Kyproula Christodoulou

Autosomal recessive spinocerebellar ataxia type 10 (SCAR10) is a rare, slowly progressive neurodegenerative disorder characterised by ataxia, cerebellar atrophy, and oculomotor abnormalities, caused by variants in the ANO10 (anoctamin 10) gene. While ANO10-mediated calcium dysregulation in Purkinje cells has been proposed to cause SCAR10, the precise pathogenic mechanism remains unclear. To investigate cellular responses to ANO10 deficiency, four ANO10-mutant neuronal cell lines - a knockout and three lines harbouring recurrent SCAR10-associated pathogenic variants - were engineered using CRISPR/Cas9 editing, expanded by clonal isolation, and characterised by flow cytometry. Mass spectrometry-based proteomics identified differentially expressed proteins between control and mutant lines, and bioinformatic analyses uncovered candidate pathways involved in disease pathogenesis. Comparative proteomic analysis revealed disruptions in synaptic function, cell cycle regulation, extracellular matrix remodelling, and immune homeostasis as candidate pathways potentially contributing to SCAR10 pathogenesis. All four processes are functionally linked to calcium signalling, aligning with previous reports implicating abnormal calcium homeostasis in spinocerebellar ataxias. This study provides, for the first time, insights into the proteomic profile of SCAR10 using cell-based models and highlights specific molecules and pathways that may contribute to disease pathogenesis. These findings offer a foundation for further investigation and experimental validation, with potential implications for the development of targeted therapeutic approaches.

Systemic Sclerosis (SSc) is an autoimmune rheumatic disease (ARD) with unclear aetiopathogenesis. Disease prognosis, diagnosis, and treatment are challengi

Background/Objectives: X-linked intellectual disability (XLID) is a highly heterogeneous disorder accounting for ~10% of all males with ID. Next-generation sequencing (NGS) has revolutionized the discovery of causal XLID genes and variants; however, many cases remain unresolved. We present a four-generation syndromic XLID family with multiple males exhibiting variable degree of ID, focal dystonia and epilepsy. Methods: Extensive cytogenetic and targeted genetic testing was initially performed, followed by whole-exome sequencing (WES) and short-read whole-genome sequencing (WGS). Apart from the routine NGS analysis pipelines, sequencing data was revisited by focusing on poorly covered/mapped regions on chromosome X (chrX), to potentially reveal unidentified clinically relevant variants. Candidate variant validation and family segregation analysis were performed with Sanger sequencing. Results: All initial diagnostic testing was negative. Subsequently, 300 previously reported “dark” chrX coding DNA sequences, overlapping 97 genes, were cross-checked against 29 chrX genes highly associated (p < 0.05) with ID and focal dystonia, according to Phenomizer. Manual inspection of the existing NGS data in two low-coverage regions, chrX:25013469-25013696 and chrX:111744737-111744820 (hg38), revealed a recurrent pathogenic ARX variant NM_139058.3:c.441_464dup p.(Ala148_Ala155dup) (ARXdup24) associated with non-syndromic or syndromic XLID, including Partington syndrome. Sanger sequencing confirmed ARXdup24 in all affected males, with carrier status in their unaffected mothers, and absence in other unaffected relatives. Conclusions: After several years of diagnostic odyssey, the pathogenic ARXdup24 variant was unmasked, supporting a genotype–phenotype correlation in the first Partington syndrome family in Cyprus. This study highlights that re-examining underrepresented genomic regions and using phenotype-driven tools can provide critical diagnostic insights in unresolved XLID cases.

Ataxia with oculomotor apraxia type 1 (AOA1) is a recently described autosomal-recessive neurodegenerative condition of childhood onset. It is caused by mutations in the APTX gene, which encodes the protein aprataxin. Clinical features include gait and limb ataxia, dysarthria, oculomotor apraxia, mild peripheral neuropathy and progression of neurological deficits.1 Some patients manifest parkinsonian symptoms or mental retardation, although the latter has been reported predominantly in Japanese patients.2 We report a patient with homozygous deletion of APTX, who presented with behavioural changes (social withdrawal), and subsequent rapid progression of neurological symptoms associated with severe cognitive decline. We suggest that complete deletion of APTX is associated with a more severe phenotype than that associated with point mutations.

Chemical analysis of the hydroethanolic Hypericum repens L. extracts was performed using the LC/Q-TοF/HRMS technique. The majority of compounds identified belonged to phenolics, particularly flavonoids. The extract was also studied for its possible bioactivities, demonstrating high antioxidant properties compared to the control (IC₅₀ values ranging from 4.6 to 9.42 μg/mL). Significant antibacterial activity was also detected against Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, and Salmonella enteritidis, with MIC values from 125 to 625 μg/mL. S. aureus presented the highest susceptibility among all bacteria tested. Additionally, the extract showed very mild anticholinesterase activity in the AChE and BChE inhibition assays. These findings provide the first insights into the phytochemical composition, as well as the antioxidant, antimicrobial, and anticholinesterase activities of H. repens extract, suggesting that the endemic Cyprus H. repens is a valuable natural rich source of bioactive compounds with a potentially broad range of bioactivities.

Dominantly inherited intronic GAA repeat expansions in the fibroblast growth factor 14 gene have recently been shown to cause spinocerebellar ataxia 27B. Currently, the pathogenic threshold of (GAA)≥300 repeat units is considered highly penetrant, while (GAA)250–299 is likely pathogenic with reduced penetrance. This study investigated the frequency of the GAA repeat expansion and the phenotypic profile in a Cypriot cohort with unresolved late-onset cerebellar ataxia. We analysed this trinucleotide repeat in 155 patients with late-onset cerebellar ataxia and 227 non-neurological disease controls. The repeat locus was examined by long-range PCR followed by fragment analysis using capillary electrophoresis, agarose gel electrophoresis and automated electrophoresis. A comprehensive comparison of all three electrophoresis techniques was conducted. Additionally, bidirectional repeat-primed PCRs and Sanger sequencing were carried out to confirm the absence of any interruptions or non-GAA motifs in the expanded alleles. The (GAA)≥250 repeat expansion was present in 12 (7.7%) patients. The average age at disease onset was 60 ± 13.5 years. The earliest age of onset was observed in a patient with a (GAA)287 repeat expansion, with ataxia symptoms appearing at 25 years of age. All patients with spinocerebellar ataxia 27B displayed symptoms of gait and appendicular ataxia. Nystagmus was observed in 41.7% of the patients, while 58.3% exhibited dysarthria. Our findings indicate that spinocerebellar ataxia 27B represents the predominant aetiology of autosomal dominant cerebellar ataxia in the Cypriot population, as this is the first dominant repeat expansion ataxia type detected in this population. Given our results and existing research, we propose including fibroblast growth factor 14 GAA repeat expansion testing as a first-tier genetic diagnostic approach for patients with late-onset cerebellar ataxia.

Euphrasia officinalis L., commonly known as eyebright, is a medicinal plant used in folk medicine for eye disorders and memory loss. Due to its abundance of compounds with proven neuroprotective properties, there has been growing interest in exploring eyebright’s potential health benefits, particularly for preventing or treating neurodegenerative diseases like Alzheimer’s disease. Here, seven distinct extracts were generated using solvents of different polarities, consecutively, from plants grown in Greece. The extracts were chemically characterized and assessed for their antioxidant, anticholinesterase, and anti-neurotoxic potentials. Our findings demonstrated eyebright’s notable antioxidant capacity with five extracts exhibiting significant anti-neurotoxic properties by enhancing cell viability by 17.5 to 22.6% in human neuroblastoma cells exposed to neurotoxic amyloid-beta peptides. The ethyl acetate and butanolic extracts were the most effective across all assays, likely due to their high concentrations of active compounds. Therefore, eyebright may be harnessed for developing functional foods, supplements, and pharmaceuticals with potential benefits against Alzheimer’s disease. This study marks the first identification of neuroprotective properties in a Euphrasia species, highlighting its broader therapeutic potential and paving the way for further research.

The purpose of this study was to identify the saponin and phenolic components in root extracts of Saponaria officinalis, a widespread species, found in Cyprus. A total of six major saponins, including gypsogenin and gypsogenic acid derivatives, as well as saponariosides C, D, and E, were identified using UHPLC/Q-TOF-MS analysis, with gypsogenin derivatives being the most common saponins detected through quantitative analysis. A total of six phenolic compounds were also identified, including rutin, quercetin galactoside, syringic acid, apigenin, protocatechuic, and vanillic acid. In addition to their saponin and phenolic contents, the root extracts were prepared through different extraction methods, and their biological activity was assessed. All samples demonstrated antioxidant capacity, as well as antibacterial activity, against four bacterial strains (Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, and Salmonella enteritidis), with the acetone extract presenting higher susceptibility. The evaluation of anticancer activity in A375 (human malignant melanoma), HeLa (human cervical epithelioid carcinoma), and HaCaT (healthy human keratinocytes) cell lines revealed that the acetone extract of S. officinalis extract demonstrated a significant inhibitory effect on the proliferation of A375 cells in a concentration-dependent manner. None of the extracts demonstrated anti-neurotoxic potential against Aβ_25–35 cytotoxic peptides. The results of this study support previous findings that reveal that the Saponaria species are an excellent natural source of biologically active compounds with antioxidant, antimicrobial, and anticancer properties.

The intronic biallelic AAGGG expansion in the replication factor C subunit 1 (RFC1) gene was recently associated with a phenotype combining cerebellar ataxia, neuropathy, and vestibular areflexia syndrome, as well as with late-onset ataxia. Following this discovery, studies in multiple populations extended the phenotypic and genotypic spectrum of this locus. Multiple benign and additional pathogenic configurations are currently known. Our main objectives were to study the prevalence of the pathogenic AAGGG expansion in the Cypriot population, to further characterize the RFC1 repeat locus allele distribution, and to search for possible novel repeat configurations. Cypriot undiagnosed patients, in the majority presenting at least with cerebellar ataxia and non-neurologic disease controls, were included in this study. A combination of conventional methods was used, including standard PCR flanking the repeat region, repeat-primed PCR, long-range PCR, and Sanger sequencing. Bioinformatics analysis of already available in-house short-read whole-genome sequencing data was also performed. A large group of undiagnosed patients (n = 194), mainly presenting with pure ataxia or with ataxia accompanied by neuropathy or additional symptoms, as well as a group of non-disease controls (n = 100), were investigated in the current study. Our findings include the diagnosis of 10 patients homozygous for the pathogenic AAGGG expansion and a high percentage of heterozygous AAGGG carriers in both groups. The benign AAAAG_n, AAAGG_n, and AAGAG_n configurations were also identified in our cohorts. We also report and discuss the identification of 2 recently reported novel and possibly benign repeat configurations, AAAGGG_n and AAGAC_n, thus confirming their existence in another distinct population, and we highlight an increased frequency of the AAAGGG_n in the patient group, including a single case of homozygosity. Our findings indicate the existence of genetic heterogeneity regarding the RFC1 repeat configurations and that the AAGGG pathogenic expansion is a frequent cause of ataxia in the Cypriot population.