Analysis Of Clinical And Genetic Characteristic Of Combined Methylmalonic Acidemia And Homocystinuria

Background:Combined methylmalonic acidemia and homocystinuria （C-MMA） is a common organic acidemia which could be classified into two types by different pathogeny:primary C-MMA and secondary C-MMA. Primary C-MMA could be classified into cb1C, cblD, cb1F, cblJ and cblX five subtypes, all of them are caused by metabolic disorders of mut and cobalamin. Among them the most common one is cblC type. The clinical manifestations is diversity and multiple systems can be involved. CblC type MMA is caused by the mutation of MMACHC gene which located on 1p34.1. There are less research about clinical phenotype, genotype of C-MMA and the relationship between them.Objective:To summarize the clinical manifestation, laboratory examination and characteristics of the genotype of 15 C-MMA cases, and further explore the relationship between clinical manifestation and genotype.Methods:This study collected 15 C-MMA patients’ information from August 2005 to February 2016 in Qilu Hospital of Shandong University. They are diagnosed by their clinical manifestations, organic acids test results and genetic testing results. In this study we use the method of blood tandem mass spectrometry and gas chromatography-mass spectrometry to analysis the level of organic acid. Using phenol-chloroform method to extract DNA, then designing MMACHC gene primer, amplifying the five exons and introns and flank area line by PCR, sequencing the PCR product, determining the form of MMACHC genetic mutation. According to its clinical manifestation and laboratory examination, we further explore the relationship between them.Results:There are 5 male patients and 10 female patient, the onset age of which rangging from 2 to 59. The mean onset age of them is 19.3, the median age is 16. Among all the patients, there are 7 patients（46.7%） mainly manifested as muscle weakness in there lower extremities, there are 7 patients mainly manifested as psychiatric disorder such as somnolence, apathy or balderdash, the last one patients mainly manifested as disequilibrium.The urine organic acid level:we tested the organic level of the urine of 14 patients among all patients, the results as follows:the MMA level of all patients elevated in different degrees, ranging from 54.5-236. luM with a average value 16.3uM.Blood tandem mass spectrometry:Except 1 case who has been treated, the ratio of C3/C2 increased in different degree, ranging from 0.34-4.8, the average value is 0.86.The Hcy level of all cases elevated, ranging from 62.8-150.1umol/l, the average value is 90.7umol/l.Mutations detection:6 different mutations were identified, including 3 missense mutations （c.482G>A,c.463G>C,c.427C>T）;1 nonsense mutation （c.609G>A）;1 deletion mutation （c.445₄46delTG） and 1 frameshift mutation （c.567dupT）, one mutation was novel （c.463G>C）.Conclusions:1.The clinical manifestations of C-MMA is diversity, multiple systems can be involved such as blood system, gastrointestinal tract and bones. The symptoms of nervous damage is remarkable, such as muscle weakness in there lower extremities, psychiatric disorder, convulsion or cognitive decline. Muscle weakness in there lower extremities, psychiatric disorder and convulsion could be the main manifestation.2.Six different mutations were identified, including 3 missense mutations （c.482G>A,c.463G>C,c.427C>T）;1 nonsense mutation （c.609G>A）;1 deletion mutation （c.445₄46delTG） and 1 frameshift mutation （c.567dupT）, one mutation was novel （c.463G>C）. The heterozygous mutation c.482G>A could lead to milder clinical symptoms, while the homozygous mutation c.609G>A could lead to more severe symptoms. In addition, this study found a novel mutation c.463G>C.3.The clinical manifestation of C-MMA showed a remarkable heterogeneity, and the genotype-phenotype correlation needed more samples analyzed to be confirmed.4.The late onset C-MMA has a better prognosis, supplement of hydroxocobalamin can obviously improve the clinical symptoms.