Intragenic loss of function mutations demonstrate the primary role of FMR1 in fragile X syndrome.

Nearly all cases of fragile X syndrome result from expansion of a CGG trinucleotide repeat found in the 5' untranslated portion of the FMR1 gene. Methylation of the expanded repeats correlates with down-regulation of transcription of FMR1; thus fragile X syndrome is postulated to be due to a loss of function of the FMR1 gene product, and this has been demonstrated at the protein level. However, the nature of the mutation offers the possibility of methylation spreading to adjacent genes with consequent loss of expression and contribution to the phenotype. Deletions of FMR1 and flanking sequence (some of substantial size) have been reported in patients with phenotypes consistent with a diagnosis of fragile X-syndrome, however, none is strictly intragenic. We report here the identification of two different intragenic loss of function mutations in FMR1: a single de novo nucleotide deletion in a young male patient (IJ) and an inherited two basepair change in an Adult male (SD), each with classical features of fragile X syndrome.

Involvement of chromosome 7 in Wilms tumor.

Cytogenetic and molecular analysis of Wilms tumors have led to the identification of two regions on the short arm of chromosome 11 (11p13 and 11p15) involved in tumor development. Recent studies have provided evidence that an additional locus on 16q is also involved. Further molecular testing may reveal additional loci associated with the development or progression of this tumor. Reports of single chromosome abnormalities in tumors generally pinpoint regions of interest that may be involved in the etiology of the tumor. We present an additional case of Wilms tumor with an isochromosome 7q as the sole cytogenetic change, resulting in loss of 7p and gain of 7q material.

A new approach in the diagnosis and follow-up of bladder cancer. FISH analysis of urine, bladder washings, and tumors.

The aim of the present study was to ascertain whether fluorescence in situ hybridization (FISH) of urine could be a useful approach in bladder cancer. Herein, we present the cytogenetic and FISH findings in patients with and without bladder cancer. The samples examined with FISH consisted of urine, bladder washings, and tumor tissue, when available. The results obtained show that the FISH technique, particularly when used on urine, is a very useful tool in the diagnosis, early detection, and management of bladder cancer.

FISH studies of urinary cells of patients with bladder cancer.

Generally, bladder cancers are characterized by complex and numerous chromosome changes that vary from tumor to tumor. Nevertheless, certain chromosome changes recur with a consistency, (eg, +7, +8, -9, and -Y). In applying fluorescence in situ hybridization (FISH) studies to urinary cells in bladder cancer we chose probes for these chromosomes as well as those for chromosomes 10 and 11. A probe for the X chromosome was used for female patients in place of the Y. In the present study, we show that FISH of urine samples can detect the presence of cancer cells in transitional cell carcinoma (TCC) of the bladder of any grade and stage, including carcinomas in situ (CIS). We analyzed 27 samples from 25 patients (three were from the same patient); 24 samples were recurrent or newly diagnosed TCC and 3 were CIS. Our results show that FISH of urine samples is a reliable test for the detection of bladder cancer cells, regardless of the grade and stage of the tumor, and that a correlation appeared to exist between invasiveness of the tumor and the number of abnormalities in such tumor.

(Over)correction of FMR1 deficiency with YAC transgenics: behavioral and physical features.

Fragile X syndrome is a common cause of mental retardation involving loss of expression of the FMR1 gene. The role of FMR1 remains undetermined but the protein appears to be involved in RNA metabolism. Fmr1 knockout mice exhibit a phenotype with some similarities to humans, such as macroorchidism and behavioral abnormalities. As a step toward understanding the function of FMR1 and the determination of the potential for therapeutic approaches to fragile X syndrome, yeast artificial chromosome (YAC) transgenic mice were generated in order to determine whether the Fmr1 knockout mouse phenotype could be rescued. Several transgenic lines were generated that carried the entire FMR1 locus with extensive amounts of flanking sequence. We observed that the YAC transgene supported production of the human protein (FMRP) which was present at levels 10 to 15 times that of endogenous protein and was expressed in a cell- and tissue-specific manner. Macro-orchidism was absent in knockout mice carrying the YAC transgene indicating functional rescue by the human protein. Given the complex behavioral phenotype in fragile X patients and the mild phenotype previously reported for the Fmr1 knockout mouse, we performed a more thorough evaluation of the Fmr1 knockout phenotype using additional behavioral assays that had not previously been reported for this animal model. The mouse displayed reduced anxiety-related responses with increased exploratory behavior. FMR1 YAC transgenic mice overexpressing the human protein did produce opposing behavioral responses and additional abnormal behaviors were also observed. These findings have significant implications for gene therapy for fragile X syndrome since overexpression of the gene may harbor its own phenotype.