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Recessive dystrophic epidermolysis bullosa is a debilitating blistering skin disorder caused by loss-of-function mutations in COL7A1, which encodes type VII collagen, the main component of anchoring fibrils at the dermal-epidermal junction. Although conventional gene therapy approaches through viral vectors have been tested in preclinical and clinical trials, they are limited by transgene size constraints and only support unregulated gene expression. Genome editing could potentially overcome some of these limitations, and CRISPR/Cas9 has already been applied in research studies to restore COL7A1 expression. The delivery of suitable repair templates for the repair of DNA cleaved by Cas9 is still a major challenge, and alternative base editing strategies may offer corrective solutions for certain mutations. We show highly targeted and efficient cytidine deamination and molecular correction of a defined recessive dystrophic epidermolysis bullosa mutation (c.425A>G), leading to restoration of full-length type VII collagen protein expression in primary human fibroblasts and induced pluripotent stem cells. Type VII collagen basement membrane expression and skin architecture were restored with de novo anchoring fibrils identified by electron microscopy in base-edited human recessive dystrophic epidermolysis bullosa grafts recovered from immunodeficient mice. The results show the potential and promise of emerging base editing technologies in tackling inherited disorders with well-defined single nucleotide mutations., (© 2023 The Authors.)

Base editing introduces precise single-nucleotide edits in genomic DNA and has the potential to treat genetic diseases such as the blistering skin disease recessive dystrophic epidermolysis bullosa (RDEB), which is characterized by mutations in the COL7A1 gene and type VII collagen (C7) deficiency. Adenine base editors (ABEs) convert A-T base pairs to G-C base pairs without requiring double-stranded DNA breaks or donor DNA templates. Here, we use ABE8e, a recently evolved ABE, to correct primary RDEB patient fibroblasts harboring the recurrent RDEB nonsense mutation c.5047 C > T (p.Arg1683Ter) in exon 54 of COL7A1 and use a next generation sequencing workflow to interrogate post-treatment outcomes. Electroporation of ABE8e mRNA into a bulk population of RDEB patient fibroblasts resulted in remarkably efficient (94.6%) correction of the pathogenic allele, restoring COL7A1 mRNA and expression of C7 protein in western blots and in 3D skin constructs. Off-target DNA analysis did not detect off-target editing in treated patient-derived fibroblasts and there was no detectable increase in A-to-I changes in the RNA. Taken together, we have established a highly efficient pipeline for gene correction in primary fibroblasts with a favorable safety profile. This work lays a foundation for developing therapies for RDEB patients using ex vivo or in vivo base editing strategies., (© 2022. The Author(s).)

Most patients with early HR+ and HER2- breast cancer receive a hormone therapy; the clinical question still open is how to identify patients who can really benefit from adjuvant chemotherapy. The accurate identification of these patients is essential to avoid an over-treatment, increasing the risk of an unnecessary toxicity; on the contrary, the omission of chemotherapy can deprive high risk patients of a potential life-saving treatment (under-treatment). Several multigene assays (MGAs), assessing the risk of relapse according to the biological characteristics of the tumor, have been developed. To date, the 21-gene assay (Oncotype DX Breast Recurrence Score ® ) is the only test developed and validated to be actionable, i.e., able to predict the benefit of adjuvant chemotherapy. The different available tests can be classified according to their clinical utility based on their prognostic and predictive value. A prognostic test gives information about the outcome of the disease, regardless of the administered therapy. When the aim of the test is to drive the treatment decisions, the predictive component, and therefore the ability to accurately identify which patients could benefit from chemotherapy, is essential. This review summarizes the clinical evidences of the Oncotype DX ® test supporting its clinical utility., Competing Interests: CONFLICTS OF INTEREST Cognetti received honoraria from Glaxo Smith Kline S.R.L., Roche S.P.A., Pfizer S.R.L., Astrazeneca S.P.A., Eli Lilly, Genomic Health S.R.L., (Copyright: © 2021 Cognetti and Naso.)