Medium-chain Acyl-CoA dehydrogenase (MCAD) mutations identified by MS/MS-Based prospective screening of newborns differ from those observed in patients with clinical symptoms: Identification and characterization of a new, prevalent mutation that results in mild MCAD deficiency

Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most frequentl y diagnosed mitochondrial beta -oxidation defect, and it is potentially fat al. Eighty percent of patients are homozygous for a common mutation, 985A - ->G, and a further 18% have this mutation in only one disease allele. In ad dition, a large number of rare disease-causing mutations have been identifi ed and characterized. There is no clear genotype-phenotype correlation. Hig h 985A -->G carrier frequencies in populations of European descent and the usual avoidance of recurrent disease episodes by patients diagnosed with MC AD deficiency who comply with a simple dietary treatment suggest that MCAD deficiency is a candidate in prospective screening of newborns. Therefore, several such screening programs employing analysis of acylcarnitines in blo od spots by tandem mass spectrometry (MS/MS) are currently used worldwide. No validation of this method by mutation analysis has yet been reported. We investigated for MCAD mutations in newborns from US populations who had be en identified by prospective MS/MS-based screening of 930,078 blood spots. An MCAD-deficiency frequency of 1/15,001 was observed. Our mutation analysi s shows that the MS/MS-based method is excellent for detection of MCAD defi ciency but that the frequency of the 985A -->G mutant allele in newborns wi th a positive acylcarnitine profile is much lower than that observed in cli nically affected patients. Our identification of a new mutation, 199T -->C, which has never been observed in patients with clinically manifested disea se but was present in a large proportion of the acylcarnitine-positive samp les, may explain this skewed ratio. Overexpression experiments showed that this is a mild folding mutation that exhibits decreased levels of enzyme ac tivity only under stringent conditions. A carrier frequency of 1/500 in the general population makes the 199T -->C mutation one of the three most prev alent mutations in the enzymes of fatty-acid oxidation.