Novel

Background and Objectives: ADP-ribosylation is a post-translational modification critical for DNA repair, chromatin remodeling, and cellular stress responses. The enzyme ARH3/ADPRHL2 (encoded by the Methods: Clinical identification and diagnosis of the patient was based primarily on MRI and whole exome sequencing. Functional assays were performed in patient fibroblast cell lines using Western blotting and immunofluorescence-based assays and compared with an ARH3 KO cell line generated by CRISPR/Cas9 technology. Results: Our data indicate that the novel p.Leu162Pro variant, combined with a known nonsense variant, c.316C>T; p.Gln106*, causes a severe reduction of ARH3 protein levels in patient fibroblasts. This leads to defective ARH3-dependent removal of ADP-ribosylation, both under basal conditions and following oxidative stress. Our results using minocycline indicate that it is a poor PARP1 inhibitor and was ineffective in reducing basal ADP-ribosylation in ARH3-deficient cells. Discussion: This study contributes to the growing body of knowledge on ARH3 deficiency associated with CONDSIAS, by identifying a new pathogenic variant and questioning the molecular rationale underlying the therapeutic use of minocycline in patients with CONDSIAS.