Missense variant c.1460 T > C (p.L487P) enhances protein degradation of ER mannosyltransferase ALG9 in two new ALG9-CDG patients presenting with West syndrome and review of the literature.

ALG9-CDG is a CDG-I defect within the group of Congenital Disorders of Glycosylation (CDG). We here describe the clinical symptoms of two new and unrelated ALG9-CDG patients, both carrying the novel homozygous missense variant c.1460 T > C (p.L487P) in the ALG9 gene which led to global developmental delay, psychomotor disability, facial dysmorphisms, brain and heart defects, hearing loss, hypotonia, as well as feeding problems. New clinical symptoms comprised West syndrome with hypsarrhythmia. Quantitative RT-PCR analysis revealed a significantly enhanced ALG9 mRNA transcript level, whereas the protein amount in fibroblasts was significantly reduced. This could be ascribed to a stronger degradation of the mutated ALG9 protein in patient fibroblasts. Lipid-linked oligosaccharide analysis showed an ALG9-CDG characteristic accumulation of Man6GlcNAc2-PP-dolichol and Man8GlcNAc2-PP-dolichol in patient cells. The clinical findings of our patients and of all previously published ALG9-CDG patients are brought together to further expand the knowledge about this rare N-glycosylation disorder. SYNOPSIS: Homozygosity for p.L487P in ALG9 causes protein degradation and leads to West syndrome.

Strong association of a common dihydropyrimidine dehydrogenase gene polymorphism with fluoropyrimidine-related toxicity in cancer patients.

BACKGROUND: Cancer patients carrying mutations in the dihydropyrimidine dehydrogenase gene (DPYD) have a high risk to experience severe drug-adverse effects following chemotherapy with fluoropyrimidine drugs such as 5-fluorouracil (5-FU) or capecitabine. The pretreatment detection of this impairment of pyrimidine catabolism could prevent serious, potentially lethal side effects. As known deleterious mutations explain only a limited proportion of the drug-adverse events, we systematically searched for additional DPYD variations associated with enhanced drug toxicity. METHODOLOGY/PRINCIPAL FINDINGS: We performed a whole gene approach covering the entire coding region and compared DPYD genotype frequencies between cancer patients with good (n = 89) and with poor (n = 39) tolerance of a fluoropyrimidine-based chemotherapy regimen. Applying logistic regression analysis and sliding window approaches we identified the strongest association with fluoropyrimidine-related grade III and IV toxicity for the non-synonymous polymorphism c.496A>G (p.Met166Val). We then confirmed our initial results using an independent sample of 53 individuals suffering from drug-adverse-effects. The combined odds ratio calculated for 92 toxicity cases was 4.42 [95% CI 2.12-9.23]; p (trend)<0.001; p (corrected) = 0.001; the attributable risk was 56.9%. Comparing tumor-type matched sets of samples, correlation of c.496A>G with toxicity was particularly present in patients with gastroesophageal and breast cancer, but did not reach significance in patients with colorectal malignancies. CONCLUSION: Our results show compelling evidence that, at least in distinct tumor types, a common DPYD polymorphism strongly contributes to the occurrence of fluoropyrimidine-related drug adverse effects. Carriers of this variant could benefit from individual dose adjustment of the fluoropyrimidine drug or alternate therapies.

Episomal vectors to monitor and induce somatic hypermutation in human Burkitt-Lymphoma cell lines.

Somatic hypermutation (SHM) occurs at a specific B-cell differentiation stage, during the germinal centre reaction, and provides a means to diversify and shape the antibody repertoire of the adaptive immune system. Burkitt-Lymphoma (BL) is a germinal centre derived B-cell malignancy. Presumably deregulation of the somatic hypermutation- and/or class switch recombination process causes a translocation between the myc-locus and one of the Ig-loci, which is characteristic for BL. We show here that we developed an episomal-based vector system to monitor and induce AID-dependent somatic hypermutation in human cell lines. Exemplarily, we utilized this system to investigate the hypermutation status of various BL cell lines. Analysis of the influence of Ig-specific cis-regulatory elements for the mutability of a green fluorescent protein (GFP) reporter transgene revealed, that the presence of Ig-enhancers is required for an efficient targeting of a constitutively transcribed GFP transgene. In contrast, a transcriptional inducible GFP transgene was hypermutated by overexpression of activation-induced cytidine deaminase (AID), even in the absence of Ig-specific sequences. This observation verifies that overexpression of AID in non hypermutating BL cell lines as well as the expression of endogenous AID in the hypermutating BL cell line Raji can overcome the target restriction of AID.