Cytosine Deaminase Base Editing to Restore COL7A1 in Dystrophic Epidermolysis Bullosa Human: Murine Skin Model.

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.

Integrin-Targeted, Short Interfering RNA Nanocomplexes for Neuroblastoma Tumor-Specific Delivery Achieve MYCN Silencing with Improved Survival.

The authors aim to develop siRNA therapeutics for cancer that can be administered systemically to target tumors and retard their growth. The efficacy of systemic delivery of siRNA to tumors with nanoparticles based on lipids or polymers is often compromised by their rapid clearance from the circulation by the liver. Here, multifunctional cationic and anionic siRNA nanoparticle formulations are described, termed receptor-targeted nanocomplexes (RTNs), that comprise peptides for siRNA packaging into nanoparticles and receptor-mediated cell uptake, together with lipids that confer nanoparticles with stealth properties to enhance stability in the circulation, and fusogenic properties to enhance endosomal release within the cell. Intravenous administration of RTNs in mice leads to predominant accumulation in xenograft tumors, with very little detected in the liver, lung, or spleen. Although non-targeted RTNs also enter the tumor, cell uptake appears to be RGD peptide-dependent indicating integrin-mediated uptake. RTNs with siRNA against MYCN (a member of the Myc family of transcription factors) in mice with MYCN-amplified neuroblastoma tumors show significant retardation of xenograft tumor growth and enhanced survival. This study shows that RTN formulations can achieve specific tumor-targeting, with minimal clearance by the liver and so enable delivery of tumor-targeted siRNA therapeutics.