Genetic and cellular studies of oxidative stress in methylmalonic aciduria (MMA) cobalamin deficiency type C (cblC) with homocystinuria (MMACHC)

Methylmalonic aciduria (MMA) cobalamin deficiency type C (cblC) with homocystinuria (MMACHC) is the most frequent genetic disorder of vitamin B₁₂ metabolism. The aim of this work was to identify the mutational spectrum in a cohort of cblC-affected patients and the analysis of the cellular oxidative stress and apoptosis processes, in the presence or absence of vitamin B₁₂. The mutational spectrum includes nine previously described mutations: c.3G>A (p.M1L), c.217C>T (p.R73X), c.271dupA (p.R91KfsX14), c.331C>T (p.R111X), c.394C>T (p.R132X), c.457C>T (p.R153X), c.481C>T (p.R161X), c.565C>A (p.R189S), and c.615C>G (p.Y205X), and two novel changes, c.90G>A (p.W30X) and c.81+2T>G (IVS1+2T>G). The most frequent change was the known c.271dupA mutation, which accounts for 85% of the mutant alleles characterized in this cohort of patients. Owing to its high frequency, a real-time PCR and subsequent high-resolution melting (HRM) analysis for this mutation has been established for diagnostic purposes. All cell lines studied presented a significant increase of intracellular reactive oxygen species (ROS) content, and also a high rate of apoptosis, suggesting that elevated ROS levels might induce apoptosis in cblC patients. In addition, ROS levels decreased in hydroxocobalamin-incubated cells, indicating that cobalamin might either directly or indirectly act as a scavenger of ROS. ROS production might be considered as a phenotypic modifier in cblC patients, and cobalamin supplementation or additional antioxidant drugs might suppress apoptosis and prevent cellular damage in these patients. Hum Mutat 30:1–9, 2009. © 2009 Wiley-Liss, Inc.