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Biomedical subjects

H Knecht

Publications and source records attributed to H Knecht.

At least 55 records · Page 3Linked to original sources
PubMed1997

Synthesis, biodistribution and renal handling of various chelate-somatostatin conjugates with metabolizable linking groups.

A series of DTPA-octreotide conjugates, with various linking groups, were synthesised to investigate the effect of different metabolizable linkers on the renal retention of radioactivity. All these newly synthesised octreotide conjugates retained the high binding affinity of octreotide for the somatostatin (SRIF) receptors either when unlabeled or radiolabeled with 111In. Some of the metabolizable linkers were rapidly degraded in vitro when incubated with a kidney homogenate. However, in vivo, all these conjugates displayed a significantly lower uptake in SRIF receptor-positive tissue compared to two conjugates with short, stable linkers. Additionally, the compounds with a potentially metabolizable linker had a higher whole-body retention of activity as opposed to the three metabolically stable compounds. Several of the linkers gave evidence of cleavage while in circulation in the blood, and it is probable that the lower tumour accumulation of most of the compounds tested was low due to the high enzymatic nature of the exocrine pancreatic tumour model used. In short, no increase in the tumour-to-kidney ratio was achieved with the analogues containing a metabolizable linker. The highest target-to-nontarget tissue ratios were obtained for the DTPA-octreotide conjugates that had short, metabolically stable linkers.

Animals↗
PubMed1997

The 30-bp deletion variant of Epstein-Barr virus-encoded latent membrane protein-1 prevails in acute infectious mononucleosis.

To assess the frequency of malignancy-associated 30-bp deletion variants of the latent membrane protein 1 (LMP-1) in benign conditions, a comparative sequence analysis was done using samples from 20 American children with acute infectious mononucleosis and 16 Swiss children with chronic tonsillar hyperplasia. The 30-bp deletion variant (LMP-1-del) was present in 66% of patients (12/20 with infectious mononucleosis and 12/16 with tonsillar hyperplasia). Two additional patients had a 3-bp deletion and an inframe insertion of 18 nucleotides, respectively. All but 1 isolate had numerous nonsilent point mutations. These data identify a hypervariable region within the C-terminus of LMP-1, in a domain required for maximal stimulation of NF-kappaB activity. These data demonstrate that LMP-1-del variants are frequent in acute infectious mononucleosis and tonsillar hyperplasia and identical to those observed in Epstein-Barr virus-associated AIDS-related lymphoma.

Acute Disease↗
PubMed1997

Deletion variants within the NF-kappaB activation domain of the LMP1 oncogene in acquired immunodeficiency syndrome-related large cell lymphomas, in prelymphomas and atypical lymphoproliferations.

The latent membrane protein 1 (LMP1) oncogene of Epstein Barr virus (EBV) is expressed in tumor cells of acquired immunodeficiency syndrome (AIDS) related lymphomas, HIV-negative, EBV-associated malignant lymphoproliferations, nasopharyngeal carcinoma, as well as in reactive immunoblasts of infectious mononucleosis. Naturally occurring LMP1 deletion variants (LMP1-del), characterized by clustered mutations and a distinct 30 base pair deletion within the carboxy terminal domain of LMP1, essential for maximal NF-kappaB stimulation, have been identified in the same conditions. These variants prevail in AIDS-related lymphomas, and are associated with clinically aggressive behaviour in HIV-negative lymphomas, and are frequent in prelymphomatous and reactive states. Functional studies showing a growth advantage of cells infected by these variants may explain the accumulation of LMP1-del in these entities. In the carboxy terminal NF-kappaB activation domain of LMP1, evidence of a hypervariable region close to the highly conserved 23 outermost amino acids essential for malignant transformation, may reflect the natural selection of growth promoting variants involved in signalling pathways. The prevalence of the same mutational pattern in AIDS-related lymphoma as well as in hyperplastic reactive states and prelymphomas supports the hypothesis that these variants confer a growth advantage manifested under impaired cellular immunity.

Amino Acid Sequence↗
PubMed1997

Molecular and functional analysis of the Epstein-Barr virus LMP1 oncogene promoter in lymphoproliferative diseases.

The expression of the Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) oncogene is tightly regulated by viral and cellular factors. LMP1 is present in the majority of nasopharyngeal carcinoma tumor cells and in Reed-Sternberg cells from Hodgkin's lymphoma, in which the only EBV nuclear antigen detected is EBNA1. The aim of this study was to test whether mutations affecting LMP1 gene expression were present in lymphoproliferative disorders, and, if so, whether their presence correlated with the clinical course of the disease. For this purpose we characterized the LMP1 promoter region from seven cases including two patients with aggressive Hodgkin's disease and two with atypical lymphoproliferative syndromes. Our results show that the sequences -298 to +29 relative to the transcription start site diverged up to 9.3% when compared with the prototype EBV strain B95-8. The cAMP responsive-like element (CRE), located at positions -37 to -44, was found to be mutated in 3 of the 7 cases. Functional analysis of transfected human embryonic kidney 293 cells using the firefly luciferase reporter gene revealed that mutations within the CRE site led to a 70% mean decrease in reporter activity. Our analysis indicates that in lymphoproliferative disorders, naturally occurring LMP1 variants that exhibit weak promoter activity are still associated with clinically progressive disease.

Base Sequence↗
PubMed1996

Expression of the LMP1 oncoprotein in the EBV negative Hodgkin's disease cell line L-428 is associated with Reed-Sternberg cell morphology.

The latent membrane protein 1 (LMP1) oncogene is one of the major proteins synthesized by the Epstein-Barr virus (EBV) and is expressed in Reed-Sternberg (RS) cells of EBV-associated Hodgkin's disease (HD). We have studied the effect of this oncoprotein on the formation of RS cells in the EBV-negative HD cell lines L-428 and KM-H2 as well as on the formation of multinucleated cells in the mononuclear human embryonic kidney cell line 293. LMP1 prototype (B95-8) and its naturally occurring carboxy terminal 30 base pair deletion variant LMP1-del were transfected into the cell lines and cytocentrifuge preparations were analysed after 24, 48, 72, 144, 216, and 240 h. While no oncoprotein expression was seen in the KM-H2 cell lines, expression of LMP1 and LMP1-del was observed in the L-428 and 293 cell lines. In the HD cell line L-428 oncoprotein expression was infrequent but when observed was very strong and preferentially associated with multinucleated RS cell morphology (71% of LMP1 positive cells). This is in contrast with the untransfected or transfected but not expressing cells where intermediate mononuclear elements predominated over multinucleated RS cells (< 3%). Frequent oncoprotein expression was observed in the 293 cells and again was associated with multinuclearity. These LMP1 expressing 293 giant cells showed strong expression of ICAM-1(CD54), not detectable in the untransfected cells. In the LMP1-del transfectants giant cells with more than four nuclei were frequently observed. However, giant cells were much less frequent in 293 cells transfected with the amino terminal deletion variant of LMP1 or the lytic form of LMP1, known to induce low NF-kappa B activation compared to the LMP1 prototype. Therefore, LMP1 mediated NF-kappa B activation appears to be involved in polycaria formation. The strong association of LMP1 expression with multinuclearity in a genetically unstable condition -the L-428 and 293 cells show multiple chromosomal abnormalities-suggests that this oncoprotein including its naturally occurring carboxy terminal deletion variant promote the formation of multinuclear cells, in particular of RS cells in EBV-associated HD.

Blotting, Western↗
PubMed1996

Deletion variants within the NF-kappa B activation domain of the LMP1 oncogene prevail in acquired immunodeficiency syndrome-related large cell lymphomas and human immunodeficiency virus-negative atypical lymphoproliferations.

This sequencing study of 17 acquired immunodeficiency syndrome-related lymphomas (9 primary brain, 8 systemic) and 8 human immunodeficiency virus-negative atypical lymphoproliferations expressing large amounts of the latent membrane protein 1 (LMP1) of Epstein-Barr virus was performed to characterize the carboxy terminal NF-kappa B activation domain of LMP1 at the molecular level in an immunocompromised host. In-frame deletions within the NF-kappa B activation domain were identified in all but 2 primary brain lymphomas, 4 systemic lymphomas, and 4 atypical lymphoproliferations, ie, in 60% of cases. In addition, non silent point mutations (range 1 to 5, mean 3.3) were detected in all cases. Although all changes occurred within the first 100 nucleotides of the carboxy terminal NF-kappa B activation domain--a critical sequence for the protein half-life--not a single point mutation was found in the remaining 62 nucleotides, necessary for malignant transformation. Such a clustering of nonrandom sequence variations, associated with a high oncoprotein expression in immunocompromised hosts, suggests that this part of the LMP1 oncogene behaves as a hypervariable region with natural selection of growth-promoting variants through prolongation of the protein half-life.

Amino Acid Sequence↗
PubMed1996

Miliary tuberculosis in a patient with Epstein-Barr virus-associated angioimmunoblastic lymphadenopathy.

A 74-year-old woman developed angioimmunoblastic lymphadenopathy (AILD) with involvement of intra-abdominal and retroperitoneal lymph nodes. Southern blot analysis showed germline configuration of the JH genes and an oligoclonal pattern of the TcR beta genes. The immunoblasts were of B-cell phenotype and often expressed the CD30 antigen and the latent membrane protein 1 (LMP1) oncogene. Six nonsilent point mutations were identified near the 3' end of the LMP1 gene, leading to a cluster of six amino acid changes within a protein domain needed for maximal NF-kappa B stimulation. After a clinical remission of 8 months the patient relapsed with generalized lymphadenopathy and died secondary to tuberculosis. The oligoclonal rearrangements of the TcR beta genes may reflect an unsuccessful cellular immune response to Mycobacterium tuberculosis or an HLA-restricted T-cell response to B-immunoblasts expressing mutated viral antigens. A positive percutaneous tuberculin test observed 6 months prior to the onset of AILD is in favor of the first possibility.

Aged↗
PubMed1996

Inflammatory pseudotumor of the liver. Evidence for follicular dendritic reticulum cell proliferation associated with clonal Epstein-Barr virus.

We describe an "inflammatory pseudotumor" of the liver that, which on detailed investigation, proved that the spindle-cell component of this lesion is derived from follicular dendritic reticulum cells (FDRC). This contention is supported by morphologic observations and by immunophenotype. The FDRC population contain Epstein-Barr virus (EBV). It is known that FDRC express the EBV receptor CD21. In this particular case, the FDRC contained clonal EBV genomes, EBV RNA (EBER) transcripts, and expressed EBV latent membrane protein (LMP1). DNA sequencing of PCR products showed three point mutations compared with the standard LMP1 sequence of the EBV strain B95-8. The findings in this case corroborate those of other investigators concerning the possible role of EBV in the development of some inflammatory pseudotumors, including the recent production of functionally active EBV-transformed FDRC-like cell lines. This association could prove instructive in delineating the histogenesis of these tumors and further assist in making prognostic and therapeutic decisions.

Aged↗
PubMed1996

Leukemia with megakaryocytic differentiation following essential thrombocythemia and myelofibrosis. Case report and review of the literature.

Leukemias of megakaryocytic lineage are rare and heterogeneous clinical entities. The nomenclature published in the literature is confusing and perhaps inappropriate to designate these primary myeloproliferative disorders. We describe a patient with essential thrombocythemia who evolved through myelofibrosis and myeloid metaplasia to a final picture of leukemia with megakaryocytic differentiation in the peripheral blood. This case illustrates different aspects of a chronic myeloproliferative disorder where myelofibrosis and myeloid metaplasia are frequent but secondary events. We have reviewed the literature focusing on the role of clonal megakaryocytic proliferation in myelofibrosis and on the clinical characterization of leukemia with megakaryocytic phenotype. We also present our interpretation of the literature which indicates that a formal review of the nomenclature is urgently needed.

Female↗
PubMed1995

Mutational hot spots within the carboxy terminal region of the LMP1 oncogene of Epstein-Barr virus are frequent in lymphoproliferative disorders.

We have recently identified in Epstein-Barr virus (EBV) positive Hodgkin's disease (HD) a variant of the latent membrane protein 1 (LMP1) oncogene characterized by four point mutations and a 30 base pair deletion. These findings led us to test whether such mutants were also present in other lymphoproliferative disorders (LPD). We analysed 98 EBV DNA positive cases (67 LPD, 15 benign conditions, 16 lymphoblastoid cell lines) by PCR for deletions within the LMP1 gene. DNA sequencing of the region coding for the carboxy terminal protein domain was performed on 24 cases. In 13 cases the same combination of 4 point mutations at positions 168,320, 168,308, 168,295 and 168,225 was identified. Of these cases, 12 had an additional point mutation at position 168,357 and eight at position 168,355, and nine had a 30 base pair deletion including nucleotides 168,285 to 168,256. These deletion mutants were identified in HD, angioimmunoblastic lymphadenopathy, B-immunoblastic lymphoma, peripheral T-cell lymphoma, and two lymphoblastoid cell lines. Our findings reveal a high frequency of non-random point mutations at preferential sites within the 3' (carboxy terminal) region of the LMP1 oncogene. The association of these mutational hot spots with LPD suggests that they are involved in EBV related lymphomagenesis and that they define a clinically relevant EBV strain.

Antigens, Viral↗
PubMed1995

EBV-associated lymphoproliferative syndrome with a distinct 69 base-pair deletion in the LMP-1 oncogene.

We describe an immunocompetent 12-year-old boy with chronic EBV infection and lymphoid interstitial pneumonitis. Lymph node biopsies showed effacement of the architecture with polymorphic cellular infiltrates, consisting predominantly of T cells and natural killer cells. No clonal rearrangement of TCR or immunoglobulin genes was seen. DNA was extracted from hilar lymph nodes; sequencing of the carboxy terminal region of the latent membrane protein 1 (LMP-1) oncogene revealed a 69 base-pair deletion and four point mutations. Immunosuppressive treatment with prednisone and cyclosporine reversed the lymphadenopathy.

Base Composition↗
PubMed1995

Chronic relapsing neuropathy associated with Castleman's disease (angiofollicular lymph node hyperplasia).

We report a 17-year-old patient who presented a chronic relapsing sensorimotor demyelinating neuropathy with 6 relapses over a 7-year period, preceding by 4 years the diagnosis of a multicentric angiofollicular lymph node hyperplasia. A role for Epstein-Barr virus (EBV) as a trigger of the neuropathy may be suggested by the presence of EBV DNA in the biopsied abdominal abdominal adenopathies. This unusual reported association seems to have a better prognosis than the known chronic progressive form of neuropathy associated with Castleman's disease and the Crow-Fukase syndrome.

Adolescent↗
PubMed1995

A deletion mutant of the LMP1 oncogene of Epstein-Barr virus is associated with evolution of angioimmunoblastic lymphadenopathy into B immunoblastic lymphoma.

The latent membrane protein 1 (LMP1) oncogene is one of the major proteins synthesized by the Epstein-Barr virus (EBV). It is expressed in Reed-Sternberg cells of Hodgkin's disease (HD), tumor cells of nasopharyngeal carcinoma (NPC), and immunoblasts of angioimmunoblastic lymphadenopathy (AILD). A particular LMP1 deletion mutant was recently identified in NPC and clinically and histologically aggressive HD. We studied two patients with AILD that subsequently progressed into immunoblastic lymphoma (IBL) in order to investigate whether the LMP1 deletion mutant was implicated in progression of AILD into IBL. Immunohistology and in situ hybridization were performed on diagnostic biopsies. DNA extracted from fresh frozen material was used for rearrangement studies and polymerase chain reaction (PCR) based amplification and sequencing of portions of the LMP1 gene. Immunohistochemistry revealed B cell origin of both cases of IBL. In the first patient clonal rearrangement of the immunoglobulin heavy-chain gene was present in IBL but not in AILD. In this patient, scattered immunoblasts of AILD and numerous tumor cells of B-IBL were shown to contain EBV transcripts (EBER1) and to express LMP1. Sequence analysis of the LMP1 gene from AILD and IBL in the first, and from IBL in the second patient, revealed identical deletions and point mutations. This LMP1 deletion mutant is identical to those which have been reported in HD and NPC. Its association with evolution of AILD into B-IBL, aggressive HD and NPC, suggests that this particular mutant is more widespread than originally thought and is clinically relevant.

Adult↗
PubMed1994

Persistence of the same viral strain in early and late relapses of Epstein-Barr virus-associated Hodgkin's disease.

Twelve cases of relapsing Hodgkin's disease were investigated for the presence of Epstein-Barr virus (EBV). Of these, 7 cases contained EBV gene products (LMP1, EBER RNA) in the diagnostic Reed-Sternberg cells and variants at first presentation and at relapse(s), whereas 5 cases were negative at both first diagnosis and relapse. Among the 7 EBV-positive cases, material for DNA extraction was available in 2 cases at both diagnosis and relapse(s). Ig and T-cell receptor gene rearrangements displayed a germline configuration in the 2 cases. However, Southern blot analysis of the terminal repeats (TR) of EBV genome showed that, in 1 of the 2 cases, the fragment was of the same size at diagnosis and in the subsequent two relapses (1 early and 1 late). The second case contained monoclonal EBV genome at diagnosis, but the Southern analysis of the TR was negative at relapse. The latent membrane protein (LMP1) sequence analysis confirmed the persistence of a distinctive viral strain in each of the 2 cases with individual abnormalities within the carboxy terminal region (5 point mutations and a 30-bp deletion for the first case and 6 point mutations for the second case). The persistence of a given strain in early and late relapses is evidence towards the view that in Hodgkin's disease such relapses are related to a single residual tumor cell clone.

Adult↗
PubMed1994

Expression of human recombination activating genes (RAG-1 and RAG-2) in lymphoma.

Two recently discovered genes, the recombination activating genes 1 and 2 (RAG-1 and RAG-2), are necessary to perform variable (V), diversity (D), and joining (J) recombination. They synergistically activate VDJ recombination to generate immunocompetent lymphocytes. Disruption of either gene results in a maturation arrest at a very early B and T cell progenitor stage. Expression and downregulation of RAG's are closely associated with interleukin 7, sIgM and TCR-CD3 complex, respectively. Assessment of RAG mRNA expression is a valuable marker in identifying the genotypic maturation status of leukemias and lymphomas. Persistent RAG expression in otherwise mature lymphoid proliferations may explain puzzling biological and clinical observations such as multiple rearrangements in lymphomas with a mature phenotype. Lack of RAG expression in Hodgkin's disease with abundant Reed-Sternberg cells is consistent with a mature phenotype of the latter. Availability of a anti-RAG-1 monoclonal antibody in the near future will facilitate RAG analysis of lymphomas.

Animals↗
PubMed1994

[Internal deletions of the latent membrane protein oncogenes of Epstein-Barr virus in Hodgkin's disease are almost identical with those of Asiatic nasopharyngeal carcinoma].

The latent membrane protein (LMP) oncogene of the Epstein-Barr virus (EBV) is 1300 base pairs (bp) long and expressed in Hodgkin and Reed-Sternberg (HRS) cells of about 50% of Hodgkin's lymphomas and in tumor cells of about 60% of nasopharyngeal carcinomas (NPC). The LMP sequences of EBV variants isolated from two NPC (NPC 1510 and NPC CAO) have recently been published. Compared to the standard EBV sequence (EBV B95-8) they both show deletions of 30 bp near the 3' end. These mutations render the LMP oncogene more aggressive in NPC. In 52 Hodgkin's lymphomas expressing the LMP oncogene was amplified the coding sequence by the polymerase chain reaction. In five tumors deletions within the coding region for the intracytoplasmic LMP domain have been found. DNA sequencing revealed three 30 bp deletions almost identical to those observed in NPC 1510 and NPC CAO. In a forth case, a 70 bp deletion encompassing the regions of the 30 bp deletions was found. Histologically, all five tumors with such deletions showed abundant HRS cells. It is likely that these mutations of the LMP oncogene in a similar way as in NPC also favour the proliferation of HRS cells in Hodgkin's disease.

Amino Acid Sequence↗