Identification of Two Novel Homozygous HAX1 Mutations in an Autosomal Recessive Jewish and Two Unrelated Sporadic British Families with Severe Congenital Neutropenia.

Severe chronic neutropenia (SCN) is a rare disorder characterized by profound neutropenia, recurrent bacterial infections and bone marrow arrest at the promyelocyte/myelocyte stage. Initially described as an autosomal recessive (AR) disease (Kostmann disease), autosomal dominant (AD), sporadic and X-linked forms have been reported. Pathogenic mutations in the neutrophil elastase ELA2 gene have been documented in both SCN and the related disorder cyclic neutropenia in a significant proportion of AD and sporadic but not AR patients, and mutations in the Wiskott-Aldrich syndrome (WAS) gene have been reported in X-linked SCN families. More recently, homozygous mutations resulting in AR disease were identified through linkage to chromosome 1p21 in Kurdish and the original Kostmann families, revealing mutations in the HAX1 gene that result in complete loss of this mitochondrial anti-apoptotic protein. In order to determine the frequency of these different mutations in SCN, we used denaturing HPLC WAVE analysis to investigate a cohort of 105 SCN kindreds seen in our institute or referred with a diagnosis of SCN of which 11 were presumed/known AD, 17 AR, 73 sporadic and 4 had X-linked disease. Overall, 25 different ELA2 mutations were detected in 31 kindreds (6 AD and 25 sporadic, 30% of the cohort). Mutations were distributed throughout the gene and were most frequent in exon 4 (42% of mutations), least frequent in exon 3 (10%). Two kindreds (2%) had WAS mutations as previously published, no new cases were identified. The remaining 72 cases were screened for all 7 HAX1 exons using mixtures of patient and control PCR products to detect homozygous substitutions. Four homozygous mutant kindreds were identified. One AR kindred of Kurdish descent had the previously reported p.Trp44X mutation. Two sisters of Jewish origin had a homozygous insertion of an extra G in a run of 5 Gs (nucleotides 121–125) in exon 2 resulting in a frameshift (p.Ser43LeufsX11); both parents were shown to be heterozygous for the mutation. Two unrelated sporadic SCN patients of Caucasian (British) origin had deletion of nucleotide G91 in exon 2 resulting in a frameshift (p.Glu31LysfsX54); to date, parents of one case have been confirmed as heterozygotes. Both frameshift mutations are predicted to result in a non-functional protein. The 4 HAX1-mutant kindreds have variable clinical severity (ranging from mild to severe) and G-CSF requirement (none to regular administration). All heterozygous parents are phenotypically normal. Our results indicate that HAX1 mutations are relatively uncommon (4% of our cohort) but not limited to patients with specific ethnic origin or apparent AR inheritance.