Novel method for detection of UBE3A protein in CSF from individuals with Angelman syndrome.

Loss of production of ubiquitin protein ligase E3A (UBE3A) in neurons leads to Angelman Syndrome (AS). There are limited methods to reliably measure UBE3A in cerebrospinal fluid (CSF), which negatively impacts therapeutic development. To overcome this gap, we developed and analytically validated a novel method for CSF UBE3A quantitation, which includes an immunoprecipitation protein capture step followed by tryptic digestion and high-resolution mass spectrometry detection of a unique UBE3A peptide. Our data suggest that we can reliably detect UBE3A at concentrations as low as 2.5 pg/mL. The assay was used to show that UBE3A could be detected in CSF samples of both healthy adults and patients with AS. As expected, CSF UBE3A levels in healthy adults (24.76 ± 6.75 pg/mL, N = 14) were significantly higher (p < 0.01) than the CSF UBE3A levels measured in two AS cohorts (5.30 ± 0.42 pg/mL, N = 19 and 5.59 ± 0.40 pg/mL, N = 10), with no significant difference in UBE3A levels observed between the two AS cohorts. There was also no significant difference in CSF UBE3A levels when comparing AS patients carrying either a mutation or chromosomal deletion in either cohort. Overall, these data demonstrate the utility of this novel CSF UBE3A assay for UBE3A quantitation in studies of AS.