Mantle cell lymphoma (MCL) displays an especially poor clinical outcome with only transient response to conventional chemotherapy, continuous relapses and a median survival of only 3 - 4 years. The Ubiquitin-proteasome pathway is known to... more
Mantle cell lymphoma (MCL) displays an especially poor clinical outcome with only transient response to conventional chemotherapy, continuous relapses and a median survival of only 3 - 4 years. The Ubiquitin-proteasome pathway is known to alter homeostasis of various oncogenes, transcription factors (e.g. NF-κB) regulators of cell cycle progression and apoptosis. In various phase II trials, proteasome inhibition with single agent bortezomib (Velcade®) achieved response rates of up to 60% in relapsed disease. Five MCL cell lines (HBL2, Granta 519, Jeko-1, NCEB-1 and Rec-1) and two hematological control cell lines (Jurkat, Karpas 422) were treated with bortezomib at a previously defined effective concentration (25 nM). Real-time RT-PCR and Western Blot analysis of cyclin D1 (CCND1), Cdk inhibitors (INK4s, KIPs), and other regulators of cell cycle and apoptosis were performed at various time points during bortezomib treatment (0 to 12 hours). Additionally, analyses of cell cycle were performed by flow cytometry. All cells lines were also exposed to different doses of bortezomib in combination with various cytostatic drugs, cell proliferation (WST-1 assay) and apoptosis (Annexin V PE/7-AAD staining) were analysed. After only 2 - 4 hours of bortezomib treatment analysis of relative RNA expression levels revealed downregulation of Cdk4 inhibitor p21CIP1, CCND1 and BCL2, thus representing early effects of proteasome inhibition in MCL cell lines. In contrast, CCND1 expression temporarily increased in cell lines with moderate sensitivity to bortezomib. Interaction between bortezomib and cytostatic drugs were evaluated applying the combination index (CI). Simultaneous exposure to bortezomib and various cytostatic drugs (cytarabine, mitoxantrone, fludarabine and gemcitabine) resulted in a significantly enhanced inhibition of proliferation and apoptosis. In order to investigate the interaction of cytostatic agents and bortezomib different incubation schedules were evelusated. Notably, pre-exposure to cytarabine and subsequent proteasome inhibition results in a strong synergism (CI = 0.5) that was not observed with prior bortezomib incubation. Interesting this pattern is chemotherapy specific, since bortezomib pre-incubation induced synergism with mitoxantrone (CI = 0.6). Finally, combination of cytarabine or gemcitabine with bortezomib was able to sensitize and inhibit cell proliferation of MCL cell lines resistant to single agent treatment. In summary, alteration of protein expression profiles and cell cycle regulators occur early after proteasome inhibition in MCL cell lines. In addition, combination of bortezomib with distinct cytostatic agents demonstrated a synergistic schedule dependent efficacy in vitro. Currently, confirmatory analyses of primary patient samples are being performed representing the rationale of a future randomized phase II/III study of the European MCL Network.
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Follicular lymphoma is a clinically and genetically heterogeneous disease, but the prognostic value of somatic mutations has not been systematically assessed. We aimed to improve risk stratification of patients receiving first-line... more
Follicular lymphoma is a clinically and genetically heterogeneous disease, but the prognostic value of somatic mutations has not been systematically assessed. We aimed to improve risk stratification of patients receiving first-line immunochemotherapy by integrating gene mutations into a prognostic model. We did DNA deep sequencing to retrospectively analyse the mutation status of 74 genes in 151 follicular lymphoma biopsy specimens that were obtained from patients within 1 year before beginning immunochemotherapy consisting of rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP). These patients were recruited between May 4, 2000, and Oct 20, 2010, as part of a phase 3 trial (GLSG2000). Eligible patients had symptomatic, advanced stage follicular lymphoma and were previously untreated. The primary endpoints were failure-free survival (defined as less than a partial remission at the end of induction, relapse, progression, or death) and overall survival calcul...
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Research Interests: Physics and Superfluidity
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Research Interests: Physics, Physical, Crust, Shell Structure, Proton, and 3 moreAstrophysique, Pairing, and neutron star
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Research Interests: Hematology, Oncology, Health Care, Health Care Policy, Medicine, and 15 moreMultidisciplinary, Cancer treatment, Multivariate Analysis, Biological Sciences, Acute Myeloid Leukemia, Humans, Female, Male, Middle Aged, Leukemias, Health Risk Analysis, Logistic Models, leukocytosis, leukocyte Count, and Cytogenetic analysis
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Hürthle thyroid tumors are characterized by frequent numerical chromosomal aberrations, including aneuploidy or polyploidy, losses and gains of some chromosomal regions and DNA fragmentation. In recent years, great attention has been paid... more
Hürthle thyroid tumors are characterized by frequent numerical chromosomal aberrations, including aneuploidy or polyploidy, losses and gains of some chromosomal regions and DNA fragmentation. In recent years, great attention has been paid to the combined analysis of morphologic and genetic features of oxyphilic tumors and to the elucidation of their pathogenesis. We analyzed for loss of heterozygosity (LOH) of the candidate regions for TCO (thyroid tumor with cell oxyphilia) and NMTC1 (nonmedullary thyroid carcinoma 1), 2 loci already mapped on chromosomes 19p13.2 and 2q21, respectively. Matched normal and tumor DNA samples from 70 patients with sporadic oxyphilic thyroid tumors and 20 with sporadic follicular tumors were subjected to microsatellite analysis using 10 markers on 19p13.2 and 6 markers on 2q21. This approach led us to the observation of a more significant LOH in oxyphilic than in follicular tumors. Allelic loss in tumor samples was evenly distributed in both 19p13.2 and 2q21 regions, in accordance with the established linkage of TCO and NMTC1 for inherited tumors. In order to investigate the possible contribution of both susceptibility loci in oxyphilic tumors, the family that led to the original mapping of TCO locus was reanalyzed for the markers in the 2q21 region. This led to the exclusion of linkage with the NMTC1 locus and to the refutation of the digenic inheritance hypothesis at least in this family.
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Although gene expression following bortezomib treatment has been previously explored, direct effects of bortezomib‐induced proteasome inhibition on protein level has not been analyzed so far. Using 2‐D PAGE in five mantle cell lymphoma... more
Although gene expression following bortezomib treatment has been previously explored, direct effects of bortezomib‐induced proteasome inhibition on protein level has not been analyzed so far. Using 2‐D PAGE in five mantle cell lymphoma cell lines, we screened for cellular protein level alterations following treatment with 25 nM bortezomib for up to 4 h. Using MS, we identified 38 of the 41 most prominent reliably detected protein spots. Twenty‐one were affected in all cell lines, whereas the remaining 20 protein spots were exclusively altered in sensitive cell lines. Western blot analysis was performed for 17 of the 38 identified proteins and 70.6% of the observed protein level alterations in 2‐D gels was verified. All cell lines exhibited alterations of the cellular protein levels of heat shock‐induced protein species (HSPA9, HSP7C, HSPA5, HSPD1), whereas sensitive cell lines also displayed altered cellular protein levels of energy metabolism (ATP5B, AK5, TPI1, ENO‐1, ALDOC, GAPDH)...