Niemann Pick disease (NPD) is an autosomal recessive disorder due to the deficit of lysosomal acid sphingomyelinase, which results in intracellular accumulation of sphingomyelin. In the present work we studied 18 patients with NPD type B, including five individuals who presented an intermediate phenotype characterised by different levels of neurological involvement. We identified nine novel mutations in the SMPD1 gene including six single base changes c.2T>G, c.96G>A, c.308T>C, c.674T>C, c.732G>C, c.841G>A (p.M1_W32del, p.W32X, p.L103P, p.L225P, p.W244C, p.A281T) and three frameshift mutations c.100delC, c.565dupC, c.575dupC (p.G34fsX42, p.P189fsX1 and p.P192fsX14). The novel c.2T>G (p.M1_W32del) mutation inactivates the first in-frame translation start site of the SMPD1 gene and in the homozygous status causes NPD type B indicating that in'vivo translation of wild type SMPD1 initiates from the first in-frame ATG. Moreover, the new c.96G>A (p.W32X) introduces a premature stop codon before the second in-frame ATG. As a consequence of either c.2T>G (p.M1_W32del) or c.96G>A (p.W32X), impaired translation from the first in-frame ATG results in a mild NPD-B phenotype instead of the severe phenotype expected for a complete deficiency of the enzyme, suggesting that when the first ATG is not functional, the second initiation codon (ATG33) still produces a fairly functional sphingomyelinase. Analysis of the patients'clinical and molecular data demonstrated that all five patients with the intermediate phenotype carried at least one severe mutation. No association between the onset of pulmonary symptoms and genotype was observed. Finally, the presence of c.96G>A (p.W32X), the most frequent allele among Italian NPD type B population, and c.1799G>C (p.R600P) as compound heterozygotes in association with severe mutations suggested a beneficial effect for both mutations.

; Web site: http://www.gaslini.org/labdppm.htm Communicated by Robert DesnickNiemann-Pick disease (NPD) results from the deficiency of lysosomal acid sphingomyelinase (SMPD1). To date, out of more than 70-disease associated alleles only a few of them have a significant frequency in various ethnic groups. In contrast, the remainder of the mutations are rare or private. In this paper we report the molecular characterization of an Italian series consisting of twenty-five NPD patients with the severe neurodegenerative A phenotype. Mutation detection identified a total of nineteen different mutations, including 14 novel mutations and five previously reported lesions. The known p.P189fs and the novel p.T542fs were the most frequent mutations accounting for 34% and 18% of the alleles, respectively. Screening the alleles for the three common polymorphisms revealed the variant c.1516G>A (exon 6) and the repeat in exon 1, but not the variant c.965C>T (exon 2). In absence of frequent mutations, the prognostic value of genotyping is limited. However, new genotype/phenotype correlations were observed for this disorder that could in the future facilitate genetic counseling and guide selection of patients for therapy.

Hunter syndrome (Mucopolysaccharidosis type II) is a rare X-linked recessive lysosomal storage disorder caused by the deficiency of the enzyme iduronate-2-sulfatase (IDS). To date, more than 200 different mutations have been reported in the IDS gene, located on Xq27.3-q28. Here, we report two new mutations (M488I and G489A) identified in hemizygosity in an Italian Hunter patient. Their "in vitro" expression by COS 7 cells was carried out in order to evaluate their functional consequence on enzyme activity as well as their possible cumulative effect on the malfunctioning of the protein. The results obtained enabled us to confirm the G489A mutation as causative. The M488I mutation, however, could not be unequivocally considered as causing disease because of its residual activity. Although a cumulative effect of the two mutations can be excluded "in vitro," we are cautious about drawing a conclusion with regard to the possible role that the two mutations could have played "in vivo" in modulating the phenotype of the patient. Finally, the knowledge of the molecular defect of the patient has enabled us to identify the carriers, providing reliable genetic counselling to the females of the family.

Hunter syndrome (Mucopolysaccharidosis type II), a rare X-linked lysosomal storage disorder, results from deleterious mutations in the iduronate-2-sulfatase (IDS) gene located on Xq27.3-q28. Partial or complete deletions and large rearrangements have been extensively reported in the IDS gene as the basis of Hunter disease. The present report, however, is the first report on a Hunter patient in which Alu-mediated recombinations are implicated. Our patient showed the skipping of exon 8 at the cDNA level, without any splice-junction defects at the genomic level, where a new large rearrangement was identified instead. This new mutant allele consisted of an extensive deletion of IDS sequence of about 3 kb, as well as an additional inserted sequence of 157 bp. Two different computer programs were necessary to elucidate the nature of the insert. NCBI-BLAST query detected a single match for 126 bp out of 157 of the fragment that aligned exactly with a specific chromosomal region, Xq25-27.1, where an AluSg sequence is adjacent to an L1. Instead, the Repeat Masker program identified only 83 bp out of 157 of the insert, which was confirmed as an AluS. The observed homology between the AluSc sequence in the IDS intron 8 and the inserted AluS element, as well as the closeness of 26 bp Alu core sequence, considered to be a recombination hotspot, made us hypothesise upon the fact that both an Alu retrotransposition and an Alu-mediated deletion underlie the disease-producing rearrangement. We, therefore, now propose a mechanism that led to the large genomic deletion causing the production of the aberrant mRNA splicing.