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1.
Haematologica ; 2019 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-31296574

RESUMO

CD38 is expressed in several types of non-Hodgkin lymphoma and constitutes a promising target for antibody-based therapy. Daratumumab (Darzalex) is a first-in-class anti-CD38 antibody approved for the treatment of relapsed/refractory multiple myeloma. It has also demonstrated clinical activity in Waldenstrom macroglobulinaemia and amyloidosis. Here, we have evaluated the activity and mechanism of action of daratumumab in preclinical in vitro and in vivo models of mantle cell lymphoma, follicular lymphoma and diffuse large B cell lymphoma, as monotherapy or in combination with standard chemo-immunotherapy. In vitro, daratumumab engages Fc-mediated cytotoxicity by antibody-dependent cell cytotoxicity and antibody-dependent cell phagocytosis in all lymphoma subtypes. In the presence of human serum, complement-dependent cell cytotoxicity was marginally engaged. We demonstrated by Selective Plane Illumination Microscopy that daratumumab fully penetrated a 3D lymphoma organoid and decreased organoid volume. In vivo, daratumumab completely prevents tumor outgrowth in models of mantle cell and follicular lymphoma, and shows comparable activity to rituximab in a disseminated in vivo model of blastic mantle cell lymphoma. Moreover, daratumumab improves overall survival in a mouse model of transformed CD20dim follicular lymphoma, where rituximab showed limited activity. Daratumumab potentiates the antitumor activity of CHOP and R-CHOP in mantle cell and follicular lymphoma xenografts. Furthermore, in a patient-derived diffuse large B cell lymphoma xenograft model, daratumumab anti-tumor activity was comparable to R-CHOP and the addition of daratumumab to either CHOP or R-CHOP led to full tumor regression. In summary, daratumumab constitutes a novel therapeutic opportunity in certain scenarios and these results warrant further clinical development.

2.
Exp Hematol Oncol ; 7: 27, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30356940

RESUMO

Background: The monoclonal antibody daratumumab, approved for treating myeloma, targets CD38, a protein on myeloma and also on CD34+ hematopoietic progenitor cells. Because mobilized CD34+ cells are critical for stem cell transplant, we investigated the in vitro activity of daratumumab on mobilized CD34+ cells from myeloma patients with no prior exposure to daratumumab. Methods: We determined the number of CD38 molecules per CD34+ cell, and whether daratumumab bound to CD34+ cells, whether C1q bound to daratumumab-coated CD34+ cells and whether daratumumab-related complement-dependent cytotoxicity (CDC) occurred. We also examined CD34+ cell progenitor cell colony capacity in assays with pre-plating incubation of CD34+ cells with daratumumab alone or with daratumumab and the CD59 inhibitory antibody BRIC229, and also assessed CD34+ cell responses to increasing doses of daratumumab in caspase 3/7 activity assays. Results: Although 75% of mobilized CD34+ cells co-express CD38, CD38 was minimally present on CD34+ cells compared to Daudi and KG-1 controls, C1q did not bind to daratumumab-coated CD34+ cells, and CDC did not occur. CD34+ cells incubated in complement-rich human serum with daratumumab alone or with daratumumab and BRIC229, and then plated in progenitor cell assays, produced similar numbers of colonies as controls. In progenitor cell assays with cryopreserved or fresh unselected or CD34-selected cells, daratumumab did not affect progenitor cell capacity, and in caspase 3/7 activity assays CD34+ cells were not affected by increasing doses of daratumumab. Conclusion: In vitro, daratumumab is not toxic to mobilized CD34+ progenitor cells from myeloma patients.

3.
Clin Lymphoma Myeloma Leuk ; 18(1): 44-51, 2018 01.
Artigo em Inglês | MEDLINE | ID: mdl-29054515

RESUMO

INTRODUCTION: Daratumumab, a human CD38 monoclonal antibody approved for multiple myeloma (MM) treatment, binds red blood cells (RBCs), resulting in panagglutination in compatibility tests. Published mitigation methods avoid additional testing, ensuring timely release of blood products. Blood transfusion management and transfusion-related outcomes of daratumumab-treated patients in the SIRIUS study are reported, with emphasis on 2 clinical sites. PATIENTS AND METHODS: Patients had MM treated with ≥ 3 prior lines of therapy, including a proteasome inhibitor and an immunomodulatory drug, or were refractory to a proteasome inhibitor and an immunomodulatory drug. RBC typing and alloantibody screening were performed in gel cards. Antibody identification using RBC panels was performed on patients with positive antibody screens. Hematology panels and serum chemistry were analyzed ≤ 2 days before each daratumumab infusion and the first daratumumab dose within each treatment cycle, respectively. Pre- and posttransfusion hemoglobin values were analyzed retrospectively. RESULTS: At clinical cutoff, patients received 236 transfusions; 47 (37.9%) of 124 patients received 147 packed RBC transfusions, and 17 (13.7%) received 89 platelet transfusions. No hemolysis was reported, and 1 platelet transfusion reaction was observed. At Mount Sinai, no transfusion adverse events were observed, no new unexpected RBC alloantibodies were identified, and transfusions increased hemoglobin values (median, 1.2 g/dL). At Levine Cancer Institute, 6 of 7 patients responded to transfusions, with a median hemoglobin change of 1.7 g/dL. CONCLUSION: In SIRIUS, no RBC transfusion reactions, including hemolysis, were observed. Observations from Mount Sinai and Levine Cancer Institute confirm that transfusions may be administered safely to daratumumab-treated patients.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Transfusão de Sangue/métodos , Imunoterapia/métodos , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/terapia , Idoso , Anticorpos Monoclonais/farmacologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/patologia , Recidiva
4.
Oncoimmunology ; 6(8): e1334744, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28919995

RESUMO

Patients with primary solid malignancies frequently exhibit signs of systemic inflammation. Notably, elevated levels of neutrophils and their associated soluble mediators are regularly observed in cancer patients, and correlate with reduced survival and increased metastasis formation. Recently, we demonstrated a mechanistic link between mammary tumor-induced IL17-producing γδ T cells, systemic expansion of immunosuppressive neutrophils and metastasis formation in a genetically engineered mouse model for invasive breast cancer. How tumors orchestrate this systemic inflammatory cascade to facilitate dissemination remains unclear. Here we show that activation of this cascade relies on CCL2-mediated induction of IL1ß in tumor-associated macrophages. In line with these findings, expression of CCL2 positively correlates with IL1Β and macrophage markers in human breast tumors. We demonstrate that blockade of CCL2 in mammary tumor-bearing mice results in reduced IL17 production by γδ T cells, decreased neutrophil expansion and enhanced CD8+ T cell activity. These results highlight a new role for CCL2 in facilitating the breast cancer-induced pro-metastatic systemic inflammatory γδ T cell - IL17 - neutrophil axis.

5.
Clin Cancer Res ; 23(6): 1493-1505, 2017 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-27637890

RESUMO

Purpose: To establish a proof-of-concept for the efficacy of the anti-CD38 antibody daratumumab in the poor prognosis CD38+ chronic lymphocytic leukemia (CLL) subtype.Experimental Design: The mechanism of action of daratumumab was assessed in CLL primary cells and cell lines using peripheral blood mononuclear cells to analyze antibody-dependent cell cytotoxicity (ADCC), murine and human macrophages to study antibody-dependent cell phagocytosis (ADCP), or human serum to analyze complement-dependent cytotoxicity (CDC). The effect of daratumumab on CLL cell migration and adhesion to extracellular matrix was characterized. Daratumumab activity was validated in two in vivo models.Results: Daratumumab demonstrated efficient lysis of patient-derived CLL cells and cell lines by ADCC in vitro and ADCP both in vitro and in vivo whereas exhibited negligible CDC in these cells. To demonstrate the therapeutic effect of daratumumab in CLL, we generated a disseminated CLL mouse model with the CD38+ MEC2 cell line and CLL patient-derived xenografts (CLL-PDX). Daratumumab significantly prolonged overall survival of MEC2 mice, completely eliminated cells from the infiltrated organs, and significantly reduced disease burden in the spleen of CLL-PDX. The effect of daratumumab on patient-derived CLL cell dissemination was demonstrated in vitro by its effect on CXCL12-induced migration and in vivo by interfering with CLL cell homing to spleen in NSG mice. Daratumumab also reduced adhesion of CLL cells to VCAM-1, accompanied by downregulation of the matrix metalloproteinase MMP9.Conclusions: These unique and substantial effects of daratumumab on CLL viability and dissemination support the investigation of its use in a clinical setting of CLL. Clin Cancer Res; 23(6); 1493-505. ©2016 AACR.


Assuntos
ADP-Ribosil Ciclase 1/genética , Anticorpos Monoclonais/administração & dosagem , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Microambiente Tumoral/efeitos dos fármacos , ADP-Ribosil Ciclase 1/imunologia , Animais , Linhagem Celular Tumoral , Citofagocitose/efeitos dos fármacos , Humanos , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/patologia , Metaloproteinase 9 da Matriz/genética , Camundongos , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Transfusion ; 56(12): 2964-2972, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27600566

RESUMO

BACKGROUND: Daratumumab (DARA) consistently interferes with routine blood bank serologic testing by directly binding to CD38 expressed on reagent red blood cells (RBCs). Treating RBCs with dithiothreitol (DTT) eliminates the DARA interference. We conducted an international, multicenter, blinded study aimed at validating the DTT method for use by blood bank laboratories worldwide. STUDY DESIGN AND METHODS: Paired plasma sample unknowns were sent to 25 participating blood bank laboratories. Sample 1 was spiked with DARA only (10 µg/mL), and Sample 2 with DARA plus a clinically significant RBC antibody (anti-D [n = 6], anti-Fya [n = 9], or anti-s [n = 10]). Sites were instructed to perform an antibody screen with and without DTT-treated RBCs and to use a DTT-treated RBC panel for antibody identification. Qualitative data about the DTT method were collected by online survey. The primary outcome was the proportion of study sites able to identify the antibody unknown in the presence of DARA. RESULTS: All sites observed the DARA interference with the antibody screen. The DARA interference was seen with all testing methods (gel, tube, or solid phase). Using the DTT method, 25 of 25 sites (100%) successfully identified the antibody unknown in the presence of DARA. Feedback on the DTT method was positive, with 17 of 19 (90%) sites responding to the survey indicating that they planned to use the DTT method to test clinical samples from DARA-treated patients. CONCLUSION: The DTT method is robust and reproducible and can be implemented by transfusion services worldwide to help provide safe blood products to patients treated with DARA.


Assuntos
Anticorpos Monoclonais/farmacologia , Ditiotreitol/farmacologia , Teste de Histocompatibilidade/normas , Anticorpos/análise , Anticorpos/sangue , Bancos de Sangue/métodos , Bancos de Sangue/normas , Segurança do Sangue , Humanos , Métodos , Controle de Qualidade , Método Simples-Cego
8.
Transfusion ; 55(6 Pt 2): 1555-62, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25988285

RESUMO

BACKGROUND: Monoclonal antibodies (MoAbs) are increasingly integrated in the standard of care. The notion that therapeutic MoAbs can interfere with clinical laboratory tests is an emerging concern that requires immediate recognition and the development of appropriate solutions. Here, we describe that treatment of multiple myeloma patients with daratumumab, a novel anti-CD38 MoAb, resulted in false-positive indirect antiglobulin tests (IATs) for all patients for 2 to 6 months after infusion. This precluded the correct identification of irregular blood group antibodies for patients requiring blood transfusion. STUDY DESIGN AND METHODS: The IAT was performed using three- and 11-donor-cell panels. Interference of daratumumab and three other anti-CD38 MoAbs was studied using fresh-frozen plasma spiked with different MoAb concentrations. Additionally it was tested whether two potentially neutralizing agents, anti-idiotype antibody and recombinant soluble CD38 (sCD38) extracellular domain, were able to inhibit the interference. RESULTS: The CD38 MoAbs caused agglutination in the IAT in a dose-dependent manner. Addition of an excess of anti-idiotype antibodies or sCD38 protein to the test abrogated CD38 MoAb interference and successfully restored irregular antibody screening and identification. DISCUSSION: CD38 MoAb therapy causes false-positive results in the IAT. The reliability of the test could be restored by adding a neutralizing agent against the CD38 MoAb to the patient's plasma. This study emphasizes that during drug development, targeted therapeutics should be investigated for potential interference with laboratory tests. Clinical laboratories should be informed when patients receive MoAb treatments and matched laboratory tests to prevent interference should be employed.


Assuntos
Anticorpos Monoclonais/imunologia , Antineoplásicos/imunologia , Tipagem e Reações Cruzadas Sanguíneas , Transfusão de Sangue , ADP-Ribosil Ciclase 1/imunologia , Idoso , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/uso terapêutico , Tipagem e Reações Cruzadas Sanguíneas/normas , Transfusão de Sangue/métodos , Teste de Coombs/normas , Reações Cruzadas , Reações Falso-Positivas , Feminino , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/sangue , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/terapia , Testes Sorológicos
9.
Transfusion ; 55(6 Pt 2): 1545-54, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25764134

RESUMO

BACKGROUND: Daratumumab (DARA), a promising novel therapy for multiple myeloma, is an IgG1κ monoclonal antibody that recognizes CD38 on myeloma cells. During routine compatibility testing, we observed that the plasma of five of five DARA-treated patients demonstrated a positive antibody screen and panreactivity on red blood cell (RBC) panel testing. We hypothesized that the observed panreactivity reflected DARA binding to CD38 on reagent RBCs, and we investigated methods to prevent this binding. STUDY DESIGN AND METHODS: DARA binding to CD38+ or CD38- HL60 cells was assessed by flow cytometry. To remove cell surface CD38, cells were incubated with dithiothreitol (DTT) or trypsin. Soluble CD38 or anti-DARA was used to neutralize DARA in solution. Routine blood bank serologic methods were used to test samples from DARA-treated patients and normal plasma samples spiked with DARA and/or alloantibodies. RESULTS: Normal plasma samples spiked with DARA (0.1-10 µg/mL) and incubated with reagent RBCs recapitulated the interference observed with samples from DARA-treated patients. Flow cytometry experiments confirmed DARA binding to CD38+ HL60 cells, but not to CD38- controls. DTT treatment of CD38+ HL60 cells reduced DARA binding by 92% by denaturing cell surface CD38. Treating DARA-containing plasma with soluble CD38 or anti-DARA idiotype also inhibited DARA binding. CONCLUSION: DARA causes panreactivity in vitro by binding to CD38 on reagent RBCs. Treating reagent RBCs with DTT is a robust method to negate the DARA interference, enabling the safe provision of blood to DARA-treated patients. Because DTT denatures Kell antigens, K- units are provided to these patients.


Assuntos
Anticorpos Monoclonais/imunologia , Tipagem e Reações Cruzadas Sanguíneas , Eritrócitos/imunologia , ADP-Ribosil Ciclase 1/genética , ADP-Ribosil Ciclase 1/metabolismo , Anticorpos Monoclonais/uso terapêutico , Reações Cruzadas/efeitos dos fármacos , Ditiotreitol/farmacologia , Relação Dose-Resposta a Droga , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Citometria de Fluxo , Células HL-60 , Humanos , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/terapia , Testes Sorológicos , Transfecção
10.
J Thorac Oncol ; 9(7): 974-82, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24922005

RESUMO

INTRODUCTION: Interleukin-6 (IL-6) can activate downstream signaling pathways in lung cancer cells, such as the STAT3 pathway, and is reported to be produced by tumor cells with activating EGFR mutations. We examined IL-6/STAT3 in lung cancer tumor tissues and the effects of siltuximab, a neutralizing antibody to human IL-6, in mouse models of lung cancer. METHODS: IL-6 and STAT3 activation levels were compared with tumor histology and presence of KRAS mutations in snap-frozen, non-small-cell lung cancer tumors. The effects of siltuximab alone or in combination with erlotinib were examined in mouse xenograft models constructed using three cell line xenograft models and one primary explant mouse model. We examined the influence of cancer-associated fibroblasts (CAFs) on tumor growth and siltuximab effects. RESULTS: IL-6 levels were higher in tumors of squamous cell versus adenocarcinoma histology and were not associated with presence of KRAS mutations. Tyrosine phosphorylation status of STAT3 did not correlate with tumor IL-6 levels. Serine phosphorylation of STAT3 was correlated with KRAS mutation status. Both tumor and stromal cells contributed to total IL-6 within tumors. Siltuximab had minimal effect as a single agent in xenografts with tumor cells alone; however, in models coadministered with CAFs, siltuximab had more potent effects on tumor inhibition. We observed no effects of combined erlotinib and siltuximab. CONCLUSIONS: IL-6 is elevated in subsets of human NSCLCs, especially with squamous cell histology. Tumors supported by stromal production of IL-6 seem to be the most vulnerable to tumor growth inhibition by siltuximab.


Assuntos
Adenocarcinoma/tratamento farmacológico , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma de Células Escamosas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Adenocarcinoma/química , Adenocarcinoma/genética , Animais , Anticorpos Monoclonais/administração & dosagem , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma Pulmonar de Células não Pequenas/química , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma de Células Escamosas/química , Carcinoma de Células Escamosas/genética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Cloridrato de Erlotinib , Feminino , Fibroblastos , Humanos , Interleucina-6/análise , Interleucina-6/imunologia , Neoplasias Pulmonares/química , Neoplasias Pulmonares/genética , Camundongos , Fosforilação , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas p21(ras) , Quinazolinas/administração & dosagem , Fator de Transcrição STAT3/metabolismo , Serina/metabolismo , Tirosina/metabolismo , Proteínas ras/genética
11.
Mol Oncol ; 8(7): 1231-9, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24816187

RESUMO

Ovarian cancer is associated with a leukocyte infiltrate and high levels of chemokines such as CCL2. We tested the hypothesis that CCL2 inhibition can enhance chemotherapy with carboplatin and paclitaxel. Elevated CCL2 expression was found in three non-MDR paclitaxel resistant ovarian cancer lines ES-2/TP, MES-OV/TP and OVCAR-3/TP, compared to parental cells. Mice xenografted with these cells were treated with the anti-human CCL2 antibody CNTO 888 and the anti-mouse MCP-1 antibody C1142, with and without paclitaxel or carboplatin. Our results show an additive effect of CCL2 blockade on the efficacy of paclitaxel and carboplatin. This therapeutic effect was largely due to inhibition of mouse stromal CCL2. We show that inhibition of CCL2 can enhance paclitaxel and carboplatin therapy of ovarian cancer.


Assuntos
Anticorpos/uso terapêutico , Antineoplásicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carboplatina/uso terapêutico , Quimiocina CCL2/antagonistas & inibidores , Neoplasias Ovarianas/tratamento farmacológico , Paclitaxel/uso terapêutico , Animais , Anticorpos/administração & dosagem , Anticorpos/imunologia , Anticorpos/farmacologia , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Carboplatina/administração & dosagem , Carboplatina/farmacologia , Linhagem Celular Tumoral , Quimiocina CCL2/imunologia , Sinergismo Farmacológico , Feminino , Humanos , Camundongos , Camundongos Nus , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/patologia , Ovário/efeitos dos fármacos , Ovário/imunologia , Ovário/patologia , Paclitaxel/administração & dosagem , Paclitaxel/farmacologia
12.
Clin Exp Metastasis ; 25(2): 139-48, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18064530

RESUMO

CNTO 95 is a fully human monoclonal antibody that recognizes alphav integrins. Previous studies have shown that CNTO 95 exhibits both anti-tumor and anti-angiogenic activities (Trikha M et al., Int J Cancer 110:326-335, 2004). In this study we investigated the biological activities of CNTO 95 on breast tumor cells both in vitro and in vivo. In vitro treatment with CNTO 95 decreased the viability of breast tumor cells adhering to vitronectin. CNTO 95 inhibited tumor cell adhesion, migration, and invasion in vitro. CNTO 95 treatment also induced tyrosine dephosphorylation of focal adhesion kinase (FAK), and the docking protein paxillin that recruits both structural and signaling molecules to focal adhesions (Turner CE, Int J Biochem Cell Biol 30:955-959, 1998; O'Neil GM et al., Trends Cell Biol 10:111-119, 2000). These results suggest that CNTO 95 inhibits breast tumor cell growth, migration and invasion by interruption of alphav integrin mediated focal adhesions and cell motility signals. In vivo studies of CNTO 95 were conducted in an orthotopic breast tumor xenograft model. Treatment with CNTO 95 resulted in significant inhibition of both tumor growth and spontaneous metastasis of MDA-MB-231 cells to the lungs. CNTO 95 also inhibited lung metastasis in a separate experimental (tail vein injection) model of metastasis. The results presented here demonstrate the anti-tumor and anti-metastatic activities of CNTO 95 in breast cancer models and provide insight into the cellular and molecular mechanisms mediating its inhibitory effects on metastasis.


Assuntos
Anticorpos Monoclonais/farmacologia , Neoplasias da Mama/metabolismo , Movimento Celular/efeitos dos fármacos , Integrina alfaV/imunologia , Animais , Anticorpos Monoclonais Humanizados , Neoplasias da Mama/patologia , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Quinase 1 de Adesão Focal/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Camundongos , Camundongos SCID , Invasividade Neoplásica , Paxilina/metabolismo , Fosforilação , Transdução de Sinais/efeitos dos fármacos , Transplante Heterólogo , Vitronectina/metabolismo
13.
Med Sci Monit ; 8(12): BR504-14, 2002 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-12503028

RESUMO

BACKGROUND: Progenipoietin-4 (ProGP-4) is an E. coli derived chimeric growth factor that activates the human Flt3 and G-CSF receptors. ProGP-4 possesses cross-species activity and treatment of mice with ProGP-4 results in increases in the number of WBC and Class II+/CD11c+ cells in both spleen and peripheral blood. Herein, we report morphologic, phenotypic and functional evaluation of Class II+/CD11c+ cells generated by in vivo administration of ProGP-4. MATERIAL/METHODS: C57BL/6 mice were injected daily with ProGP for 7 to 18 days. Leukocytes from spleen and peripheral blood were analyzed by flow cytometry to enumerate and characterize changes in DC populations. Spleens from ProGP treated mice were evaluated by immunocytochemistry and enriched CD11c+ populations were functionally assessed in a mixed lymphocyte assay and in an antigen dependent CTL assay. RESULTS: Administration of this dual receptor agonist to mice resulted in dose-dependent increases in the numbers of total white blood cells and Class II+/CD11c+ cells in spleen and peripheral blood. CD11c+ cells from ProGP-4 treated mice co-expressed DEC205 and also expressed CD80, CD86 and CD40, albeit at lower levels as compared to Class II+/CD11c+ cells from untreated animals. Despite lower co-stimulatory molecule expression, ProGP-4-generated Class II+/CD11c+ cells stimulated proliferation of allogeneic T cells and an antigen-specific T cell hybridoma as efficiently as bone marrow derived dendritic cells from untreated mice. CONCLUSIONS: The data presented in this report highlight the ability of E. coli derived ProGP-4 to expand large numbers of functional DC in the peripheral blood and lymphoid organs in vivo using a rodent model of hematopoiesis. E. coli derived chimeric receptor agonists such as ProGP-4 may enable further investigations of immunotherapeutic approaches to the treatment of diseases such as cancer and autoimmunity.


Assuntos
Células Dendríticas/efeitos dos fármacos , Substâncias de Crescimento/farmacologia , Proteínas Proto-Oncogênicas/agonistas , Receptores de Fator Estimulador de Colônias de Granulócitos/agonistas , Animais , Antígeno CD11c/metabolismo , Fatores Estimuladores de Colônias/farmacologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Escherichia coli/genética , Feminino , Antígenos de Histocompatibilidade Classe II/metabolismo , Humanos , Imunoterapia , Proteínas de Membrana/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Receptores Proteína Tirosina Quinases , Proteínas Recombinantes de Fusão/farmacologia , Proteínas Recombinantes , Tirosina Quinase 3 Semelhante a fms
14.
Int J Mol Med ; 10(4): 385-94, 2002 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12239583

RESUMO

Repopulating hematopoietic cell compartments after myeloablative chemotherapy remains a key factor in a successful chemotherapy program. Modified and chimeric cytokines have been developed to help reduce inflammation, fever and hospitalization time for patients. A chimeric cytokine, progenipoietin-1 (ProGP-1), containing the G-CSF and FL receptor agonists binds both the G-CSF receptor and FLT-3. It also stimulates the growth of dendritic cells, which play an important role in immunotherapy. While in vivo effects of ProGP-1 are well described, the mechanisms by which it stimulates growth are not well understood. We have investigated the effects of ProGP-1 on prevention of apoptosis in the human hematopoietic cell line OCI-AML.5. ProGP-1 promoted cellular proliferation better than G-CSF or FL separately but stimulated proliferation similar to their co-addition as demonstrated by growth curves and [3H]-thymidine incorporation. ProGP-1 prevented apoptosis to a greater degree than G-CSF or FL alone as determined by annexin V/propidium iodide binding and TUNEL assays. ProGP-1 promoted maintenance of the mitochondrial membrane potential better than G-CSF or FL alone. In addition, Pro-GP promoted a lower redox potential as higher levels of free radicals were detected after cytokine treatment than in cytokine-deprived cells implying increased respiration. These data indicate that ProGP-1 promotes the proliferation and prevents the apoptosis of human hematopoietic cells better than FL or G-CSF alone, and to a similar extent as their co-addition. Thus, ProGP-1 can be used to repopulate certain hematopoietic cells as a single entity rather than the introduction of two different cytokines.


Assuntos
Apoptose/fisiologia , Fatores Estimuladores de Colônias/metabolismo , Células-Tronco Hematopoéticas/metabolismo , Laranja Acridina , Anexina A5 , Apoptose/efeitos dos fármacos , Fatores Estimuladores de Colônias/farmacologia , Etídio , Fator Estimulador de Colônias de Granulócitos/farmacologia , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Marcação In Situ das Extremidades Cortadas , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Propídio , Proteínas Recombinantes
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