Dipeptidylpeptidase IV (CD26) defines leukemic stem cells (LSC) in chronic myeloid leukemia

Chronic myeloid leukemia (CML) is a stem cell (SC) neoplasm characterized by the BCR/ABL1 oncogene. Although mechanisms of BCR/ABL1-induced transformation are well-defined, little is known about effector-molecules contributing to malignant expansion and the extramedullary spread of leukemic SC (LSC) in CML. We have identified the cytokine-targeting surface enzyme dipeptidylpeptidase-IV (DPPIV/CD26) as a novel, specific and pathogenetically relevant biomarker of CD34+/CD38─ CML LSC. In functional assays, CD26 was identified as target enzyme disrupting the SDF-1-CXCR4-axis by cleaving SDF-1, a chemotaxin recruiting CXCR4+ SC. CD26 was not detected on normal SC or LSC in other hematopoietic malignancies. Correspondingly, CD26+ LSC decreased to low or undetectable levels during successful treatment with imatinib. CD26+ CML LSC engrafted NOD-SCID-IL-2Rγ−/− (NSG) mice with BCR/ABL1+ cells, whereas CD26─ SC from the same patients produced multilineage BCR/ABL1– engraftment. Finally, targeting of CD26 by gliptins suppressed the expansion of BCR/ABL1+ cells. Together, CD26 is a new biomarker and target of CML LSC. CD26 expression may explain the abnormal extramedullary spread of CML LSC, and inhibition of CD26 may revert abnormal LSC function and support curative treatment approaches in this malignancy

Mast cells as a unique hematopoietic lineage and cell system:From Paul Ehrlich's visions to precision medicine concepts

The origin and functions of mast cells (MCs) have been debated since their description by Paul Ehrlich in 1879. MCs have long been considered 'reactive bystanders' and 'amplifiers' in inflammatory processes, allergic reactions, and host responses to infectious diseases. However, knowledge about the origin, phenotypes and functions of MCs has increased substantially over the past 50 years. MCs are now known to be derived from multipotent hematopoietic progenitors, which, through a process of differentiation and maturation, form a unique hematopoietic lineage residing in multiple organs. In particular, MCs are distinguishable from basophils and other hematopoietic cells by their unique phenotype, origin(s), and spectrum of functions, both in innate and adaptive immune responses and in other settings. The concept of a unique MC lineage is further supported by the development of a distinct group of neoplasms, collectively referred to as mastocytosis, in which MC precursors expand as clonal cells. The clinical consequences of the expansion and/or activation of MCs are best established in mastocytosis and in allergic inflammation. However, MCs have also been implicated as important participants in a number of additional pathologic conditions and physiological processes. In this article, we review concepts regarding MC development, factors controlling MC expansion and activation, and some of the fundamental roles MCs may play in both health and disease. We also discuss new concepts for suppressing MC expansion and/or activation using molecularly-targeted drugs

Novel markers of leukemic stem cells (LSC) in chronic myeloid leukemia (CML)

Die chronisch myeloische Leukämie (CML) ist eine Stammzellerkrankung, die durch die Expression des Philadelphia Chromosoms (Ph) und dem entsprechenden Onkoprotein, BCR-ABL1 gekennzeichnet ist. Die Ph+ CML ist durch eine Akkumulation von unreifen und reifen myeloischen Zellen im peripheren Blut (PB) und Knochenmark (KM) charakterisiert. Reifere myeloische Zellen im CML Klon haben nur eine limitierte Teilungsfähigkeit und sind, ähnlich wie normale myeloische Zellen, dazu programmiert in den gerichteten Zelltod (Apoptose) überzugehen. Eine kleine Fraktion der CML Zellen, die sogenannten CML Stammzellen, besitzt jedoch die Eigenschaft zur Selbsterneuerung (self-renewal) und propagiert somit die unlimitierte Expansion der Erkrankung. Bis heute ist wenig über den Phänotyp, die Funktion und das Target-Expressionsmuster von leukämischen Stammzellen (LSZ) in der CML bekannt. Während meiner Doktorarbeit habe ich den Phänotyp und die funktionellen Eigenschaften von CML LSZ untersucht und mich dabei speziell auf neue diagnostische Marker und potentielle Targets konzentriert. In diesen Studien konnte ich zeigen, dass CML LSZ die Interleukin-2 Rezeptor Alpha Kette und das Zytokin-degradierende Zelloberflächen-Enzym Dipeptidylpeptidase IV (DPPIV=CD26) aberrant exprimieren. In fast allen Patienten mit CML, die im Zuge dieser Arbeit getestet wurden, konnte die Expression beider Marker bestätigt werden. CD25 konnte auch auf LSZ in Patienten mit akuter myeloischer Leukämie (AML) detektiert werden, während CD26 als spezifischer Marker der CML LSZ fungiert, was unserer Meinung nach von diagnostischer Signifikanz ist. Funktionell wurde CD25 als ein negativer Wachstumsregulator auf CML LSZ identifiziert, während CD26 den CXCR4-Liganden Stromal Cell-Derived Factor-1 (SDF-1) enzymatisch degradiert und somit die Interaktion zwischen LZS und der Stammzell-Nische im Knochenmark unterbricht, was die hochregulierte extramedulläre Expansion der Stamm- und Progenitorzellen in der CML erklären könnte. Im Zuge meiner Arbeit haben wir noch mehrere andere Marker und Targets, wie zum Beispiel CD44 und CD52 auf den CML LSZ identifiziert. Die phänotypische Charakterisierung der CML LSZ sollte dazu beitragen, diese Zellen besser quantifizieren und besser anreichern zu können, was sowohl in der Praxis als auch in der Forschung nützlich sein könnte. Außerdem sollten meine Analysen dazu beitragen, eine diagnostische Phänotypisierung der CML LSZ zu etablieren und neue LSZ-eradizierende Behandlungsstrategien in der CML zu entwickeln.Chronic myeloid leukemia (CML) is a stem cell-derived leukemia in which neoplastic cells exhibit the Philadelphia (Ph) chromosome and the related oncoprotein, BCR-ABL1. Ph+ CML is characterized by an accumulation of immature and mature myeloid progenitor cells in the peripheral blood (PB) and bone marrow (BM). Whereas the more mature myeloid cells in the CML clone have only a limited capacity to divide and are programmed to undergo apoptosis in a similar way compared to normal myeloid cells, a small fraction of neoplastic cells in the CML clone are considered to have self-renewal and thus long-term disease-propagating ability. However, so far, little is known about the phenotype, function, and target expression-profiles of these leukemic stem cells (LSC) in patients with CML. During my doctoral thesis, I have examined phenotypic and functional properties of CML LSC, with special focus on new diagnostic markers and potential targets of therapy. In these studies, I found that CML LSC aberrantly display the interleukin-2 receptor alpha chain CD25 and the cytokine-targeting cell surface enzyme dipeptidylpeptidase IV (DPPIV=CD26). Both markers were found to be expressed on LSC in almost all CML patients tested. However, whereas CD25 was also detectable in LSC in patients with acute myeloid leukemia (AML), CD26 was identified as a specific marker of CML LSC, which may be of diagnostic significance. Functionally, CD25 was identified as a negative growth regulator of CML (stem) cells, whereas CD26 was identified as a cytokine-targeting LSC antigen that disrupts LSC-niche interactions by degrading the CXCR4 ligand stromal cell derived factor-1 (SDF-1), which may explain the massive extramedullary spread of myeloid stem- and progenitor cells in CML. During my thesis, we also identified a number of additional markers and targets on CML LSC, such as CD44 and CD52. The characterization of CML LSC should assist in their identification and enumeration in daily practice and in CML research, and should contribute to the development of diagnostic LSC phenotyping and of new LSC-eradicating treatment strategies in CML.submitted by Mag. rer. Irina SadovnikZusammenfassung in deutscher SpracheMedizinische Universität Wien, Dissertation, 2016OeB

Eosinophils and eosinophil-associated disorders: immunological, clinical, and molecular complexity.

Eosinophils and their mediators play a crucial role in various reactive states such as bacterial and viral infections, chronic inflammatory disorders, and certain hematologic malignancies. Depending on the underlying pathology, molecular defect(s), and the cytokine- and mediator-cascades involved, peripheral blood and tissue hypereosinophilia (HE) may develop and may lead to organ dysfunction or even organ damage which usually leads to the diagnosis of a HE syndrome (HES). In some of these patients, the etiology and impact of HE remain unclear. These patients are diagnosed with idiopathic HE. In other patients, HES is diagnosed but the etiology remains unknown - these patients are classified as idiopathic HES. For patients with HES, early therapeutic application of agents reducing eosinophil counts is usually effective in avoiding irreversible organ damage. Therefore, it is important to systematically explore various diagnostic markers and to correctly identify the disease elicitors and etiology. Depending on the presence and type of underlying disease, HES are classified into primary (clonal) HES, reactive HES, and idiopathic HES. In most of these patients, effective therapies can be administered. The current article provides an overview of the pathogenesis of eosinophil-associated disorders, with special emphasis on the molecular, immunological, and clinical complexity of HE and HES. In addition, diagnostic criteria and the classification of eosinophil disorders are reviewed in light of new developments in the field

What are the challenges in 2016 regarding resistance to tyrosine kinase inhibitors in chronic myeloid leukemia and cancer?

In the past decade, the treatment of chronic myeloid leukemia (CML) has undergone a drastic evolution. The discovery and success of imatinib and second-generation tyrosine kinase inhibitors have substantially increased the outcome for CML patients. The next step in medical and scientific research is to better understand the malignancy so as to eventually find a cure to eliminate all leukemic cells from patients. One of the key issues is about the resistance of the leukemic stem cells to tyrosine kinase inhibitors. Here, we briefly describe our current studies on CML resistance, and leukemic stem cell modeling and characterization

Immunotherapy-Based Targeting and Elimination of Leukemic Stem Cells in AML and CML

The concept of leukemic stem cells (LSC) has been developed with the idea to explain the clonal hierarchies and architectures in leukemia, and the more or less curative anti-neoplastic effects of various targeted drugs. It is now widely accepted that curative therapies must have the potential to eliminate or completely suppress LSC, as only these cells can restore and propagate the malignancy for unlimited time periods. Since LSC represent a minor cell fraction in the leukemic clone, little is known about their properties and target expression profiles. Over the past few years, several cell-specific immunotherapy concepts have been developed, including new generations of cell-targeting antibodies, antibody–toxin conjugates, bispecific antibodies, and CAR-T cell-based strategies. Whereas such concepts have been translated and may improve outcomes of therapy in certain lymphoid neoplasms and a few other malignancies, only little is known about immunological targets that are clinically relevant and can be employed to establish such therapies in myeloid neoplasms. In the current article, we provide an overview of the immunologically relevant molecular targets expressed on LSC in patients with acute myeloid leukemia (AML) and chronic myeloid leukemia (CML). In addition, we discuss the current status of antibody-based therapies in these malignancies, their mode of action, and successful examples from the field

Identification of Ponatinib as a potent inhibitor of growth, migration and activation of neoplastic eosinophils carrying FIP1L1-PDGFRA

In chronic eosinophilic leukemia (CEL), the transforming oncoprotein FIP1L1-PDGFRA is a major target of therapy. In most patients, the tyrosine kinase inhibitor (TKI) imatinib induces complete remission. For patients who are intolerant or resistant, novel TKI have been proposed. We examined the in vitro effects of 14 kinase blockers on growth and function of EOL-1 cells, a FIP1L1-PDGFRA+ eosinophil cell line. Major growth-inhibitory effects were seen with all PDGFR-blocking agents, with IC50 values in the low nM-range: ponatinib: 0.1-0.2 nM, sorafenib: 0.1-0.2 nM, masitinib: 0.2-0.5 nM, nilotinib: 0.2-1 nM, dasatinib: 0.5-2 nM, sunitinib: 1-2 nM, midostaurin: 5-10 nM. These drugs were also found to block activation of PDGFR-downstream signaling molecules, including Akt, S6, and STAT5 in EOL-1 cells. All effective TKI produced apoptosis in EOL-1 cells as determined by microscopy, Annexin-V/PI, and caspase-3-staining. In addition, PDGFR-targeting TKI were found to inhibit cytokine-induced migration of EOL-1 cells. In all bioassays employed, ponatinib was found to be the most potent compound in EOL-1 cells. In addition, ponatinib was found to downregulate expression of the activation-linked surface antigen CD63 on EOL-1 cells, and to suppress growth of primary neoplastic eosinophils. We also examined drug effects on Ba/F3 cells expressing two clinically relevant imatinib-resistant mutant-forms of FIP1L1-PDGFRA, namely T674I and D842V. Strong inhibitory effects on both mutants were only seen with ponatinib. In summary, novel PDGFR-targeting TKI may be alternative agents for the treatment of patients with imatinib-resistant CEL. Although several different PDGFR-targeting agents are effective, the most potent drug appears to be ponatinib.status: publishe

Journal of Hematology & Oncology / Cancer stem cells in basic science and in translational oncology : can we translate into clinical application?

Since their description and identification in leukemias and solid tumors, cancer stem cells (CSC) have been the subject of intensive research in translational oncology. Indeed, recent advances have led to the identification of CSC markers, CSC targets, and the preclinical and clinical evaluation of the CSC-eradicating (curative) potential of various drugs. However, although diverse CSC markers and targets have been identified, several questions remain, such as the origin and evolution of CSC, mechanisms underlying resistance of CSC against various targeted drugs, and the biochemical basis and function of stroma cell-CSC interactions in the so-called ‘stem cell niche. Additional aspects that have to be taken into account when considering CSC elimination as primary treatment-goal are the genomic plasticity and extensive subclone formation of CSC. Notably, various cell fractions with different combinations of molecular aberrations and varying proliferative potential may display CSC function in a given neoplasm, and the related molecular complexity of the genome in CSC subsets is considered to contribute essentially to disease evolution and acquired drug resistance. In the current article, we discuss new developments in the field of CSC research and whether these new concepts can be exploited in clinical practice in the future.(VLID)486718

Identification of CD203c as a New Basophil-Specific Flow-Marker in Ph⁺ Chronic Myeloid Leukemia

Basophilia is a crucial prognostic variable in Ph-chromosome-positive chronic myeloid leukemia (CML). The ectoenzyme CD203c is an activation-linked surface antigen that is expressed specifically on basophil-committed progenitor cells and mature basophils. We examined the expression of CD203c on progenitors and/or basophils in 21 healthy donors and 44 patients with CML. As expected, the numbers of CD203c+ blood leukocytes were significantly higher in CML patients compared to controls (percentage of CD203c+ cells among viable cells in CML at diagnosis: 4.19 ± 3.68% vs. controls: 0.53 ± 0.23%, p + cells at diagnosis correlate with the disease-related risk-profile. Incubation of CML basophils with an anti-IgE-antibody resulted in further upregulation of CD203c. After successful treatment with imatinib and/or other BCR::ABL1 inhibitors leading to major or complete molecular responses, the numbers of CD203c+ basophils decreased substantially in our CML patients compared to pre-treatment values. Together, CD203c is overexpressed on CML basophils, is further upregulated by IgE receptor cross-linking, and may serve as a biomarker to quantify basophilia in patients with CML at diagnosis and during therapy