OBJECTIVE3-Methylcrotonyl-coenzyme A carboxylase deficiency (MCCD) is an autosomal recessive inborn error of leucine catabolism. The cases suspected as MCCD detected by neonatal screening are not rare. The aim of the study was to investigate the clinical outcomes in cases suspected as MCCD by neonatal screening. The second aim was to investigate the mutation spectrum of MCC gene in Chinese population and hotspot mutation.

METHODForty-two cases (male 33, female 9) , who had higher blood 3-hydroxy-isovalerylcarnitine (C5-OH) levels(cut-off <0.6 µmol/L) detected by neonatal screening using MS/MS, were recruited to this study during Sept.2011 to Mar.2013. The C5-OH concentrations were [0.84 (0.61-20.15) µmol/L] in 42 cases at the screening recall. Five cases were firstly diagnosed as maternal MCCD, 6 cases as benign MCCD and 31 cases were suspected as MCCD. To follow up the height, weight, mental development, blood C5-OH concentrations and urinary 3-methylcrotonyl-glycine (3-MCG) and 3-hydroxy isovalerate (3-HIVA) in order to investigate the clinical outcome. The MCCC1 and MCCC2 gene mutation were analyzed for some cases. The novel gene variants were evaluated, and the influence of novel missense variants on the protein structure and function were predicted by PolyPhen-2, SIFT, UniProt and PDB software.

RESULT(1) Forty-two cases had no symptoms, their physical and mental development were normal in the last visit at the median ages of 29 months, the oldest age of follow up was nearly 9 years. (2) Gene mutation analysis was performed for 29 cases with informed consent signed by parents.Fourteen different mutations were identified in 19 cases. The mutations in MCCC1 gene accounted for 86%, the most common mutation was c.ins1680A, (accounted for 40%). Nine kinds of novel variant were detected including 211AG>CC/p.Q74P, c.295G>A/p.G99S, c.764A>C/p.H255P, c.964G>A/p. E322K, c.1331G>A/p.R444H, c.1124delT, c.39_58del20, c.1518delG, c.639+2T>A.Other 3 kinds of mutation in MCCC1 gene and 2 kinds of mutation in MCCC2 gene have been reported previously; the amino acid of mutant positions of five kinds of novel missense variant are almost highly conserved. These missense variants were predicted to cause change of human MCC protein side chain structure by changing hydrogen bonding, size of amino acid residue and electric charge, and predicted to damage the protein function possibly according to PolyPhen-2 and PDB analysis. So these novel variants may be disease-causing mutations. No mutation were detected in 10 cases. (3) Blood concentrations of C5-OH when screening, recall and end of follow-up in maternal MCCD was 3.50 (1.63-11.43), 1.84 (1.00-9.30), 0.27 (0.26-5.81) µmol/L. There was a significant downward trend.In contrast, benign MCCD group was 8.20 (3.60-9.60), 9.67 (3.88-20.15), 23.0 (5.87-49.10) µmol/L.It showed a rising trend. Children's urinary 3-MCG of benign MCCD group was found abnormally elevated in 4 cases (100%) when they were recalled.

CONCLUSIONA certain number of cases with MCCD or suspected as MCCD in this study had no symptoms and normal physical and mental development after follow-up to oldest age of nearly 9 years. The mutation in MCCC1 gene is common, nine novel mutations were found, c.ins1680A may be a hotspot mutation in Chinese population. The urinary GC/MS analysis and blood MS/MS analysis for mother should be routinely performed for all cases with high blood C5-OH level detected by neonatal screening.