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Annals of hematology 102(2), 349-358 (2022). doi:10.1007/s00277-022-05080-7, Published by Springer, Berlin ; Heidelberg ; New York

Additional file 1. Supplemental methods, supplemental figures, and supplemental tables S1, S2, and S4.

Blood cancer journal 11(2), 21 (2021). doi:10.1038/s41408-021-00417-3, Published by Nature Publishing Group, London [u.a.]

We studied a subset of hematopoietic stem cells (HSCs) that are defined by elevated expression of CD41 (CD41hi) and showed bias for differentiation toward megakaryocytes (Mks). Mouse models of myeloproliferative neoplasms (MPNs) expressing JAK2-V617F (VF) displayed increased frequencies and percentages of the CD41hi vs CD41lo HSCs compared with wild-type controls. An increase in CD41hi HSCs that correlated with JAK2-V617F mutant allele burden was also found in bone marrow from patients with MPN. CD41hi HSCs produced a higher number of Mk-colonies of HSCs in single-cell cultures in vitro, but showed reduced long-term reconstitution potential compared with CD41lo HSCs in competitive transplantations in vivo. RNA expression profiling showed an upregulated cell cycle, Myc, and oxidative phosphorylation gene signatures in CD41hi HSCs, whereas CD41lo HSCs showed higher gene expression of interferon and the JAK/STAT and TNF��/NF��B signaling pathways. Higher cell cycle activity and elevated levels of reactive oxygen species were confirmed in CD41hi HSCs by flow cytometry. Expression of Epcr, a marker for quiescent HSCs inversely correlated with expression of CD41 in mice, but did not show such reciprocal expression pattern in patients with MPN. Treatment with interferon-�� further increased the frequency and percentage of CD41hi HSCs and reduced the number of JAK2-V617F+ HSCs in mice and patients with MPN. The shift toward the CD41hi subset of HSCs by interferon-�� provides a possible mechanism of how interferon-�� preferentially targets the JAK2 mutant clone.

Blood cancer journal 11(6), 115 (2021). doi:10.1038/s41408-021-00509-0, Published by Nature Publishing Group, London [u.a.]

Figure S6. Confirmation of successful STAT1 or STAT2 knockout. Western blotting of several 32D-BCR-ABL or 32D-JAK2V617F STAT1 or STAT2 knockout clones. STAT2 antibody was used to confirm the knockout, and GAPDH served as the loading control. 32D cells were WEHI starved for 24 h before starting the experiment. wt – wild-type clones, ko – knockout clones, het – presumed heterozygous clones (PDF 134 kb)

Supplementary Material & Methods. Table S1. Primer for cloning STAT1 and STAT2 with included restriction sites. Table S2. STAT1 and STAT2 primers for mutagenesis. Table S3. RT-qPCR primer for gene expression analysis. Table S4. Western blot antibodies. Table S5. gRNA Oligonucleotides. Table S6. ChIP antibodies. Table S7. ChIP-PCR Primer. Table S8. GSEA of a set of 45 interferon-stimulated genes (ISG). Table S9. Statistics for the enrichment analysis of transcription factor binding sites (TFBS) in acetylation (H3K9ac) peaks. (DOCX 44 kb)

Figure S9. Full RT-qPCR panels of tested ISGs. Illustration of the RT-qPCR results of 32D-BCR-ABL- and 32D-JAK2V617F-WT or -STATko or -STAT1(Y/F) and STAT2(Y/F) reconstituted cell clones treated with IFNa (100 U/ml) or left untreated (triplicate), corresponding to the data given in Figs. 3f and 4d. (a) Stat1 and Stat2, (b) Irf7 and Irf9. Stat2 qPCR primer detected the ectopically expressed Stat2 mRNA, explaining the strong upregulation, and endogenous Stat2 can thus not be evaluated in the reconstituted experiments (gray bars). Independent experiments were performed three times. (PDF 56 kb)

Figure S8. Illustration of pY-STAT1 in 32D-JAK2V617F cells in the absence of IFNa. STAT1 and pY-STAT1 stainings of Fig. 4c, shown with increased contrast. The arrows are indicating the pY-STAT1 bands in 32D-BCR-ABL- and 32D-JAK2V617F-STATko cells. (PDF 203 kb)

Figure S7. Comparison of CRISPR/Cas9 manipulated 32D cell lines treated with 100 U IFNa in survival and titration of lower IFNa dosages. Indicated (A) 32D-BCR-ABL and (B) 32D-JAK2V617F cell lines were analyzed in an MTT assay and treated with 100 U IFNa for 72 h (abstracted from Fig. 4a, b). Absorption was normalized to untreated control cells and statistically analyzed using a t test. Mean values ± SD are indicated. *p

Figure S2. MTT assay of 32D-BCR-ABL and 32D-JAK2V617F cells treated with IFNa. 32D-BCR-ABL-(blue) and 32D-JAK2V617F-(red) positive cells were treated with IFNa (0–104 U/ml) alone (continuous lines) or in combination with 0.1 μM imatinib (IM) or ruxolitinib (Rux) (dotted lines) for 72 h and the viability was measured by MTT. Viability was normalized to the untreated control and mean values ± SD are depicted. The respective 32D cells were WEHI starved for 24 h before starting the experiments. Experiments were performed in triplicate and conducted three times. (PDF 27 kb)

Figure S1. Knockout of STAT1 or STAT2 with CRISPR/Cas9n technology. Four guide RNAs have been generated for STAT1 or STAT2 knockout in 32D-BCR-ABL and 32D-JAK2V617F cells, respectively. Excised exons are given. (PDF 19 kb)

Figure S4. Effect of extrinsic soluble factors on gene expression in 32D-EV- or 32D-JAK2V617F-positive cells. Supernatant of WEHI-starved 32D-EV- or 32D-JAK2V617F-positive cells was generated overnight, and after removal of the cells, fresh EV (green) or JAK2V617F-(red) positive cells were incubated with the supernatant for 2 h prior to RNA extraction to analyze the expression of IFN target genes. Mean ± SD values are shown as % of Gapdh. Independent experiments were performed three times and in triplicate, respectively. (PDF 25 kb)

Figure S3. BCR-ABL reduces ISG expression in 32D cells. Gene expression microarray analysis of 32D-EV, 32D-BCR-ABL, or 32D-JAK2V617F cells. Fold change of gene expression is shown, depicting downregulation of the analyzed gene in blue and upregulation in red. (PDF 134 kb)

Figure S5. Correlation of ISG expression and JAK2V617F allelic burden and Western blot of 32D EV, BCR-ABL, or JAK2V617F cells. A, ISG expression (% of GAPDH) and JAK2V617F allelic burden (in %) were plotted against each other and a significant correlation was only found for IRF9 (p = 0.0492). Samples with additional mutations (TET2, ASXL1, or EZH2) were highlighted with a red circle. B, Western blot analysis of pY-STAT1 and overall STAT1 protein after TKI and/or IFNa treatment (4 h) of 32D EV (only −/+ IFNa), BCR-ABL, or JAK2V617F cells. All cell lines were starved of WEHI (source of IL-3) for 24 h before treatment. GAPDH served as the loading control. The same Western blot is shown in Fig. 2c lacking 32D EV cells. (PDF 74 kb)

Figure S10. H3K9 acetylation profile at ISGs. The ChIP-seq acetylation was visualized with the Integrative Genomics Viewer (IGV). Shown are the H3K9 acetylation peaks at the genomic regions of Irf1, Irf17, Irf9, and Stat1 in 32D-JAK2V7F (JAK2V617F) (red), 32D-BCR-ABL (blue), and 32D-EV (green). (PDF 108 kb)

The onset of global epigenetic changes in chromatin that drive tumor proliferation and heterogeneity is a hallmark of many forms cancer. Identifying the epigenetic mechanisms that govern these changes and developing therapeutic approaches to modulate them, is a well-established avenue pursued in translational cancer medicine. Chronic myeloid leukemia (CML) arises clonally when a hematopoietic stem cell (HSC) acquires the capacity to produce the constitutively active tyrosine kinase BCR-ABL1 fusion protein which drives tumor development. Treatment with tyrosine kinase inhibitors (TKI) that target BCR-ABL1 has been transformative in CML management but it does not lead to cure in the vast majority of patients. Thus novel therapeutic approaches are required and these must target changes to biological pathways that are aberrant in CML − including those that occur when epigenetic mechanisms are altered. These changes may be due to alterations in DNA or histones, their biochemical modifications and requisite ‘writer’ proteins, or to dysregulation of various types of non-coding RNAs that collectively function as modulators of transcriptional control and DNA integrity. Here, we review the evidence for subverted epigenetic mechanisms in CML and how these impact on a diverse set of biological pathways, on disease progression, prognosis and drug resistance. We will also discuss recent progress towards developing epigenetic therapies that show promise to improve CML patient care and may lead to improved cure rates.

RWTH-Themen : Berichte aus d. Rheinisch-Westfälischen Technischen Hochschule Aachen SS 2018, 80-85 (2018). special issue: "Profilbereich „Molecular Science & Engineering“ (MSE) / Redaktion: Rauke Xenia Bornefeld ; Angelika Hamacher ; Nives Sunara", Published by RWTH, Aachen

Supplementary methods. Table S1. Bone marrow fibrosis grade. Table S2. Treatment-emergent AEs of any grade that occurred in ≥ 2 patients. Table S3. AEs leading to treatment interruption or modification. Table S4. Mean (± SD) total and unbound steady-state vismodegib plasma concentration. (DOCX 24 kb)

Clinical Case Reports 5(5), 664-670 (2017). doi:10.1002/ccr3.890, Published by Wiley, Chichester

Scientific reports 6, 35454 (2016). doi:10.1038/srep35454, Published by Nature Publishing Group, London

Supplementary material & methods.

Regulation of megakaryocytic factors by CALR mutants in murine and human cell lines. a Detection of Fli-1 and Ets-1 or b Gata1 mRNA expression by RT-qPCR in the indicated 32D and HL60e cells. The experiments were performed in triplicates. Mean and SD are indicated. ***P

Expression of the C-terminally YFP-tagged CALR del52 mutant leads to STAT5 phosphorylation and cytokine-independent growth of 32D MPL cells. a 32D cells stably transduced with WT CALR-flag-YFP, CALR del52-flag-YFP, or empty vector (EV) −/+ MPL were WEHI-starved for 16 h and protein lysates were prepared. SDS-PAGE and Western blotting was performed and the indicated antibodies were used for immunodetection. YFP was detected using a GFP-specific antibody, and GAPDH served as loading control. b The indicated 32D MPL cells were cultured in WEHI-free medium (2 × 105 cells/ml). The cells were counted every 24 h for 3 days. The cell counts are mean values of triplicates. (PDF 53 kb)

Mutant CALR grants factor-independence, protects from apoptosis, and activates downstream signaling in an MPL-dependent manner. a A proliferation assay was performed with the indicated cell lines (2 × 105 cells/ml). 32D MPL cells were counted every 24 h for 4 days. The cell counts are mean values of triplicates. b Empty vector, WT CALR, CALR del52, and ins5 expressing 32D cells were seeded in a density of 5 × 105 cells/ml and grown for 48 h in WEHI-free medium. Apoptosis was analyzed by flow cytometry after staining with Annexin V-APC and 7-AAD. Mean and SD are indicated. *P

Missing CALR-mutant detection in Western blotting is not due to loss of the antibody epitope or an SDS-PAGE artifact. a HEK293T cells were transiently transfected with expression vectors for flag-tagged WT CALR, CALR del52, a C-terminal truncation mutant (ΔC; aa R366) and empty vector. Twenty-four hours after, transfection lysates were generated and SDS-PAGE followed by Western blotting were performed. The indicated antibodies were used for immunostaining. The used CALR antibody bound to the CALR ΔC mutant. b Native page followed by Western blotting was performed with non-denatured protein lysates of 32D CALR cells. The PVDF membrane was stained with CALR antibody, and the membrane was stripped and stained with a flag-specific antibody. (PDF 67 kb)

Gunhild Keller-von Amsberg,1 Steffen Koschmieder21Department of Hematology and Oncology, University Cancer Center Hamburg, University Hospital Hamburg Eppendorf, 2Department of Medicine (Hematology, Oncology, and Stem Cell Transplantation), University Medical Center of Aachen and RWTH Aachen University, Aachen, GermanyAbstract: Bosutinib (SKI-606) is an orally available, once-daily, dual Src and Abl kinase inhibitor with promising clinical potential in first-, second-, and third-line treatment of chronic myeloid leukemia (CML). Bosutinib effectively inhibits wild-type BCR-ABL and most imatinib-resistant BCR-ABL mutations except for V299L and T315I. Low hematologic toxicity is a remarkable characteristic of this novel second-generation tyrosine kinase inhibitor, and this has been ascribed to its minimal activity against the platelet-derived growth factor receptor and KIT. Low-grade, typically self-limiting diarrhea, which usually appears within the first few weeks after treatment initiation, represents the predominant toxicity of bosutinib. Other treatment-associated adverse events are mostly mild to moderate. Bosutinib has been approved by the US Food and Drug Administration for the treatment of chronic, accelerated, or blast phase Philadelphia chromosome-positive CML in adult patients with resistance or intolerance to prior therapy. This review summarizes the main properties of bosutinib and the currently available data on its clinical potential in the treatment of CML.Keywords: bosutinib, chronic myeloid leukemia, BCR-ABL, Src/Abl kinase inhibitor, point mutation, imatinib resistance

Introduction: Older patients with acute myeloid leukemia (AML) experience short survival despite intensive chemotherapy. Azacitidine has promising activity in patients with low proliferating AML. The aim of this dose-finding part of this trial was to evaluate feasibility and safety of azacitidine combined with a cytarabine- and daunorubicin-based chemotherapy in older patients with AML. Trial Design: Prospective, randomised, open, phase II trial with parallel group design and fixed sample size. Patients and Methods: Patients aged 61 years or older, with untreated acute myeloid leukemia with a leukocyte count of ,20,000/ml at the time of study entry and adequate organ function were eligible. Patients were randomised to receive azacitidine either 37.5 (dose level 1) or 75 mg/sqm (dose level 2) for five days before each cycle of induction (7+3 cytarabine plus daunorubicine) and consolidation (intermediate-dose cytarabine) therapy. Dose-limiting toxicity was the primary endpoint. Results: Six patients each were randomised into each dose level and evaluable for analysis. No dose-limiting toxicity occurred in either dose level. Nine serious adverse events occurred in five patients (three in the 37.5 mg, two in the 75 mg arm) with two fatal outcomes. Two patients at the 37.5 mg/sqm dose level and four patients at the 75 mg/sqm level achieved a complete remission after induction therapy. Median overall survival was 266 days and median event-free survival 215 days after a median follow up of 616 days. Conclusions: The combination of azacitidine 75 mg/sqm with standard induction therapy is feasible in older patients with AML and was selected as an investigational arm in the randomised controlled part of this phase-II study, which is currently halted due to an increased cardiac toxicity observed in the experimental arm. Trial Registration: This trial is registered at clinical trials.gov (identifier: NCT00915252).

The transcription factor RUNX1 is essential to establish the haematopoietic gene expression programme; however, the mechanism of how it activates transcription of haematopoietic stem cell (HSC) genes is still elusive. Here, we obtained novel insights into RUNX1 function by studying regulation of the human CD34 gene, which is expressed in HSCs. Using transgenic mice carrying human CD34 PAC constructs, we identified a novel downstream regulatory element (DRE), which is bound by RUNX1 and is necessary for human CD34 expression in long-term (LT)-HSCs. Conditional deletion of Runx1 in mice harbouring human CD34 promoter-DRE constructs abrogates human CD34 expression. We demonstrate by chromosome conformation capture assays in LT-HSCs that the DRE physically interacts with the human CD34 promoter. Targeted mutagenesis of RUNX binding sites leads to perturbation of this interaction and decreased human CD34 expression in LT-HSCs. Overall, our in vivo data provide novel evidence about the role of RUNX1 in mediating interactions between distal and proximal elements of the HSC gene CD34.

Mouse models of human malignancy have greatly enhanced our understanding of disease pathophysiology and have led to novel therapeutic approaches, some with extraordinary success, one such example being inhibition of the BCR-ABL1 oncogene in chronic myeloid leukaemia (CML). Here, we review aspects of the biology of CML that have been uncovered at least in part through the generation and analysis of retroviral and transgenic mouse models of BCR-ABL1 disease. It can be expected that these models will also serve as important tools in the future, especially in the rational design of strategies to eradicate leukemic stem cells and potentially cure CML as well as other cancers.

Medicine 95(20), e3355 (2016). doi:10.1097/MD.0000000000003355, Published by Lippincott Williams & Wilkins, Baltimore, Md.

Journal of experimental medicine : JEM 212(10), 1551-1569 (2015). doi:10.1084/jem.20141661, Published by Rockefeller Univ. Press, New York, NY

Myeloproliferative neoplasms (MPNs) often carry JAK2(V617F), MPL(W515L), or CALR(del52) mutations. Current treatment options for MPNs include cytoreduction by hydroxyurea and JAK1/2 inhibition by ruxolitinib, both of which are not curative. We show here that cell lines expressing JAK2(V617F), MPL(W515L), or CALR(del52) accumulated reactive oxygen species-induced DNA double-strand breaks (DSBs) and were modestly sensitive to poly-ADP-ribose polymerase (PARP) inhibitors olaparib and BMN673. At the same time, primary MPN cell samples from individual patients displayed a high degree of variability in sensitivity to these drugs. Ruxolitinib inhibited 2 major DSB repair mechanisms, BRCA-mediated homologous recombination and DNA-dependent protein kinase-mediated nonhomologous end-joining, and, when combined with olaparib, caused abundant accumulation of toxic DSBs resulting in enhanced elimination of MPN primary cells, including the disease-initiating cells from the majority of patients. Moreover, the combination of BMN673, ruxolitinib, and hydroxyurea was highly effective in vivo against JAK2(V617F)+ murine MPN-like disease and also against JAK2(V617F)+, CALR(del52)+, and MPL(W515L)+ primary MPN xenografts. In conclusion, we postulate that ruxolitinib-induced deficiencies in DSB repair pathways sensitized MPN cells to synthetic lethality triggered by PARP inhibitors.

Scientific reports 5, 1-9 (2015). doi:10.1038/srep08190, Published by Nature Publishing Group, London

Summary Functional contributions of individual cellular components of the bone-marrow microenvironment to myelofibrosis (MF) in patients with myeloproliferative neoplasms (MPNs) are incompletely understood. We aimed to generate a comprehensive map of the stroma in MPNs/MFs on a single-cell level in murine models and patient samples. Our analysis revealed two distinct mesenchymal stromal cell (MSC) subsets as pro-fibrotic cells. MSCs were functionally reprogrammed in a stage-dependent manner with loss of their progenitor status and initiation of differentiation in the pre-fibrotic and acquisition of a pro-fibrotic and inflammatory phenotype in the fibrotic stage. The expression of the alarmin complex S100A8/S100A9 in MSC marked disease progression toward the fibrotic phase in murine models and in patient stroma and plasma. Tasquinimod, a small-molecule inhibiting S100A8/S100A9 signaling, significantly ameliorated the MPN phenotype and fibrosis in JAK2V617F-mutated murine models, highlighting that S100A8/S100A9 is an attractive therapeutic target in MPNs.

Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2022; Aachen : RWTH Aachen University 1 Online-Ressource : Illustrationen, Diagramme (2022). = Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2022, According to WHO, Polycythemia vera (PV) and Myelofibrosis (MF) are classified as BCR-Abl-negative myeloproliferative neoplasm (MPN). Transformation to acute myeloid leukemia and high rates of thrombosis are major complications of MPN leading to a reduced life expectancy. Previous studies have already contributed to a better understanding of the underlying hemostaseological changes, but further investigations are needed to complement these findings. The initial aim of this study was to complete the descriptive picture of altered coagulation using established and easily accessible methods and to seek for potential markers related to thrombosis risk by means of correlation analyses. Samples of Plasma from 53 patients, 28 suffering from MF, and 18 controls were analyzed and compared by rotation thrombelastometry (TEM) and thrombin generation (TG). Further influences on patients laboratory findings caused by clinical chemistry, medication, previous thrombosis and variant risk scores were evaluated. Due to the high proportion of MF patients, the focus of the study was shifted toward this condition, making it quite different from previous studies. Results in thrombin generation also confirm already known characteristics of coagulation from PV and Essential Thrombocythemia for a collective with a higher stake of MF.TEM results support statements of other studies that both Clot Formation Time and Maximum Clot Firmness are significantly increased in MPN. A prothrombotic influence of platelets has also been demonstrated. Furthermore, Hydroxyurea intake led to normalization of TEM values. In addition, LDH and Urea could be identified as potential markers to assess current thrombosis risk based on their highly significant correlations in TEM. The new and most important observation is a significantly reduced lysis especially in MF patients, which might contribute to increased risk of thrombosis. These results may provide clinical consequences to help reduce morbidity and mortality in MPN patients, e.g. by means of individually controlled anticoagulation therapy., Published by RWTH Aachen University, Aachen

Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2021; Aachen : RWTH Aachen University 1 Online-Ressource : Illustrationen, Diagramme (2021). = Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2021, Acute lymphoblastic leukemia (ALL) is a disease of lymphoid progenitor cells with an often aggressive course and is frequently driven by the BCR-ABL fusion gene t(9;22) in adults. The fusion gene results from the Philadelphia translocation t(9;22) and codes for a constitutively active tyrosine kinase which can be efficiently inhibited by Tyrosine kinase inhibitors like imatinib or dasatinib. However, BCR-ABL+ ALL cells rapidly develop mutations against these targeted agents, leading to relapse and an overall still dismal prognosis for patients with this disease. To date, allogeneic stem cell transplantation is the only known curative option. Older patients can still often not undergo this strenuous procedure because of comorbidities or frailty. In this work, we show that the combination of TKIs with proteasome inhibitors efficiently kills Ph+ ALL cells by induction of apoptosis and thereby broadens the spectrum of therapeutic options for patients with Ph+ ALL. We show the efficacy of proteasome inhibitors in cell lines generated from primary refractory cases of this disease in the study at hand. The single agent as well as combined bortezomib or ixazomib and dasatinib treatment had greater effects on cell viability in both cell lines than the dasatinib single agent treatment. Outstanding we did not observe an antagonistic effect between TKI and proteasome inhibitor in all our experiments. Proteasome inhibition as well as TKI treatment and the combination of both inhibitor classes mainly results in apoptosis in our Ph+ ALL cell lines, as shown by the results of the annexin V staining und the western blot. In addition, proteasome inhibitor treatment affects the unfolded protein response of Ph+ ALL cells and cause significant cell stress. To summarize, our results demonstrate a significant pro-apoptotic effect of proteasome inhibitors on Ph+ ALL cell lines. Treatment of BCR-ABL1 positive acute lymphoblastic leukemia with a combined therapeutic regime including the current standard therapy dasatinib together with proteasome inhibitors appears to be an encouraging approach but needs further validation in vivo and in the clinical setting., Published by RWTH Aachen University, Aachen

Dissertation, RWTH Aachen University, 2020; Aachen 1 Online-Ressource (146 Seiten) : Illustrationen, Diagramme (2020). = Dissertation, RWTH Aachen University, 2020, Classical BCR-ABL-negative myeloproliferative neoplasms (MPN) are a heterogeneous group of hematologic malignancies including polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF). The JAK2V617F mutation plays a central role in these disorders and can be found in 90% of PV and approximately 50-60% of ET and PMF. Hypoxia-inducible factor 1 (HIF-1) is a master transcriptional regulator of the response to decreases in cellular oxygen levels, also referred to as hypoxia. Within this thesis, the role of HIF-1 in JAK2V617F-positive MPN was examined. HIF-1α protein levels were significantly elevated in Jak2V617F expressing 32D cells compared to Jak2WT control cells. Pharmacological inhibition of HIF-1 binding to hypoxia response elements (HREs) with echinomycin impaired growth and survival by inducing apoptosis and cell cycle arrest in Jak2V617F transduced 32D cells, but not Jak2WT controls. Attenuation of DNA binding was verified by chromatin immunoprecipitation experiments. Findings from the pharmacologic intervention were corroborated by shRNA-mediated knockdown (KD) of HIF-1α in JAK2V617F-positive cells upon simultaneous inhibition of HIF-2, which was upregulated in HIF-1α KD cells in a compensatory manner. Echinomycin selectively abrogated growth, survival and clonogenic potential of bone marrow and peripheral blood mononuclear cells from JAK2V617F-positive patients, while cells from healthy donors were unaffected by the treatment. The efficacy was confirmed in JAK2V617F-positive iPS cell-derived progenitors from PV patients, whereas JAK2WT cells were unaffected by the treatment. HIF-1 target genes involved in modulating the energy metabolism to aerobic glycolysis were identified as a possible underlying mechanism, with increased expression of Pdk1, Glut1, Pfkfb3 and others. These findings were substantiated by transcriptome analysis of CD34+ cells from primary MPN patients. Collectively, the findings obtained during this doctoral thesis provide a greater understanding of the role of HIF-1 in JAK2V617F-positive cells and indicate that HIF-1 is a new potential therapeutic target in classical BCR-ABL negative MPN., Published by Aachen

Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2019; Aachen 1 Online-Ressource (iii, 155 Seiten) : Illustrationen, Diagramme (2019). = Dissertation, Rheinisch-Westfälische Technische Hochschule Aachen, 2019, This study focuses on the modeling of the disease progression in Myeloproliferative Neoplasms (MPNs), which are a group of chronic blood cancers with the possibility of leukemic transformation. For modeling disease progression in MPNs and by considering cancer progression as a multistep process which engages a hierarchy of different levels of functionalities, we developed hybrid models using datasets from heterogeneous sources such as gene expression, simulated data from population dynamics models and clinical laboratory results. Based on the different genetic origin of different MPNs, we divided our research into two main parts: Philadelphia positive MPNs and Philadelphia negative MPNs. In the first part and for the Philadelphia positive MPN, with the help of systems theory concepts, we developed a hybrid model by integrating data-driven models and a population-based-mechanistic model. In the second part and for the Philadelphia negative MPNs, we developed a hybrid classification model by the integration of smaller sub-models on separate subspaces of the available feature space. We showed that the use of the concept of hybrid modeling for tracking disease progression in the Philadelphia positive MPN enables patient-specific risk assessment to avoid leukemic transformation. Based on our results in the Philadelphia negative MPN part, we showed that hybrid models not only result in more accurate classification and diagnosis of MPNs but also help reduction of overfitting problem of pure data-driven models. Our developed models for MPNs classification can be improved by the integration of more data types such as omics data, which is at the current time unfortunately not available. More data on different levels can add more information about the involved mechanisms at different levels of cancer progression hierarchy., Published by Aachen