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J Amos

Publications and source records attributed to J Amos.

At least 19 recordsLinked to original sources

Testing environment for single-gene disorders in U.S. reference laboratories.

Molecular diagnosis is the detection of pathogenic mutations in DNA and RNA samples prepared from at-risk patients. Principles underlying DNA-based diagnosis originate from localization and identification of genes responsible for human disease and the characterization of pathogenic mutations; new diagnostic tests are in constant development. Indications for testing include diagnosis, carrier risk revision, prenatal diagnosis, presymptomatic diagnosis, and population screening. The majority of DNA diagnoses are performed in a reference laboratory in which the referring physician, rather that the patient, is the client of the laboratory. Thus, the role of the genetic counselor in a reference laboratory is quite different from traditional clinical practice. As with all clinical laboratory tests, molecular diagnostics are interpreted within the context of the patient's overall clinical presentation and are overlaid with regulatory programs of quality assurance, quality improvement, and quality control. The greatest challenges facing the clinical laboratory are genetic heterogeneity, cost control, regulatory and patent compliance, technology transfer, and the genetic education of the referring physician.

Genetic Counseling↗

Diminished support for linkage between manic depressive illness and X-chromosome markers in three Israeli pedigrees.

The hypothesis that chromosomal region Xq27-28 harbours a gene for manic-depression has been a focus of interest in human genetics. X-linked inheritance of manic depressive illness has been re-examined in 3 multigeneration Israeli kindreds. Extension and re-evaluation of pedigree data, including new individuals, diagnostic follow-up, and analysis with DNA markers, shows greatly diminished support for linkage to Xq28. The peak lod scores in two of the pedigrees have dropped several lod units to clearly negative values at the RCP-F8-G6PD gene cluster. On the other hand, positive lod scores (Zmax = 2.09) are sustained in another pedigree at the same map location. None of the pedigrees show linkage to more proximal markers, including the Xq27 locus DXS98. Our analysis underscores the uncertainties in studying complex disorders.

Adult↗

Waardenburg syndrome (WS) type I is caused by defects at multiple loci, one of which is near ALPP on chromosome 2: first report of the WS consortium.

Previous studies have localized the gene for Waardenburg syndrome (WS) type I to the distal portion of chromosome 2q, near the ALPP locus. We pooled linkage data obtained from 41 WS type I and 3 WS type II families which were typed for six polymorphic loci on chromosome 2q in order to refine the location of the WS locus (WS1) and evaluate the extent of genetic heterogeneity. In the course of this work, we developed diagnostic criteria for genetic and phenotypic studies. Our findings, based on two-locus and multilocus analysis using a linkage map established from reference pedigrees, suggest that there are two or more mutations causing WS, one of which (i.e., WS1) is located on chromosome 2q, between the ALPP and FN1 loci, at distances of 7.8 cM and 11.2 cM for each marker, respectively. The results also indicate that WS1 is responsible for the illness in approximately 45% of all families in this sample. However, the odds favoring this position over a location between ALPP and SAG are only 2:1 when alternate assumptions about the proportion of linked families are considered. We conclude that a more saturated map of this region of chromosome 2q, including highly polymorphic markers, will be needed to accurately distinguish linked families and, ultimately, isolate the mutant gene.

Alkaline Phosphatase↗

The value of deletion analysis for carrier detection in Duchenne muscular dystrophy (DMD).

We performed genetic analysis for carrier detection for several at-risk females in a four-generation Duchenne muscular dystrophy (DMD) pedigree using deletion analysis. We demonstrated that dosage analysis is a suitable alternative method to determine the carrier status of female relatives of DMD patients shown to have a deletion within the DMD gene. Subsequently, we diagnosed an affected male fetus for an at-risk female shown to be a DMD carrier by deletion analysis. The usefulness of deletion and linkage analysis are compared. In this family, linkage analysis was complicated by the unavailability of key family members, two recombination events and by previously undisclosed nonpaternity. We found that dosage analysis was more efficient than linkage for carrier evaluation in this family.

Adult↗

Genetic heterogeneity in tuberous sclerosis. Study of a large collaborative dataset.

Tuberous sclerosis (TSC) is a multisystem autosomal dominant hamartosis whose genetics is complicated by reduced penetrance and widely varying clinical expression. Results of linkage analyses have variously suggested two different locations for a TSC gene. A collaborative dataset has been assembled to clarify the issue of genetic heterogeneity. We have now analyzed the data from a combined sample of 111 families. Using Ott's HOMOG programs, we completed three tests of homogeneity: (1) for chromosome 9q, (2) for chromosome 11q, and (3) for the combined 9q and 11q data. For test 1 the chi-square (1 df) was 21.54 (p less than 0.001), for test 2 the chi-square (1 df) was 0.13 (p greater than 0.35), and for test 3 the chi-square (2 df) was 37.61 (p less than 0.0001). Additionally, we examined the combined data for evidence that a third, as yet unlinked locus exists. Results of this last test were suggestive but not significant. Clearly loci for TSC are present on both chromosomes 9q and 11q. The maximum likelihood estimate of the proportion of chromosome 9q-linked families is 0.38, for chromosome 11q-linked families is 0.47, and for the unlinked type 0.15. Alternative explanations for these latter families include chance sampling of recombinants, nongenetic phenocopies, or misclassification.

Chromosomes, Human, Pair 11↗

Multiple mutations in highly conserved residues are found in mildly affected cystic fibrosis patients.

We have identified three different point mutations in the coding region of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Each mutation segregates with the disease in two- or three-generation pedigrees and is not found on the normal chromosome of any documented cystic fibrosis carrier. One of the mutations is found in two independent families that contain at least one individual with a mild course of disease. All of these alterations replace charged amino acids with less polar residues and are found in the putative transmembrane sections of the molecule. The mutated amino acids are found to be conserved in both rodents and amphibians and lie in a region of CFTR that is believed to form a channel in the membrane. Although these alterations are rare, they provide important clues to functionally important regions of the molecule.

Adenosine Triphosphate↗

A frame-shift mutation in the cystic fibrosis gene.

Cystic fibrosis (CF) is a common recessive lethal genetic disorder, affecting 1 in 1,600 Caucasians. The disease causes defective regulation of chloride-ion transport in exocrine cells. Although in all CF families the disease is linked to a locus on chromosome 7q31, there is clinical heterogeneity in the severity of the disease and the age at which it is diagnosed. CF is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. A three-nucleotide deletion (delta F508) causing the loss of a phenylalanine residue in the tenth exon of the CFTR gene has been found on 70% of CF chromosomes. We have now characterized a CF family in which neither parent of the affected individual carries the common mutation, and identified a two-nucleotide insertion in the CF allele of the mother. The mutation introduces a termination codon in exon 13 of the CFTR gene at residue 821, and is predicted to result in the production of a severely truncated nonfunctional protein.

Alleles↗

[Polymorphism of restriction fragment length in the detection of the precise status of monosomy 21 in a deformed retarded girl].

The authors used genomic single copy DNA fragments cloned from chromosome 21 to study cytogenetic abnormalities in patients not easily defined by conventional cytogenetic means. Ten restriction fragment length polymorphisms (RLFP) detected by 8 independent probes were used to detect homologous sequences from chromosome 21 in genomic digests of DNA from one patient and her parents. The proband is a 3 1/2-year-old girl who was referred to us at 1 month of age because of hypertonia, hirsutism, flattened nasal bridge, antimongoloid slant of palpebral fissures, high arched palate and bilateral hip dysplasia. The karyotype of the proband was: 46, XX, -3, -21, + ? del (3) (3 pter----3q1:) +? (3qter----3q1:: 21q21----21 pter). GTG banding and the karyotype of her parents were normal (in peripheral blood and skin fibroblasts). She was re-examined by us every three months, because she showed physical and psychomotor retardation. We traced the inheritance of RFLPs from her parents, and familial molecular studies showed in contrast to the cytogenetic analysis that the patient is disomic for all regions of 21q tested by our collection of probes. The use of molecular technology has resulted in a more precise definition of 21 chromosome abnormalities and especially the "complete" monosomy 21 which is extremely rare in live born infants.

Abnormalities, Multiple↗

Inhibition of metabolic cooperation between mammalian cells in culture by tumor promoters.

The influence of phorbol-related tumour promoters and non-promoters on metabolic cooperation between wild-type and mutant Chinese hamster cells has been studied. The recovery, in medium containing 8-azaguanine, of hypoxanthine phosphoribosyl transferase-deficient (HPRT-) V79 cells co-cultured with an excess (2 x 10(6) per 9 cm petri-dish) of wild-type cells was determined in the presence and absence of each compound. Under the latter conditions (solvent treatment only) metabolic cooperation consistently reduced the cloning efficiency of HPRT- cells to approximately 10% of that in cultures without wild-type cells. However, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), the most potent known tumour promoter, almost totally reversed the effect of wild-type cells on mutant recovery when included in the medium at concentrations as low as 1 nM. TPA also markedly enhanced the expression, in crowded cultures, of HPRT- mutants induced by the carcinogen N-methyl-N-nitrosourea. This property was not shared by other phorbol esters which are inactive as tumour promoters, or by polycyclic aromatic hydrocarbons, but was exhibited to a lesser degree by mezerein, a diterpene ester of significant but weaker promoting activity than TPA. Confirmation that the effect of TPA on mutant expression is the result of inhibition of metabolic cooperation was obtained in experiments using autoradiography, which showed that low doses are able to block the transfer of [3H]-uridine nucleotides from prelabelled V79 cells to unlabelled V79 cells in contact. These findings have prompted us to formulate a working hypothesis for the mode of action of TPA in vivo as a tumour promoter based on its interference with this type of intercellular communication.

Carcinogens↗

Mutagenicity of carcinogenic methylating agents is associated with a specific DNA modification.

The carcinogenic potency of simple aliphatic alkylating agents such as the alkylnitrosamides and the alkylmethanesulphonates is positively correlated with their ability to alkylate the relatively weakly nucleophilic oxygen atoms in DNA, particularly the O6-atom of guanine. Differences in the spectrum of DNA alkylations produced by these agents can be rationalised on chemical grounds in that the electrophilic reactivity of the alkylating species determines the extent to which it will react at sites of weaker nucleophilicity. Alkylation of the more strongly nucleophilic ring nitrogen atoms of the purine bases, which is the main site of reaction with all these agents, appears to be much less important in alkylation carcinogenesis. O6-alkylation of guanine is likely to interfere with DNA base-pair hydrogen bonding and is possibly the major DNA modification responsible for the induction of GC yields AT transition mutations in bacteria and bacteriophage by alkylating agents. Here, we have studied the effects of three methylating agents of contrasting carcinogenic potency on mammalian (V79 Chinese hamster) cells in in culture. We report that the mutagenicity but not the cytotoxicity of each agent reflects its carcinogenicity and, furthermore, that the marked differences in mutagenicity are closely paralleled by differences in levels of O6-guanine methylation.

Alkylating Agents↗

Cell-mediated mutagenesis in cultured Chinese hamster cells by polycyclic hydrocarbons: mutagenicity and DNA reaction related to carcinogenicity in a series of compounds.

Three polycyclic hydrocarbons, benz(a)anthracene, 3-methylcholanthrene and 7,12-dimethylbenz(a)anthracene, have been studied in a cell-mediated mutagenesis system using BHK 21 cells to metabolize the hydrocarbons and V-79 cells as targets for detecting induced cytotoxicity and mutation. In large-scale experiments, the DNA of V-79 cells was analyzed by column chromatography to determine the nature and true extent of reaction of hydrocarbons with dexoyribonucleosides. Products with DNA formed by the two carcinogenic compounds were qualitatively very similar to those reported to occur in vivo and in primary cell cultures. Binding indices were calculated from the tritium content of DNA-hydrocarbon products, related to overall metabolism, for these two compounds together with benzo(a)pyrene and 7-methylbenz(a)anthracene using data from a previous study. These values reflected differences in carcinogenic potency between the compounds. Induced mutation frequencies were related to the extent of DNA reaction with each compound. At equivalent extents of DNA reaction with hydrocarbon products, levels of induced mutation were not significantly different.

9,10-Dimethyl-1,2-benzanthracene↗

The cytotoxic, mutagenic and clastogenic effects of chromium-containing compounds on mammalian cells in culture.

Examples of chromic and chromate salts have been examined for their effects on a cultured Chinese hamster cell line. The responses studied were cytotoxicity, mutagenesis and clastogenesis. Chromate (hexavalent chromium) salts of both high and medium water solubility were active in producing all three classes of response, whereas an insoluble chromate salt and a soluble chromic (trivalent chromium) salt were inactive. In addition to illustrating the value of using mammalian cells in culture for screening chemicals for biological activity, the results of this study reinforce current views regarding the genotoxic properties of chromates.

Animals↗

Different tumours induced by benzo(a)pyrene and its 7,8-dihydrodiol injected into adult mouse salivary gland.

A comparison has been made between the carcinogenic activities of benzo(a)pyrene and the proposed proximate carcinogen, benzo(a)pyrene 7,8-dihydrodiol, in the adult C57BL mouse submandibular salivary gland. In preliminary studies using a range of doses, the dihydrodiol was slightly less active than the parent hydrocarbon in this system. There was a difference in the type of tumour induced by the 2 compounds. Benzo(a)pyrene induced tumours of the salivary glands at the site of injection, whereas the dihydrodiol induced malignant lymphosarcomas, particularly of the thymus, which were often metastatic to other orgnas. Possible reasons for the different sites of action of the 2 compounds are discussed.

Animals↗

The nature of the deoxyribonucleosides involved in the binding of carcinogenic hydrocarbons to the DNA of mouse embryo cells.

The DNA of mouse embryo cells was specifically labelled in the purine moieties with (G(3)H)-deoxyadenosine or in the cytosine moieties with (5-(3)H)-deoxycytidine. These cells were then treated with 7-methylbenz (a) anthracene (7MBA) or benzo (a)-pyrene (B(a)P) and the DNA isolated, degraded and fractionated by LH20 Sephadex column chromatography. When the purines of the DNA were tritium-labelled, radioactive hydroccarbondeoxyribonucleoside products were obtained. No such products were found with deoxycytidine pre-labelled DNA. Contrary to an earlier suggestion, these results indicate that it is the purine moieties of DNA which react with the metabolically activated hydrocarbon derivative in vivo.

Animals↗