Biomedical subjects

Adrian Wiestner

Publications and source records attributed to Adrian Wiestner.

15 recordsLinked to original sources

PubMed2006

Schedule-dependent synergy between the heat shock protein 90 inhibitor 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin and doxorubicin restores apoptosis to p53-mutant lymphoma cell lines.

PURPOSE: Loss of p53 function impairs apoptosis induced by DNA-damaging agents used for cancer therapy. Here, we examined the effect of the heat shock protein 90 (HSP90) inhibitor 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (DMAG) on doxorubicin-induced apoptosis in lymphoma. We aimed to establish the optimal schedule for administration of both drugs in combination and the molecular basis for their interaction. EXPERIMENTAL DESIGN: Isogenic lymphoblastoid and nonisogenic lymphoma cell lines differing in p53 status were exposed to each drug or combination. Drug effects were examined using Annexin V, active caspase-3, cell cycle, and cytotoxicity assays. Synergy was evaluated by median effect/combination index. Protein expression and kinase inhibition provided insight into the molecular mechanisms of drug interaction. RESULTS: Presence of mutant p53 conferred increased survival to single agents. Nevertheless, DMAG showed synergistic toxicity with doxorubicin independently of p53 status. Synergy required exposure to doxorubicin before DMAG. DMAG-mediated down-regulation of CHK1, a known HSP90 client, forced doxorubicin-treated cells into premature mitosis followed by apoptosis. A CHK1 inhibitor, SB-218078, reproduced the effect of DMAG. Administration of DMAG before doxorubicin resulted in G1-S arrest and protection from apoptosis, leading to additive or antagonistic interactions that were exacerbated by p53 mutation. CONCLUSIONS: Administration of DMAG to doxorubicin-primed cells induced premature mitosis and had a synergistic effect on apoptosis regardless of p53 status. These observations provide a rationale for prospective clinical trials and stress the need to consider schedule of exposure as a critical determinant of the overall response when DMAG is combined with chemotherapeutic agents for the treatment of patients with relapsed/refractory disease.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed2006

Flow cytometry for ZAP-70: New colors for chronic lymphocytic leukemia.

ZAP-70 has become one of the most studied prognostic markers in Chronic Lymphocytic Leukemia (CLL). ZAP-70 is remarkable in many ways: ZAP-70 has been identified as the best discriminating gene between prognostically distinct CLL subtypes using large scale gene expression profiling; ZAP-70 has been shown to enhance signal transduction in CLL B-cells and therefore could contribute to disease progression; and ZAP-70 is one of the rare examples of an intracellular target considered for clinical flow cytometry. This issue attests to the enormous effort and the steady progress made in overcoming technical challenges of testing for ZAP-70 expression and sets the foundation for a successful translation of this important marker into clinical practice. Despite the best effort, one will likely have to accept that not all cases can be clearly assigned to one or the other group, given that ZAP-70 expression between CLL patients falls along a continuum from absent to high. Nevertheless, ZAP-70 expression could become a key parameter to guide patients towards risk adapted treatment strategies in prospective clinical trials.

Biomarkers, Tumor↗

PubMed2005

Four human t(11;14)(q13;q32)-containing cell lines having classic and variant features of Mantle Cell Lymphoma.

The objectives of this study were foremost to further characterize pre-existing cell lines containing the t(11;14)(q13;q32) translocation. This translocation along with cyclin D1 overexpression is characteristic of Mantle Cell Lymphoma (MCL), an aggressive B cell neoplasm. Considerable variation in the abundance of cyclin D1 expression was observed. mRNA levels were examined by RT-PCR as differences in cyclin D1 mRNA abundance have been shown to synergize with INK4A/Arf deletions to dictate proliferation rate and survival in MCL patient samples. In this study, the cell lines, Z-138 and HBL-2, which exhibited the fastest growth rates and the shortest survival times in Rag2-M mice, had high expression of either one or both cyclin D1 mRNA isoforms and had negligible expression of p16. On the other hand, NCEB-1 and JVM-2 had low expression of both mRNA isoforms, retained p16 expression, and had slower growth rates and exhibited longer survival times in Rag2-M mice. Furthermore, JVM-2, which was found to have the lowest expression of cyclin D1, was the only cell line that expressed cyclin D2. The results of the characterization of Z-138, HBL-2, NCEB-1 and JVM-2 reveal that this group of cell lines represents both classic and variant features of MCL.

Animals↗

PubMed2005

More ZAP for chronic lymphocytic leukemia (CLL).

Antigen stimulation of the leukemic clone is increasingly implicated in the pathogenesis of CLL. Chen and colleagues provide evidence that expression of ZAP-70 in CLL B cells renders IgM signaling more effective and thereby could contribute to the more rapidly progressive clinical course of ZAP-70-positive CLL.

Animals↗

PubMed2004

Imatinib inhibits T-cell receptor-mediated T-cell proliferation and activation in a dose-dependent manner.

The tyrosine kinase inhibitor imatinib (imatinib, STI571, Glivec, and Gleevec) is increasingly used in patients undergoing allogeneic transplantation for leukemia. However, little is known regarding its potential immunoregulatory effects. Here, we investigate the effect of imatinib on T-cell receptor (TCR)-mediated activation of human T cells. Following stimulation with the anti-CD3 antibody 12F6, proliferation of activated T cells was almost completely inhibited by 10 microM imatinib. Furthermore, antigen-triggered expansion of CD8+ T cells in response to immunodominant cytomegalovirus (CMV) and Epstein-Barr virus (EBV) peptides was significantly reduced. Up-regulation of the activation markers CD25 and CD69 in response to TCR cross-linking was suppressed by imatinib at a mean inhibitory concentration 50% (IC50) of 5.4 microM and 7.3 microM, respectively; interleukin 2 (IL-2) production was also impaired. Analysis of the TCR-induced signaling cascade showed that imatinib substantially reduced tyrosine phosphorylation of ZAP70 and LAT in response to activation through the TCR. Sequence comparisons of all 90 tyrosine kinase genes in the human genome for homology in the adenosine triphosphate (ATP) binding pocket identified LCK, which is required for ZAP70 activation, as a likely target for imatinib. The IC50 for LCK inhibition by imatinib was 0.6 microM to 0.8 microM in an in vitro tyrosine kinase assay. In summary, imatinib can interfere with T-cell activation in vitro, and its impact on the frequency of opportunistic infections and graft-versus-host or graft-versus-leukemia reactions after transplantation should be investigated in clinical trials.

Adaptor Proteins, Signal Transducing↗

PubMed2004

Fludarabine treatment of patients with chronic lymphocytic leukemia induces a p53-dependent gene expression response.

Fludarabine, the current standard treatment for B-cell chronic lymphocytic leukemia (CLL), can induce apoptosis in CLL cells in vitro, and a number of molecular mechanisms contribute to its cytotoxicity. Using gene expression profiling, we investigated the molecular consequences of fludarabine treatment of patients with CLL in vivo. In 7 patients with CLL, a consistent gene expression signature of in vivo fludarabine exposure was identified. Many of the fludarabine signature genes were known p53 target genes and genes involved in DNA repair. In vitro treatment of CLL cells with fludarabine induced the same set of genes as observed in vivo, and many of these genes were also induced by in vitro exposure of CLL cells to ionizing radiation. Using isogenic p53 wild-type and null lymphoblastoid cell lines, we confirmed that many of the fludarabine signature genes were also p53 target genes. Because in vivo treatment with fludarabine induces a p53-dependent gene expression response, fludarabine treatment has the potential to select p53-mutant CLL cells, which are more drug resistant and associated with an aggressive clinical course. These considerations suggest that fludarabine treatment should be given in strict accordance to the current National Cancer Institute (NCI) guidelines that have established criteria of disease activity that warrant treatment.

Adult↗

PubMed2004

Autoreactive, cytotoxic T lymphocytes specific for peptides derived from normal B-cell differentiation antigens in healthy individuals and patients with B-cell malignancies.

PURPOSE: To investigate potential immunotherapeutic strategies in B lymphocytic malignancies we looked for CTLs recognizing CD19 and CD20 epitopes. EXPERIMENTAL DESIGN: Three CD19 and CD20 peptides binding to HLA-A*0201 were identified and used to detect peptide specific CTLs by a quantitative real-time PCR to measure IFN-gamma mRNA expression in 23 healthy individuals and 28 patients (18 chronic lymphocytic leukemia (CLL), 7 follicular lymphoma, 2 acute lymphocytic leukemia, and 1 large cell lymphoma). Peptide-specific CTLs were expanded in culture with CD40-activated B cells to test lytic activity in three patients. RESULTS: In healthy individuals, CD8+ T-cell responses were detected in one to CD19(74-82), in three to CD20(127-135), and three to CD20(188-196). Seven of 27 patients (6 with CLL) had CD8+ T cells recognizing CD19(74-82). Seven patients responded to CD20(127-135) and three to CD20(188-196). All were CLL patients. CD19(74-82)-specific CTLs from three patients were expanded over 4 weeks. These cells were HLA-A*0201 specific and lytic for peptide-loaded antigen-presenting cells but not to malignant or unpulsed B cells. CONCLUSIONS: CTLs that recognize CD19 and CD20 epitopes exist in healthy individuals and may be increased in CLL patients. They are of low avidity and require high doses of peptide for activation. Strategies to increase T-cell avidity would be necessary for T-cell immunotherapeutic approaches using the peptides studied.

Adult↗

PubMed2004

ZAP-70 expression and prognosis in chronic lymphocytic leukaemia.

BACKGROUND: Chronic lymphocytic leukaemia (CLL) is a heterogeneous disease; many patients never need treatment, whereas some have poor outcomes. New treatments, which can induce complete remissions, allow patients with poor outlook to be treated while they are still asymptomatic. Whether or not the IgVH gene is mutated is the best predictor of clinical outcome, but this assay is unsuited to the routine laboratory. The gene coding for ZAP-70, a tyrosine kinase protein normally expressed in T and NK cells, has been shown by gene-expression profiling to be differentially expressed between patients with mutated and unmutated IgVH genes. We assessed whether ZAP-70 could be used as a prognostic marker in CLL. METHODS: We developed a flow cytometry assay for ZAP-70 protein expression and investigated its concordance with ZAP-70 mRNA expression, IgVH gene mutational status, and clinical outcome in 167 patients with CLL. FINDINGS: We showed high concordance between ZAP-70 protein expression and IgVH gene mutations. 108 patients (65%) had mutated IgVH genes and were ZAP-70 negative; 46 (28%) had unmutated IgVH genes and were ZAP-70 positive. Findings were discordant in 13 patients: six (4%) had mutated IgVH genes but were ZAP-70 positive, and seven (4%) had unmutated IgVH genes and were ZAP-70 negative. Expression of mRNA showed 97% concordance with ZAP-70 protein. Median survival was 24.4 years (95% CI 15.1-33.8) in ZAP-70 negative patients and 9.3 years (7.0-11.5) in those who were ZAP-70 positive (hazard ratio 5.5, 2.8-.8). INTERPRETATION: ZAP-70 protein, which can be measured by flow cytometry in the general laboratory, is a reliable prognostic marker in CLL, equivalent to that of IgVH gene mutational status.

Aged↗

PubMed2004

Overexpression of c-maf is a frequent oncogenic event in multiple myeloma that promotes proliferation and pathological interactions with bone marrow stroma.

The oncogene c-maf is translocated in approximately 5%-10% of multiple myelomas. Unexpectedly, we observed c-maf expression in myeloma cell lines lacking c-maf translocations and in 50% of multiple myeloma bone marrow samples. By gene expression profiling, we identified three c-maf target genes: cyclin D2, integrin beta7, and CCR1. c-maf transactivated the cyclin D2 promoter and enhanced myeloma proliferation, whereas dominant inhibition of c-maf blocked tumor formation in immunodeficient mice. c-maf-driven expression of integrin beta7 enhanced myeloma adhesion to bone marrow stroma and increased production of VEGF. We propose that c-maf transforms plasma cells by stimulating cell cycle progression and by altering bone marrow stromal interactions. The frequent overexpression of c-maf in myeloma makes it an attractive target for therapeutic intervention.

Animals↗

PubMed2003

Late presentation of dyskeratosis congenita as apparently acquired aplastic anaemia due to mutations in telomerase RNA.

Aplastic anaemia in adults is usually acquired, but rarely constitutional types of bone marrow failure can occur late in life. We assessed two families with onset of pancytopenia in adults and detected two novel point mutations in the telomerase RNA gene (TERC) in each family. This gene is abnormal in some kindreds with dyskeratosis congenita. Individuals in our families with mutated TERC did not have physical signs of dyskeratosis congenita, and their blood counts were nearly normal, but all had severely shortened telomeres, reduced haemopoietic function, and raised serum erythropoietin and thrombopoietin. Bone marrow failure of variable severity due to dyskeratosis congenita, historically characterised by associated physical anomalies and early pancytopenia, may be present in otherwise phenotypically normal adults, and can masquerade as acquired aplastic anaemia.

Adult↗

PubMed2003

A gene expression-based method to diagnose clinically distinct subgroups of diffuse large B cell lymphoma.

To classify cancer specimens by their gene expression profiles, we created a statistical method based on Bayes' rule that estimates the probability of membership in one of two cancer subgroups. We used this method to classify diffuse large B cell lymphoma (DLBCL) biopsy samples into two gene expression subgroups based on data obtained from spotted cDNA microarrays. The germinal center B cell-like (GCB) DLBCL subgroup expressed genes characteristic of normal germinal center B cells whereas the activated B cell-like (ABC) DLBCL subgroup expressed a subset of the genes that are characteristic of plasma cells, particularly those encoding endoplasmic reticulum and golgi proteins involved in secretion. We next used this predictor to discover these subgroups within a second set of DLBCL biopsies that had been profiled by using oligonucleotide microarrays [Shipp, M. A., et al. (2002) Nat. Med. 8, 68-74]. The GCB and ABC DLBCL subgroups identified in this data set had significantly different 5-yr survival rates after multiagent chemotherapy (62% vs. 26%; P < or = 0.0051), in accord with analyses of other DLBCL cohorts. These results demonstrate the ability of this gene expression-based predictor to classify DLBCLs into biologically and clinically distinct subgroups irrespective of the method used to measure gene expression.

B-Lymphocytes↗

PubMed2003

ZAP-70 expression identifies a chronic lymphocytic leukemia subtype with unmutated immunoglobulin genes, inferior clinical outcome, and distinct gene expression profile.

The presence or absence of somatic mutations in the expressed immunoglobulin heavy chain variable regions (IgVH) of chronic lymphocytic leukemia (CLL) cells provides prognostic information. Patients whose leukemic cells express unmutated IgVH regions (Ig-unmutated CLL) often have progressive disease, whereas patients whose leukemic cells express mutated IgVH regions (Ig-mutated CLL) more often have an indolent disease. Given the difficulty in performing IgVH sequencing in a routine diagnostic laboratory, this prognostic distinction is currently unavailable to most patients. Pilot gene expression profiling studies in patients with CLL identified genes that were differentially expressed between the Ig-unmutated and Ig-mutated CLL subtypes. Here, we have profiled an expanded cohort of 107 patients and show that ZAP-70 is the gene that best distinguishes the CLL subtypes. Ig-unmutated CLL expressed ZAP-70 5.54-fold more highly than Ig-mutated CLL (P < 10(-21)). ZAP-70 expression correctly predicted IgVH mutation status in 93% of patients. ZAP-70 expression and IgVH mutation status were comparable in their ability to predict time to treatment requirement following diagnosis. In 7 patients, ZAP-70 expression and IgVH mutation status were discordant: 4 Ig-mutated CLLs had high ZAP-70 expression and 3 Ig-unmutated CLLs had low ZAP-70 expression. Among these ZAP-70 "outliers," those with Ig-mutated CLL had clinical features that are uncharacteristic of this CLL subtype: 2 required early treatment and 2 used a mutated VH3-21 gene, an IgVH gene that has been associated with progressive disease. We developed reverse transcriptase-polymerase chain reaction and immunohistochemical assays for ZAP-70 expression that can be applied clinically and would yield important prognostic information for patients with CLL.

ADP-ribosyl Cyclase↗

PubMed2003

Towards molecular diagnosis and targeted therapy of lymphoid malignancies.

Gene expression profiling of cancer began as a research tool but is rapidly moving towards clinical application. The diagnostic category of diffuse large B-cell lymphoma (DLBCL) can now be viewed as an amalgam of several different diseases that have distinct gene expression profiles, oncogenic mechanisms, and clinical outcomes. Other diagnostic categories such as chronic lymphocytic leukemia (CLL) have a single gene expression signature that distinguishes them from other lymphoid malignancies. Nevertheless, elevated expression of a single gene, ZAP-70, is characteristic of a more aggressive subtype of CLL that may require novel treatment approaches. In mantle cell lymphoma (MCL), a quantitative measurement of gene expression associated with tumor proliferation is a powerful predictor of survival. Ultimately, the molecular diagnosis of these malignancies will identify molecular pathways that can be exploited for therapy. For example, one of the gene expression subgroups of DLBCL, termed activated B-cell like DLBCL, is characterized by constitutive activation of the nuclear factor-kappaB (NF-kappaB) signaling pathway, and interference with this pathway selectively kills these lymphoma cells. The full benefit of gene expression profiling can only be realized if we incorporate this technology into prospective clinical trials.

Drug Delivery Systems↗

PubMed2003

The proliferation gene expression signature is a quantitative integrator of oncogenic events that predicts survival in mantle cell lymphoma.

We used gene expression profiling to establish a molecular diagnosis of mantle cell lymphoma (MCL), to elucidate its pathogenesis, and to predict the length of survival of these patients. An MCL gene expression signature defined a large subset of MCLs that expressed cyclin D1 and a novel subset that lacked cyclin D1 expression. A precise measurement of tumor cell proliferation, provided by the expression of proliferation signature genes, identified patient subsets that differed by more than 5 years in median survival. Differences in cyclin D1 mRNA abundance synergized with INK4a/ARF locus deletions to dictate tumor proliferation rate and survival. We propose a quantitative model of the aberrant cell cycle regulation in MCL that provides a rationale for the design of cell cycle inhibitor therapy in this malignancy.

ADP-Ribosylation Factors↗

PubMed2002

Rituximab in the treatment of acquired factor VIII inhibitors.

Autoantibodies against factor VIII (FVIII) are rare but can cause life-threatening bleeding requiring costly factor replacement and prolonged immunosuppression. We report 4 consecutively treated patients whose acquired FVIII inhibitors responded rapidly to immunosuppressive regimens that included rituximab, a monoclonal antibody against CD20(+) B cells. Three patients had spontaneously occurring inhibitors. The fourth, a patient with mild hemophilia A, developed both an autoantibody and an alloantibody following recombinant FVIII treatment. Pretreatment FVIII activities ranged from less than 1% to 4% and inhibitor titers from 5 to 60 Bethesda units (BU). One patient with polymyalgia rheumatica who developed the inhibitor while receiving prednisone responded to single agent rituximab. The hemophilia patient had rapid resolution of the autoantibody, whereas the alloantibody persisted for months. Responses continue off treatment from more than 7 to more than 12 months. This report adds to the growing evidence that rituximab has efficacy in immune disorders resulting from autoantibody formation.

Adult↗