Notch signaling is involved in tumorigenesis, but its role in B–chronic lymphocytic leukemia (B-CLL) pathogenesis is not completely defined. This study examined the expression and activation of Notch ...receptors in B-CLL cells and the role of Notch signaling in sustaining the survival of these cells. Our results show that B-CLL cells but not normal B cells constitutively express Notch1 and Notch2 proteins as well as their ligands Jagged1 and Jagged2. Notch signaling is constitutively activated in B-CLL cells, and its activation is further increased in B-CLL cells, which resist spontaneous apoptosis after 24-hour ex vivo culture. Notch stimulation by a soluble Jagged1 ligand increases B-CLL cell survival and is accompanied by increased nuclear factor–kappa B (NF-κB) activity and cellular inhibitor of apoptosis protein 2 (c-IAP2) and X-linked inhibitor of apoptosis protein (XIAP) expression. In contrast, Notch-signaling inhibition by the γ-secretase inhibitor I (GSI; z-Leu-Leu-Nle-CHO) and the specific Notch2 down-regulation by small-interfering RNA accelerate spontaneous B-CLL cell apoptosis. Apoptotic activity of GSI is accompanied by reduction of NF-κB activity and c-IAP2 and XIAP expression. Overall, our findings show that Notch signaling plays a critical role in B-CLL cell survival and apoptosis resistance and suggest that it could be a novel potential therapeutic target.
Establishment of hematopoietic stem-cell (HSC) transplantation from mismatched relatives is feasible for patients with acute leukemia. As our original method of graft processing was unsuitable for ...large-scale clinical studies, we use automated devices for CD34+ cell purification.
Sixty-seven patients with acute myeloid leukemia (AML; 19 complete remission CR 1, 14 CR 2, nine CR > 2, 25 in relapse) and 37 with acute lymphoid leukemia (ALL; 14 CR 1, eight CR 2, two CR > 2, 13 in relapse) were conditioned with total-body irradiation, thiotepa, fludarabine, and antithymocyte globulin. Peripheral-blood progenitor cells were mobilized with recombinant human granulocyte colony-stimulating factor and depleted of T-cells using CD34+ cell immunoselection. No post-transplantation graft-versus-host disease (GvHD) prophylaxis was administered.
Primary engraftment was achieved in 94 of 101 assessable patients. Six of the seven patients who rejected the primary graft, engrafted after a second transplantation. Overall, 100 of 101 patients engrafted. Acute GvHD developed in eight of 100 patients, and chronic GvHD, in five of 70 assessable patients. Thirty-eight patients died of nonleukemic causes. Relapse occurred in nine of 66 patients receiving transplantation in remission and in 17 of 38 receiving transplantation in relapse. Median follow-up of the 40 patients who survived event-free was 22 months (range, 1 to 65 months). Event-free survival (+/- standard deviation) rate was 48% +/- 8% and 46% +/- 10%, respectively, for the 42 AML and 24 ALL patients receiving transplantation in remission.
Our transplantation procedure provides reliable, reproducible CD34+ cell purification, high engraftment rates, and prevention of GvHD. The mismatched-related transplant emerges as a viable, alternative source of stem cells for acute leukemia patients without matched donors and/or those who urgently need transplantation.
The proteasome inhibitor bortezomib may increase osteoblast-related markers in multiple myeloma (MM) patients; however, its potential osteoblastic stimulatory effect is not known. In this study, we ...show that bortezomib significantly induced a stimulatory effect on osteoblast markers in human mesenchymal cells without affecting the number of osteoblast progenitors in bone marrow cultures or the viability of mature osteoblasts. Consistently we found that bortezomib significantly increased the transcription factor Runx2/Cbfa1 activity in human osteoblast progenitors and osteoblasts without affecting nuclear and cytoplasmatic active β-catenin levels. Consequently a stimulatory effect of bortezomib on bone nodule formation was also demonstrated in osteoblast progenitors. These in vitro observations were confirmed in vivo by the finding of a significant increase in the number of osteoblastic cells × mm2 of bone tissue and in the number of Runx2/Cbfa1-positive osteoblastic cells that was observed in MM patients who responded to bortezomib. Our in vitro and in vivo observations support the hypothesis that a direct stimulatory effect on bone formation process could occur during bortezomib treatment.
The GIMEMA ALL 0288 trial was designed to evaluate the impact of a 7-day prednisone (PDN) pretreatment on complete remission (CR) achievement and length, the influence of the addition of ...cyclophosphamide (random I) to a conventional 4-drug induction on CR rate and duration, and whether an early post-CR intensification (random II) by an 8-drug consolidation could improve CR duration. Median follow-up of this study was 7.3 years. From January 1988 to April 1994, among 794 adult (> 12 but < 60 years) patients registered, 778 were eligible. Their median age was 27.5 years; 73% had B-lineage acute lymphoblastic leukemia (ALL) and 22% had T-lineage disease; 18% showed associated myeloid markers; 47 of 216 analyzed patients (22%) had Philadelphia chromosome–positive ALL. Response to PDN pretreatment was observed in 65% of cases. CR was achieved in 627 patients (82%). Resistant patients and induction death rates were 11% and 7%, respectively. Random II was applied to 388 patients with CR; 201 had maintenance alone and 187 had consolidation followed by maintenance. The relapse rate was 60%; isolated central nervous system relapses were 8% of all CRs and 13% of all relapses. Median survival (overall survival OS), continuous complete remission (CCR), and disease-free survival (DFS) were 2.2, 2.4, and 2 years, respectively. PDN pretreatment response resulted the main independent factor influencing CR achievement, OS, CCR, and DFS; the addition of cyclophosphamide in induction significantly influenced CR achievement in a multivariate analysis. Neither induction intensification nor early consolidation appeared to influence CCR and DFS duration. For the first time PDN pretreatment response proved to be a powerful factor predicting disease outcome in adult ALL patients.
The hypoxia-inducible factor (HIF) transcriptional system enables cell adaptation to limited O
2 availability, transducing this signal into patho-physiological responses such as angiogenesis, ...erythropoiesis, vasomotor control, and altered energy metabolism, as well as cell survival decisions. However, other factors beyond hypoxia are known to activate this pleiotropic transcription factor. The aim of this study was to characterize HIF in human hematopoietic stem cells (HSCs) and evidence is provided that granulocyte colony stimulating factor-mobilized CD34+- and CD133+-HSCs express a stabilized cytoplasmic form of HIF-1α under normoxic conditions. It is shown that HIF-1α stabilization correlates with down-regulation of the tumour suppressor von Hippel-Lindau protein (pVHL) and is positively controlled by NADPH-oxidase-dependent production of reactive oxygen species, indicating a specific O
2-independent post-transcriptional control of HIF in mobilized HSCs. This novel finding is discussed in the context of the proposed role of HIF as a mediator of progenitor cell recruitment to injured ischemic tissues and/or in the control of the maintenance of the undifferentiated state.
Purpose: Vasculogenesis is a physiologic process typical of fetal development in which new blood vessels develop from undifferentiated
precursors (or angioblasts). In tumors, near angiogenesis, ...vasculogenesis contributes to the formation of the microvascular
plexus that is important for diffusion. Here, we show that hematopoietic stem and progenitor cells (HSPC) of multiple myeloma
(MM) patients are able to differentiate into cells with endothelial phenotype on exposure to angiogenic cytokines.
Experimental Design: Circulating HSPCs were purified with an anti-CD133 antibody from patients with newly diagnosed MM before autologous transplantation
and exposed to vascular endothelial growth factor (VEGF), fibroblast growth factor-2 and insulin-like growth factor in a 3-week
culture.
Results: HSPCs gradually lost CD133 expression and acquired VEGF receptor-2, factor VIII–related antigen, and vascular endothelial-cadherin
expression. The expression pattern overlapped with paired MM endothelial cells (MMEC). During culture, cells adhered to fibronectin,
spread, and acquired an endothelial cell shape. Differentiated HSPCs also became capillarogenic in the Matrigel assay with
maximal activity at the third week of culture. Bone marrow biopsies revealed HSPCs inside the neovessel wall in patients with
MM but not in those with monoclonal gammopathy of undetermined significance.
Conclusions: In patients with MM, but not in those with monoclonal gammopathy of undetermined significance, HSPCs contribute to the neovessel
wall building together with MMECs. Therefore, besides angiogenesis, HSPC-linked vasculogenesis contributes to neovascularization
in MM patients. Tentatively, we hypothesize that in HSPC cultures a multipotent cell population expressing low VEGF receptor-2
levels corresponds to the endothelial progenitor cell precursor and seems to be the MMEC precursor.
Stem cell–dose escalation is one way to overcome immune rejection of incompatible stem cells. However, the number of hematopoietic precursors required for overcoming the immune barrier in recipients ...pretreated with sublethal regimens cannot be attained with the state-of-the-art technology for stem cell mobilization. This issue was addressed by the observation that cells within the human CD34+ population are endowed with veto activity. In the current study, we demonstrated that it is possible to harvest about 28- to 80-fold more veto cells on culturing of purified CD34+cells for 7 to 12 days with an early-acting cytokine mixture including Flt3-ligand, stem cell factor, and thrombopoietin. Analysis of the expanded cells with fluorescence-activated cell-sorter scanning revealed that the predominant phenotype of CD34+CD33− cells used at the initiation of the culture was replaced at the end of the culture by cells expressing early myeloid phenotypes such as CD34+CD33+ and CD34−CD33+. These maturation events were associated with a significant gain in veto activity as exemplified by the minimal ratio of veto to effector cells at which significant veto activity was detected. Thus, whereas purified unexpanded CD34+ cells exhibited veto activity at a veto-to-effector cell ratio of 0.5, the expanded cells attained an equivalent activity at a ratio of 0.125. The availability of novel sources of veto cells such as those in this study might contribute to the realization of immunologic tolerance in “minitransplants,” without any risk of graft-versus-host disease.
Primary myelofibrosis (PMF) is a chronic myeloproliferative neoplasm characterized by progressive anemia, massive splenomegaly, leukoerythroblastosis, extramedullary hematopoiesis and in about 50% of ...cases the presence of
JAK2V617F
mutation. Curative therapy in PMF is currently possible only with allogeneic haematopoietic stem cell transplantation which is, unfortunately, associated with relatively high risks of mortality and morbidity which undermine its broad applications. Non-transplant treatment modalities are used for palliative purposes. Recently, anti-angiogenic drugs such as thalidomide have been used to treat these patients on the basis of the prominent bone marrow angiogenesis. Here, we report the case of a patient suffering from
JAK2V617F
-positive PMF with marked bone marrow neo-angiogenesis. The patient was treated with thalidomide but after 20 days developed life-threatening toxic epidermal necrolysis (TEN). To the best of our knowledge this is the first case of TEN in a patient with PMF under thalidomide therapy.
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