High throughput parallel analysis of hundreds of patient samples for more than 100 mutations in multiple disease genes

As more mutations are identified in genes of known sequence, there is a crucial need in the areas of medical genetics and genome analysis for rapid, accurate and cost-effective methods of mutation detection. We have developed a multiplex allele-specific diagnostic assay (MASDA) for analysis of large numbers of samples (> 500) simultaneously for a large number of known mutations (> 100) in a single assay. MASDA utilizes oligonucleotide hybridization to interrogate DNA sequences. Multiplex DNA samples are immobilized on a solid support and a single hybridization is performed with a pool of allele-specific oligonucleotide (ASO) probes. Any probes complementary to specific mutations present in a given sample are in effect affinity purified from the pool by the target DNA. Sequence-specific band patterns (fingerprints), generated by chemical or enzymatic sequencing of the bound ASO(s), easily identify the specific mutation(s). Using this design, in a single diagnostic assay, we tested samples for 66 cystic fibrosis (CF) mutations, 14 beta-thalassemia mutations, two sickle cell anemia (SCA) mutations, three Tay-Sachs mutations, eight Gaucher mutations, four mutations in Canavan disease, four mutations in Fanconi anemia, and five mutations in BRCA1. Each mutation was correctly identified. Finally, in a blinded study of 106 of these mutations in > 500 patients, all mutations were properly identified. There were no false positives or false negatives. The MASDA assay is capable of detecting point mutations as well as small insertion or deletion mutations. This technology is amenable to automation and is suitable for immediate utilization for high-throughput genetic diagnostics in clinical and research laboratories.      

Progressive osseous heteroplasia in the face of a child

We describe a rare case of progressive osseous heteroplasia of the face in a child. Biopsy showed osteoma cutis superficially with ectopic bone formation in the deeper tissues including skeletal muscle. Analysis of DNA from peripheral blood leukocytes showed mutations in the gene encoding the alpha subunit of the stimulatory G protein of adenylyl cyclase (GNAS1), confirming the diagnosis of progressive osseous heteroplasia.     

Familial Similarities of Changes in Cognitive, Behavioral, and Physiological Variables in a Cardiovascular Health Promotion Program

A number of studies have demonstrated that physiological andbehavioral cardiovascular disease (CVD) risk factors aggregatewithin families. This fact, and the potential mediating rolethat the family plays in behavior change, have led to the developmentof family-based CVD risk reduction programs, including the SanDiego Family Health Project. The aggregation of behavioral,physiological, and cognitive changes within families was assessedduring a 1-year intervention. We found evidence of modest butsignificant aggregation of change. There was more aggregationof change in behavioral variables than in physiological or cognitivevariables. More significant correlations were found among 3-dayfood record measures than among 24-hour recall dietary measures,suggesting an influence of assessment method. Aggregation ofchange within families was stronger within generations thanacross generations. These data point to the importance of involvingall age groups in health promotion programs.     

Variable expression of autosomal recessive polycystic kidney disease and congenital hepatic fibrosis within a family

Blyth and Ockenden [1971] assigned patients with autosomal recessive polycystic kidney disease (ARPCKD) to 4 discrete groups (perinatal, neonatal, infantile, juvenile) on the basis of the age of presentation. They and others speculated that at least 4 genes were responsible for what they considered to be closely related, but different conditions. These views have gained wide but not universal acceptance. Some workers have insisted that the perinatal and neonatal "forms" of ARPCKD differ fundamentally from the juvenile "form." However, others have proposed that ARPCKD-CHF (congenital hepatic fibrosis) and CHF-ARPCKD are manifestations of the same disease with variation of expression in a kindred. We report on a patient who presented at birth (1979) with ARPCKD and respiratory distress. He died at 18 hr. An older sib presented at 16 yr in 1984. She had no symptoms, but her mother wanted reassurance that the daughter did not have a condition similar to that of the deceased sib. Blood pressure was 120/80 mm Hg and there was hepatosplenomegaly. A diagnosis of renal tubular ectasia and CHF was made by ultrasonography, radiologic studies, and a liver biopsy. The evidence from families such as this favors the concept that ARPCKD and CHF presenting as Blyth and Ockenden's perinatal form, and CHF and renal tubular ectasia as their juvenile form, are manifestations of the same genetic disorder, and that the different manifestations are more likely variations in expression than the results of different mutant genes. The manifestations in this family add weight to the growing body of evidence that intrafamilial variability may occur, not only in autosomal dominant conditions, but also in autosomal recessive disorders.     

Inconsistent expression of both centromeres of a dicentric Y chromosome in a child with ambiguous external genitalia.

A newborn child with ambiguous external genitalia had evidence of internal female development on the left and internal male development on the right. Blood chromosome analysis showed three cell types: 45,X; 46,XY with the Y being submetacentric and about twice the usual size with two 'centromeric' C bands; and 46,X,dic(Y). Chromosome studies from the skin, uterus, and Fallopian tube showed almost exclusively 45,X cells. This represents the second reported patient in whom two centromeres are inconsistently expressed though present as shown by two 'centromeric' C bands.     

Genetic counseling in ornithine carbamoyltransferase deficiency

Ornithine transcarbamylase (OTC) deficiency is an inborn error of urea cycle metabolism, responsible for lethal hyperammonemia in males and for severe symptoms in at least 20% of heterozygous females. The authors provide here additional data on the informativity of the protein loading test (PLT) for the detection of heterozygotes. They show that the risk of being a carrier for the mother of an affected boy falls from 2/3 a priori to only 1/8 if her PLT is negative. The risk for the mother of heterozygote girl falls from 1/2 a priori to 1/16 if her PLT is negative.