nature Communications, 13 (1), ISSN:2041-1723

Self-rated health (SRH) is an independent determinant for all-cause mortality. We aimed to examine the independent and combined associations of components of physical fitness with SRH at baseline (cross-sectional) and two years later (longitudinal) in children and adolescents. Spanish youth (N = 1378) aged 8 to 17.9 years participated at baseline. The dropout rate at 2-year follow-up was 19.5% (n = 270). Participants were categorized as either children (8 to 11.9 years age) or adolescents (12 to 17.9 years age). The ALPHA health- related fitness test battery for youth was used to assess physical fitness, and SRH was measured by a single-item question. Cumulative link, ANOVA and ANCOVA models were fitted to analyze the data. Cardiorespiratory fitness, relative upper body isometric muscular strength, muscular strength score, and global physical fitness were positively associated with SRH in children (OR, 1.048, 95% CI, 1.020&ndash, 1.076, OR, 18.921, 95% CI, 3.47&ndash, 104.355, OR, 1.213, 95% CI, 1.117&ndash, 1.319, and OR, 1.170, 95% CI, 1.081&ndash, 1.266, respectively, all p &lt, 0.001) and adolescents (OR, 1.057, 95% CI, 1.037&ndash, OR, 5.707, 95% CI, 1.122&ndash, 29.205, OR, 1.169, 95% CI, 1.070&ndash, 1.278, and OR, 1.154 95% CI, 1.100&ndash, 1.210, respectively, 0.001), and motor fitness was positively associated with SRH only in adolescents at baseline (OR, 1.192, 95% CI, 1.066&ndash, 1.309, p &lt, 0.01). Cardiorespiratory fitness and global physical fitness were positively associated with SRH in children two years later (OR, 1.056, 95% CI, 1.023&ndash, 1.091, 0.001, and OR, 1.082, 95% CI, 1.031&ndash, 1.136, 0.01, respectively). Only cardiorespiratory fitness was independently associated with SRH in children and adolescents at baseline (OR, 1.059, 95% CI, 1.029&ndash, 1.090, and OR, 1.073, 95% CI, 1.050&ndash, 1.097, respectively, both p &lt, 0.001) and two years later (OR, 1.075, 95% CI, 1.040&ndash, 1.112, and OR, 1.043, 95% CI, 1.014&ndash, 1.074, 0.01, respectively). A high level of cardiorespiratory fitness at baseline or maintaining high levels of cardiorespiratory fitness from the baseline to 2-year follow-up were associated with a higher level of SRH at 2-year follow-up in children (p &lt, 0.01) and adolescents (p &lt, 0.05). These findings emphasize the importance of cardiorespiratory fitness as strong predictor of present and future SRH in youth. Intervention programs to enhance cardiorespiratory fitness level of the youth population are urgently needed for present and future youth&rsquo, s health.

Gene targeting constitutes a new step in the development of gene therapy for inherited diseases. Although previous studies have shown the feasibility of editing fibroblasts from Fanconi anemia (FA) patients, here we aimed at conducting therapeutic gene editing in clinically relevant cells, such as hematopoietic stem cells (HSCs). In our first experiments, we showed that zinc finger nuclease (ZFN)‐mediated insertion of a non‐therapeutic EGFP‐reporter donor in the AAVS1 “safe harbor” locus of FA‐A lymphoblastic cell lines (LCLs), indicating that FANCA is not essential for the editing of human cells. When the same approach was conducted with therapeutic FANCA donors, an efficient phenotypic correction of FA‐A LCLs was obtained. Using primary cord blood CD34+ cells from healthy donors, gene targeting was confirmed not only in in vitro cultured cells, but also in hematopoietic precursors responsible for the repopulation of primary and secondary immunodeficient mice. Moreover, when similar experiments were conducted with mobilized peripheral blood CD34+ cells from FA‐A patients, we could demonstrate for the first time that gene targeting in primary hematopoietic precursors from FA patients is feasible and compatible with the phenotypic correction of these clinically relevant cells.