Biology subjects

Samarut, E.

Publications and source records attributed to Samarut, E..

4 recordsLinked to original sources

bioRxiv2026

Repurposing the HMG-CoA Reductase Inhibitor Atorvastatin for SRD5A3-CDG

SRD5A3-CDG is a rare autosomal recessive congenital disorder of glycosylation characterized by multisystemic dysfunction, including neurological, psychomotor, cognitive, and visual impairments. Approximately 60 cases have been reported, with treatment limited to symptomatic management. SRD5A3 encodes a polyprenal reductase enzyme essential for synthesizing dolichol, a lipid carrier of the oligosaccharide precursor in N-glycosylation. To address the lack of effective treatments and disease models suitable for high-throughput screening, we developed the first C. elegans model of SRD5A3-CDG, harboring the homozygous W19X nonsense mutation commonly observed in patients. This model recapitulates disease-relevant phenotypes, including developmental delays, neurological dysfunction, and mevalonate pathway dysregulation. Using this model, we conducted a high-throughput motility-based drug repurposing screen and identified atorvastatin, an FDA-approved HMG-CoA reductase inhibitor, as a repurposing candidate. Atorvastatin rescued disease-relevant phenotypes in the worm model and restored polyprenol-to-dolichol ratios in patient fibroblasts. These findings highlight atorvastatin as a promising drug repurposing candidate for SRD5A3-CDG.

cell biology↗

bioRxiv2025

Optimized CRISPR/Cas9 Electroporation and Single Cell Cloning Protocol for Generating Pure Cellular Models in Human Immortalized Myoblasts

BackgroundGenome editing in human skeletal muscle research requires protocols that maximize delivery while preserving viability and clonal outgrowth. We sought to develop a reagent-free workflow for CRISPR/Cas9 editing in human immortalized myoblasts and to demonstrate its performance in two use cases, an IARS1 knockout and an MLIP homozygous knock-in. MethodsWe optimized electroporation parameters using a green fluorescent protein reporter to compare three electrical settings for transfection and survival in E6/E7 myoblasts, then applied ribonucleoprotein delivery for editing. We evaluated the effect of confluency at electroporation, performed single-cell cloning without antibiotics or fluorescence-activated sorting, and validated edits by high-resolution melting pre-screen followed by Sanger sequencing. ResultsElectroporation optimization identified one parameter set that maximized delivery while preserving viability. Performing electroporation at low confluency increased clonal outgrowth and editing rates. The workflow yielded an 84% success rate for IARS1 knockout and a 3.3% success rate for MLIP homozygous knock-in. High-resolution melting provided a very sensitive pre-screen, detecting 96% to 100% of actual edits, reducing the number of Sanger sequencing needed. Performance was reproducible across runs and myoblast lines and increasing single-cell seeding scaled yields without compromising purity. ConclusionsThis work provides a practical and reproducible selection-free protocol that couples electroporation optimization, low confluency editing, single-cell cloning, and high-resolution melting sorting to generate pure edited myoblast lines. The approach is applicable to disease modeling in neuromuscular research and clarifies feasibility boundaries for essential genes and homology-directed repair in these cells.

cell biology↗

bioRxiv2024

PARP inhibitor counteracts Temozolomide Resistance in Glioblastoma Multiforme

BackgroundGlioblastoma multiforme (GBM) is the most common malignant primary brain tumour in adults and is invariably associated with poor prognosis. Resistance to Temozolomide (TMZ), the standard chemotherapeutic agent, remains a major clinical challenge, particularly due to DNA mismatch repair (MMR) deficiencies. The aim of this study was to determine whether combining TMZ with the poly(ADP-ribose) polymerase inhibitor Olaparib (OLA) could overcome TMZ resistance in GBM. MethodsWe conducted in vitro experiments using U251 cell-line, including a TMZ-resistant derivative, and primary GBM cultures derived from patient tumours. A CRISPR/Cas9 knockout screen was employed to identify genes involved in TMZ resistance. Cell viability, proliferation, and morphology were assessed following treatment with TMZ, OLA, or their combination. ResultsThe CRISPR screen identified inactivation of MMR pathway genes as key mediators of TMZ resistance. Co-treatment with OLA and TMZ demonstrated synergistic cytotoxicity in both parental and TMZ-resistant U251 cells, as well as in primary GBM cultures at diagnosis or relapse. Notably, OLA restored sensitivity to TMZ in MMR-deficient contexts and in tumours expressing O6-methylguanine-DNA-methyltransferase (MGMT). The combination treatment induced persistent DNA damage, cell cycle disruption, and cell death. ConclusionsThese findings provide strong preclinical evidence that combining TMZ with OLA can effectively overcome key mechanisms of TMZ resistance in GBM. This approach offers a promising therapeutic strategy warranting further clinical investigation. IMPORTANCE OF THE STUDYTemozolomide (TMZ) resistance remains a major therapeutic obstacle in glioblastoma (GBM), often driven by MMR deficiency or MGMT expression. While poly(ADP-ribose) polymerase (PARP) inhibitors have shown potential in other cancers, their role in overcoming TMZ resistance in GBM has remained unclear. In this study, a CRISPR screen identified MMR deficiency as a key driver of TMZ resistance. We further demonstrate that co-treatment with the PARP inhibitor Olaparib (OLA) restores TMZ sensitivity in both MMR-deficient and MGMT-expressing GBM cells and patient-derived cultures. These findings provide strong preclinical evidence supporting PARP inhibition as a promising therapeutic strategy to overcome chemoresistance in GBM and justify further clinical investigation. KEY POINTSO_LIPARP inhibitor Olaparib restores temozolomide sensitivity in resistant GBM cells C_LIO_LICombination therapy overcomes resistance driven by MMR deficiency or MGMT expression C_LIO_LIDual treatment induces persistent DNA damage and apoptosis in glioblastoma primary cultures C_LI

cancer biology↗

bioRxiv2024

RFC1 regulates the expansion of neural progenitors in the developing zebrafish cerebellum

DNA replication and repair are basic yet essential molecular processes for all cells. RFC1 encodes the largest subunit of the Replication Factor C (RFC), which is a clamp-loader during DNA replication and repair. Intronic repeat expansion in RFC1 has recently been associated with so-called RFC1-related disorders, which mainly encompass late-onset cerebellar ataxias. However, the mechanisms that make certain tissues more susceptible to defects in these universal pathways remain mysterious. In this study, we provide the first investigation of RFC1 gene function in vivo using zebrafish. We showed that RFC1 is expressed in neural progenitor cells within the developing cerebellum and that it is necessary to maintain these cells genomic integrity during neurogenic maturation. Accordingly, RFC1 loss-of-function leads to a severe cerebellar phenotype due to impaired neurogenesis of both Purkinje and granule cells. Our data thus point to a specific role of RFC1 in the developing cerebellum, paving the way for a better understanding of the pathogenic mechanisms underlying RFC1-related disorders.

developmental biology↗