bluuebunny/crossref_metadata_2025_split huggingface dataset:
Degrasyn-Induced Trafficking of BCR-ABL as a Novel Mechanism of Kinase Inactivation.
Authors: Donato Nicholas , Kapuria Vaibhav , Sun Hanshi , Maxwell David , Bartholomeusz Geoffrey , Bornmann William , Peng Zhenghong , Pal Ashutosh , Talpaz Moshe 
Published: October 2019

Abstract
Inhibitors that inactivate specific tyrosine kinases have proven to be a very effective form of therapy of many leukemias and hematopoetic disorders. Most inhibitors function by competing for the ATP-binding pocket or by preventing association with protein substrates. However, clinical and molecular studies have shown that small changes in the structure of the target kinase (point mutations, post-translational modification) affect inhibitor binding affinities, resulting in resistance to this class of inhibitor. Therefore, development of agents that reduce the activity of leukemogenic kinases through alternate mechanisms are of great interest. We previously described a novel class of compounds, termed degrasyns, which reduced BCR-ABL (and Jak2) kinase activity through a unique mechanism. Degrasyn treatment of CML or BaF3 cells expressing wild-type or mutant (T315I) BCR-ABL resulted in a reduction of cytoplasmic BCR-ABL protein levels and loss of downstream signaling without a direct effect on BCR-ABL enzymatic activity. Biochemical studies demonstrated that degrasyn induced a rapid translocation of BCR-ABL from the cytosol to the cytoskeletal fraction (complete within 60 min) and this event correlated with loss of BCR-ABL signaling and initiation of apoptosis. A GFP-fusion protein composed of wild-type or T315I mutant BCR-ABL demonstrated that degrasyn induced high density translocation to the cytoskeletal fraction, as determined by direct fluorescence imaging. Translocation was specific for BCR-ABL and not other kinases (except Jak2) or signaling proteins. Translocation correlated with degrasyn-mediated tyrosine phosphorylation of a subset of specific proteins, including the tyrosine kinase Lyn, in the insoluble cellular fraction. CML cells expressing high levels of Lyn were more sensitive to degrasyn-mediated apoptosis suggesting that Lyn plays a role in degrasyn activity. Together these results suggest that degrasyn inactivates BCR-ABL by inducing its translocation to a cellular compartment that prevents its participation in oncogenic signaling. Degrasyn-induced BCR-ABL translocation was not effected by mutations that block dasatinib and imatinib activity and shows greater activity against some forms of imatinib resistance (Lyn overexpression). Animal studies demonstrated that degrasyn has anti-leukemic activity and functions through a unique mechanism of action. Development of inhibitors with this mode of action may be of significance for CML patients that fail therapy with ATP-binding pocket-directed tyrosine kinase inhibitors.
Upregulation of CXCR4 Is Essential for the Migration and Survival of Quiescent CML Progenitor Cells in the Bone Marrow Microenvironment after Imatinib Treatment.
Authors: Tabe Yoko , Jin Linhua , Konoplev Sergej , Hongbo Lu , Kimura Shinya , Rios Mary-Beth , Kantarjian Hagop M. , Andreeff Michael , Konopleva Marina 
Published: October 2019

Abstract
Chronic myeloid leukemia (CML) is driven by constitutively activated Bcr-Abl tyrosine kinase, which causes the defective adhesion of CML cells to bone marrow (BM) stroma. The overexpression of p210Bcr-Abl was reported to down-regulate CXCR4 expression, resulting in cell migration defects in CML. We proposed that the tyrosine kinase inhibitors imatinib or INNO-406 may restore CXCR4 expression and cause the migration of CML cells to BM microenvironment niches which results in acquisition of stroma-mediated chemoresistance of CML progenitor cells. In KBM5 and K562 cells, imatinib or INNO-406 increased CXCR4 expression and migration. Imatinib induced a G0/1 cell cycle block in CML cells, which was further enhanced in a bone marrow-derived stromal cell (MSC) co-culture system. Quiescent KBM5 cells migrating to stromal cells were resistant to Imatinib-induced cell death. These anti-apoptotic effects were abrogated by inhibition of CXCR4 (AMD3465) or Integrin-linked kinase (QLT0267). BM CD34+ cells from newly diagnosed (n=15) or blast crisis CML patients (n=5) expressed markedly lower CXCR4 levels compared with normal CD34+ BM progenitor cells (n=12) (p<0.05), and was associated with deficient migration of CD34+ CML cells. A longitudinal analysis of peripheral blood samples from CML patients in blast crisis treated with imatinib (n=5) revealed induction of CXCR4 in peripheral blood CD34+ progenitor cells within the first few days after initiation of treatment, which coincided with decreased WBC and absolute blast counts. After prolonged (3–10 day) exposure to imatinib, however, CXCR4 levels of CD34+ PB CML cells decreased, frequently below the starting levels. CXCR4 immunostaining on blasts/immature cells in BM biopsy sections from patients demonstrated that 5 of 7 patient samples were CXCR4-negative prior to imatinib therapy; 4 of these 5 became CXCR4 positive after imatinib treatment. While patients in blast-phase had short-lived hematologic responses to imatinib, Ph+ clone remained predominant in the bone marrow, and all patients ultimately relapsed with recurrence of blasts in the BM. Altogether, these findings suggest that the up-regulation of CXCR4 by imatinib promotes migration of CML cells to BM stroma, causing G0/1 cell-cycle arrest and enhancing the survival of quiescent CML progenitor cells within bone marrow microenvironment. This provides the rationale for interfering with the protective effects of BM stroma cells by inhibiting CXCR4 and/or integrin signaling, which could be of benefit in eradicating residual quiescent CML cells.
Grb10 Mediated Akt Activation Is Required for Induction of CML Like Myeloproliferative Disease in Mice by BCR-ABL.
Authors: Albers Corinna , Illert Anna L. , Miething Cornelius , Peschel Christian , Duyster Justus 
Published: October 2019

Abstract
Chronic myelogenous leukaemia (CML) results from the neoplastic transformation of hematopoietic stem cells (HSC) and is characterized by a chromosomal translocation t(9;22)(q34;q11). This aberration leads to the expression of the oncogenic tyrosine kinase BCR-ABL, which mediates signals for proliferation, transformation and anti-apoptosis via various signalling pathways. Grb10, a member of the growth factor bound proteins, is known to bind activated tyrosine kinases like BCR-ABL and might be involved in the activation of the Akt signalling pathway. Here we report the impact of Grb10 for BCR-ABL mediated transformation. We exerted a siRNA based approach in combination with a murine bone marrow transplantation model. To this end we designed a MSCV based retrovirus encoding both a Grb10 microRNA and the BCR-ABL oncogene on a single construct. This approach ensured knockdowns of more than 90% in every BCR-ABL transformed cell. Methylcellulose assays demonstrated that bone marrow coexpressing Grb10 microRNA and BCR-ABL had a 4-fold decreased colony forming ability compared to control cells. We then transduced bone marrow (BM) with retrovirus coexpressing Grb10 microRNA and p185 BCR-ABL and transplanted lethally irradiated recipient Balb/C mice. The onset and progression of leukaemia was significantly delayed in mice transplanted with Grb10 microRNA and BCR-ABL compared with the BCR-ABL transduced control microRNA group. However, we were not able to completely avoid the development of leukaemia by Grb10 knockdown. Mice transplanted with the Grb10 knockdown construct showed a delayed lymphoblastic disease, positive for B220, whereas the control group developed a rapid myeloproliferative disease, characterized by CD11b and Gr-1. In vitro analysis of BaF/3 and 32D cells showed that Grb10 knockdown in combination with BCR-ABL expression leads to a reduced phosphorylation of Akt. Taken together our data demonstrate that Grb10 is required for the development of a myeloproliferative disease by BCR-ABL in mice. Hereby, Grb10 seems to be critical for the BCR-ABL induced activation of the Akt pathway. In addition, this study describes a novel approach to express an oncogene and a microRNA using a single retroviral construct. This tool can be used to systematically screen for drugable signalling targets involved in oncogenesis.
Specific MiRNA Silencing by Lentivirus Mediated Antagomir Expression in Chronic Myeloid Leukemia (CML) Cells.
Authors: Scherr Michaela , Venturini Letizia , Battmer Karin , Schaller-Schoenitz Michael , Schaefer Daniel , Dallmann Iris , Ganser Arnold , Eder Matthias 
Published: October 2019

Abstract
Micro RNAs (miRNA) are small non-coding RNAs that regulate gene expression by specific hybridization to complementary sequences in the 3′ untranslated region of corresponding mRNAs. Concomitant recruitment of specific multi-protein complexes results either in inhibition of mRNA translation or mRNA degradation. miRNAs are processed in a regulated multi-step process from primary transcripts into mature miRNAs by cellular components which are also at least partially involved in the process of RNA interference (RNAi). Aberrant expression of specific miRNAs has recently been described in human lymphoma and leukemia. In particular, BCR-ABL and c-MYC dependent over-expression of the polycistronic and oncogenic miR-17-92 cluster (encoding miR-17, miR-18a, miR-19a, miR-20a, miR-19b, and miR-92) has been described in chronic myeloid leukemia (CML) cell lines, primary CD34+ cells from CML patients (Venturini et al. 2007), and in lung cancer. In BCR-ABL positive K562 cells, miR-17-92 encoded miRNAs repress luciferase activity in miRNA-specific reporter assays. In addition, lentivirus-mediated over-expression of miR-17-92 increases both cell proliferation and sensitivity to imatinib induced cell death. To analyse the function of individual miRNAs of the miR-17-92 polycistron, we generated lentivirus-based strategies to induce stable miRNA-specific loss- and gain-of function phenotypes for miR-18a, miR-19b, and miR-20a, respectively. Over-expression of miRNAs embedded within miR-30-derived sequences from an internal SFFV-LTR promoter allows isolation of K562 cells with increased miRNA expression. In contrast, expression of complementary oligonucleotides (antagomirs) from a H1 promoter located in the lentiviral 3′LTR can induce stable hypomorphic miRNA-phenotypes. In lentivirally transduced K562 cells, individual silencing of miR-18a, miR-19b, and miR-20a by the corresponding antagomirs (ant-miR-18a, ant-miR-19b, ant-miR-20a) specifically relieves miRNA-mediated reporter gene repression. Correspondingly, inhibition of miRNA-function correlates to reduced ‘miRNA’-amplification by miRNA-specific quantitative RT-PCR. Furthermore, protein expression of E2F-1, a known miR-20 target, is enhanced by lentivirally expressed anti-miR-20 in a dose-dependent manner, whereas over-expression of miR-20a reduces E2F-1 levels. Finally, combined over-expression of specific miRNAs and antagomirs reveals specific induction of cell proliferation by miR-18a but strong inhibition by miR-20a in K562 cells, respectively. In contrast, anti-miR-18a, but not anti-miR-19b, anti-miR-20a, or control antagomirs inhibits proliferation of K562 cells. These data demonstrate individual and complementary functions of miR-17-92 encoded miRNAs in CML and identify potential targets for specific therapeutic intervention on the miRNA level.
Cbl-b Deficiency Enhances Motility and Impairs Leukemogenesis by Bcr-Abl.
Authors: Badger-Brown Karla M. , Bailey Monica L. , Penninger Josef , Barber Dwayne L. 
Published: October 2019

Abstract
One of the prominent tyrosine phosphorylated substrates of BCR-ABL identified in cells from chronic-phase Chronic Myeloid Leukemia (CML) patients is the Cbl ubiquitin ligase and adaptor protein. Bone marrow transplantation studies have revealed that Cbl is dispensable for CML development, as animals receiving Cbl-deficient bone marrow cells expressing BCR-ABL develop a myeloproliferative disease (MPD) with a similar latency and phenotype as wild type bone marrow recipients (Dinulescu et al. 22: 8852–60, 2003). Cbl belongs to a family of E3 ubiquitin ligases that also contains Cbl-b and Cbl-3. In contrast to the equivalent Cbl protein expression displayed in hematopoietic cell lines expressing BCR-ABL, addition of the oncogene leads to a decreased expression of the related family member, Cbl-b. The objective of this study was to determine whether Cbl-b is a critical signaling intermediate downstream of BCR-ABL. To investigate this objective, we performed retroviral transduction of wild type- and Cbl-b- deficient bone marrow with BCR-ABL and utilized these cells for bone marrow transplantation of lethally irradiated mice. Recipients of wild type donor cells transduced with the p210 isoform of BCR-ABL, succumb to a rapidly fatal CML-like MPD, while the lack of Cbl-b produces a significant delay in morbidity (average latency of 32 days as compared with 18 days for recipients of wild-type marrow). Expansion of differentiated neutrophils, as visualized by histopathological analysis, is evident in the peripheral blood, bone marrow and spleen of all transplanted animals. However, circulating white blood cells are augmented in number (average wbc count for Cbl-b(+/+), 150 × 106 cells / mL; Cbl-b(−/−), 83 × 106 cells / mL) and enhanced spleen size is observed in Cbl-b(+/+) mice (35 mg / g of body mass) as compared to Cbl-b(−/−) (19 mg/g) animals. Liver masses were comparable between the two transplants. In depth flow cytometric studies of splenic cells reveals the characteristic decrease in B and T cells and enhanced myeloid lineages. Notably, an increased number of granulocytes (Gr-1+/Mac-1+ cells) is observed in the Cbl-b-knockout animals, suggesting enhanced migration but decreased survival of leukemic cells in the Cbl-b−/− spleen. In support of this hypothesis, fibroblasts deficient in Cbl-b display enhanced chemotactic migration toward Fetal Calf Serum. As migration is a key determinant of cellular retention in the bone marrow, the loss of Cbl-b could ultimately be affecting cell localization. Therefore, our data supports a continued study of Cbl-b as a motility regulator in BCR-ABL-dependent CML progression.
Growth Factor Independent 1b (Gfi1b) Is Highly Expressed in Human CML and Accelerates p210BCR-ABL Induced Leukemia in Mice.
Authors: Vassen Lothar , Tenen Daniel G. , van der Reijden Bert A. , Duehrsen Ulrich , Moroy Tarik , Mahlmann Stefan , Ebeling Peter , Jansen Joop H. 
Published: October 2019

Abstract
Chronic myeloid leukemia (CML) is one of the most frequent leukemic diseases and invariably associated with a reciprocal t(9;22) translocation which creates a juxtaposition of the BCR and ABL genes to form the p230-, p190- or p210BCR-ABL constitutively active tyrosine kinases. This constitutive tyrosine kinase activity is the cause of CML and other leukemic diseases. CML is very efficiently treated with small molecule tyrosine kinase inhibitors, but CML is a stem cell initiated disease and surprisingly the leukemic stem cells cannot be eradicated by tyrosine kinase inhibitor treatment, resulting in a high risk of the development of therapy resistance. To design more successful therapies, it is primordial to understand the molecular mechanisms underlying this disease. Here we show that growth factor independent 1b (Gfi1b), also translocated in CML, is strongly induced in peripheral blood mononuclear cells (PBMCs) in patients with chronic- or acute myeloid leukemia (AML) as well as in patients suffering from myeloprolifarative syndrome (MPS) or B-lymphoblastic leukemia (B-ALL). Gfi1b is a transcriptional repressor essential for erythroid- and megacaryocytic cells, but also expressed in hematopoietic stem cells, early myeloid precursors and mature myeloid cells in peripheral blood. Interestingly, the expression of Gfi1b is further induced by long term treatment with tyrosine kinase inhibitors like Glivec, although the patients are phenotypically in remission. Additionally we describe a new splice variant of Gfi1b which is overrepresented in CML, AML and B-ALL, but not in MPS. Using a mouse model system we demonstrate, that Gfi1b strongly accelerates p210BCR-ABL induced leukemogenesis. We further show that Gfi1b inhibits the activity of PU.1, a key molecule in hematopoiesis and repressor of leukemogenesis. Our findings bear considerable significance with regard to the role of Gfi1b as an oncogene in human leukemia and to its possible value as a new target for leukemia therapy.
Long-Term Efficacy of Dasatinib in Chronic-Phase CML: Results from the Phase I Trial (CA180002).
Authors: Cortes Jorge , Sawyers Charles L. , Kantarjian Hagop M. , Shah Neil P. , Paquette Ronald , Paliwal Prashni , Bleickardt Eric , Talpaz Moshe 
Published: October 2019

Abstract
Imatinib resistance in CML is frequently associated with BCR-ABL mutations. Such kinase domain mutations are emerging as a common mechanistic theme in cases of acquired resistance to kinase inhibitor therapy. Five-year follow-up from the IRIS trial indicates that the incidence of hematologic resistance to imatinib is 17% and >30% of patients have discontinued imatinib therapy on study. Preclinical studies reveal that dasatinib (SPRYCEL®), a novel BCR-ABL inhibitor, is 325-fold more potent than imatinib in inhibiting the kinase activity of BCR-ABL, as well as demonstrating activity against all imatinib-resistant BCR-ABL mutations tested to date, with the exception of T315I. Here we present an update of a Phase-I dose-escalation study, with enrollment between November 2003 and April 2005, of dasatinib in imatinib-resistant or -intolerant patients with CML or Ph+ ALL; this report focuses on the long-term follow-up of the 45 patients (56% male) with late chronic-phase disease. Dasatinib was administered at doses of 15–180 mg/day on a QD or BID schedule. Median duration of CML prior to entry was 8 years (range 1–17). Median age was 63 years (range 28–79). Prior therapy included interferon-α in 91% of patients and stem-cell transplantation in 4%; 62% had received prior imatinib doses >600 mg. With a minimum follow-up of 27 months, the rate of complete hematologic response (CHR) was 91% (41/45). Major cytogenetic responses (MCyR) were attained in 51% of patients (23/45), with complete CyR (CCyR) in 44% (20/45). The rates of CCyR for the QD and BID schedules were 45% (10/22) and 43% (10/23), respectively. The median duration of CHR and MCyR have not been reached with a median duration of treatment of 28 months. In a landmark analysis, the 36-month progression-free survival (PFS) rate was 87% and overall survival (OS) was 94% for patients who achieved a MCyR within the first year of dasatinib therapy. For patients who did not achieve a MCyR within the first year, the 36-month PFS figure was 28% and OS was 68%. These long-term data confirm the durability and depth of hematologic and cytogenetic response associated with dasatinib in patients who have failed treatment with imatinib. Evidence is also provided to substantiate that the significant response rates observed with dasatinib therapy in this population translate into prolonged progression-free and overall survival. Dasatinib therefore has the potential to positively impact on the natural history of CML.
Imatinib Dose Can Be Safely Reduced after Complete Cytogenetic Response (CCyR) in Patients (pts) with Chronic Myeloid Leukemia (CML) in Early Chronic Phase (CP) Treated with High-Dose Imatinib.
Authors: Jain Nitin , Kantarjian Hagop M. , Fava Carmen , Thomas Deborah , Burger Jan A. , Borthakur Gautam , Pate Odeal , Cortes Jorge 
Published: October 2019

Abstract
Background Imatinib (standard dose, SD, 400mg/d) is standard therapy in CP CML. Several observations suggest that initial therapy with high-dose (HD) imatinib might be more effective. We previously reported that HD imatinib (800mg/d) leads to significantly better rates of complete cytogenetic response (CCyR), major molecular response (MMR) and complete molecular response (CMR), and improved event-free survival compared to SD. However, HD imatinib is associated with more frequent grade 3–4 hematological toxicity compared to SD. In addition, 36% pts required dose reduction due to toxicities. We investigated the outcome of pts who were maintained on 800mg/d compared to those whose dose was reduced from 800mg/d.
Methods Pts enrolled in 3 sequential trials using imatinib for frontline therapy for CP CML were analyzed: 208 treated with HD and 50 with SD. Primary end points were event-free survival (EFS) and transformation-free survival (TFS).
Results The median age was 48 years (range 17–84 years). Median follow up was 49 months. 4 year event-free survival (EFS) and transformation-free survival (TFS) were significantly better for pts treated with HD imatinib compared to SD (EFS: 92% vs. 77%, p=0.01; TFS: 98% vs. 89%, p=0.0045). Among pts treated with HD, 100 (48%) had a dose reduction at any time [600mg/d (n=50), 400mg/d (n=38), 300mg/d (n=12)]. Of them, 45 (45%) were dose-reduced after achieving CCyR including 26 (26%) after MMR. The median time to dose reduction was 3 months (range 1–60 months). For pts starting with HD imatinib, there was no difference between pts continuing with HD and those with a dose reduction in terms of EFS at 4 years (92% in both groups, p=0.65) and TFS at 4 years (98% vs. 97%, respectively, p=0.36). However, EFS at 4 years was significantly worse if the dose reduction was done prior to achievement of CCyR (84% vs. 100%, p=0.02). Similarly, patients who were dose-reduced before MMR had significantly worse 4 year EFS (88% vs. 100%, p=0.02). EFS curve for patients who had dose reduction done prior to CCyR was similar to that of pts treated with SD imatinib (p=0.59).
Conclusions These results indicate that pts treated with HD imatinib can be safely dose reduced once they have achieved CCyR (preferably MMR) without having any impact on EFS or TFS. Such dose reduction may help decrease side effects as well as cost of the treatment. A prospective evaluation of this approach is warranted.