1/10. Analysis of tumor cell evolution in a melanoma: evidence of mutational and selective pressure for loss of p16ink4 and for microsatellite instability.Tumorigenesis and tumor progression can be considered an evolutionary process. In order to deduce information on the mutational and selective pressures during melanoma progression we performed microsatellite analysis at 42 autosomal and two X-linked loci in a microdissected primary melanoma and its nine metastases. Loss of heterozygosity at locus D9S259 was the only genetic change observed in all metastases. The pattern of loss of heterozygosity at loci D9S162 and D9S171 within the region of common loss on chromosome 9p21 which encompasses the tumor suppressor gene p16ink4 enabled the distinction of four genetically different tumor cell populations. Three cell lineages showed homozygous loss of the p16ink4 gene, which evolved independently in each tumor cell population within the primary tumor. Additional allele losses could be demonstrated at markers D14S53 and DXS998. The fourth lineage did not demonstrate loss of heterozygosity at loci D9S162 and D9S171 and contained the wild type p16ink4 gene but was characterized by abundant microsatellite instability. The evolutionary approach towards tumorigenesis and tumor progression used in this study thus confirms the role of p16ink4 inactivation for melanoma progression but not for melanoma initiation; it suggests the existence of additional putative tumor suppressor genes located on 9p as well as on the long arm of chromosome 14 and shows that microsatellite instability may represent an alternative pathway of tumor cell evolution in malignant melanoma.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
2/10. Progression in a case of kearns-sayre syndrome.The quantitative relationship between deleted mitochondrial dna and the clinical course of patients with kearns-sayre syndrome is poorly understood. We investigated this point using tissue samples obtained at age 10 years when the patient was diagnosed as kearns-sayre syndrome and at age 20 years when he died of disseminated intravascular coagulation. By long polymerase chain reaction, a shortened mitochondrial genome (8.8 kb; normal, 16.6 kb) was detected in the patient. By quantitative competitive polymerase chain reaction, the percentage of deletion-carrying mitochondrial dna was not increased as expected and did not differ significantly by tissue type or sampling time or correlate with clinical course. Although we could not demonstrate that the amounts of wild-type mitochondrial dna decreased with accelerating progression, it was emphasized that such a reduction of mitochondrial dna in various tissues, including those of the central nervous system, could play a significant pathogenetic role, since only wild-type mitochondrial dna is functional in patients with large-scale deletions of mitochondrial dna.- - - - - - - - - - ranking = 2keywords = wild (Clic here for more details about this article) |
3/10. An Epstein-Barr virus deletion mutant associated with fatal lymphoproliferative disease unresponsive to therapy with virus-specific CTLs.There is a growing interest in using antigen-specific T cells for the treatment of human malignancy. For example, adoptive transfer of Epstein-Barr virus (EBV)-specific cytotoxic T lymphocytes (CTLs) has been effective prophylaxis and treatment of EBV-associated lymphoproliferative disease in immunocompromised patients. For all immunotherapies, however, there has been a hypothetical concern that mutations in tumor-specific antigens may lead to tumor escape. We now demonstrate that such events may indeed occur, with lethal outcome. A patient who developed lymphoma after marrow transplantation received donor-derived, EBV-specific CTLs but died with progressive disease. The tumor cells proved substantially less sensitive to cytolysis than the EBV-transformed B-cell line used for CTL generation. The major cytolytic activity of the donor CTL was directed against 2 HLA-A11-restricted epitopes in the viral EBNA-3B antigen. sequence analysis of this gene in the tumor virus revealed a 245-base pair deletion, which removed these 2 CTL epitopes. Hence, the viral antigen in the tumor had mutated in a way that allowed escape from CTLs. Analysis of EBV polymorphisms demonstrated that before CTL infusion, more than one virus was present, including a virus with wild-type EBNA-3B. After CTL infusion, only the virus with the EBNA-3B deletion could be detected, suggesting that the infused CTLs had selected a resistant strain in vivo. Such an occurrence, even when polyclonal CTL lines are used against genetically stable virus antigens, suggests that escape mutants may be a serious problem when CTL therapy is directed against more unstable tumor cell-derived targets.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
4/10. Recruitment of nonexpanded polyglutamine proteins to intranuclear aggregates in neuronal intranuclear hyaline inclusion disease.Recruitment of polyglutamine-containing proteins into nuclear inclusions (NIs) was investigated in neuronal intranuclear hyaline inclusion disease (NIHID). Some polyglutamine-containing proteins, ataxin-2, ataxin-3, and tata box binding protein (TBP), as well as unidentified proteins with expanded polyglutamine tracts were recruited into NIs with different frequencies. Ataxin-3 was incorporated into most of the NIs and disappeared from its normal cytoplasmic localization, whereas only a small fraction of NIs contained ataxin-2 and TBP. The consistent presence of ataxin-3 in NIs could reflect a biological feature of wild-type ataxin-3, which is translocated into the nucleus under pathological conditions and participates in the formation of aggregates. Ataxin-2 also accumulated in the nucleus, but was not necessarily incorporated into NIs, suggesting that transport of these cytoplasmic proteins into the nucleus and their recruitment into NIs are not wholly explained by an interaction with a polyglutamine stretch and must be regulated in part by other mechanisms. The prevalence of ubiquitin-immunopositive NIs was inversely correlated to neuronal loss in all cases examined. This correlation could be explained if NI formation is a protective mechanism involving the ubiquitin-proteasome pathway. This hypothesis is supported by the finding that the polyglutamine epitope in the center of NIs was surrounded by ubiquitin.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
5/10. Full-length sequence and functional analysis of hepatitis b virus genome in a virus carrier: a case report suggesting the impact of pre-S and core promoter mutations on the progression of the disease.In chronic hepatitis b virus (HBV) infection, the quiescent immunotolerant phase evolves into the immunoactive phase. The aim of the present study was to clarify the virological alterations relevant to progression. Serial serum samples obtained from a patient with HBV during long-term follow-up were analysed by sequencing of the full-length HBV-dna using polymerase chain reaction (PCR). In addition, PCR products of HBV genome from each serum sample were transfected into HuH-7 human hepatoma cells for the functional analysis of the transfected viral genomes. Based on the HBV-dna sequence analysis, the patient had the genotype C virus, and the mutant HBV with common core promoter mutations (T(1762)A(1764)) and deletion of the pre-S region responsible for large surface protein transcription emerged before the onset of hepatitis. When the vigorous host immune response developed (indicated by the flare-up of hepatitis), the mutant HBV containing common core promoter mutations and another pre-S deletion causing lack of the surface protein promoter became predominant. The HBV-dna sequences, other than pre-S and core promoter regions were identical to the wild-type sequence throughout the study. transfection of PCR products containing the mutant HBV sequences resulted in increased amounts of intracellular replicative intermediates but the decreased secretion of HBsAg and HBeAg into culture media, suggesting accumulation of nonenveloped viral core particles within the cells. These results indicate that pre-S deletion and core promoter mutations may participate cooperatively in progression of the disease.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
6/10. Progressive onset of adrenal insufficiency and hypogonadism of pituitary origin caused by a complex genetic rearrangement within DAX-1.DAX-1 [dosage-sensitive sex reversal, adrenal hypoplasia congenital (AHC) critical region on the x chromosome, gene 1] is a transcription factor expressed in the adrenal gland and at all levels of the gonadotrope axis. Inactivating mutations of DAX1 result in the X-linked form of AHC with associated hypogonadotropic hypogonadism. AHC usually reveals itself as adrenal failure in early infancy, although a wide range of phenotypic expression has been reported. We describe a patient who was diagnosed with adrenal failure at 6 wk of age, but who experienced recovery of adrenal function of several months' duration later in infancy. He subsequently failed to undergo puberty because of hypogonadotropic hypogonadism of pituitary origin, and he was also diagnosed with schizophrenia in early adulthood. Molecular genetic analyses revealed a complex rearrangement in DAX1, including a 2.2-kb deletion spanning the entire second exon and a small 27-bp insertion. The putative protein encoded by this mutated gene is 429 amino acids long. The initial 389 residues probably correspond to the wild-type DAX-1 sequence, whereas the last 40 amino acids are presumably completely unrelated, being transcribed from the intronic sequence adjacent to exon 1. in vitro functional analyses confirm the absence of repressor activity exerted by such mutant protein. These studies expand the genotypic and phenotypic spectrum of DAX-1 insufficiency in humans.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
7/10. Prolonged retention of drug resistance mutations and rapid disease progression in the absence of therapy after primary hiv infection.We report two separate, unrelated instances of the transmission of hiv-containing mutations associated with high levels of resistance to protease inhibitors or reverse transcriptase inhibitors. In the absence of antiretroviral drugs, these mutations persisted almost unchanged in the newly infected index cases, whereas most mutations reverted to wild type in the source patients upon discontinuation of therapy. Furthermore, a rapid loss of CD4 cells was observed in the newly infected individuals.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
8/10. Slow progression of ataxia-telangiectasia with double missense and in frame splice mutations.ataxia-telangiectasia (A-T) is caused by mutations of the ATM gene, the product of which is involved in the regulation of cellular responses to radiation damage. ataxia usually starts in early childhood but a delayed age at onset and slower rate of neurological deterioration has been found for some patients with variant A-T. Only few patients have been documented to survive into the 4th decade. We report on a patient with an attenuated form of A-T who was diagnosed as having A-T by the age of 52 years and died by the age of 60 years. He was found to be a compound heterozygote for a double missense mutation (D2625E and A2626P) and a novel splicing mutation (496 5G --> A) of the ATM gene. Cytogenetic studies of the patient's lymphoblastoid cells revealed modest levels of bleomycin-induced chromosomal instability. Residual ATM protein was found at a level of 10-20% of wildtype. Low residual ATM kinase activity could be demonstrated towards p53, whereas it was poorly detectable towards nibrin. Our results corroborate the view that the clinical variability of A-T is partly determined by the mutation type and indicate that A-T can extend to late adulthood disease.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
9/10. Intentional overdose of Large Animal Immobilon.We describe a case of voluntary self-injection with Large Animal Immobilon, a veterinary anaesthesia product containing etorphine, a very strong opioid, and acepromazine, a phenothiazine. This resulted in cardiorespiratory arrest and the need for sustained haemodynamic support after resuscitation. Large Animal Immobilon is used under specific conditions only, mainly in zoo and wildlife medicine. Primary toxicological analysis, although guided by the presumed toxin, could only detect a metabolite of acepromazine in the urine. Further analysis was able to show some traces of etorphine. A number of topics are treated, including the apparent potency of the etorphine and the selection of a suitable antidote, taking into account the different properties of the respective agents.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
10/10. Somatic mosaicism in fanconi anemia: molecular basis and clinical significance.Approximately 25% of patients with fanconi anemia (FA) have evidence of spontaneously occurring mosaicism as manifest by the presence of two subpopulations of lymphocytes, one of which is hypersensitive to cross-linking agents (e.g. mitomycin C) while the other behaves normally in response to these agents. The molecular basis of this phenotypic reversion has not yet been determined. We have investigated 8 FA patients with evidence of mosaicism. Epstein-Barr virus-immortalized lymphoblastoid cell lines established from these patients exhibited an IC50 for mitomycin C of 25 to > 100 nM compared to a mean of 2 /- 2 nM for 20 nonmosaic FA patients and 49 /- 11 nM for 8 healthy controls. In 3 patients who were compound heterozygotes for pathogenic FAC gene mutations the molecular mechanism of the mosaicism was investigated by haplotype analysis. The results indicated that an intragenic mitotic recombination must have occurred leading to a segregation of a wild-type allele in the reverted cells and suggested two patterns of recombination. In 1 patient a single intragenic crossover between the maternally and paternally inherited mutations occurred associated with markers located distally to the FAC gene; in the other 2 patients (sibs) the mechanism appears to have been gene conversion resulting in segregants which have lost one pathogenic mutation. In 6 of the 8 patients the hematological symptoms were relatively mild despite an age range of 9-30 years.- - - - - - - - - - ranking = 1keywords = wild (Clic here for more details about this article) |
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