71. A high-throughput amplicon screen for somatic UBA1 variants in Cytopenic and Giant Cell Arteritis cohorts
Creators: name: Poulter2, James
name: Martin9, Stephen
name: Grey3, Mark
name: Bosko Andjelic4
name: Nga5, Emma
name: Triantafyllopoulou6, Diana
name: Khan7, Alesia
name: Evans7, Paul
name: Sarah Mackie1, 8
name: Ann Morgan8, 9
name: Cargo7, Catherine
name: Sinisa Savic1, 8
Publisher: name: Zenodo
Year: 2022

description: <strong>Background:</strong> Somatic mutations in the gene encoding the major E1 ubiquitin ligase, <em>UBA1 </em>(X-linked), were recently identified as a cause of VEXAS, a late-onset acquired auto-inflammatory syndrome. Patients with VEXAS are typically male, aged over 60 years with a severe systemic inflammatory disorder, cytopenia and dysplastic features in their bone marrow. Differential diagnoses for patients subsequently found to have VEXAS include relapsing polychondritis, Sweets syndrome, myelodysplastic syndrome (MDS), giant cell arteritis (GCA) and undifferentiated systemic autoinflammatory syndrome (uSAID). We therefore sought to screen DNA from individuals with a non-diagnostic cytopenia or GCA, for known VEXAS-associated mutations. <br> <strong>Methods: </strong>We developed a multiplexed UBA1 amplicon sequencing assay, allowing quick screening of large cohorts while also providing sufficient sequencing depth to identify somatic mutations to an allele frequency &lt;1%. Using this assay, we screened genomic DNA from 612 males diagnosed with GCA (patients recruited into UKGCA consortium), and bone marrow derived DNA from 1,055 cases with an undiagnosed cytopenia. <br> <strong>Results</strong>: 1,667 DNA samples were screened for variants in <em>UBA1</em> exon 3 by amplicon sequencing. In total, 1,650 (98.9%) of samples had a read depth &gt;100 and all 1,667 (100%) had a read depth &gt;30. Seven individuals were identified with a UBA1 mutation (Table 1), all present in the cytopenic cohort (n=7/1,055, 0.66%). No variants were identified in the GCA cohort. All variants identified have been published as a cause of VEXAS and either lead to substitution of Met41 or affect the canonical splice acceptor site. On average, the variant allele frequency (VAF) of the mutation was 44.1% (range:29.5%-59.3%). All variants were confirmed by Sanger sequencing. Six of the patients were identified from the 595 male cytopenic cases (1.00%). Interestingly, 1 patient was from the 460 cytopenic female cases. (0.02%). A whole genome SNP array on bone marrow extracted genomic DNA of the female case (P2) revealed a normal X chromosome dosage, with no indication of loss of heterozygosity over <em>UBA1</em>. The VAF of P2 was 33%, within the range observed in the male cases (range: 19%-52%).<br> <strong>Conclusions</strong>: Our study suggests that, despite the overlap in clinical features, VEXAS is rarely misdiagnosed as GCA but identified 1.0% of males with an undiagnosed cytopenia have VEXAS. The identification of a <em>UBA1</em> variant in a female case adds further evidence that VEXAS should not be ruled out as a differential diagnosis in females with VEXAS-like symptoms.<br> <br> <strong>Disclosures:</strong> None<br> <strong>Table 1</strong>.