Methylmalonic Acidemia Panel

Methylmalonic acidemia (MMA) is typically a severe disease with onset in infancy. Patients may present with lethargy, vomiting, hepatomegaly, acidosis, hypoglycemia and neutropenia. Many patients die in childhood; those that survive often experience neurological and renal complications. Milder forms of the disease are also known. Today, many cases are detected through routine neonatal screening with tandem mass spectrometry (Fenton et al. 2014; Watkins and Rosenblatt 2014; Manoli et al. 2016).

The MMADHC, MMAA, MMAB, MCEE, MMUT and CD320 genes encode proteins directly related to cobalamin transport or metabolism, or methylmalonyl-CoA metabolism. Defects in these genes have been shown to lead to the development of isolated MMA. Defects in each gene are associated with a specific type of MMA disorder (cblD, cblA, cblB, MCEE, MUT or CD320 deficiency, respectively). The majority of MMA patients have defects in the MMUT gene, followed by the MMAA and then MMAB genes (Manoli et al. 2016). Defects in the MMADHC, MCEE and CD320 genes are rare, having only been reported in a small number of families (Bikker et al. 2006; Dobson et al 2006; Carrillo et al. 2013; Manoli et al. 2016). It should be noted that pathogenic variants in the CD320 gene lead to transient methylmalonic aciduria and/or homocystinuria, which has not been associated with the neurological or hematological symptoms of the cblD, cblA, cblB, mcee or mut deficiency disorders. However, CD320 deficient individuals may be detected via newborn screening (Quadros et al. 2010; Karth et al. 2012; Watkins and Rosenblatt 2014).

This sequencing panel also includes coverage for several genes that do not encode proteins directly involved in cobalamin or methylmalonyl-CoA metabolism. Defects in the ALDH6A1 gene lead to methylmalonate semialdehyde dehydrogenase deficiency (MMSDH), which is an inborn error of valine and thymine metabolism (Chambliss et al. 2000; Marcadier et al. 2013). Defects in the ACSF3 and MLYCD genes lead to combined malonic and methylmalonic aciduria (CMAMMA) (FitzPatrick et al. 1999; Alfares et al. 2011; Sloan et al. 2011; Baertling et al. 2014). Defects in the SUCLA2 and SUCLG1 genes lead to mitochondrial DNA depletion syndromes (MDSs) (Elpeleg et al. 2005; Carrozzo et al. 2007; Ostergaard et al. 2007; Carrozzo et al. 2016). These five genes are included in this panel as defects in all of them have been shown to result in methylmalonic acidemia in at least some patients. Phenotypically, these disorders may be similar or quite different from the cblD, cblA, cblB, mcee or mut types of methylmalonic acidemia.

More details regarding each disorder are available on individual gene test pages.

Isolated methylmalonic acidemia (MMA) can result from pathogenic variants in at least six different genes directly associated with inborn errors of cobalamin metabolism or transport, or methylmalonyl-CoA metabolism (MMADHC, MMAA, MMAB, MCEE, MMUT and CD320). In addition, defects in an additional five genes (ACSF3, ALDH6A1, MLYCD, SUCLA2 and SUCLG1) may also lead to methylmalonic acidemia, although these genes are not directly related to cobalamin metabolism or transport or methylmalonyl-CoA metabolism. All of these disorders are inherited in an autosomal recessive manner.

Massively parallel sequencing plus Sanger confirmation will detect the vast majority of sequence variants in the MMADHC, MMAA, MMAB, MCEE, MMUT, CD320, ACSF3, ALDH6A1, MLYCD, SUCLA2 and SUCLG1 genes. It should be noted that large deletions that may not be detectable via sequencing have been reported in the MMAA, MMUT, ACSF3 and SUCLA2 genes (Human Gene Mutation Database).

See individual gene test descriptions for information on molecular biology of gene products.

The overall clinical sensitivity of this sequencing panel is expected to be high. Based on enzymatic studies of patients with isolated methylmalonic acidemia, Hörster et al. (2007) reported that ~63% of patients can be classified as mut0 or mut -, ~24% as cblA type, and ~13% as cblB type. Two molecular studies of patients diagnosed biochemically, enzymatically and via complementation analysis with isolated methylmalonic acidemia have been fairly consistent with these results. First, Martinez et al. (2005) evaluated 25 MMA patients and found MMUT pathogenic variants in 13 (52%), MMAA pathogenic variants in 7 (28%), and MMAB pathogenic variants in 2 (8%). Merinero et al. (2008) studied 32 patients with isolated MMA and found MMUT causative variants in 19 (59%), MMAA causative variants in 9 (28%), and MMAB causative variants in 4 (13%).

Pathogenic variants in the MMADHC, MCEE, ACSF3, ALDH6A1, CD320, SUCLA2, SUCLG1 and MLYCD genes are expected to make up some fraction of the remaining patients. In one recent study of 131 methylmalonic acidemia/aciduria patients without a diagnosis based on testing of the more commonly involved genes, 8 patients were found to have pathogenic variants in less commonly mutated genes (2 patients with SUCLG1 variants and 5 patients with ACSF3 variants) (Pupavac et al. 2016). Of note, the ALDH6A1 and MLYCD genes were not included in this study.

Nearly all reported pathogenic variants in the eleven genes in this panel (MMUT, MMAA, MMAB, MMADHC, MCEE, ACSF3, ALDH6A1, CD320, MLYCD, SUCLA2 and SUCLG1) are detectable via direct sequencing, so the analytical sensitivity of this sequencing panel is expected to be high.

To date, no gross deletions or insertions have been reported in the MMAB, MMADHC or SUCLG1 genes, and only 1-2 gross deletions have been reported in the MMAA and ACSF3 genes (Nizon et al. 2013; Pupavac et al. 2016). Deletions and duplications appear to be somewhat more common in the MMUT and SUCLA2 genes (Acquaviva et al. 2005; Chu et al. 2016; Forny et al. 2016), though it is difficult to precisely estimate the fraction of patients carrying such large copy number variants. In summary, the sensitivity of duplication/deletion testing for these rare disorders, although not precisely known, is low.

This test is performed using Next-Gen sequencing with additional Sanger sequencing as necessary.

This panel provides 100% coverage of all coding exons of the genes plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define coverage as ≥20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

In addition to the regions described above, this testing includes coverage of the following deep intronic variant: MCEE c.379-644A>G.

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).