Félix Prieto

Mutational analysis of the MPZ, PMP22 and Cx32 genes in patients of Spanish ancestry with Charcot-Marie-Tooth disease and hereditary neuropathy with liability to pressure palsies

Abstract Charcot-Marie-Tooth disease (CMT) and hereditary neuropathy with liability to pressure palsies (HNPP) are two inherited peripheral neuropathies. The most prevalent mutations are a reciprocal 1.5-Mb duplication and 1.5-Mb deletion, respectively, at the CMT1A/HNPP locus on chromosome 17p11.2. Point mutations in the coding region of the myelin genes, peripheral myelin protein 22 (PMP22), myelin protein zero (MPZ) or connexin 32 (Cx32) have been reported in CMT patients, including CMT type 1 (CMT1), CMT type 2 (CMT2) and Déjérine-Sottas neuropathy (DS) patients, and only in the coding region of PMP22 in HNPP families lacking a deletion. We have investigated point and small mutations in the MPZ, PMP22 and Cx32 genes in a series of patients of Spanish ancestry: 47 CMT patients without duplications, and 5 HNPP patients without deletions. We found 15 different mutations in 16 CMT patients (34%). Nine different mutations in ten patients were detected in the Cx32 gene, this being the most frequently involved gene in this series, whereas five mutations involved the MPZ gene and only one the PMP22 gene. Six out of nine nucleotide substitutions in the Cx32 gene involved two codons encoding arginine at positions 164 and 183, suggesting that these two codons may constitute two Cx32 regions prone to mutate in the Spanish population. Analysis of HNPP patients revealed a 5′ splicing mutation in intron 1 of the PMP22 gene in a family with autosomal dominance, which confirms allelic heterogeneity in HNPP. Ectopic mRNA analysis on leukocytes suggests that this mutation might behave as a null allele.

Mutation of the XNP/ATR-X gene in a family with severe mental retardation, spastic paraplegia and skewed pattern of X inactivation: demonstration that the mutation is involved in the inactivation bias.

We would like to thank the family members for their cooperation and Dr. Charles Schwartz for helpful discussion. This work was supported by the INSERM program PARMIFR and by the PROGRES network, as well as by Spanish Ministry of Health project FIS98/0170 (Fondo de Investigaciones Sanitarias).

11q23 abnormalities in children with acute nonlymphocytic leukemia (M4–M5): Association with previous chemotherapy

Cytogenetic studies of 12 patients aged less than 14 years with acute nonlymphoblastic leukemia (ANLL) (M4-M5) showed structural abnormalities on chromosome 11 at band q23-q24 in five cases (41.8%). Four of these 12 patients had ANLL (M4-M5) after treatment with cytostatics for non-Hodgkin lymphoma in one case and for an acute lymphoblastic leukemia (ALL) in the other three. Three of these four cases had 11q23 abnormalities [one [one 46,XY,t(11;17)(q23;25); another 47,XY,+8,-15,del(11)(q23),+der(15)t(15;?)(p11;?); the third 47,XX,+8,t(3;17) (p11;q25),t(4;11)(q21;q23)] and one had a normal karyotype on being diagnosed ANLL (M4-M5). The notable increase of ANLL (M4-M5) in our patients who had received cytostatics as treatment for a previous neoplasia makes evaluation of our results timely in comparison with those of other groups who use these therapeutic protocols.

Chromosome banding patterns in patients with chronic myelocytic leukemia

One hundred and nine patients with Ph1-positive chronic myelocytic leukemia were cytogenetically studied with banding methods. Seventy-eight patients were studied in the chronic phase and 39 patients in the blastic phase. The standard translocation was present in 107 cases. Two patients showed complex translocations involving chromosomes No. 6, 9, 22, 11 and No. 9, 22, 11, respectively. Ph1-negative cells were detected in 8 cases (7%). Chromosome aberrations in addition to the Ph1 chromosome were observed in 6 cases (8%) during the chronic phase. The karyotypic findings during the blastic phase were similar to those reported in the past [trisomy 8, iso(17q), and a second Ph1]. The significance of Ph1-negative cells, the geographic heterogeneity of the chromosomal aberrations, the effect of chemotherapy on the appearance of new clones, and the importance of the materials and methods used for the comparison of cytogenetic patterns at different laboratories are discussed.

Translocations involving chromosomes #3 and #12: Hematologic diseases associated with abnormalities of these chromosomes

Two hematologic cases with translocations involving chromosomes #3 and #12 are described. The first case is that of a myeloproliferative disorder (preleukemia?) associated with a (3;12)(q29;q24) translocation in the bone marrow cells. No evidence of leukemic transformation has appeared to date. The second case is that of acute leukemia (AL) (M4 type) in which leukemic cells with t(3;12)(p14;q24) were seen. The roles of chromosomes #3 and #12 in hematopoiesis are considered, and the abnormalities affecting these chromosomes in various hematologic disorders have been tabulated and correlated. Abnormalities in chromosomes #3 and #12 appear to be common and nonrandom in hematologic diseases of a premalignant and a malignant type.

Trisomy 4: Another specific anomaly in acute nonlymphocytic leukemia

We present herein a case of acute nonlymphocytic leukemia, type M2 of the FAB classification, in which the cytogenetic study of the bone marrow cells showed a trisomy 4 as the only alteration. This case was detected from among 118 cases of acute nonlymphocytic leukemia under cytogenetic study.

A two base pair deletion in the PQBP1 gene is associated with microphthalmia, microcephaly, and mental retardation

X-linked mental retardation has been traditionally divided into syndromic (S-XLMR) and non-syndromic forms (NS-XLMR), although the borderlines between these phenotypes begin to vanish and mutations in a single gene, for example PQBP1, can cause S-XLMR as well as NS-XLMR. Here, we report two maternal cousins with an apparently X-linked phenotype of mental retardation (MR), microphthalmia, choroid coloboma, microcephaly, renal hypoplasia, and spastic paraplegia. By multipoint linkage analysis with markers spanning the entire X-chromosome we mapped the disease locus to a 28-Mb interval between Xp11.4 and Xq12, including the BCOR gene. A missense mutation in BCOR was described in a family with Lenz microphthalmia syndrome, a phenotype showing substantial overlapping features with that described in the two cousins. However, no mutation in the BCOR gene was found in both patients. Subsequent mutation analysis of PQBP1, located within the delineated linkage interval in Xp11.23, revealed a 2-bp deletion, c.461_462delAG, that cosegregated with the disease. Notably, the same mutation is associated with the Hamel cerebropalatocardiac syndrome, another form of S-XLMR. Haplotype analysis suggests a germline mosaicism of the 2-bp deletion in the maternal grandmother of both affected individuals. In summary, our findings demonstrate for the first time that mutations in PQBP1 are associated with an S-XLMR phenotype including microphthalmia, thereby further extending the clinical spectrum of phenotypes associated with PQBP1 mutations.

Intronic L1 insertion and F268S, novel mutations in RPS6KA3 (RSK2) causing Coffin-Lowry syndrome

Two novel mutations of the ribosomal S6 kinase 2 gene (also known as RSK2) have been identified in two unrelated patients with Coffin–Lowry syndrome. The first mutation consists of a de novo insertion of a 5′‐truncated LINE‐1 element at position −8 of intron 3, which leads to a skipping of exon 4, leading to a shift of the reading frame and a premature stop codon. The L1 fragment (2800 bp) showed a rearrangement with a small deletion, a partial inversion of the ORF 2, flanked by short direct repeats which duplicate the acceptor splice site. However, cDNA analysis of the patient shows that both sites are apparently not functional.

Pediatric Brain Tumors: Loss of Heterozygosity at 17p and TP53 Gene Mutations

Cytogenetic and molecular analyses of primitive neuroectodermal tumors (PNETs) of the central nervous system (CNS) have demonstrated material losses of 17p, the region that contains the TP53 gene, as the most frequent abnormality. Mutations in the TP53 gene are, however, very rare in these tumors. These findings strongly suggest that another, as yet unidentified, gene on 17p may be involved. We performed a search for loss of heterozygosity (LOH) on 17p by microsatellite markers on 26 childhood CNS tumors as well as TP53 gene mutations (exons 5-8) by single-strand conformational polymorphism analysis on 41 pediatric brain tumor samples of distinct histologic types. LOH was detected in 10 cases: 7 PNET, 2 astrocytomas, and 1 glioblastoma multiforme. In 4 of the PNETs the losses were limited to more distal markers. On the other hand, TP53 mutations were detected in 6 of 41 samples studied. Our results not only confirm the low penetrance of the TP53 gene on pediatric CNS tumors, but also provide further evidence of a putative tumor suppressor gene distal to TP53, between markers (D17S938, D17S926) and 17pter, specifically taking part in the development of PNET.

Molecular characterisation of partial chromosome 21 aneuploidies by fluorescent PCR.

Although trisomy of chromosome 21 is the most prevalent human genetic disorder, data from partial 21 aneuploidies are very scanty. Eight different partial aneuploidies for chromosome 21 were characterised by fluorescence quantitative PCR. Allelic dosage analysis was performed for each patient using 25 CHLC STRs covering the entire q arm. The length of the corresponding trisomies and monosomies was ascertained for five partial trisomics and three partial monosomics. All trisomic patients carried unbalanced translocations involving chromosome 21, whereas one of the monosomic patients bore a ring chromosome 21 and another showed an interstitial deletion of chromosome 21. The chromosomal breakpoints of two partial trisomy patients could be clearly delimited. However, the other three trisomies involved most of the 21 q arm as three allelic doses were detected for each marker. Although these latter patients do not show all the features of Down syndrome, genotype/phenotype correlations agree with previously reported data. The chromosomal breakpoints observed in two partially monosomic patients helped further to define the region involved in different phenotypic features associated with chromosome 21 monosomy. Telomeric material loss was also detected in a patient bearing a ring 21 chromosome. The parental origin of the aneuploidy was assigned for each case, which allowed us to conclude that two of the monosomic cases originated from de novo chromosomal rearrangements. There was no correlation with parental sex in contrast to trisomic patients originating from meiotic non-disjunction.