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Biomedical subjects

D Y Mason

Publications and source records attributed to D Y Mason.

At least 37 records · Page 2Linked to original sources
PubMed2000

Functional characterization of HLA-F and binding of HLA-F tetramers to ILT2 and ILT4 receptors.

HLA-F is a human non-classical MHC molecule. Recombinant HLA-F heavy chain was refolded with 2-microglobulin to form a stable complex. This complex was used as an immunogen to produce a highly specific, high-affinity monoclonal antibody (FG1) that was used to study directly the cellular biology and tissue distribution of HLA-F. HLA-F has a restricted pattern of tissue expression in tonsil, spleen, and thymus. HLA-F could be immunoprecipitated from B cell lines and from HUT-78, a T cell line. HLA-F binds TAP, but unlike the classical human class I molecules, was undetected at the cell surface. HLA-F tetramers stain peripheral blood monocytes and B cells. HLA-F tetramer binding could be conferred on non-binding cells by transfection with the inhibitory receptors ILT2 and ILT4. Surface plasmon resonance studies demonstrated a direct molecular interaction of HLA-F with ILT2 and ILT4. These results, together with structural predictions based on the sequence of HLA-F, suggest that HLA-F may be a peptide binding molecule and may reach the cell surface under favorable conditions, which may include the presence of specific peptide or peptides. At the cell surface it would be capable of interacting with LIR1 (ILT2) and LIR2 (ILT4) receptors and so altering the activation threshold of immune effector cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Expression of the ALK tyrosine kinase gene in neuroblastoma.

ALK (anaplastic lymphoma kinase) is a tyrosine kinase receptor, expressed as part of the chimeric NPM-ALK protein, in anaplastic large cell lymphomas (ALCLs) exhibiting the t(2;5)(p23;q35) translocation. As a result of this translocation, the NPM (nucleophosmin) gene is fused to the portion of the ALK gene encoding its intracytoplasmic segment. In normal mouse tissues, mRNA encoding the Alk receptor has been found only in neural cells, suggesting involvement of this receptor in the development of the nervous system. The purpose of the present study was to examine the presence of ALK transcripts and protein in normal human tissues and a variety of cell lines and human tumors. Emphasis was placed on neuroblastomas because other tyrosine kinase receptors are expressed in human neuroblastomas. Fifty-six cell lines, including 29 lines of neural origin, and lymphoid and nonlymphoid tissue specimens, including 24 neuroblastomas, were investigated for ALK expression, using reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemistry. The results confirmed that mRNA encoding ALK protein was not detectable in any normal or neoplastic hematopoietic tissue tested, except for t(2;5)-positive ALCL. The salient finding was that 13 of the 29 cell lines of neural origin and 22 of 24 neuroblastomas were found to express ALK transcripts and ALK protein. However, no correlation was evident between any known prognostic factors and the level of ALK expression.

Anaplastic Lymphoma Kinase↗
PubMed2000

Anaplastic large-cell lymphomas of B-cell phenotype are anaplastic lymphoma kinase (ALK) negative and belong to the spectrum of diffuse large B-cell lymphomas.

There is controversy in the literature as to whether anaplastic large-cell lymphoma of B-cell phenotype is related to the t(2;5)-positive T- or 'null' cell lymphoma of the same morphology. We report a study of 24 lymphomas with morphological features of anaplastic large-cell lymphoma which expressed one or more B-cell markers and lacked T-lineage markers. Clinical features were more in keeping with large B-cell lymphoma than with classical t(2;5)-positive anaplastic large-cell lymphoma, and immunostaining for anaplastic lymphoma kinase (ALK) protein provided no evidence for the (2;5) translocation (or one of its variants). The staining patterns for CD20 and CD79 were typical of diffuse large B-cell lymphoma, CD30 expression was variable, and most cases (15/22) lacked epithelial membrane antigen (EMA). These findings support the view that 'B-cell anaplastic large-cell lymphoma' is unrelated to t(2;5)-positive (ALK-positive) lymphoma, and that it represents a morphological pattern occasionally encountered among diffuse large B-cell lymphomas. By the same reasoning, most tumours diagnosed as 'ALK-negative anaplastic large-cell lymphoma of T-cell or null phenotype' probably belong to the spectrum of peripheral T-cell lymphomas.

Adolescent↗
PubMed1999

Lymphomas expressing ALK fusion protein(s) other than NPM-ALK.

The tumor cells in ALK-positive lymphoma ("ALKoma") usually express the product of the NPM-ALK chimeric gene, generated by the t(2;5) chromosomal translocation. However, 10% to 20% of ALK-positive lymphomas express ALK fusion protein(s) other than NPM-ALK, and in this report, we describe the immunohistologic and clinicopathologic features of 15 such cases. The absence of the NPM-ALK fusion gene was confirmed by reverse transcriptase-polymerase chain reaction (RT-PCR) in 8 cases and by fluorescence in situ hybridization (FISH) analysis in a further 2 cases. In each case, ALK staining was restricted to the cytoplasm and the N-terminus of NPM to the nucleus (contrasting with lymphomas expressing NPM-ALK in which cytoplasmic as well as nuclear labeling is seen). However, in the course of screening 53 ALK-positive lymphomas, 2 biopsies were found that had a "cytoplasm-only" ALK staining pattern but that nevertheless were shown to carry the (2;5) (by NPM staining and RT-PCR). The 15 cases resembled typical NPM-ALK-positive lymphomas in that all were of T or null phenotype, usually occurred in young male patients, and frequently presented with advanced disease associated with systemic symptoms and extranodal involvement. Moreover, their prognosis was excellent and indistinguishable from that of classical t(2;5)-positive tumors, but was clearly different from that of ALK-negative anaplastic large-cell lymphomas. These results suggest that lymphomas carrying variants of the NPM-ALK fusion protein can be detected by immunostaining for ALK and NPM and also that they can be grouped with classical t(2;5)-positive tumors as a single entity (ALK-positive lymphoma or "ALKoma") that shows a better prognosis than ALK-negative anaplastic large-cell lymphoma.

Adolescent↗
PubMed1999

TRK-fused gene (TFG) is a new partner of ALK in anaplastic large cell lymphoma producing two structurally different TFG-ALK translocations.

Anaplastic large cell lymphoma (ALCL) is associated with the t(2;5)(p23;q35), which generates the NPM-ALK fusion gene encoding an 80-kD protein. Several studies have suggested that genes other than NPM may be fused to the ALK gene. Here we have identified TRK-fused gene (TFG) as a new ALK partner in 2 ALCL, 1 of which exhibited a t(2;3)(p23;q21). In these cases, TFG was involved in 2 different fusion genes, TFG-ALK(S) and TFG-ALK(L), coding respectively 85-kD and 97-kD chimeric proteins. The ALK breakpoint in these translocations was the same as in the classic t(2;5) translocation. These 2 proteins were both active in an in vitro tyrosine kinase assay showing that the new cloned cDNA sequences are translated into chimeric proteins with functional activity. These findings indicate that TFG can provide an alternative to NPM as a fusion partner responsible for activation of the ALK and the pathogenesis of ALCL.

Adult↗
PubMed1999

t(1;2)(q21;p23) and t(2;3)(p23;q21): two novel variant translocations of the t(2;5)(p23;q35) in anaplastic large cell lymphoma.

Cytogenetic investigations in two cases of anaplastic large cell lymphoma (ALCL) showed novel variants of the classical (2;5)(p23;q35) translocation, namely a t(1;2)(q21;p23) and a t(2;3)(p23;q21). The tumor cells in both cases gave positive immunohistochemical labeling for ALK protein (with both monoclonal and polyclonal antibodies), demonstrating that these translocations induce aberrant expression of this kinase and suggesting that genes other than NPM can activate the ALK gene in ALCL. These two cases were shown by an in vitro kinase assay to express ALK kinases (104 kD and 97 kD, respectively), which differed in size from the classical NPM-ALK fusion product (80 kD). Moreover, ALK expression was confined to the cytoplasm of the tumor cells in each case, supporting the hypothesis that the observed nuclear localization of NPM-ALK in classical ALCL is not the site of oncogenic activity of the ALK kinase.

Adolescent↗
PubMed1999

BTG1: a triiodothyronine target involved in the myogenic influence of the hormone.

The product of the B-cell translocation gene 1 (BTG1), a member of an antiproliferative protein family including Tis-21/PC3 and Tob, is thought to play an important role in the regulation of cell cycle progression. We have shown in a previous work that triiodothyronine (T3) stimulates quail myoblast differentiation, partly through a cAMP-dependent mechanism involved in the stimulation of cell cycle withdrawal. Furthermore, we found that T3 or 8-Br-cAMP increases BTG1 nuclear accumulation in confluent myoblast cultures. In this study, we report that BTG1 is essentially expressed at cell confluence and in differentiated myotubes. Whereas neither T3 nor cAMP exerted a direct transcriptional control upon BTG1 expression, we found that AP-1 activity, a crucial target involved in the triiodothyronine myogenic influence, repressed BTG1 expression, thus probably explaining the low BTG1 expression level in proliferating myoblasts. In transient transfection studies, we demonstrated that an AP-1-like sequence located in the BTG1 promoter was involved in this negative regulation. Our present data also bring evidence that the stimulation of BTG1 nuclear accumulation by T3 or 8-Br-cAMP probably results from an increased nuclear import or retention in the nucleus. Lastly, BTG1 overexpression in quail myoblasts mimicked the T3 or 8-Br-cAMP myogenic influence: (i) inhibition of myoblast proliferation due to an increased rate of myoblast withdrawal from the cell cycle; and (ii) stimulation of terminal differentiation. These data suggest that BTG1 is probably involved in T3 and cAMP myogenic influences. In conclusion, BTG1 is a T3 target involved in the regulation of myoblast differentiation.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1999

Detection of normal and chimeric nucleophosmin in human cells.

In anaplastic large-cell lymphoma (ALCL), the (2;5) chromosomal translocation creates a fusion gene encoding the 80-kD NPM-ALK hybrid protein. This report describes three new monoclonal antibodies, two of which recognize, by Western blotting, the N-terminal portion of NPM present in the NPM-ALK fusion protein and also in two other NPM fusion proteins (NPM-RARalpha and NPM-MLF1). The third antibody recognizes the C-terminal portion (deleted in NPM-ALK) and reacts only with wild-type NPM. The three antibodies immunostain wild-type NPM (in paraffin-embedded normal tissue samples) in cell nuclei and in the cytoplasm of mitotic cells. Cerebral neurones, exceptionally, show diffuse cytoplasmic labeling. In contrast to normal tissues, the two antibodies against the N-terminal portion of NPM labeled the cytoplasm of neoplastic cells, in four ALK-positive ALCL, reflecting their reactivity with NPM-ALK fusion protein, whereas the antibody to the C-terminal NPM epitope labeled only cell nuclei. Immunocytochemical labeling with these antibodies can therefore confirm that an ALK-positive lymphoma expresses NPM-ALK (rather than a variant ALK-fusion protein) and may also provide evidence for chromosomal anomalies involving the NPM gene other than the classical (2;5) translocation.

Animals↗
PubMed1999

Immunohistochemical screening for oncogenic tyrosine kinase activation.

Tyrosine kinases causing the abnormal phosphorylation of intracellular proteins have been shown to contribute to oncogenic transformation in a number of human neoplasms. Immunohistological staining of routine biopsy sections for increased levels of phosphotyrosine may therefore provide a simple means of screening for tumours containing activated tyrosine kinases. In this study, monoclonal antibodies to phosphotyrosine were used to immunostain a cell line and tumour biopsies from lymphomas known to contain the activated anaplastic-lymphoma-kinase (ALK) tyrosine kinase. A range of normal and other neoplastic tissues were also immunostained for comparison. An anaplastic large cell lymphoma (ALCL) cell line carrying the (2;5) translocation, which creates the activated nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) tyrosine kinase, was strongly labelled. Routine tissue biopsies from five cases of ALK-positive ALCL were also strongly positive for phosphotyrosine. The characteristic granular cytoplasmic labelling pattern for phosphotyrosine observed in a B-cell lymphoma (expressing full length ALK kinase) was identical to that obtained using an ALK-specific antibody, thus confirming that labelling for phosphotyrosine in lymphoma cells reflects the presence of an activated kinase. When normal lymphoid tissues were stained, there was little or no labelling for phosphotyrosine, but stronger labelling was seen in other cells and tissues; for example, endothelial cells and some carcinoma samples. Whilst the strong labelling for phosphotyrosine observed in the lymphoma cells is due to the presence of activated ALK, the strong staining of some normal cells presumably represents physiologically active kinases and this should be taken into account when interpreting the immunostaining of non-lymphoid tumours. The simplicity of this method, however, means that it offers a new rapid approach to the screening of large numbers of tumours for the presence of aberrant tyrosine kinase activation, particularly if they arise from tissues which normally contain only background levels of phosphotyrosine.

Antibodies, Monoclonal↗
PubMed1999

Identification of the CD85 antigen as ILT2, an inhibitory MHC class I receptor of the immunoglobulin superfamily.

The CD85 molecule was originally defined at the Fifth Workshop on Leucocyte Antigens in 1993 by two monoclonal antibodies, VMP55 and GHI/75. This cell-surface glycoprotein is expressed on B cells, monocytes, and subpopulations of T and natural killer (NK) cells, and particularly high levels are expressed by normal and neoplastic plasma cells and by hairy cell leukemia B cells. We affinity purified the CD85 antigen and obtained tryptic peptide sequence which indicated that this molecule might be ILT2, a recently described inhibitory major histocompatibility complex class I receptor of the immunoglobulin superfamily. This was confirmed by showing that both of the original anti-CD85 mAbs stained ILT2 transfectants. The cell signaling role demonstrated for ILT2 is consistent with the previously reported involvement of CD85 in T cell activation.

Antibodies, Monoclonal↗
PubMed1999

Biochemical detection of novel anaplastic lymphoma kinase proteins in tissue sections of anaplastic large cell lymphoma.

The (2;5) translocation, found in many T-cell and null cell anaplastic large cell lymphomas (ALCLs), creates a hybrid gene encoding the 80-kd NPM-ALK protein. Typically neoplastic cells show labeling of both nucleus and cytoplasm for anaplastic lymphoma kinase (ALK) and for the N-terminus of nucleophosmin (NPM). However, 10-20% of cases exhibit cytoplasmic labeling only for ALK, indicating the probable presence of variants of the classical (2;5) translocation that do not involve the NPM gene. We report the detection (using Western blotting and an in vitro kinase assay) in seven such ALCL cases, of ALK proteins with molecular masses of 85 kd, 97 kd (one case exhibiting a (2;3)(p23;q21) translocation), 104 kd (one case carried a (1;2)(q21;p23) translocation), and 113 kd. Tyrosine kinase activity was detected in four of these proteins, but the N-terminal portion of NPM could not be detected. These results show how ALCL cases that express ALK proteins other than NPM-ALK can be detected by sensitive biochemical techniques using routine cryostat sections.

Animals↗
PubMed1999

Lymphocyte-specific protein 1: a specific marker of human leucocytes.

While both murine and human homologues of the LSP1 gene (lymphocyte-specific gene 1) and its protein products have been identified, studies on human LSP1 have been limited. The present report describes a detailed immunocytochemical study of the distribution and localization of human LSP1 in both normal and neoplastic cells and tissues. The specificity of the monoclonal anti-LSP1 reagent was confirmed by expression cloning and transfection studies. The intracellular 60 000 MW LSP1 protein was found to be present in peripheral blood B cells, monocytes and granulocytes but absent in a subpopulation of circulating T cells (10-15% of CD3-positive T cells). The presence of LSP1 protein in medullary thymocytes, but only in scattered cortical thymocytes, provided additional evidence for heterogeneity of expression in T cells. Novel observations also included the presence of LSP1 in plasma cells, dendritic cells and Langerhans' cells. The leucocyte-restricted distribution of LSP1 protein means that it may play an important role in haematopathology. LSP1 protein was detected in a wide range of leukaemias and lymphomas, particularly of B-cell origin, and in tumour cells in classical Hodgkin's disease. Of interest was the indication of a reciprocal relationship in the expression of LSP1 and ALK (anaplastic lymphoma kinase) proteins in patients with anaplastic large cell lymphoma. As the anti-LSP1 reagent used in the present study recognizes a formalin-resistant epitope it should be of considerable value in the diagnosis of routinely fixed material.

Anaplastic Lymphoma Kinase↗
PubMed1998

ALK-positive lymphoma: a single disease with a broad spectrum of morphology.

The t(2;5)(p23;q35) translocation, associated with anaplastic large-cell lymphoma (ALCL), results in the expression of a chimeric NPM-ALK protein that can be detected by the ALK1 monoclonal antibody. This report describes the morphologic and phenotypic spectrum of 123 cases of lymphoma that all express ALK protein. The results provide strong evidence that the morphologic patterns of ALCL described in previous reports as representing possible subtypes of ALCL, eg, common type, lymphohistiocytic, or small cell patterns, are morphologic variants of the same disease entity. All of these morphologic patterns could be found within this series, and in some patients different subtypes coexisted in a single biopsy or were found in successive biopsies from a single patient. The link between these morphologic subtypes is further reinforced by the presence in all cases of a highly characteristic large cell, with an eccentric nucleus and an eosinophilic paranuclear region. We suggest that this cell can be considered as a major distinguishing feature of ALK-positive lymphomas. Another characteristic of these tumors was the perivascular pattern of neoplastic cell infiltration seen in a significant number of cases. In addition to ALK protein, all tumors expressed epithelial membrane antigen and lacked CD15, features that may be of value in differentiating ALCL from Hodgkin's disease. In the majority of cases (84%), malignant cells showed both a cytoplasmic and nuclear staining for ALK1 and thus presumably carried the 2;5 translocation, but staining was restricted to the cytoplasm in a few cases, suggesting that translocations other than t(2;5) may induce expression of ALK protein. We conclude from this study that ALK-positive neoplasms represent a distinct entity. Because their morphology is often neither anaplastic nor large cell, we suggest that they should henceforward be referred to as ALK lymphomas.

Adolescent↗
PubMed1998

Nucleolar localization of the nucleophosmin-anaplastic lymphoma kinase is not required for malignant transformation.

The (2;5)(p23;q35) lymphoma-associated chromosomal translocation creates a novel fusion gene that incorporates parts of the anaplastic lymphoma kinase (ALK) receptor tyrosine kinase and nucleophosmin genes. We report here that the product of this fusion gene accumulates within the nucleoli of neoplastic cells, and that previous reports of a predominantly cytoplasmic localization for the protein represent a tissue-processing artifact. However, nucleolar accumulation of nucleophosmin-ALK may not be necessary for its oncogenic action, because an ALK protein expressed in a lymphoma carrying a variant (1;2) chromosomal translocation did not accumulate in nucleoli. Furthermore, an engineered hybrid TPR-ALK protein can transform rodent fibroblasts and produce lymphomas in mice while remaining confined to the cytoplasm. We propose that the transforming action of ALK may not be reliant on its nucleolar localization, a hypothesis that may have implications for studies of other proteins involved in oncogenesis that are relocalized after the creation of fusion genes.

Anaplastic Lymphoma Kinase↗
PubMed1998

Kupffer cell staining by an HFE-specific monoclonal antibody: implications for hereditary haemochromatosis.

Hereditary haemochromatosis is an inherited disorder of iron absorption that leads to excessive iron storage in the liver and other organs. A candidate disease gene HFE has been identified that encodes a novel MHC class I like protein. We report the development of a monoclonal antibody (HFE-JB1) specific for recombinant refolded HFE protein. The antibody immunoprecipitates a 49 kD protein from the cell line U937, a histiocytic lymphoma. It binds HFE but does not recognize other recombinant non-classic MHC class I proteins (HLA-E, F and G), nor does it react with a variety of recombinant classic class I MHC molecules. COS cells transfected with HFE in culture are stained specifically. The immunohistochemical staining pattern in human tissues is unique and can be defined as a subset of the transferrin receptor positive cells. In the liver HFE protein was shown to be present on Kupffer cells and endothelium (sinusoidal lining cells), but absent from the parenchyma. Kupffer cells from an untreated C282Y HH patient failed to stain with the antibody. In the normal gut scattered cells in the crypts are stained. HFE was also present on capillary endothelium in the brain (a site of high levels of transferrin receptor) and on scattered cells in the cerebellum and cortex. These results raise interesting questions concerning the function of HFE in the control of body iron content and distribution.

Animals↗
PubMed1998

Statistical evaluation of diagnostic and prognostic features of CD30+ cutaneous lymphoproliferative disorders: a clinicopathologic study of 65 cases.

Several clinical and histopathologic features of 65 CD30+ cutaneous lymphoproliferations were evaluated for their diagnostic value between CD30+ primary versus secondary cutaneous lymphomas and for their prognostic significance. Primary cutaneous disease, spontaneous regression, and absence of extracutaneous spreading (but not age < or =60 years) were associated with a better prognosis. Epithelial membrane antigen, BNH9, CD15 or CBF.78 antigen were expressed in all types of cutaneous lymphoproliferations. However, epithelial membrane antigen immunoreactivity was more frequently expressed in CD30+ secondary cutaneous large-cell lymphoma. Among CD30+ primary cutaneous large-cell lymphoma, CD15 expression was only seen in localized skin lesions. P53 expression was not associated with spontaneous regression, extracutaneous spreading, or survival. Nested reverse transcriptase-polymerase chain reaction allowed the detection of NPM-ALK transcripts in 10 of 26 CD30+ primary and in 3 of 11 secondary cutaneous large-cell lymphomas. The ALK protein was detected in only 1 of 50 primary and in 4 of 15 secondary cutaneous CD30+ lymphoproliferations. In CD30+ primary cutaneous lymphoproliferation, NPM-ALK transcripts might be expressed by very rare normal or tumoral cells that are undetectable by immunohistochemistry. However, the expression of either NPM-ALK transcripts or ALK-protein was not correlated with prognosis or age in CD30+ cutaneous lymphoproliferations.

Biomarkers, Tumor↗
PubMed1997

Retrovirus-mediated gene transfer of NPM-ALK causes lymphoid malignancy in mice.

Approximately 5% to 10% of all non-Hodgkin's lymphomas contain a t(2;5)(p23;q35) chromosomal rearrangement, which we have previously shown results in the generation of the fusion protein nucleophosmin-anaplastic lymphoma kinase (NPM-ALK). To assess the transforming potential of NPM-ALK in an animal model, we infected 5-fluorouracil-treated murine bone marrow using retroviral stocks and transplanted this infected marrow into lethally irradiated BALB/cByJ mice. Male mice were transplanted with bone marrow from female donors at 10 weeks of age, with 7 of the animals receiving marrow infected with a retroviral construct, pSR alphaMSVtkneo-NPM-ALK, that contains the human NPM-ALK cDNA, and 4 serving as a control group, receiving "empty" pSR alphaMSVtkneo-infected marrow. Whereas all mice in the control group were alive and well up to 11 months after transplantation, 4 of the 7 mice transplanted with marrow containing the NPM-ALK construct developed lymphoma within 4 to 6 months. Tumors arose in the mesenteric lymph nodes, with metastases to the lungs, kidneys, liver, spleen, and the paraspinal area. When cells from the tumors and bone marrow were transplanted into sublethally irradiated secondary recipients, 10 of these 13 mice developed tumors within 9 months. Immunoblot analysis of cell lysates using an ALK polyclonal antibody showed NPM-ALK expression in all tumors examined. Histologically, the tumors were composed of a uniform population of large immunoblastic cells with basophilic cytoplasm, centrally placed nuclei, and distinct nucleoli. Genotypic analysis showed that the tumors were B-lineage and clonal, with rearrangements of the Ig heavy- and kappa light-chain loci and no rearrangements of the T-cell receptor beta locus. Immunocytochemical studies confirmed the presence of IgM heavy chains and kappa light chains within the tumor cells. Thus, in this retroviral gene transfer model, NPM-ALK expression in mice causes B-lineage large-cell lymphoma, suggesting a direct causative role for this activated fusion tyrosine kinase in human lymphoma.

3T3 Cells↗