RESUMO
Cell cycle quiescence is a critical feature contributing to haematopoietic stem cell (HSC) maintenance. Although various candidate stromal cells have been identified as potential HSC niches, the spatial localization of quiescent HSCs in the bone marrow remains unclear. Here, using a novel approach that combines whole-mount confocal immunofluorescence imaging techniques and computational modelling to analyse significant three-dimensional associations in the mouse bone marrow among vascular structures, stromal cells and HSCs, we show that quiescent HSCs associate specifically with small arterioles that are preferentially found in endosteal bone marrow. These arterioles are ensheathed exclusively by rare NG2 (also known as CSPG4)(+) pericytes, distinct from sinusoid-associated leptin receptor (LEPR)(+) cells. Pharmacological or genetic activation of the HSC cell cycle alters the distribution of HSCs from NG2(+) periarteriolar niches to LEPR(+) perisinusoidal niches. Conditional depletion of NG2(+) cells induces HSC cycling and reduces functional long-term repopulating HSCs in the bone marrow. These results thus indicate that arteriolar niches are indispensable for maintaining HSC quiescence.
Assuntos
Arteríolas/citologia , Células-Tronco Hematopoéticas/citologia , Nicho de Células-Tronco , Animais , Medula Óssea/irrigação sanguínea , Divisão Celular , Separação Celular , Feminino , Citometria de Fluxo , Células-Tronco Hematopoéticas/metabolismo , Masculino , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Endogâmicos C57BL , Nestina/metabolismoRESUMO
Bcl-2/IgH rearrangements can be quantified in follicular lymphoma (FL) from peripheral blood (PB) by polymerase chain reaction (PCR). The prognostic value of Bcl-2/IgH levels in FL remains controversial. We therefore prospectively studied PB Bcl-2/IgH levels from 173 first-line FL patients who were consecutively enrolled, randomized, and treated within the multicenter phase 3 clinical trial NHL1-2003 comparing bendamustine-rituximab (B-R) with rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone. From April 2005 to August 2008, 783 pre- and posttreatment PB samples were quantified by quantitative PCR. At inclusion, 114 patients (66%) tested positive and 59 (34%) were negative for Bcl-2/IgH. High pretreatment Bcl-2/IgH levels had an adverse effect on progression-free survival (PFS) compared with intermediate or low levels (high vs intermediate: hazard [HR], 4.28; 95% confidence interval [CI], 1.70-10.77; P = .002; high vs low: HR, 3.02; 95% CI, 1.55-5.86; P = .001). No PFS difference between treatment arms was observed in Bcl-2/IgH-positive patients. A positive posttreatment Bcl-2/IgH status was associated with shorter PFS (8.7 months vs not reached; HR, 3.15; 95% CI, 1.51-6.58; P = .002). By multivariate analysis, the pretreatment Bcl-2/IgH level was the strongest predictor for PFS. Our data suggest that pre- and posttreatment Bcl-2/IgH levels from PB have significant prognostic value for PFS in FL patients receiving first-line immunochemotherapy. This trial was registered at www.clinicaltrials.gov as #NCT00991211 and at the German Federal Institute for Drugs and Medical Devices as #BfArM-4021335.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Linfoma Folicular/sangue , Linfoma Folicular/tratamento farmacológico , Proteínas de Fusão Oncogênica/sangue , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Cloridrato de Bendamustina/administração & dosagem , Cloridrato de Bendamustina/uso terapêutico , Ciclofosfamida/administração & dosagem , Ciclofosfamida/uso terapêutico , Intervalo Livre de Doença , Doxorrubicina/administração & dosagem , Doxorrubicina/uso terapêutico , Feminino , Seguimentos , Humanos , Estimativa de Kaplan-Meier , Linfoma Folicular/diagnóstico , Linfoma Folicular/genética , Masculino , Pessoa de Meia-Idade , Prednisona/administração & dosagem , Prednisona/uso terapêutico , Prognóstico , Rituximab/administração & dosagem , Rituximab/uso terapêutico , Vincristina/administração & dosagem , Vincristina/uso terapêuticoRESUMO
New drugs for the treatment of multiple myeloma (MM) comprise immunomodulatory substances such as lenalidomide and related compounds. While lenalidomide has found its way into first-line treatment as well as into relapse therapy, little is known about lenalidomide effects on normal hematopoietic stem and progenitor cells (HSPCs). In this study, we investigated whether HSPCs are influenced by lenalidomide on a phenotypic, functional and gene expression level. For that purpose, samples from patients with MM were obtained who underwent equivalent first-line treatment including induction therapy, cytotoxic stem cell mobilization and high-dose melphalan therapy followed by autologous blood stem cell transplantation and a subsequent uniform lenalidomide consolidation treatment within a prospective clinical trial. We found that after six months of lenalidomide therapy, the number of CD34(+) HSPCs decreased. Additionally, lenalidomide affects the numerical composition of hematopoietic cells in the bone marrow while it does not affect long-term HSPC proliferation in vitro. We found a significant amplification of fetal hemoglobin (HbF) expression on a transcriptional level and can confirm a stimulated erythropoiesis on a phenotypic level. These effects were accompanied by silencing of the TGF-ß signaling pathway on the gene expression and protein level that is known to be amplified in active MM. However, these pleiotropic effects gave no evidence for mutagenic potential. In conclusion, lenalidomide does not exert long-term effects on proliferation of HSPCs but instead promotes erythropoiesis by shifting hemoglobin expression toward HbF and by silencing the TGF-ß signaling pathway.
Assuntos
Eritropoese/efeitos dos fármacos , Mieloma Múltiplo/sangue , Mieloma Múltiplo/tratamento farmacológico , Mielopoese/efeitos dos fármacos , Talidomida/análogos & derivados , Inibidores da Angiogênese/uso terapêutico , Medula Óssea/efeitos dos fármacos , Quimioterapia de Consolidação , Hemoglobina Fetal/metabolismo , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Lenalidomida , Pessoa de Meia-Idade , Talidomida/uso terapêuticoRESUMO
Bleeding complications are a significant clinical problem in patients with myelodysplastic syndromes even at sufficient platelet counts (>50,000/µl). However, the underlying pathology of this hemorrhagic diathesis is still unknown. Here, we analyzed the platelet proteome of patients with myelodysplastic syndromes by quantitative two-dimensional difference gel electrophoresis followed by mass spectrometric protein identification. Proteins identified with lower concentrations, such as Talin-1, Vinculin, Myosin-9, Filmain-A, and Actin play critical roles in integrin αIIbß3 signaling and thus platelet aggregation. Despite normal agonist receptor expression, calcium flux, and granule release upon activation, the activation capacity of integrin αIIbß3 was diminished in myelodysplastic syndrome platelets. Förster resonance energy transfer analysis showed a reduced co-localization of Talin-1 to the integrin's ß3-subunit, which is required for receptor activation and fibrinogen binding. In addition, platelet spreading on immobilized fibrinogen was incomplete, and platelet aggregation assays confirmed a general defect in integrin-dependent platelet aggregation in patients with myelodysplastic syndromes. Our data provide novel aspects on the molecular pathology of impaired platelet function in myelodysplastic syndromes and suggest a mechanism of defective integrin αIIbß3 signaling that may contribute to the hemorrhagic diathesis observed in these patients.
Assuntos
Plaquetas/metabolismo , Integrinas/fisiologia , Síndromes Mielodisplásicas/fisiopatologia , Agregação Plaquetária , Proteoma/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Ácido Araquidônico/fisiologia , Adesão Celular , Células Cultivadas , Colágeno/fisiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/metabolismo , Mapas de Interação de Proteínas , Receptores de Superfície Celular/metabolismo , Transdução de Sinais , Adulto JovemRESUMO
Knockdown of the transcription factor PU.1 (encoded by Sfpi1) leads to acute myeloid leukemia (AML) in mice. We examined the transcriptome of preleukemic hematopoietic stem cells (HSCs) in which PU.1 was knocked down (referred to as 'PU.1-knockdown HSCs') to identify transcriptional changes preceding malignant transformation. Transcription factors c-Jun and JunB were among the top-downregulated targets. Restoration of c-Jun expression in preleukemic cells rescued the PU.1 knockdown-initiated myelomonocytic differentiation block. Lentiviral restoration of JunB at the leukemic stage led to loss of leukemic self-renewal capacity and prevented leukemia in NOD-SCID mice into which leukemic PU.1-knockdown cells were transplanted. Examination of human individuals with AML confirmed the correlation between PU.1 and JunB downregulation. These results delineate a transcriptional pattern that precedes leukemic transformation in PU.1-knockdown HSCs and demonstrate that decreased levels of c-Jun and JunB contribute to the development of PU.1 knockdown-induced AML by blocking differentiation and increasing self-renewal. Therefore, examination of disturbed gene expression in HSCs can identify genes whose dysregulation is essential for leukemic stem cell function and that are targets for therapeutic interventions.
Assuntos
Células-Tronco Hematopoéticas/patologia , Leucemia Mieloide Aguda/genética , Proteínas Proto-Oncogênicas c-jun/fisiologia , Proteínas Proto-Oncogênicas/genética , Transativadores/genética , Animais , Diferenciação Celular/genética , Transformação Celular Neoplásica/genética , Regulação para Baixo , Granulócitos/citologia , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Leucemia Mieloide Aguda/metabolismo , Leucemia Mieloide Aguda/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Monócitos/citologia , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-jun/genética , Proteínas Proto-Oncogênicas c-jun/metabolismo , Transativadores/metabolismo , Transcrição Gênica , Transdução GenéticaRESUMO
The mechanisms mediating hematopoietic stem and progenitor cell (HSPC) mobilization by G-CSF are complex. We have found previously that G-CSF-enforced mobilization is controlled by peripheral sympathetic nerves via norepinephrine (NE) signaling. In the present study, we show that G-CSF likely alters sympathetic tone directly and that methods to increase adrenergic activity in the BM microenvironment enhance progenitor mobilization. Peripheral sympathetic nerve neurons express the G-CSF receptor and ex vivo stimulation of peripheral sympathetic nerve neurons with G-CSF reduced NE reuptake significantly, suggesting that G-CSF potentiates the sympathetic tone by increasing NE availability. Based on these data, we investigated the NE reuptake inhibitor desipramine in HSPC mobilization. Whereas desipramine did not by itself elicit circulating HSPCs, it increased G-CSF-triggered mobilization efficiency significantly and rescued mobilization in a model mimicking "poor mobilizers." Therefore, these data suggest that blockade of NE reuptake may be a novel therapeutic target to increase stem cell yield in patients.
Assuntos
Movimento Celular/fisiologia , Mobilização de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Norepinefrina/metabolismo , Animais , Células Cultivadas , Imunofluorescência , Fator Estimulador de Colônias de Granulócitos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Sistema Nervoso Simpático/citologia , Sistema Nervoso Simpático/metabolismoRESUMO
Multiple myeloma (MM) is a clonal plasma cell disorder frequently accompanied by hematopoietic impairment. We show that hematopoietic stem and progenitor cells (HSPCs), in particular megakaryocyte-erythrocyte progenitors, are diminished in the BM of MM patients. Genomic profiling of HSPC subsets revealed deregulations of signaling cascades, most notably TGFß signaling, and pathways involved in cytoskeletal organization, migration, adhesion, and cell-cycle regulation in the patients. Functionally, proliferation, colony formation, and long-term self-renewal were impaired as a consequence of activated TGFß signaling. In accordance, TGFß levels in the BM extracellular fluid were elevated and mesenchymal stromal cells (MSCs) had a reduced capacity to support long-term hematopoiesis of HSPCs that completely recovered on blockade of TGFß signaling. Furthermore, we found defective actin assembly and down-regulation of the adhesion receptor CD44 in MM HSPCs functionally reflected by impaired migration and adhesion. Still, transplantation into myeloma-free NOG mice revealed even enhanced engraftment and normal differentiation capacities of MM HSPCs, which underlines that functional impairment of HSPCs depends on MM-related microenvironmental cues and is reversible. Taken together, these data implicate that hematopoietic suppression in MM emerges from the HSPCs as a result of MM-related microenvironmental alterations.
Assuntos
Antígenos CD34/metabolismo , Biomarcadores/metabolismo , Medula Óssea/patologia , Células-Tronco Hematopoéticas/patologia , Células Progenitoras de Megacariócitos e Eritrócitos/patologia , Mieloma Múltiplo/metabolismo , Mieloma Múltiplo/patologia , Animais , Western Blotting , Medula Óssea/metabolismo , Estudos de Casos e Controles , Adesão Celular , Ciclo Celular , Diferenciação Celular , Movimento Celular , Proliferação de Células , Células Cultivadas , Feminino , Citometria de Fluxo , Imunofluorescência , Perfilação da Expressão Gênica , Células-Tronco Hematopoéticas/metabolismo , Humanos , Técnicas Imunoenzimáticas , Masculino , Células Progenitoras de Megacariócitos e Eritrócitos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/patologia , Camundongos , Camundongos Endogâmicos NOD , Mieloma Múltiplo/genética , Análise de Sequência com Séries de Oligonucleotídeos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de SinaisRESUMO
PURPOSE: 4-1BB (CD137) is a costimulatory immune receptor expressed on activated T cells, activated B cells, natural killer cells and tumor-infiltrating lymphocytes, making it a promising target for cancer immunotherapy. Cinrebafusp alfa, a monoclonal antibody-like bispecific protein targeting HER2 and 4-1BB, aims to localize 4-1BB activation to HER2-positive tumors. This study evaluated the safety, tolerability, and preliminary efficacy of cinrebafusp alfa in patients with previously treated HER2-positive malignancies. EXPERIMENTAL DESIGN: This was a multi-center dose escalation study involving patients with HER2-positive malignancies who had received prior treatment. The study assessed the safety and efficacy of cinrebafusp alfa across various dose levels. Patients were assigned to different cohorts, and antitumor responses were evaluated. The study aimed to determine the maximum tolerated dose (MTD) and to observe any clinical activity at different dose levels. RESULTS: Out of 40 evaluable patients in the 'active dose' efficacy cohorts, 5 showed an antitumor response, resulting in an overall response rate (ORR) of 12.5% and a disease control rate of 52.5%. Clinical activity was observed at the 8 mg/kg and 18 mg/kg dose levels, with confirmed objective response rates of 28.6% and 25.0%, respectively. Cinrebafusp alfa was safe and tolerable, with Grade ≤2 infusion-related reactions being the most frequent treatment-related adverse event. MTD was not reached during the study. CONCLUSION: Cinrebafusp alfa demonstrates promising activity in patients with HER2-positive malignancies who have progressed on prior HER2-targeting regimens. Its acceptable safety profile suggests it could be a treatment option for patients not responding to existing HER2-directed therapies.
RESUMO
The prognostic relevance of minimal residual disease (MRD) in patients with multiple myeloma is still an open question. We measured MRD levels in bone marrow (BM) samples of 53 patients treated with high-dose therapy (HDT) and autologous peripheral blood stem cell transplantation using real-time quantitative (RQ)-IgH-PCR with allel-specific oligonucleotide probes. We identified a prognostically relevant threshold level of 0.2% 2IgH/ß-actin ratio in the BM before HDT. Twenty-six patients with MRD levels below this value were termed as the "low-MRD group," whereas 27 patients with levels above this threshold were allocated to the "high-MRD group." Median event-free-survival (EFS) in the low-MRD group was significantly (P = .001) longer than in the high-MRD group with 35 versus 20 months, respectively. Overall survival (OS) within the low-MRD group was also significantly longer with 70 versus 45 months (P = .04). Using multivariate analysis, we found that the pretransplantation MRD level was an independent prognostic factor for EFS (P = .003) and OS (P = .05). Further, EFS of patients in the high-MRD could be improved (P = .005) when they achieved a low MRD level after HDT. In conclusion, measuring MRD is of prognostic relevance in patients with MM, and low MRD levels should be a goal of treatment.
Assuntos
Medula Óssea/patologia , Mieloma Múltiplo/patologia , Mieloma Múltiplo/terapia , Transplante de Células-Tronco de Sangue Periférico , Adulto , Idoso , Terapia Combinada , Feminino , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/cirurgia , Neoplasia Residual , Prognóstico , Reação em Cadeia da Polimerase em Tempo Real/métodos , Análise de Sobrevida , Resultado do TratamentoRESUMO
Patients suffering from high-risk myelodysplastic syndrome (MDS) or acute myelogenous leukemia (AML) secondary to MDS (sAML) are characterized by poor response to conventional cytotoxic chemotherapy. The purpose of our prospective single-center study was to examine the safety and efficacy of an allogeneic hematopoietic stem cell transplantation (HSCT) following a sequential conditioning regimen as first-line therapy for previously untreated patients with high-risk MDS or sAML. Between November 2003 and June 2010, 30 patients (20 high-risk MDS, 10 sAML) received fludarabine (4 × 30 mg/m(2)), amsacrine (4 × 100 mg/m(2)), and Ara-C (4 × 2 g/m(2), FLAMSA). After 2 to 3 days of rest, patients received high-dose melphalan alone (200 mg/m(2) for patients with an age <50 years, 150 mg/m(2) for patients with an age between 50 and 60 years, and 100 mg/m(2) for patients with an age >60 years; n = 24) or melphalan and thiotepa (10 mg/kg, Mel/Thio, n = 6). Following these high-dose conditioning regimens, a median number of 7.7 × 10(6) CD34(+) cells/kg body weight (range: 2.9 × 10(6)-17.2 × 10(6)) were transplanted from 13 related or 17 unrelated donors. Antithymocyte globulin (Fresenius 30-60 mg/kg) as well as tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. All patients except 1 with primary graft failure achieved complete remission after HSCT. After a median follow-up time of 28 months (range: 7-81), 21 patients (70%) were alive and free of disease. Overall, 4 patients relapsed. At 2 years, overall survival, event-free survival, and treatment-related mortality were 70%, 63%, and 30%, respectively. Because of undue toxicity, thiotepa is no longer part of the conditioning regimen. Our results add to the body of evidence that a FLAMSA-based sequential conditioning therapy is effective for previously untreated patients with high-risk MDS or sAML.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Transplante de Células-Tronco Hematopoéticas/métodos , Leucemia Mieloide Aguda/terapia , Síndromes Mielodisplásicas/terapia , Condicionamento Pré-Transplante/métodos , Adulto , Idoso , Amsacrina/administração & dosagem , Bussulfano/administração & dosagem , Ciclofosfamida/administração & dosagem , Citarabina/administração & dosagem , Intervalo Livre de Doença , Feminino , Doença Enxerto-Hospedeiro/etiologia , Doença Enxerto-Hospedeiro/prevenção & controle , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/cirurgia , Masculino , Melfalan/administração & dosagem , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/tratamento farmacológico , Síndromes Mielodisplásicas/cirurgia , Condicionamento Pré-Transplante/efeitos adversos , Transplante Homólogo , Doadores não Relacionados , Vidarabina/administração & dosagem , Vidarabina/análogos & derivadosRESUMO
BACKGROUND: Few data are available on therapy-related myelodysplastic syndromes and acute myeloid leukemia developing after radioiodine treatment. DESIGN AND METHODS: We retrospectively analyzed 39 patients with myeloid neoplasms following radioiodine treatment, whose data were reported to the Duesseldorf Myelodysplastic Syndromes Register (8 of 3814 patients) and five other German Myelodysplastic Syndromes centers (n=31) between 1982 and 2011. These data were compared with those from 165 patients from our Myelodysplastic Syndromes Register with therapy-related myeloid neoplasms following chemotherapy (n=90), radiation (n=30), or radiochemotherapy (n=45). RESULTS: With a median latency of 79 months, 18 patients developed therapy-related acute myeloid leukemia and 21 presented with therapy-related myelodysplastic syndromes (8 refractory anemia with excess blasts I/II, 6 refractory anemia with multilineage dysplasia, 3 myelodysplastic syndromes with del(5q), 1 refractory anemia, 1 refractory anemia with ring sideroblasts, 1 chronic myelomonocytic leukemia II, 1 myelodysplastic/myeloproliferative neoplasm unclassifiable). Risk assessment according to the International Prognostic Scoring System was low-risk in 23%, intermediate-1 in 29%, intermediate-2 in 35%, and high-risk in 13%. Karyotype was abnormal in 68%, with chromosomes 7 (30%), 5 (26%), 8 (26%) and 3 (17%) being most frequently affected. No differences in the distribution of gender, World Health Organization subtype, acute myeloid leukemia progression, International Prognostic Scoring System score, and cytogenetic risk were observed between patients with therapy-related myeloid neoplasms following radioiodine or other treatment modalities. Of 17 patients who received induction chemotherapy, 71% were refractory to this treatment or died from treatment-related toxicity. The median overall survival in the entire group was 21.7 months (95%-CI 10.5-33 months) and did not differ significantly in comparison to the survival of patients with therapy-related myeloid neoplasms following other cytotoxic treatments. Patients with therapy-related acute myeloid leukemia had significantly inferior overall survival (12.4 versus 28.7 months, P=0.002). CONCLUSIONS: Patients developing a therapy-related myeloid neoplasm after radioiodine treatment usually present with biological characteristics similar to those seen in patients with therapy-related myeloid neoplasms following other cytotoxic treatment modalities, associated with a low response rate to induction chemotherapy and poor prognosis.
Assuntos
Leucemia Mieloide Aguda/epidemiologia , Síndromes Mielodisplásicas/epidemiologia , Neoplasias Induzidas por Radiação/epidemiologia , Segunda Neoplasia Primária/epidemiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Radioisótopos do Iodo/efeitos adversos , Leucemia Mieloide Aguda/mortalidade , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/mortalidade , Neoplasias Induzidas por Radiação/mortalidade , Segunda Neoplasia Primária/mortalidade , Estudos Retrospectivos , Taxa de Sobrevida , Resultado do Tratamento , Adulto JovemRESUMO
Acute myeloid leukemia (AML) is a heterogeneous hematological malignancy. Treatment of patients suffering from high-risk AML as defined by clinical parameters, cytogenetics, and/or molecular analyses is often unsuccessful. OSI-461 is a pro-apoptotic compound that has been proposed as a novel therapeutic option for patients suffering from solid tumors like prostate or colorectal carcinoma. But little is known about its anti-proliferative potential in AML. Hence, we treated bone marrow derived CD34(+) selected blast cells from 20 AML patients and the five AML cell lines KG-1a, THP-1, HL-60, U-937, and MV4-11 with the physiologically achievable concentration of 1 µM OSI-461 or equal amounts of DMSO as a control. Following incubation with OSI-461, we found a consistent induction of apoptosis and an accumulation of cells in the G2/M phase of the cell cycle. In addition, we demonstrate that the OSI-461 mediated anti-proliferative effects observed in AML are associated with the induction of the pro-apoptotic cytokine mda-7/IL-24 and activation of the growth arrest and DNA-damage inducible genes (GADD) 45α and 45γ. Furthermore, OSI-461 treated leukemia cells did not regain their proliferative potential for up to 8 days after cessation of treatment following the initial 48 h treatment period with 1 µM OSI-461. This indicates sufficient targeting of the leukemia-initiating cells in our in vitro experiments through OSI-461. The AML samples tested in this study included samples from patients who were resistant to conventional chemotherapy and/or had FLT3-ITD mutations demonstrating the high potential of OSI-461 in human AML.
Assuntos
Apoptose/efeitos dos fármacos , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/patologia , Sulindaco/análogos & derivados , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral/efeitos dos fármacos , Ensaios Clínicos como Assunto , Expressão Gênica/efeitos dos fármacos , Humanos , Inibidores de Fosfodiesterase/farmacologia , Inibidores de Fosfodiesterase/uso terapêutico , Sulindaco/farmacologia , Sulindaco/uso terapêuticoRESUMO
Acute myeloid leukemia is a heterogeneous disease with varying genetic and molecular pathologies. Non-steroidal anti-inflammatory drugs (NSAIDs) have been proven to possess significant anti-proliferative potential in various cancer cells in vitro and in vivo. Hence, treatment with these agents can be utilized to study disease specific anti-proliferative pathways. In this study, a total number of 42 bone marrow derived CD34(+) selected de novo AML patient samples and the AML cell lines THP-1 and HL-60 were treated with the NSAIDs Sulindac sulfide and Diclofenac. We analyzed viability, apoptosis, differentiation and addressed the molecular mechanisms involved. We found a consistent induction of apoptosis and to some extent an increased myeloid differentiation capacity in NSAID treated AML cells. Comprehensive protein and gene expression profiling of Diclofenac treated AML cells revealed transcriptional activation of GADD45α and its downstream MAPK/JNK pathway as well as increased protein levels of the caspase-3 precursor. This pointed towards a role of the c-Jun NH(2)-terminal kinase (JNK) in NSAID mediated apoptosis that we found indeed to be dependent on JNK activity as addition of a specific JNK-inhibitor abrogated apoptosis. Furthermore, the AP-1 transcription factor family members' c-Jun, JunB and Fra-2 were transcriptionally activated in NSAID treated AML cells and re-expression of these transcription factors led to activation of GADD45α with induction of apoptosis. Mechanistically, we demonstrate that NSAIDs induce apoptosis in AML through a novel pathway involving increased expression of AP-1 heterodimers, which by itself is sufficient to induce GADD45α expression with consecutive activation of JNK and induction of apoptosis.
Assuntos
Apoptose , Diclofenaco/uso terapêutico , Leucemia Mieloide Aguda/tratamento farmacológico , Sulindaco/análogos & derivados , Fator de Transcrição AP-1/metabolismo , Anti-Inflamatórios não Esteroides/uso terapêutico , Caspase 3/metabolismo , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular , Sobrevivência Celular , Clonagem Molecular , Citometria de Fluxo , Antígeno 2 Relacionado a Fos/metabolismo , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Vetores Genéticos , Células HL-60 , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Interferência de RNA , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Sulindaco/uso terapêutico , Ativação TranscricionalRESUMO
Thrombocytopenia is a frequent symptom and clinical challenge in patients with myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). Eltrombopag is a small molecule thrombopoietin receptor agonist that might be a new option to treat thrombocytopenia in these diseases, provided that it does not stimulate malignant hematopoiesis. In this work, we studied the effects of Eltrombopag on proliferation, apoptosis, differentiation, colony formation, and malignant self-renewal of bone marrow mononuclear cells of patients with AML and MDS. Malignant bone marrow mononuclear cells did not show increased proliferation, or increased clonogenic capacity at concentrations of Eltrombopag ranging from 0.1 to 30 microg/mL. On the contrary, we observed a moderate, statistically nonsignificant (P = .18), decrease of numbers of malignant cells (mean, 56%; SD, 28%). Eltrombopag neither led to increased 5-bromo-2-deoxyuridine incorporation, decreased apoptosis, an increase of malignant self-renewal, nor enhanced in vivo engraftment in xenotransplantations. Furthermore, we found that Eltrombopag was capable of increasing megakaryocytic differentiation and formation of normal megakaryocytic colonies in patients with AML and MDS. These results provide a preclinical rationale for further testing of Eltrombopag for treatment of thrombocytopenia in AML and MDS.
Assuntos
Benzoatos/farmacologia , Hidrazinas/farmacologia , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia Mieloide Aguda/metabolismo , Síndromes Mielodisplásicas/tratamento farmacológico , Síndromes Mielodisplásicas/metabolismo , Pirazóis/farmacologia , Receptores de Trombopoetina/agonistas , Trombocitopenia/tratamento farmacológico , Trombocitopenia/metabolismo , Animais , Apoptose/efeitos dos fármacos , Benzoatos/uso terapêutico , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Diferenciação Celular , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Hematopoese/efeitos dos fármacos , Humanos , Hidrazinas/uso terapêutico , Leucemia Mieloide Aguda/patologia , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Masculino , Megacariócitos/metabolismo , Megacariócitos/patologia , Camundongos , Camundongos Endogâmicos NOD , Síndromes Mielodisplásicas/patologia , Pirazóis/uso terapêutico , Receptores de Trombopoetina/metabolismo , Trombocitopenia/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Since its introduction in 1983, high-dose therapy followed by autologous peripheral blood stem cell transplantation is a pillar of the treatment of patients with multiple myeloma. In the last decades, a multitude of clinical trials helped to improve strategies based on high-dose therapy and autologous stem cell transplantation resulting in a continuously prolongation of overall survival of patients. In this chapter we will review the progress, which has been made in order to enhance the mobilisation of autologous stem cells and increase the effectiveness of this treatment.
Assuntos
Antineoplásicos/uso terapêutico , Mieloma Múltiplo/terapia , Transplante de Células-Tronco de Sangue Periférico , Antineoplásicos/administração & dosagem , Moléculas de Adesão Celular/fisiologia , Terapia Combinada , Ciclofosfamida/uso terapêutico , Filgrastim , Fator Estimulador de Colônias de Granulócitos/uso terapêutico , Fatores de Crescimento de Células Hematopoéticas/fisiologia , Humanos , Dose Máxima Tolerável , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/cirurgia , Polietilenoglicóis , Proteínas Recombinantes , Transplante AutólogoRESUMO
Cytochrome P450 epoxygenases (CYP450) have been recently shown to promote malignant progression. Here we investigated the mRNA and protein expression and potential clinical relevance of CYP2C9 in esophageal cancer. Highest expression was detected in esophageal adenocarcinoma (EAC; n=78) and adjacent esophageal mucosa (NEM; n=79). Levels of CYP2C9 in EAC and NEM were significantly higher compared to esophageal squamous cell carcinoma (ESCC; n=105). Early tumor stages and well-differentiated tumors showed a significantly higher CYP2C9 expression compared to progressed tumors. Moreover, CYP2C9 expression was correlated to high Ki-67 labeling indices in EAC and Ki-67 positive tumor cells in EAC and ESCC. Selective inhibition of CYP2C9 decreased tumor cell proliferation (KYSE30, PT1590 and OE19) in vitro, which was abolished by 11,12-epoxyeicosatrienoic acid (11,12-EET). Cell-cycle analysis using FACS revealed that inhibition of CYP2C9 leads to a G0/G1 phase cell-cycle arrest. CYP2C9 seems to be relevant for early esophageal cancer development by promoting tumor cell proliferation. Pharmacological inhibition of CYP2C9 might contribute to a more efficient therapy in CYP2C9 highly expressing esophageal cancers.
Assuntos
Hidrocarboneto de Aril Hidroxilases/metabolismo , Carcinoma de Células Escamosas/enzimologia , Carcinoma de Células Escamosas/patologia , Proliferação de Células , Neoplasias Esofágicas/enzimologia , Neoplasias Esofágicas/patologia , Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Ácido 8,11,14-Eicosatrienoico/farmacologia , Hidrocarboneto de Aril Hidroxilases/genética , Linhagem Celular Tumoral , Citocromo P-450 CYP2C9 , Progressão da Doença , Fase G1 , Humanos , Imuno-Histoquímica , Fase de Repouso do Ciclo CelularRESUMO
PURPOSE: Monopolar spindle 1 (MPS1) kinase inhibitor, BAY 1217389 (BAY) synergizes with paclitaxel. This phase I study assessed the combination of BAY with paclitaxel using a novel randomized continuous reassessment method (rCRM) to improve dose determination. PATIENTS AND METHODS: Patients with solid tumors were randomized to receive oral BAY (twice daily 2-days-on/5-days-off) with weekly paclitaxel (90 mg/m2) or paclitaxel monotherapy in cycle 1. Dose escalation was guided by CRM modeling. Primary objectives were to assess safety, establish the MTD of BAY, and to evaluate the pharmacokinetic profiles for both compounds. Simulations were performed to determine the contribution of the rCRM for dose determination. RESULTS: In total, 75 patients were enrolled. The main dose-limiting toxicities were hematologic toxicities (55.6%). The MTD of BAY was established at 64 mg twice daily with paclitaxel. Inclusion of a control arm enabled the definitive attribution of grade ≥3 neutropenia to higher BAY exposure [AUC0-12 (P< 0.001)]. After determining the MTD, we included 19 patients with breast cancer at this dose for dose expansion. Other common toxicities were nausea (45.3%), fatigue (41.3%), and diarrhea (40.0%). Overall confirmed responses were seen in 31.6% of evaluable patients. Simulations showed that rCRM outperforms traditional designs in determining the true MTD. CONCLUSIONS: The combination of BAY with paclitaxel was associated with considerable toxicity without a therapeutic window. However, the use of the rCRM design enabled us to determine the exposure-toxicity relation for BAY. Therefore, we propose that the rCRM could improve dose determination in phase I trials that combine agents with overlapping toxicities.
Assuntos
Neoplasias da Mama , Neoplasias , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Neoplasias da Mama/patologia , Relação Dose-Resposta a Droga , Feminino , Humanos , Dose Máxima Tolerável , Neoplasias/patologia , PaclitaxelRESUMO
To further clarify the role of ribosomal protein S14 (RPS14) in myelodysplastic syndrome, we examined RPS14 transcription in bone marrow derived CD34+ cells from patients with non-5q- myelodysplastic syndrome and found a reduced expression of RPS14 in 51 of 72 (71%) patients. MDS patients with an intermediate-1 risk (INT-1) score according to the international prognostic scoring system and low RPS14 expression had a superior median overall survival of not reached versus 25 months compared to INT-1 patients with high RPS14 expression (p=0.0249). Using multivariate analysis, the RPS14 expression status was confirmed as an independent predictor for survival in INT-1 patients.
Assuntos
Aberrações Cromossômicas , Cromossomos Humanos Par 5/genética , Regulação Neoplásica da Expressão Gênica , Síndromes Mielodisplásicas/genética , Síndromes Mielodisplásicas/mortalidade , Proteínas Ribossômicas/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/classificação , Sistema de Registros , Taxa de Sobrevida/tendênciasRESUMO
BACKGROUND: Pegylated granulocyte colony-stimulating factor (G-CSF) has recently been introduced as a new compound for mobilization of CD34(+) hematopoietic stem and progenitor cells. In this study, we compared the molecular and functional characteristics of CD34(+) cells mobilized by pegylated G-CSF with those mobilized by unconjugated G-CSF. DESIGN AND METHODS: Gene expression of immunomagnetically enriched CD34(+) cells from leukapheresis products of patients who were given pegylated-G-CSF or unconjugated G-CSF was analyzed using Affymetrix HG Focus microarrays and quantitative reverse transcriptase polymerase chain reaction. Flow cytometry and fluorescence activated cell sorting was conducted to assess the CD34(+) subset composition and to obtain Lin(-), CD34(+), CD38(-) hematopoietic stem cells. Cell cycle assays and clonogenic assays were performed for functional corroboration. RESULTS: Pegylated G-CSF and unconjugated G-CSF mobilized CD34(+) and hematopoietic stem cells with different molecular phenotypes and functional properties. The CD34(+) cells mobilized by pegylated G-CSF had higher expression levels of genes indicative of early hematopoiesis, including HOXA9, MEIS1 and GATA3. We found lower expression of genes characteristic of erythroid and later stages of myeloid differentiation and a lower functional burst-forming unit erythroid/colony-forming unit-granulocyte-macrophage ratio. Consistently, greater numbers of hematopoietic stem cells and common myeloid progenitors and fewer megakaryocyte-erthrocyte progenitors were found in the pegylated-G-CSF-mobilized CD34(+) cells. Additionally, sorted pegylated-G-CSF-mobilized hematopoietic stem cells displayed higher expression of HOXA9 in comparison to G-CSF-mobilized hematopoietic stem cells. In line with the gene expression data, CD34(+) cells mobilized by pegylated G-CSF, as well as sorted hematopoietic stem cells, showed a significantly greater cell cycle activity. CONCLUSIONS: Stimulation with pegylated-G-CSF or G-CSF results in different expression of key regulatory genes and different functional properties of mobilized hematopoietic stem cells as well as their progeny, a finding that might be relevant for the application of these cells in blood stem cell transplantation.