Publications by authors named "Christian Sillaber"

52 Publications

Attenuation of canonical NF-κB signaling maintains function and stability of human Treg.

FEBS J 2021 Jan 28;288(2):640-662. Epub 2020 May 28.

Department of Laboratory Medicine, Medical University of Vienna, Vienna, Austria.

Nuclear factor 'κ-light-chain-enhancer' of activated B cells (NF-κB) signaling is a signaling pathway used by most immune cells to promote immunostimulatory functions. Recent studies have indicated that regulatory T cells (Treg) differentially integrate TCR-derived signals, thereby maintaining their suppressive features. However, the role of NF-κB signaling in the activation of human peripheral blood (PB) Treg has not been fully elucidated so far. We show that the activity of the master transcription factor forkhead box protein 3 (FOXP3) attenuates p65 phosphorylation and nuclear translocation of the NF-κB proteins p50, p65, and c-Rel following activation in human Treg. Using pharmacological and genetic inhibition of canonical NF-κB signaling in FOXP3-transgenic T cells and PB Treg from healthy donors as well as Treg from a patient with a primary NFKB1 haploinsufficiency, we validate that Treg activation and suppressive capacity is independent of NF-κB signaling. Additionally, repression of residual NF-κB signaling in Treg further enhances interleukin-10 (IL-10) production. Blockade of NF-κB signaling can be exploited for the generation of in vitro induced Treg (iTreg) with enhanced suppressive capacity and functional stability. In this respect, dual blockade of mammalian target of rapamycin (mTOR) and NF-κB signaling was accompanied by enhanced expression of the transcription factors FOXP1 and FOXP3 and demethylation of the Treg-specific demethylated region compared to iTreg generated under mTOR blockade alone. Thus, we provide first insights into the role of NF-κB signaling in human Treg. These findings could lead to strategies for the selective manipulation of Treg and the generation of improved iTreg for cellular therapy.
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http://dx.doi.org/10.1111/febs.15361DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7891634PMC
January 2021

Inhibition of complement C1s in patients with cold agglutinin disease: lessons learned from a named patient program.

Blood Adv 2020 03;4(6):997-1005

Division of Hematology, Department of Internal Medicine I, Medical University of Vienna, Vienna, Austria.

Cold agglutinin disease (CAD) causes predominantly extravascular hemolysis and anemia via complement activation. Sutimlimab is a novel humanized monoclonal antibody directed against classical pathway complement factor C1s. We aimed to evaluate the safety and efficacy of long-term maintenance treatment with sutimlimab in patients with CAD. Seven CAD patients treated with sutimlimab as part of a phase 1B study were transitioned to a named patient program. After a loading dose, patients received biweekly (once every 2 weeks) infusions of sutimlimab at various doses. When a patient's laboratory data showed signs of breakthrough hemolysis, the dose of sutimlimab was increased. Three patients started with a dose of 45 mg/kg, another 3 with 60 mg/kg, and 1 with a fixed dose of 5.5 g every other week. All CAD patients responded to re-treatment, and sutimlimab increased hemoglobin from a median initial level of 7.7 g/dL to a median peak of 12.5 g/dL (P = .016). Patients maintained near normal hemoglobin levels except for a few breakthrough events that were related to underdosing and which resolved after the appropriate dose increase. Four of the patients included were eventually treated with a biweekly 5.5 g fixed-dose regimen of sutimlimab. None of them had any breakthrough hemolysis. All patients remained transfusion free while receiving sutimlimab. There were no treatment-related serious adverse events. Overlapping treatment with erythropoietin, rituximab, or ibrutinib in individual patients was safe and did not cause untoward drug interactions. Long-term maintenance treatment with sutimlimab was safe, effectively inhibited hemolysis, and significantly increased hemoglobin levels in re-exposed, previously transfusion-dependent CAD patients.
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http://dx.doi.org/10.1182/bloodadvances.2019001321DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7094024PMC
March 2020

Treatment with brentuximab vedotin plus bendamustine in unselected patients with CD30-positive aggressive lymphomas.

Eur J Haematol 2020 Mar 10;104(3):251-258. Epub 2020 Jan 10.

Division of Hematology and Hemostaseology, Department of Medicine I, Medical University of Vienna, Vienna, Austria.

Objectives: A treatment regimen consisting of bendamustine and brentuximab vedotin (BV) has been described as a highly potent salvage therapy and as an effective induction therapy leading to high response rates before autologous stem cell transplantation (ASCT) in patients with classical Hodgkin lymphoma (cHL). In this retrospective analysis, we aimed to assess this therapy's efficacy in unselected patients with cHL and CD30+ peripheral T-cell lymphoma (PTCL).

Patients And Methods: Data of 28 patients with cHL and five patients with PTCL treated with a combination of bendamustine and BV at three Austrian tertiary cancer centers were analyzed.

Results: In patients with cHL, the ORR was 100% (78.6% CR, 21.4% PR). After 17 months median follow-up, median survival times were not reached; 1-year PFS was 81.9%, and 1-year OS was 95.7%. Thirteen eligible patients (46.4%) successfully underwent planned ASCT after salvage therapy with bendamustine and BV and subsequent high-dose chemotherapy. Three of the five PTCL patients achieved CR, while two did not respond and died during or shortly after therapy.

Conclusion: A combination of bendamustine and BV is an effective salvage and induction therapy before ASCT in patients with relapsed/refractory cHL. Further research is warranted to evaluate the use in patients with PTCL.
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http://dx.doi.org/10.1111/ejh.13368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7065019PMC
March 2020

Nivolumab for Newly Diagnosed Advanced-Stage Classic Hodgkin Lymphoma: Safety and Efficacy in the Phase II CheckMate 205 Study.

J Clin Oncol 2019 08 21;37(23):1997-2007. Epub 2019 May 21.

17Mayo Clinic, Rochester, MN.

Purpose: Nivolumab, an anti-programmed death-1 monoclonal antibody, has demonstrated frequent and durable responses in relapsed/refractory classic Hodgkin lymphoma (cHL). We report results from Cohort D of the CheckMate 205 trial, which assessed nivolumab monotherapy followed by nivolumab plus doxorubicin, vinblastine, and dacarbazine (N-AVD) for newly diagnosed cHL.

Methods: Patients 18 years of age or older with untreated, advanced-stage (defined as III to IV and IIB with unfavorable risk factors) cHL were eligible for Cohort D of this multicenter, noncomparative, phase II trial. Patients received nivolumab monotherapy for four doses, followed by 12 doses of N-AVD; all doses were every 2 weeks, and nivolumab was administered at 240 mg intravenously. The primary end point was safety. Efficacy end points included objective response rate and modified progression-free survival, defined as time to disease progression/relapse, death, or next therapy. Chromosome 9p24.1 alterations and programmed death-ligand 1 expression were assessed in Hodgkin Reed-Sternberg cells in evaluable patients.

Results: A total of 51 patients were enrolled and treated. At diagnosis, 49% of patients had an International Prognostic Score of 3 or greater. Overall, 59% experienced a grade 3 to 4 treatment-related adverse event. Treatment-related febrile neutropenia was reported in 10% of patients. Endocrine immune-mediated adverse events were all grade 1 to 2 and did not require high-dose corticosteroids; all nonendocrine immune-mediated adverse events resolved (most commonly, rash; 5.9%). At the end of therapy, the objective response rate (95% CI) per independent radiology review committee was 84% (71% to 93%), with 67% (52% to 79%), achieving complete remission (five patients [10%] were nonevaluable and counted as nonresponders). With a minimum follow-up of 9.4 months, 9-month modified progression-free survival was 92%. Patients with higher-level Hodgkin Reed-Sternberg programmed death-ligand 1 expression had more favorable responses to N-AVD ( = .041).

Conclusion: Nivolumab followed by N-AVD was associated with promising efficacy and safety profiles for newly diagnosed, advanced-stage cHL.
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http://dx.doi.org/10.1200/JCO.19.00315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6688776PMC
August 2019

Inhibition of complement C1s improves severe hemolytic anemia in cold agglutinin disease: a first-in-human trial.

Blood 2019 02 17;133(9):893-901. Epub 2018 Dec 17.

Department of Clinical Pharmacology and.

Cold agglutinin disease is a difficult-to-treat autoimmune hemolytic anemia in which immunoglobulin M antibodies bind to erythrocytes and fix complement, resulting in predominantly extravascular hemolysis. This trial tested the hypothesis that the anti-C1s antibody sutimlimab would ameliorate hemolytic anemia. Ten patients with cold agglutinin disease participated in the phase 1b component of a first-in-human trial. Patients received a test dose of 10-mg/kg sutimlimab followed by a full dose of 60 mg/kg 1 to 4 days later and 3 additional weekly doses of 60 mg/kg. All infusions were well tolerated without premedication. No drug-related serious adverse events were observed. Seven of 10 patients with cold agglutinin disease responded with a hemoglobin increase >2 g/dL. Sutimlimab rapidly increased hemoglobin levels by a median of 1.6 g/dL within the first week, and by a median of 3.9 g/dL (interquartile range, 1.3-4.5 g/dL; 95% confidence interval, 2.1-4.5) within 6 weeks ( = .005). Sutimlimab rapidly abrogated extravascular hemolysis, normalizing bilirubin levels within 24 hours in most patients and normalizing haptoglobin levels in 4 patients within 1 week. Hemolytic anemia recurred when drug levels were cleared from the circulation 3 to 4 weeks after the last dose of sutimlimab. Reexposure to sutimlimab in a named patient program recapitulated the control of hemolytic anemia. All 6 previously transfused patients became transfusion-free during treatment. Sutimlimab was safe, well tolerated, and rapidly stopped C1s complement-mediated hemolysis in patients with cold agglutinin disease, significantly increasing hemoglobin levels and precluding the need for transfusions. This trial was registered at www.clinicaltrials.gov as #NCT02502903.
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http://dx.doi.org/10.1182/blood-2018-06-856930DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6396179PMC
February 2019

Ultra-early response assessment in lymphoma treatment: [F]FDG PET/MR captures changes in glucose metabolism and cell density within the first 72 hours of treatment.

Eur J Nucl Med Mol Imaging 2018 06 26;45(6):931-940. Epub 2018 Feb 26.

Department of Biomedical Imaging and Image-guided Therapy, Division of Nuclear Medicine, Medical University of Vienna, Waehringer Guertel 18-20, 1090, Vienna, Austria.

Purpose: To determine whether, in patients with Hodgkin lymphoma (HL) or non-Hodgkin lymphoma (NHL), [F]FDG PET/MR can capture treatment effects within the first week after treatment initiation, and whether changes in glucose metabolism and cell density occur simultaneously.

Methods: Patients with histologically proven HL or NHL were included in this prospective IRB-approved study. Patients underwent [F]FDG PET/MR before, and then 48-72 h after (follow-up 1, FU-1) and 1 week after (FU-2) initiation of the first cycle of their respective standard chemotherapy (for HL) or immunochemotherapy (for NHL). Standardized [F]FDG uptake values (SUVmax, SUVmean) and apparent diffusion coefficients (ADCmin, ADCmean) based on diffusion-weighted MRI, and metabolic and morphological tumour volumes (MTV, VOL) were assessed at each time-point. Multilevel analyses with an unstructured covariance matrix, and pair-wise post-hoc tests were used to test for significant changes in SUVs, ADCs, MTVs and VOLs between the three time-points.

Results: A total of 58 patients (11 with HL and 47 with NHL) with 166 lesions were analysed. Lesion-based mean rates of change in SUVmax, SUVmean, ADCmin, ADCmean, MTV and VOL between baseline and FU-1 were -46.8%, -33.3%, +20.3%, +14%, -46% and -12.8%, respectively, and between baseline and FU-2 were -65.1%, -49%, +50.7%, +32.4%, -61.1% and -24.2%, respectively. These changes were statistically significant (P < 0.01) except for the change in VOL between baseline and FU-1 (P = 0.079).

Conclusion: In lymphoma patients, [F]FDG PET/MR can capture treatment-induced changes in glucose metabolism and cell density as early as 48-72 h after treatment initiation.
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http://dx.doi.org/10.1007/s00259-018-3937-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5915494PMC
June 2018

Probability of remaining in unsustained complete remission after steroid therapy withdrawal in patients with primary warm-antibody reactive autoimmune hemolytic anemia.

Wien Klin Wochenschr 2016 Apr 24;128(7-8):234-7. Epub 2015 Sep 24.

Division of Hematology and Hemostaseology, Department of Medicine I, Medical University of Vienna, Währinger Gürtel 18-20, 1090, Vienna, Austria.

Background: Primary warm autoimmune hemolytic anemia (WAIHA) is a rare autoimmune disorder frequently responding to corticosteroid first-line treatment and effective second-line treatment options such as splenectomy or anti-CD20 antibody therapy. Disease management is frequently hampered by a lack of evidence.

Methods: We have investigated the probability of sustained treatment-free remission after steroid induction to facilitate clinical decision making regarding timing and necessity of second-line treatments. Response data from 31 patients with primary WAIHA initially treated with steroids were retrospectively analyzed. All patients responded by achieving a hemoglobin of at least 10 mg/dl.

Results: After steroid tapering and final withdrawal, 9 of 30 patients remained in unsustained complete remission (CR). The probability of remaining in CR after steroid treatment only was 38.2 % (2 SD 20.6 %) at 15 months. The median remission duration was 100 + months with a range of 12 + to 163 + months. Of note, none of the remaining patients still on steroids achieved CR beyond 15 + months.

Conclusion: These data indicate that a considerable proportion of patients do not need further treatment and that relapses will not occur after 15 months in CR.
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http://dx.doi.org/10.1007/s00508-015-0863-yDOI Listing
April 2016

Transformed mycosis fungoides: bridging to allogeneic stem cell transplantation with brentuximab vedotin.

Leuk Lymphoma 2016 19;57(1):206-8. Epub 2015 Jun 19.

a Department of Internal Medicine I , Division of Hematology and Hemostaseology, Medical University of Vienna , Vienna , Austria.

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http://dx.doi.org/10.3109/10428194.2015.1044748DOI Listing
October 2016

Evaluation of Diffusion-Weighted Magnetic Resonance Imaging for Follow-up and Treatment Response Assessment of Lymphoma: Results of an 18F-FDG-PET/CT-Controlled Prospective Study in 64 Patients.

Clin Cancer Res 2015 Jun 2;21(11):2506-13. Epub 2015 Mar 2.

Department of Internal Medicine I, Medical University of Vienna, Vienna, Austria.

Purpose: To determine the value of diffusion-weighted MRI (DWI-MRI) for treatment response assessment in 2-[18F]fluoro-2-deoxy-D-glucose (FDG)-avid lymphoma.

Experimental Design: Patients with FDG-avid Hodgkin (HL) or non-Hodgkin lymphoma (NHL) at pretherapeutic 18F-FDG-PET/CT, who had also undergone pretherapeutic whole-body DWI-MRI, were included in this prospective study. Depending on the histologic lymphoma subtype, patients received different systemic treatment regimens, and follow-up DWI-MRI and 18F-FDG-PET/CT were performed at one or more time points, depending on the clinical course. For each follow-up DWI-MRI, region-based rates of agreement, and rates of agreement in terms of treatment response (complete remission, partial remission, stable disease, or progressive disease), relative to the corresponding 18F-FDG-PET/CT, were calculated.

Results: Sixty-four patients were included: 10 with HL, 22 with aggressive NHL, and 32 with indolent NHL. The overall region-based agreement of DWI-MRI with 18F-FDG-PET/CT was 99.4%. For the 51 interim examinations (performed after 1-3 therapy cycles), region-based agreement of DWI-MRI with 18F-FDG-PET/CT was 99.2%, and for the 48 end-of-treatment examinations, agreement was 99.8%. No significant differences, in terms of region-based agreement between DWI-MRI and 18F-FDG-PET/CT, were observed between the three lymphoma groups (HL, aggressive NHL, indolent NHL; P = 0.25), or between interim and end-of-treatment examinations (P = 0.21). With regard to treatment response assessment, DWI-MRI agreed with 18F-FDG-PET/CT in 99 of 102 follow-up examinations (97.1%), with a κ value of 0.94 (P < 0.0001).

Conclusions: In patients with FDG-avid lymphoma, DWI-MRI may be a feasible alternative to 18F-FDG-PET/CT for follow-up and treatment response assessment.
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http://dx.doi.org/10.1158/1078-0432.CCR-14-2454DOI Listing
June 2015

Serum-tryptase at diagnosis: a novel biomarker improving prognostication in Ph(+) CML.

Am J Cancer Res 2015 15;5(1):354-62. Epub 2014 Dec 15.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna Vienna, Austria ; Ludwig Boltzmann Cluster Oncology, Medical University of Vienna Vienna, Austria.

Basophilia is an established prognostic variable in Ph-chromosome+ chronic myeloid leukemia (CML). However, in CML, basophils are often immature and thus escape microscopic quantification. We have previously shown that tryptase is produced and secreted by immature CML basophils. In the current study, serum samples of 79 CML patients (chronic phase=CP, n=69; accelerated/blast phase=AP/BP, n=10) treated with BCR/ABL inhibitors, were analyzed for their tryptase content. Serum-tryptase levels at diagnosis were found to correlate with basophil counts and were higher in AP/BP patients (median tryptase: 29.9 ng/mL) compared to patients with CP (11.7 ng/mL; p<0.05). In 20/69 patients with CP, progression occurred. The progression-rate was higher in patients with tryptase >15 ng/mL (31%) compared to those with normal tryptase levels (9%, p<0.05). To validate tryptase as new prognostic variable, we replaced basophils by tryptase levels in the EUTOS score. This modified EUTOS-T score was found to predict progression-free and event-free survival significantly better, with p values of 0.000064 and 0.00369, respectively, compared to the original EUTOS score (progression-free survival: p=0.019; event-free survival: p=0.156). In conclusion, our data show that the serum-tryptase level at diagnosis is a powerful prognostic biomarker in CML. Inclusion of tryptase in prognostic CML scores may improve their predictive value.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4300686PMC
January 2015

Cold antibody autoimmune hemolytic anemia and lymphoproliferative disorders: a retrospective study of 20 patients including clinical, hematological, and molecular findings.

Wien Klin Wochenschr 2014 Jun 20;126(11-12):376-82. Epub 2014 May 20.

Division of Hematology and Hemostaseology, Depatment of Internal Medicine I, Medical University of Vienna, Währinger Gürtel 18-20, 1097, Vienna, Austria.

A total of 20 patients with cold antibody hemolytic anemia were evaluated in a retrospective study of them, 15 had a monoclonal gammopathy of unknown significance (MGUS): 14 with MGUS of immunoglobulin M (IgM) subtype and 1 with immunoglobulin G subtype. One patient had smoldering Waldenström's macroglobulinemia, but four patients had no monoclonal protein and no evidence of lymphoma. However, in three of these patients, we were able to demonstrate a (mono-)clonal rearrangement of their immunoglobulin heavy and/or light chains. Of the 20 patients, 5 had IgHV34 nucleotide sequence indicating that the antibody was directed against the "I" antigen. Two patients exhibited a progressive increase of IgM over time, however without increasing hemolytic activity. Moreover, in two patients with long-term follow-up, we were able to correlate recurrent hemolytic activity with low environmental temperatures. Among four patients treated with rituximab, all four responded to treatment. However, treatment effect was only transient in all of them.
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http://dx.doi.org/10.1007/s00508-014-0547-zDOI Listing
June 2014

Evaluation of diffusion-weighted MRI for pretherapeutic assessment and staging of lymphoma: results of a prospective study in 140 patients.

Clin Cancer Res 2014 Jun 2;20(11):2984-93. Epub 2014 Apr 2.

Authors' Affiliations: Departments of Biomedical Imaging and Image-guided Therapy, Internal Medicine I, Internal Medicine III, and Opthalmology and Optometry, and Institute of Pathology, Medical University of Vienna, Vienna, Austria.

Purpose: To determine the value of diffusion-weighted MRI (DWI-MRI) for pretherapeutic imaging of fluorodeoxyglucose (FDG)-avid lymphoma and lymphoma with variable FDG avidity.

Experimental Design: Treatment-naïve patients with lymphoma who were referred for whole-body staging were included in this prospective study. Group A included patients with FDG-avid lymphoma (e.g., Hodgkin, diffuse large B-cell, and follicular lymphoma), whereas Group B included patients with lymphoma of variable FDG avidity [e.g., extranodal marginal zone B-cell lymphoma of the mucosa-associated lymphoid tissue (MALT)]. All patients underwent DWI-MRI and 18F-FDG- positron emission tomography/computed tomography (PET/CT). Region-based sensitivity and agreement with Ann Arbor staging, relative to the reference standard, were calculated for DWI-MRI, and, in Group B, also 18F-FDG-PET/CT and contrast-enhanced (CE-) CT.

Results: In Group A (100 patients), DWI-MRI had a region-based sensitivity of 97%, and with regard to staging, agreed with the reference standard in 94 of 100 patients (κ, 0.92). In Group B (40 patients; 38 MALT lymphomas and 2 small lymphocytic lymphomas/chronic lymphocytic leukemias), DWI-MRI, 18F-FDG-PET/CT, and CE-CT had region-based sensitivities of 94.4%, 60.9%, and 70.7%, respectively. With regard to staging in Group B, DWI-MRI, 18F-FDG-PET/CT, and CE-CT agreed with the reference standard in 37 of 40, 26 of 40, and 24 of 40 patients, with κ values of 0.89, 0.52, and 0.43, respectively.

Conclusions: In patients with FDG-avid lymphoma, DWI-MRI seems to be only slightly inferior to 18F-FDG-PET/CT with regard to pretherapeutic regional assessment and staging. In patients with lymphoma subtypes that show a variable FDG avidity (e.g., MALT lymphoma), DWI-MRI seems to be superior to both 18F-FDG-PET/CT and CE-CT.
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http://dx.doi.org/10.1158/1078-0432.CCR-13-3355DOI Listing
June 2014

Endogenous erythroid colony formation in chronic myeloid leukemia: a recurrent finding associated with persistent minimal residual disease under imatinib.

Stem Cells Dev 2013 Dec 4;22(23):3043-51. Epub 2013 Sep 4.

1 Department of Laboratory Medicine, Medical University of Vienna , Vienna, Austria .

In vitro endogenous erythroid colony (EEC) formation is a common finding in BCR-ABL-negative myeloproliferative neoplasms. The aim of the present study was to determine the prevalence and the clinical significance of EEC growth in chronic myeloid leukemia (CML). Results of clonogeneic progenitor cell assays from 52 patients with newly diagnosed CML were correlated with disease characteristics at presentation and molecular response to imatinib. EECs (median 7 per dish, range 1-39) were detectable in 16 patients (31%). The proportion of patients with a high-risk Sokal score was lower in the EEC group (7% vs. 30%, respectively). The cumulative incidence of achieving a major molecular response after 2 years of imatinib was similar for both groups. However, patients with EECs were less likely to achieve a more profound decline of BCR-ABL transcripts. After 6 years of imatinib, the cumulative probability [95% CI] of reaching a ≥4 log reduction of BCR-ABL was 48% [16%; 92%] for patients of the EEC group and 84% [63%; 97%] for patients of the No EEC group. The probability [95% CI] of achieving a >4.5 log reduction of BCR-ABL after 7 years was 13% [2%; 61%] for patients with EECs and 52% [30%; 78%] for patients without EECs. In vitro EECs disappeared after achievement of a major molecular response in all evaluable patients. The data indicate that EEC formation is a recurrent finding in patients with CML which deserves further attention as a possible biomarker predicting the degree of molecular response to imatinib.
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http://dx.doi.org/10.1089/scd.2013.0096DOI Listing
December 2013

Identification of basophils as a major source of hepatocyte growth factor in chronic myeloid leukemia: a novel mechanism of BCR-ABL1-independent disease progression.

Neoplasia 2012 Jul;14(7):572-84

Ludwig Boltzmann Cluster Oncology, Vienna, Austria.

Chronic myeloid leukemia (CML) is a hematopoietic neoplasm characterized by the Philadelphia chromosome and the related BCR-ABL1 oncoprotein. Acceleration of CML is usually accompanied by basophilia. Several proangiogenic molecules have been implicated in disease acceleration, including the hepatocyte growth factor (HGF). However, little is known so far about the cellular distribution and function of HGF in CML. We here report that HGF is expressed abundantly in purified CML basophils and in the basophil-committed CML line KU812, whereas all other cell types examined expressed only trace amounts of HGF or no HGF. Interleukin 3, a major regulator of human basophils, was found to promote HGF expression in CML basophils. By contrast, BCR-ABL1 failed to induce HGF synthesis in CML cells, and imatinib failed to inhibit expression of HGF in these cells. Recombinant HGF as well as basophil-derived HGF induced endothelial cell migration in a scratch wound assay, and these effects of HGF were reverted by an anti-HGF antibody as well as by pharmacologic c-Met inhibitors. In addition, anti-HGF and c-Met inhibitors were found to suppress the spontaneous growth of KU812 cells, suggesting autocrine growth regulation. Together, HGF is a BCR-ABL1-independent angiogenic and autocrine growth regulator in CML. Basophils are a unique source of HGF in these patients and may play a more active role in disease-associated angiogenesis and disease progression than has so far been assumed. Our data also suggest that HGF and c-Met are potential therapeutic targets in CML.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3421954PMC
http://dx.doi.org/10.1593/neo.12724DOI Listing
July 2012

Progressive peripheral arterial occlusive disease and other vascular events during nilotinib therapy in CML.

Am J Hematol 2011 Jul 27;86(7):533-9. Epub 2011 Apr 27.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Austria.

The second generation BCR/ABL kinase inhibitor nilotinib is increasingly used for the treatment of imatinib-resistant chronic myeloid leukemia (CML). So far, nilotinib is considered a well-tolerated drug with little if any side effects, although an increase in the fasting glucose level has been reported. We examined a series of 24 consecutive CML patients treated with nilotinib in our center for the development of non-hematologic adverse events. Three of these 24 CML patients developed a rapidly progressive peripheral arterial occlusive disease (PAOD) during treatment with nilotinib. In all three cases, PAOD required repeated angioplasty and/or multiple surgeries within a few months. No PAOD was known before nilotinib-therapy in these patients, although all three had received imatinib. In two patients, pre-existing risk factors predisposing for PAOD were known, and one of them had developed diabetes mellitus during nilotinib. In the other 21 patients treated with nilotinib in our center, one less severe PAOD, one myocardial infarction, one spinal infarction, one subdural hematoma, and one sudden death of unknown etiology were recorded. In summary, treatment with nilotinib may be associated with an increased risk of vascular adverse events, including PAOD development. In a subgroup of patients, these events are severe or even life-threatening. Although the exact mechanisms remain unknown, we recommend screening for pre-existing PAOD and for vascular risk factors such as diabetes mellitus in all patients before starting nilotinib and in the follow up during nilotinib-therapy.
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http://dx.doi.org/10.1002/ajh.22037DOI Listing
July 2011

High STAT5 levels mediate imatinib resistance and indicate disease progression in chronic myeloid leukemia.

Blood 2011 Mar 10;117(12):3409-20. Epub 2011 Jan 10.

Center of Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.

In BCR-ABL1(+) leukemia, drug resistance is often associated with up-regulation of BCR-ABL1 or multidrug transporters as well as BCR-ABL1 mutations. Here we show that the expression level of the transcription factor STAT5 is another parameter that determines the sensitivity of BCR-ABL1(+) cells against tyrosine kinase inhibitors (TKIs), such as imatinib, nilotinib, or dasatinib. Abelson-transformed cells, expressing high levels of STAT5, were found to be significantly less sensitive to TKI-induced apoptosis in vitro and in vivo but not to other cytotoxic drugs, such as hydroxyurea, interferon-β, or Aca-dC. The STAT5-mediated protection requires tyrosine phosphorylation of STAT5 independent of JAK2 and transcriptional activity. In support of this concept, under imatinib treatment and with disease progression, STAT5 mRNA and protein levels increased in patients with Ph(+) chronic myeloid leukemia. Based on our data, we propose a model in which disease progression in BCR-ABL1(+) leukemia leads to up-regulated STAT5 expression. This may be in part the result of clonal selection of cells with high STAT5 levels. STAT5 then accounts for the resistance against TKIs, thereby explaining the dose escalation frequently required in patients reaching accelerated phase. It also suggests that STAT5 may serve as an attractive target to overcome imatinib resistance in BCR-ABL1(+) leukemia.
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http://dx.doi.org/10.1182/blood-2009-10-248211DOI Listing
March 2011

Polo-like kinase 1 (Plk1) as a novel drug target in chronic myeloid leukemia: overriding imatinib resistance with the Plk1 inhibitor BI 2536.

Cancer Res 2010 Feb 9;70(4):1513-23. Epub 2010 Feb 9.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Institute of Immunology, Clinical Institute of Medical and Chemical Laboratory Diagnostics, Medical University of Vienna, A-1090 Vienna, Austria.

In most patients with chronic myeloid leukemia (CML), the disease can be kept under control using the BCR/ABL kinase inhibitor imatinib. Nevertheless, resistance or intolerance to imatinib and other BCR/ABL inhibitors may occur during therapy. Therefore, CML research is focusing on novel targets and targeted drugs. Polo-like kinase 1 (Plk1) is a serine/threonine kinase that plays an essential role in mitosis. In this study, we examined the expression of Plk1 in CML cells and its potential role as a therapeutic target. Plk1 was found to be expressed in phosphorylated form in the CML cell line K562 as well as in primary CML cells in all patients tested. Inhibition of BCR/ABL by imatinib or nilotinib (AMN107) led to decreased expression of the Plk1 protein in CML cells, suggesting that BCR/ABL promotes Plk1 generation. Silencing of Plk1 in CML cells by a small interfering RNA approach was followed by cell cycle arrest and apoptosis. Furthermore, the Plk1-targeting drug BI 2536 was found to inhibit proliferation of imatinib-sensitive and imatinib-resistant CML cells, including leukemic cells, carrying the T315 mutation of BCR/ABL with reasonable IC(50) values (1-50 nmol/L). The growth-inhibitory effects of BI 2536 on CML cells were found to be associated with cell cycle arrest and apoptosis. Moreover, BI 2536 was found to synergize with imatinib and nilotinib in producing growth inhibition in CML cells. In conclusion, Plk1 is expressed in CML cells and may represent a novel, interesting target in imatinib-sensitive and imatinib-resistant CML.
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http://dx.doi.org/10.1158/0008-5472.CAN-09-2181DOI Listing
February 2010

Expression of activated STAT5 in neoplastic mast cells in systemic mastocytosis: subcellular distribution and role of the transforming oncoprotein KIT D816V.

Am J Pathol 2009 Dec 5;175(6):2416-29. Epub 2009 Nov 5.

Department of Internal Medicine I, Medical University of Vienna, Vienna, Austria.

Recent data suggest that the signal transducer and activator of transcription (STAT)5 contributes to differentiation and growth of mast cells. It has also been described that constitutively phosphorylated STAT5 (pSTAT5) plays a pro-oncogenic role in various myeloid neoplasms. We examined the expression of pSTAT5 in neoplastic mast cells in systemic mastocytosis and asked whether the disease-related oncoprotein KIT D816V is involved in STAT5 activation. As assessed by immunohistochemistry using the anti-pSTAT5 antibody AX1, neoplastic mast cells were found to display pSTAT5 in all SM patients examined (n = 40). Expression of pSTAT5 was also demonstrable in the KIT D816V-positive mast cell leukemia cell line HMC-1. Using various staining-protocols, pSTAT5 was found to be located in both the cytoplasmic and nuclear compartment of mast cells. To define the functional role of KIT D816V in STAT5-activation, Ba/F3 cells with doxycycline-inducible expression of KIT D816V were used. In these cells, induction of KIT D816V resulted in an increased expression of pSTAT5 without substantial increase in total STAT5. Moreover, the KIT D816V-targeting kinase-inhibitor PKC412 was found to counteract expression of pSTAT5 in HMC-1 cells as well as doxycycline-induced expression of pSTAT5 in Ba/F3 cells. Finally, a dominant negative STAT5-construct was found to inhibit growth of HMC-1 cells. Together, our data show that neoplastic mast cells express cytoplasmic and nuclear pSTAT5, that KIT D816V promotes STAT5-activation, and that STAT5-activation contributes to growth of neoplastic mast cells.
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http://dx.doi.org/10.2353/ajpath.2009.080953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2789628PMC
December 2009

Evaluation of in vivo antineoplastic effects of rapamycin in patients with chemotherapy-refractory AML.

Eur J Intern Med 2009 Dec 7;20(8):775-8. Epub 2009 Oct 7.

Department of Internal Medicine I, Division of Hematology & Hemostaseology, Medical University of Vienna, Austria.

Background: The mammalian target of rapamycin (mTOR) has recently been identified as a potential target in acute myeloid leukemia (AML).

Methods: We treated 5 patients with chemotherapy-refractory AML with the mTOR-inhibitor rapamycin at 2mg per os daily for 14 days, with dose adjustment allowed to reach a target serum rapamycin concentration of 10-20 ng/mL. Four of five patients received additional hydroxyurea at constant dose during treatment with rapamycin.

Results: Two patients achieved a leukocyte response, in one of them, a prolonged response was seen. In the other patients, blast counts remained stable or increased during rapamycin therapy. We did not observe severe hematologic or non-hematologic side effects of rapamycin.

Conclusion: Rapamycin at 2mg per day acts mildly cytoreductive in a subgroup of patients with refractory AML. Higher doses and drug combinations may be required to obtain long lasting anti-leukemic effects in these patients.
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http://dx.doi.org/10.1016/j.ejim.2009.09.007DOI Listing
December 2009

Identification of proapoptotic Bim as a tumor suppressor in neoplastic mast cells: role of KIT D816V and effects of various targeted drugs.

Blood 2009 Dec 22;114(26):5342-51. Epub 2009 Oct 22.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Systemic mastocytosis (SM) is a myeloid neoplasm involving mast cells (MCs) and their progenitors. In most cases, neoplastic cells display the D816V-mutated variant of KIT. KIT D816V exhibits constitutive tyrosine kinase (TK) activity and has been implicated in increased survival and growth of neoplastic MCs. Recent data suggest that the proapoptotic BH3-only death regulator Bim plays a role as a tumor suppressor in various myeloid neoplasms. We found that KIT D816V suppresses expression of Bim in Ba/F3 cells. The KIT D816-induced down-regulation of Bim was rescued by the KIT-targeting drug PKC412/midostaurin. Both PKC412 and the proteasome-inhibitor bortezomib were found to decrease growth and promote expression of Bim in MC leukemia cell lines HMC-1.1 (D816V negative) and HMC-1.2 (D816V positive). Both drugs were also found to counteract growth of primary neoplastic MCs. Furthermore, midostaurin was found to cooperate with bortezomib and with the BH3-mimetic obatoclax in producing growth inhibition in both HMC-1 subclones. Finally, a Bim-specific siRNA was found to rescue HMC-1 cells from PKC412-induced cell death. Our data show that KIT D816V suppresses expression of proapoptotic Bim in neoplastic MCs. Targeting of Bcl-2 family members by drugs promoting Bim (re)-expression, or by BH3-mimetics such as obatoclax, may be an attractive therapy concept in SM.
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http://dx.doi.org/10.1182/blood-2008-08-175190DOI Listing
December 2009

Dasatinib in the treatment of chronic myeloid leukemia in accelerated phase after imatinib failure: the START a trial.

J Clin Oncol 2009 Jul 1;27(21):3472-9. Epub 2009 Jun 1.

Hammersmith Hospital, Imperial College School of Medicine, Du Cane Rd, London W12 0NN, United Kingdom.

PURPOSE Patients with chronic myelogenous leukemia in accelerated phase (CML-AP) that is resistant or intolerant to imatinib have limited therapeutic options. Dasatinib, a potent inhibitor of BCR-ABL and SRC-family kinases, has efficacy in patients with CML-AP who have experienced treatment failure with imatinib. We now report follow-up data from the full patient cohort of 174 patients enrolled onto a phase II trial to provide a more complete assessment of the efficacy and safety of dasatinib in this population. PATIENTS AND METHODS Patients with imatinib-resistant (n = 161) or -intolerant (n = 13) CML-AP received dasatinib 70 mg orally twice daily. Results At a median follow-up of 14.1 months (treatment duration, 0.1 to 21.7 months), major and complete hematologic responses were attained by 64% and 45% of patients, respectively, and major and complete cytogenetic responses were achieved in 39% and 32% of patients, respectively. Responses were achieved irrespective of imatinib status (resistant or intolerant), prior stem-cell transplantation, or the presence of prior BCR-ABL mutation. The 12-month progression-free survival and overall survival rates were 66% and 82%, respectively. Dasatinib was generally well tolerated; the most frequent nonhematologic severe treatment-related adverse event was diarrhea (52%; grade 3 to 4, 8%). Cytopenias were common, including grade 3 to 4 neutropenia (76%) and thrombocytopenia (82%). Pleural effusion occurred in 27% of patients (grade 3 to 4, 5%). CONCLUSION Dasatinib is effective in patients with CML-AP after imatinib treatment failure.
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http://dx.doi.org/10.1200/JCO.2007.14.3339DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4979080PMC
July 2009

Diagnostic algorithms, monitoring, prognostication, and therapy in chronic myeloid leukemia (CML): a proposal of the Austrian CML platform.

Wien Klin Wochenschr 2008 ;120(21-22):697-709

Department of Internal Medicine I, Division of Haematology & Hemostaseology, Medical University of Vienna, Austria.

Chronic myeloid leukemia (CML) is a stem cell disease characterized by the BCR/ABL oncoprotein. The ABL kinase inhibitor imatinib is effective in most patients and considered standard first-line therapy. However, not all patients show a long-lasting response to this drug. In fact, resistance against imatinib has been described and is an emerging clinical problem in CML. For these patients, novel multi-kinase inhibitors such as nilotinib or dasatinib as well as stem cell transplantation, represent alternative treatment options. The decision concerning second-line therapies and selection of drugs is usually based on the presence and type of BCR/ABL mutations, the phase of disease, other disease-related factors as well as patient-related factors including age, co-morbidity, and pharmacologic determinants. The current article provides an overview on diagnostic and therapeutic strategies for patients with treatment-naïve and imatinib-resistant CML, together with proposed algorithms that were discussed and approved by members of the CML platform of the Austrian Society for Hematology and Oncology (OGHO) in 2007 and 2008. The resulting recommendations should assist in diagnosis and prognostication in CML, follow-up and disease-monitoring, patient selection for interventional therapies, and in the preparation and conduct of clinical trials.
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http://dx.doi.org/10.1007/s00508-008-1100-8DOI Listing
February 2010

Dasatinib inhibits the growth and survival of neoplastic human eosinophils (EOL-1) through targeting of FIP1L1-PDGFRalpha.

Exp Hematol 2008 Oct 10;36(10):1244-53. Epub 2008 Jul 10.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Objective: Chronic eosinophilic leukemia (CEL) is a myeloproliferative disorder characterized by molecular and/or cytogenetic evidence of clonality of eosinophils, marked eosinophilia, and organ damage. In many patients, the transforming mutation FIP1L1-PDGFRalpha and the related CHIC2 deletion are found. The respective oncoprotein, FIP1L1-PDGFRalpha, is considered to play a major role in malignant cell growth in CEL. The tyrosine kinase (TK) inhibitor imatinib (STI571) has been described to counteract the TK activity of FIP1L1-PDGFRalpha in most patients. However, not all patients with CEL show a response to imatinib. Therefore, several attempts have been made to identify other TK inhibitors that counteract growth of neoplastic eosinophils.

Materials And Methods: We provide evidence that dasatinib, a multi-targeted kinase inhibitor, blocks the growth and survival of EOL-1, an eosinophil leukemia cell line carrying FIP1L1-PDGFRalpha.

Results: The effects of dasatinib on proliferation of EOL-1 cells were dose-dependent, with an IC50 of 0.5 to 1 nM, which was found to be in the same range when compared to IC50 values produced with imatinib. Dasatinib was also found to induce apoptosis in EOL-1 cells in a dose-dependent manner (IC50: 1-10 nM). The apoptosis-inducing effects of dasatinib on EOL-1 cells were demonstrable by light microscopy, flow cytometry, and in a TUNEL assay. In Western blot experiments, dasatinib completely blocked the phosphorylation of FIP1L1-PDGFRalpha in EOL-1 cells.

Conclusions: Dasatinib inhibits the growth of leukemic eosinophils through targeting of the disease-related oncoprotein FIP1L1-PDGFRalpha. Based on this observation, dasatinib may be considered as a new interesting treatment option for patients with CEL.
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http://dx.doi.org/10.1016/j.exphem.2008.04.017DOI Listing
October 2008

Diagnostic and prognostic value of new biochemical and immunohistochemical parameters in chronic myeloid leukemia.

Leuk Lymphoma 2008 Apr;49(4):635-8

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Chronic myeloid leukemia (CML) is a stem-cell disease characterized by multilineage expansion of clonal BCR/ABL+ cells. Transformation from chronic into accelerated and blast phase of CML is usually associated with drug resistance and is accompanied by typical clinical and/or laboratory features, such as splenomegaly, increase in precursor cells, disturbed megakaryopoiesis, basophilia or marrow fibrosis. Because of new treatment options, early recognition of disease-acceleration is of importance. In this article, we review most recent developments in diagnostic procedures employing basophil-related biochemical and histopathological markers. These tests are useful to quantitate basophil-lineage cells in the peripheral blood in CML, to determine and quantify basophilia in the bone marrow, and to detect focal accumulations of blast cells and megakaryocytes as well as increased angiogenesis and fibrosis in bone marrow sections. Application of these markers may assist in determining the phase of disease and may help to better predict the prognosis in individual patients.
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http://dx.doi.org/10.1080/10428190701858849DOI Listing
April 2008

Unique effects of KIT D816V in BaF3 cells: induction of cluster formation, histamine synthesis, and early mast cell differentiation antigens.

J Immunol 2008 Apr;180(8):5466-76

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Oncogenic tyrosine kinases (TK) usually convert growth factor-dependent cells to factor independence with autonomous proliferation. However, TK-driven neoplasms often are indolent and characterized by cell differentiation rather than proliferation. A prototype of an indolent TK-driven neoplasm is indolent systemic mastocytosis. We found that the D816V-mutated variant of KIT, a TK detectable in most patients with systemic mastocytosis, induces cluster formation and expression of several mast cell differentiation and adhesion Ags, including microphthalmia transcription factor, IL-4 receptor, histamine, CD63, and ICAM-1 in IL-3-dependent BaF3 cells. By contrast, wild-type KIT did not induce cluster formation or mast cell differentiation Ags. Additionally, KIT D816V, but not wild-type KIT, induced STAT5 activation in BaF3 cells. However, despite these intriguing effects, KIT D816V did not convert BaF3 cells to factor-independent proliferation. Correspondingly, BaF3 cells with conditional expression of KIT D816V did not form tumors in nude mice. Together, the biologic effects of KIT D816V in BaF3 cells match strikingly with the clinical course of indolent systemic mastocytosis and with our recently established transgenic mouse model, in which KIT D816V induces indolent mast cell accumulations but usually does not induce a malignant mast cell disease. Based on all these results, it is hypothesized that KIT D816V as a single hit may be sufficient to cause indolent systemic mastocytosis, whereas additional defects may be required to induce aggressive mast cell disorders.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2976849PMC
http://dx.doi.org/10.4049/jimmunol.180.8.5466DOI Listing
April 2008

Targeting of heat shock protein 32 (Hsp32)/heme oxygenase-1 (HO-1) in leukemic cells in chronic myeloid leukemia: a novel approach to overcome resistance against imatinib.

Blood 2008 Feb 16;111(4):2200-10. Epub 2007 Nov 16.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Resistance toward imatinib and other BCR/ABL tyrosine kinase inhibitors remains an increasing clinical problem in the treatment of advanced stages of chronic myeloid leukemia (CML). We recently have identified the heat shock protein 32 (Hsp32)/heme oxygenase-1 (HO-1) as a BCR/ABL-dependent survival molecule in CML cells. We here show that silencing Hsp32/HO-1 in CML cells by an siRNA approach results in induction of apoptosis. Moreover, targeting Hsp32/HO-1 by either pegylated zinc protoporphyrine (PEG-ZnPP) or styrene maleic acid-micelle-encapsulated ZnPP (SMA-ZnPP) resulted in growth inhibition of BCR/ABL-transformed cells. The effects of PEG-ZnPP and SMA-ZnPP were demonstrable in Ba/F3 cells carrying various imatinib-resistant mutants of BCR/ABL, including the T315I mutant, which exhibits resistance against all clinically available BCR/ABL tyrosine kinase inhibitors. Growth-inhibitory effects of PEG-ZnPP and SMA-ZnPP also were observed in the CML-derived human cell lines K562 and KU812 as well as in primary leukemic cells obtained from patients with freshly diagnosed CML or imatinib-resistant CML. Finally, Hsp32/HO-1-targeting compounds were found to synergize with either imatinib or nilotinib in producing growth inhibition in imatinib-resistant K562 cells and in Ba/F3 cells harboring the T315I mutant of BCR/ABL. In summary, these data show that HO-1 is a promising novel target in imatinib-resistant CML.
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http://dx.doi.org/10.1182/blood-2006-11-055723DOI Listing
February 2008

Synergistic growth-inhibitory effects of two tyrosine kinase inhibitors, dasatinib and PKC412, on neoplastic mast cells expressing the D816V-mutated oncogenic variant of KIT.

Haematologica 2007 Nov;92(11):1451-9

Department of Internal Medicine I, Division of Hematology & Hemostaseology, Medical University of Vienna, Vienna, Austria.

Background And Objectives: In a majority of all patients with systemic mastocytosis (SM) including those with mast cell leukemia (MCL), neoplastic mast cells (MC) display the D816V-mutated variant of KIT. The respective oncoprotein, KIT D816V, exhibits constitutive tyrosine kinase (TK) activity and has been implicated in malignant cell growth. Therefore, several attempts have been made to identify KIT D816V-targeting drugs.

Design And Methods: We examined the effects of the novel TK-inhibitor dasatinib alone and in combination with other targeted drugs on growth of neoplastic MC.

Results: Confirming previous studies, dasatinib was found to inhibit the TK activity of wild type (wt) KIT and KIT-D816V as well as growth and survival of neoplastic MC and of the MCL cell line, HMC-1. The growth-inhibitory effects of dasatinib in HMC-1 cells were found to be associated with a decrease in expression of CD2 and CD63. In addition, we found that dasatinib blocks KIT D816V-induced cluster-formation and viability in Ba/F3 cells. In drug combination experiments, dasatinib was found to co-operate with PKC412, AMN107, imatinib, and 2CdA in producing growth-inhibition and apoptosis in neoplastic MC. In HMC-1.1 cells lacking KIT D816V, all drug interactions were found to be synergistic in nature. By contrast, in HMC-1.2 cells exhibiting KIT D816V, only the combinations dasatinib+PKC412 and dasatinib+2CdA were found to produce synergistic effects.

Interpretation And Conclusions: Combinations of targeted drugs may represent an interesting pharmacologic approach for the treatment of aggressive SM or MCL.
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http://dx.doi.org/10.3324/haematol.11339DOI Listing
November 2007

Myeloid leukemias express a broad spectrum of VEGF receptors including neuropilin-1 (NRP-1) and NRP-2.

Leuk Lymphoma 2007 Oct;48(10):1997-2007

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Vascular endothelial growth factor (VEGF) is produced in neoplastic cells in various myeloid neoplasms and may act as an autocrine growth-regulator. We have examined the expression of five VEGF receptors (VEGR1/Flt-1, VEGFR2/KDR, Flt-4, neuropilin-1 = NRP-1, NRP-2) in leukemic cells obtained from patients with acute myeloid leukemia (n = 28), chronic myeloid leukemia (n = 14), chronic eosinophilic leukemia (n = 3), chronic myelomonocytic leukemia (n = 9), or mast cell leukemia/systemic mastocytosis (n = 3) as well as in respective cell lines. Expression of VEGFR mRNA was analyzed by RT-PCR, and expression of VEGFR protein by immunocytochemistry. In most patients, leukemic cells expressed NRP-1 mRNA and NRP-2 mRNA independent of the type of disease. By contrast, transcripts for Flt-1, KDR, and Flt-4 were expressed variably without a clear correlation to the type of leukemia. Expression of VEGF receptors was also demonstrable at the protein level in all cases tested. In conclusion, neoplastic cells in myeloid leukemias frequently express VEGFR including NRP-1 and NRP-2.
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http://dx.doi.org/10.1080/10428190701534424DOI Listing
October 2007

Plasma levels of P-selectin are determined by platelet turn-over and the P-selectin Thr715Pro polymorphism.

Thromb Res 2008 14;121(4):573-9. Epub 2007 Aug 14.

Clinic for Blood Group Serology, Department of Internal Medicine I, Medical University of Vienna, Währinger Gürtel 18-20, 1090 Vienna, Austria.

Background: Plasma levels of soluble P-selectin (sP-selectin) are often used to demonstrate platelet activation.

Methods: We determined sP-selectin in a variety of disorders characterized by high or low platelet counts and compared their levels with those in healthy subjects. Furthermore, we determined the Thr715Pro polymorphism in all subjects.

Results: Total concentrations of sP-selectin were clearly associated with levels of platelet counts. Thus, calculation of sP-selectin per platelet showed that these levels in patients with thrombocytopenia due to marrow failure and in patients with increased platelet counts were similar to those in controls. Only patients with an increased platelet turn-over had elevated sP-selectin per platelet. While carriers of the Pro715 polymorphism had lower sP-selectin levels than non-carriers, this genetic disposition was over-ruled in patients with increased platelet turn-over.

Conclusion: For the demonstration of platelet activation it is preferable to define sP-selectin based on platelet counts under the consideration of the Pro715Thr polymorphism.
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http://dx.doi.org/10.1016/j.thromres.2007.05.025DOI Listing
April 2008

Synergistic antiproliferative effects of KIT tyrosine kinase inhibitors on neoplastic canine mast cells.

Exp Hematol 2007 Oct 2;35(10):1510-21. Epub 2007 Aug 2.

Department of Internal Medicine I, Division of Hematology and Hemostaseology, Medical University of Vienna, Vienna, Austria.

Aggressive mast cell (MC) tumors are hematopoietic neoplasms characterized by uncontrolled growth of MC and resistance to conventional drugs. In most cases, the tyrosine kinase (TK) receptor KIT is involved in malignant cell growth. Therefore, several KIT TK-targeting drugs are currently being tested for their ability to block growth of neoplastic MC. We examined the effects of four TK inhibitors (imatinib, midostaurin, nilotinib, and dasatinib) on C2 canine mastocytoma cells, as well as primary neoplastic canine MC. As assessed by (3)H-thymidine incorporation experiments, all TK inhibitors produced dose-dependent inhibition of proliferation in C2 cells with the following IC(50) values: imatinib: 269 +/- 180 nM, midostaurin: 157 +/- 35 nM, nilotinib: 55 +/- 24 nM, dasatinib: 12 +/- 3 nM. Growth-inhibitory effects of TK inhibitors were also observed in primary neoplastic mast cells, although IC(50) values for each drug varied from patient to patient, with midostaurin being the most potent agent in all samples tested. In consecutive experiments, we were able to show that TK inhibitors cooperate with each other in producing growth inhibition in C2 cells with synergistic effects observed with most drug combinations. In flow cytometry and TUNEL assay experiments, growth-inhibitory effects of TK inhibitors were found to be associated with cell-cycle arrest and apoptosis. Together, these data show that several TK-targeting drugs induce apoptosis and inhibit proliferation in canine mastocytoma cells in vitro, and that synergistic drug interactions can be obtained. Clinical trials are now warranted to explore whether these TK inhibitors also counteract growth of neoplastic cells in vivo in patients with aggressive MC tumors.
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http://dx.doi.org/10.1016/j.exphem.2007.06.005DOI Listing
October 2007