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Biomedical subjects

M Greaves

Publications and source records attributed to M Greaves.

At least 55 records · Page 3Linked to original sources
PubMed1999

Structure and possible mechanisms of TEL-AML1 gene fusions in childhood acute lymphoblastic leukemia.

TEL-AML1 gene fusion derived by chromosomal translocation is a common acquired genetic lesion in pediatric cancer that is present in approximately 25% of B-cell precursor acute lymphoblastic leukemias, and recent evidence suggests that this recombination event may initiate leukemogenesis prenatally during fetal hemopoiesis. Analysis of the DNA sequence and structure surrounding the breakpoints may reveal clues to their formation. A long-distance inverse PCR strategy was used to amplify TEL-AML1 genomic fusion sequences from diagnostic DNA from nine patients. Breakpoints were scattered within the 14 kb of intronic DNA between exons 5 and 6 of TEL and in two putative cluster regions within AML1 intron 1. Fusion sequences exhibited characteristic signs of nonhomologous end joining, including microhomologies at the end points, and small deletions and duplications. DNA sequences near the breakpoints did not reveal any consistent characteristic signal sequences of the V(D)J recombinase, topoisomerase II consensus sites, or other sequence motifs associated with recombination. However, several translocations occurred near a repeat region of TEL that was found to be highly polymorphic. This region was cloned and found in nuclease sensitivity assays to exhibit paranemic structures, which may have contributed to DNA breakage or illegitimate recombination. The data are compatible with the possibility that TEL-AML1 translocations occur by nonhomologous recombination involving imprecise, constitutive repair processes following DNA double-strand breaks.

Base Sequence↗
PubMed1999

Protracted and variable latency of acute lymphoblastic leukemia after TEL-AML1 gene fusion in utero.

We report a pair of identical twins with concordant acute lymphoblastic leukemia (ALL). Unusually, their diagnoses were spaced 9 years apart at ages 5 and 14. Leukemic cells in both twins had a TEL-AML1 rearrangement, which was characterized at the DNA level by an adaptation of a long distance polymerase chain reaction (PCR) method. The genomic fusion sequence was identical in the two leukemias, indicative of a single cell origin in one fetus, in utero. At the time twin 1 was diagnosed (aged 5 years), the bone marrow of twin 2 was hematologically normal. However, retrospective scrutiny of the DNA from an archived slide with clonotypic TEL-AML1 primers showed that the presumptive preleukemic clone was present and disseminated 9 years before a clinical diagnosis. These data provide novel insight into the natural history of childhood leukemia and suggest that consequent to a prenatal initiation of a leukemic clone, most probably by TEL-AML fusion itself, the latency of ALL can be both extremely variable and protracted. This, in turn, is likely to reflect the timing of critical secondary events.

Adolescent↗
PubMed1999

Antiphospholipid antibodies and thrombosis.

Antiphospholipid antibodies are associated with arterial and venous thrombosis, recurrent pregnancy loss, and thrombocytopenia. Although the antibodies have not been conclusively shown to be causal in thrombosis and miscarriage, they are useful laboratory markers for the antiphospholipid syndrome. The identification of the syndrome is clinically important because of the risk of recurrent thrombosis and the need for antithrombotic therapy in many cases. Diagnosis and treatment of antiphospholipid syndrome is difficult, however, because of the protean clinical manifestations and associations, limitations of existing laboratory tests for antiphospholipid antibodies, and the absence of evidence-based guidance on best management.

Abortion, Habitual↗
PubMed1999

Molecular genetics, natural history and the demise of childhood leukaemia.

The patterns of genetic change, clonal evolution, natural history and latency are very different in the paediatric leukaemias compared with adult epithelial cancers but are similar to those in other childhood cancers of mesenchymal stem cell origin. This distinction has a biological logic in the context of the selective pressures for clonal emergence in different developmental and cellular contexts and has a major impact on curability. Most childhood leukaemias and some other mesenchymal stem cell tumours are of fetal origin and can metastasize without corruption of restraints on cell proliferation or bypassing apoptosis. In marked contrast to most invasive or metastatic epithelial carcinomas in adults, these former cancers then retain sensitivity to therapeutic apoptosis. Moreover, their abbreviated and less complex evolutionary status is associated with less genetic diversity and instability, minimising opportunity for clonal selection for resistance. A minority of leukaemias in children and a higher fraction in adults do, however, have genetic alterations that bypass cell cycle controls and apoptosis imposition. These are the 'bad news' genotypes. The cellular and molecular diversity of acute leukaemia impacts also on aetiology. Paediatric acute leukaemias can be initiated prenatally by illegitimate recombination and fusion gene formation in fetal haemopoiesis. For acute lymphoblastic leukaemia (ALL) in children, twin studies suggest that a secondary postnatal molecular event is also required. This may be promoted by an abnormal or delayed response to common infections. Even for a classic case of a cancer that is intrinsically curable by systematic chemotherapy i.e. childhood ALL, prevention may turn out to be the preferred option.

Apoptosis↗
PubMed1999

Molecular genetics, natural history and the demise of childhood leukaemia.

The patterns of genetic change, clonal evolution, natural history and latency are very different in the paediatric leukaemias compared with adult epithelial cancers but are similar to those in other childhood cancers of mesenchymal stem cell origin. This distinction has a biological logic in the context of the selective pressures for clonal emergence in different developmental and cellular contexts and has a major impact on curability. Most childhood leukaemias and some other mesenchymal stem cell tumours are of fetal origin and can metastasize without corruption of restraints on cell proliferation or bypassing apoptosis. In marked contrast to most invasive or metastatic epithelial carcinomas in adults, these former cancers then retain sensitivity to therapeutic apoptosis. Moreover, their abbreviated and less complex evolutionary status is associated with less genetic diversity and instability, minimising opportunity for clonal selection for resistance. A minority of leukaemias in children and a higher fraction in adults do, however, have genetic alterations that bypass cell cycle controls and apoptosis imposition. These are the 'bad news' genotypes. The cellular and molecular diversity of acute leukaemia impacts also on aetiology. Paediatric acute leukaemias can be initiated prenatally by illegitimate recombination and fusion gene formation in fetal haemopoiesis. For acute lymphoblastic leukaemia (ALL) in children, twin studies suggest that a secondary postnatal molecular event is also required. This may be promoted by an abnormal or delayed response to common infections. Even for a classic case of a cancer that is intrinsically curable by systematic chemotherapy i.e. childhood ALL, prevention may turn out to be the preferred option.

Adolescent↗
PubMed1999

JC and BK virus sequences are not detectable in leukaemic samples from children with common acute lymphoblastic leukaemia.

Epidemiological evidence suggests that childhood leukaemia, and possibly common acute lymphoblastic leukaemia in particular, may have an infectious aetiology. Smith (1997 J Immunother 20: 89-100) recently suggested that the critical infectious event occurs during pregnancy, and identified the polyoma virus JC as a candidate agent. In the present study we investigated whether genomes from the JC virus, and closely related BK virus, could be detected in leukaemic cells. No positive results were obtained suggesting that JC virus is unlikely to play a direct role in leukaemogenesis.

BK Virus↗
PubMed1999

Expression of interleukin-1beta and tumour necrosis factor-alpha in plasma cells from patients with multiple myeloma.

Interleukin-1beta(IL-1beta) and tumour necrosis factor-alpha (TNF-alpha) are potent bone resorbing cytokines that may contribute to the development of the osteolytic bone disease observed in patients with multiple myeloma (MM). Although these factors have been identified in cultures of bone marrow mononuclear cells isolated from patients, the identity of the cells responsible for producing IL-1beta and TNFalpha remains unclear. Using a sensitive dual-colour fluorescence in situ hybridization (FISH) technique and a two-colour immunofluorescence method we have investigated the expression of the mRNA and protein, for IL-1beta and TNFalpha, by individual bone marrow plasma cells from patients with MM and monoclonal gammopathy of undetermined significance (MGUS). The mRNA for IL-1beta and TNFalpha was identified in all cells expressing the immunoglobulin light chain from all patients with MM and MGUS. However, the IL-1beta protein could not be detected in cytoplasmic light chain positive cells in any of the patients examined. In contrast, the TNFalpha protein was detected in clonal plasma cells from patients with both MM and MGUS. Interestingly, the IL-1beta and TNFalpha mRNA and proteins were readily detected within a small proportion of the non-plasma cells from patients with both MM and MGUS. These data suggest that myeloma cells in vivo are able to produce TNFalpha but not IL-1beta. In addition, a small proportion of accessory cells are likely to be able to contribute to the production of both ILbeta and TNFalpha.

Aged↗
PubMed1999

Platelet glycoprotein IIIa polymorphism HPA 1b (PlA2): no association with platelet fibrinogen binding.

The role of the platelet glycoprotein (GP) IIIa polymorphism HPA 1b (PlA2) in the risk of arterial thrombosis is controversial. We investigated the effect of the 1b allele on platelet fibrinogen binding by flow cytometry. Samples from 35 healthy platelet and plasma donors possessing the 1b allele were compared with 35 1a/1a donors. We found no allele-dependent difference in percentage of platelets binding fibrinogen (P = 0.6), nor in mean cell fluorescence (P = 0.3) following stimulation with ADP. These results render it unlikely that any relationship between the 1b polymorphism and arterial thrombosis is mediated by a significant effect on fibrinogen binding.

Adult↗
PubMed1999

Haemochromatosis mutations in North-East Scotland.

The HFE gene and its mutations C282Y and H63D cause hereditary haemochromatosis (HH). Among 54 affected individuals from North-East Scotland, 91% were homozygous for C282Y and 5.5% were compound heterozygotes for C282Y and H63D. The general population allele frequencies were high (8% and 15.7% for C282Y and H63D respectively). Although it is likely that HH is under diagnosed, these figures suggest that disease expression is variable, and many of those with the genetic predisposition HH will never develop the clinical consequences of iron overload. This has implications for diagnosis and predictive testing.

Female↗
PubMed1999

Thrombosis, markers of thrombotic risk, indwelling central venous catheters and antithrombotic prophylaxis using low-dose warfarin in subjects with malignant disease.

Markers of thrombotic risk--fibrinogen, factor VIII and immunoglobulin G (IgG) anticardiolipin titres--were measured, and the presence of lupus anticoagulant and factor V Leiden were assessed in 84 patients with a solid or haematological malignancy. These patients were monitored, following the insertion of an indwelling venous catheter, for thrombosis. Fifty-five were given prophylactic low-dose warfarin. Over a mean of 15 weeks there were 10 (12%) thrombotic events in 10 patients. Seven were on warfarin. Haemorrhagic problems occurred in three (5%), all on warfarin. Of the 84 patients, 86% had raised fibrinogen levels, 37% elevated factor VIII and 44% raised anticardiolipin levels. Lupus anticoagulant was present in five and three were heterozygous for factor V Leiden. A high prevalence of a range of prothrombotic changes was confirmed and the frequent presence of low-titre anticardiolipin antibody in subjects with malignancy demonstrated. However, none of these parameters predicted the development of thrombosis (P > 0.05).

Anticoagulants↗
PubMed1999

Marked variation in antiphospholipid antibodies during pregnancy: relationships to pregnancy outcome.

Variations in blood concentrations of antiphospholipid antibodies (APA) were investigated through the course of pregnancy in women who had a history of recurrent pregnancy loss, and were related to changes in pregnancy outcome. Serial measurements of APA were made in 16 women with antiphospholipid syndrome (APS) and 16 with negative APA tests pre-pregnancy. There was considerable intraindividual variation in test results through pregnancy. There was a significantly higher ratio of dilute Russell's viper venom time and IgG ACA titre in the first trimester compared with results pre-pregnancy in women with APS. Furthermore, transiently positive APA results were noted in the control group during pregnancy and some women with antiphospholipid syndrome tested negative for APA in mid- and late pregnancy. We have demonstrated clinically important variations in the results of tests for APA during pregnancy in women with APS.

Abortion, Habitual↗
PubMed1999

Inadequacies in the postnatal management of fetomaternal alloimmune thrombocytopenia (FMAIT).

93 (31%) of the 301 Haematology Departments in the U.K. responded to a questionnaire about the postnatal management of fetomaternal alloimmune thrombocytopenia (FMAIT). The number of reported cases of FMAIT was less than half than that estimated from its known incidence, suggesting that the condition is under-recognized. There was a consensus that the optimal approach to postnatal management is to transfuse compatible platelets promptly. However, a number of problems in the delivery of treatment were identified, including an apparent lack of awareness of the potential seriousness of the condition amongst clinical staff, and the availability of HPA-1a negative platelets.

Antibody Specificity↗
PubMed1998

Fetal origins of the TEL-AML1 fusion gene in identical twins with leukemia.

The TEL (ETV6)-AML1 (CBFA2) gene fusion is the most common reciprocal chromosomal rearrangement in childhood cancer occurring in approximately 25% of the most predominant subtype of leukemia- common acute lymphoblastic leukemia. The TEL-AML1 genomic sequence has been characterized in a pair of monozygotic twins diagnosed at ages 3 years, 6 months and 4 years, 10 months with common acute lymphoblastic leukemia. The twin leukemic DNA shared the same unique (or clonotypic) but nonconstitutive TEL-AML1 fusion sequence. The most plausible explanation for this finding is a single cell origin of the TEL-AML fusion in one fetus in utero, probably as a leukemia-initiating mutation, followed by intraplacental metastasis of clonal progeny to the other twin. Clonal identity is further supported by the finding that the leukemic cells in the two twins shared an identical rearranged IGH allele. These data have implications for the etiology and natural history of childhood leukemia.

Amino Acid Sequence↗
PubMed1998

Regulation of the apoptotic response to radiation damage in B cell development.

B lymphocyte precursor cells are ultrasensitive to DNA damage induced by irradiation and drugs and die by apoptosis at very low levels of exposure. Previous studies have shown that this high level sensitivity is p53-dependent, associated with very low level expression of Bcl-2 protein and can be reversed by expression of a bcl-2 transgene. We show here that transition from the pro-B to pre-B and then mature B cell stages of murine lymphopoiesis is accompanied by changes in proliferating cells in sensitivity to X-irradiation induced apoptosis and that this is paralleled by variation in the ratio of anti-(Bcl-2/Bcl-chiL) to pro-(Bax) apoptotic proteins. These are however not fixed or invariant features of developmental stage as they can be modulated by interactions via adhesive interactions with stromal cells, stromal proteins and growth factors. We interpret these data in the context of the stringent developmental regulation of clonal lymphopoiesis and the contingency programming of cells that have extensive proliferative potential with a very low threshold for apoptosis following DNA damage.

Animals↗
PubMed1998

Interleukin-6 is expressed by plasma cells from patients with multiple myeloma and monoclonal gammopathy of undetermined significance.

Interleukin-6 (IL-6) is an important growth factor for human myeloma cells in vitro and in vivo. However, the identity of the cells producing IL-6 in vivo in patients with multiple myeloma (MM) remains the subject of debate. We have developed a sensitive dual-colour fluorescence in situ hybridization (FISH) technique to investigate the expression of IL-6 mRNA by individual bone marrow plasma cells from patients with multiple myeloma, monoclonal gammopathy of undetermined significance (MGUS) and healthy subjects. IL-6 mRNA could be identified in all immunoglobulin light chain (IgLC) expressing cells from all patients with MM and MGUS. The IL-6 protein could also be detected by direct immunofluorescence in all plasma cells (cytoplasmic light chain positive) from all patients with MM and MGUS. Furthermore, it was also possible to demonstrate cytoplasmic IL-6 staining of plasma cells from patients with MM by flow cytometric analysis. In contrast, neither the IL-6 mRNA or protein could be detected in normal plasma cells from healthy bone marrow donors. These data demonstrate that plasma cells from patients with MM and MGUS express the IL-6 mRNA and synthesize the IL-6 protein and support the hypothesis that autocrine synthesis of IL-6 is of importance in patients with MM.

Aged↗
PubMed1998

IgG from patients with antiphospholipid syndrome binds to platelets without induction of platelet activation.

The clinical manifestations of the antiphospholipid syndrome (APLS) include arterial and venous thrombosis, thrombocytopenia and fetal loss, but the pathogenic mechanisms remain unclear. It has been hypothesized that platelet activation by autoantibody may be a pathogenic mechanism. We studied IgG binding, microparticle (mp) formation and P-selectin expression by flow cytometry in normal platelets after incubation in serum from 11 patients with antiphospholipid antibodies and that from 10 normal healthy subjects. Levels of platelet-associated IgG were significantly higher after incubation in patient sera (mean 17.2, range 2.0-75.0%) compared with normal sera (mean 2.0, range 1.2-3.7%, P<0.05). Incubation of normal platelets in serum led to increased microparticle formation (P<0.01) and P-selectin expression (P < 0.05), compared with unstimulated platelets. There was no significant difference, however, between microparticle formation nor P-Selectin expression induced by patient serum (mp 3.0 (1.6-5.0)%; P-selectin 8.0 (4.0-16.6)%) versus normal serum (mp 3.2 (2.1-4.5)%; P-selectin 10.1 (4.0-15.6); median (range)). Pre-activation of platelets with sub-threshold ADP concentrations or thrombin receptor activator peptide resulted in a small increase in microparticle formation, but there was still no significant difference between the effects of patient and control sera. Despite the presence of platelet membrane binding IgG in serum from 5/11 patients with antiphospholipid antibodies, there was no evidence for associated enhanced platelet-activating ability. This study supports antiplatelet reactivity in antiphospholipid syndrome, but not a direct platelet-activating role for platelet-directed autoantibodies.

Antibodies, Antiphospholipid↗