Combined Use of Dendritic Cells Enhances Specific Anti-Leukemia Effect of Leukemia Cell-Derived-HSP70 in a Mouse Model with Minimal Residual Leukemia Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3065-3065
Author(s):  
Kazuya Sato ◽  
Yoshihiro Torimoto ◽  
Yasuyuki Iuchi ◽  
Yasuaki Tamura ◽  
Junko Jimbo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
Bone Marrow Cells ◽  
Leukemia Cell ◽  
Class I ◽  
Biochemical Data ◽  
Leukemia Cells ◽  
Combined Use ◽  
Minimal Residual

Abstract Background: Heat shock proteins (HSPs) are molecular chaperones binding a broad repertorie of endogenous antigenic peptides and carrying them to the MHC. Because the identification of each tumor specific antigen is not necessary, the immunotherapy using HSPs is more practical than other immunological procedures. Meanwhile, relapse due to minimal residual disease (MRD) is a big problem of autologous stem cell transplantation (SCT) against leukemia. We previously reported that immunotherapy using leukemia cell-derived HSPs is effective against MRD after syngeneic bone marrow transplantation (BMT) in mice (Sato et al. Blood, 2001). However, patients receiving SCT are usually immunocompromised due to repeated anti-cancer therapies. Accordingly, it is important to enhance the cytotoxicities (CTXs) against leukemia. Dendritic cells (DCs) are known as professional antigen-presenting cells with a specific receptor for HSPs and are expected to play a major role in immunotherapy. In this study, we evaluated whether the vaccination of DCs pulsed with HSP70 enhances the anti-leukemia effect induced by leukemia cell-derived HSP70 after syngeneic BMT and evaluated the safety of this immunotherapy. Methods: Three class-I-identical mouse tumor cell lines (A20: B-cell leukemia; T27A: myeloid leukemia; colo26: colonic carcinoma) and syngeneic balb/c mice were used in this study. HSP70 was purified from tumor cells. DCs were generated from bone marrow cells cultured with GM-CSF. DCs were pulsed with HSP70 (HSP70-pulsed-DCs) in vitro. Mice were received total body irradiation (TBI) and transplanted bone marrow cells after TBI, then inoculated 2.5 x 104 A20 cells intravenously. HSP70 or HSP70-pulsed-DCs was subcutaneously administrated. Survival days of immunized mice were compared using Kaplan and Meier methods. CTXs of splenocytes against A20 cells were determined by 51Cr release assay. Histological findings of liver and knee joint and biochemical data of serum of immunized mice were investigated. Results: All mice without immunization or immunized with DCs alone died from leukemic dissemination within 90 days after A20 inoculation, whereas mice immunized with A20-derived HSP70 (A20-HSP70) or A20-HSP70-pulsed-DCs survived long significantly. Additionally, although only 60% of the mice immunized with A20-HSP70 survived on day 120, all the mice immunized with A20-HSP70-pulsed-DCs survived with no residual leukemia cells over 120 days. Moreover, splenocytes of mice immunized with A20-HSP70-pulsed-DCs showed significantly higher CTXs against A20 cells in vitro compare to those with A20-HSP70 alone. However, no CTXs against A20 cells were induced by immunization with colo26-or T27A-HSP70-pulsed-DCs. These CTXs against A20 cells were significantly blocked by anti-CD8 and anti-MHC class-I, but not by anti-CD4. Additionally, no abnormal findings were detected either in biochemical data of serum or in histopathology of liver and joint tissue in long term immunized mice. Conclusions: Combined use of dendritic cells with leukemia cell-derived HSP70 enhances anti-leukemia effect by inducing specific cytotoxic activities against leukemia cells, and eradicates MRD effectively and safely even for immunocompromised status after syngeneic BMT. This approach would be useful for a further application of HSP in leukemia-patients after autologous SCT.

scholarly journals The Effect of Influenza Vaccines on Maturation of Dendritic Cells Generated from Bone Marrow

2021 ◽  
Vol 5 (1) ◽  
Author(s):  
Kostinova AM ◽  
◽  
Yukhacheva DV ◽  
Akhmatova EA ◽  
Akhmatova NK ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
Bone Marrow Cells ◽  
Human Bone ◽  
Influenza Vaccines ◽  
Human Bone Marrow ◽  
Vitro Model ◽  
Cytokine Stimulation ◽  
Marrow Cells

Background: Possibility to control immune system by regulating the activity of Dendritic Cells (DC) with the help of vaccines or other immunobiological drugs opens great prospects for infectious, oncological and autoimmune control. The aim of this study was to evaluate in vitro the effect of adjuvant subunit and non-adjuvant split influenza vaccines on maturation of DCs from human bone marrow. Methods: From bone marrow cells of healthy volunteers, DCs were obtained using rGM-CSF and IL-4. On the 8th day of cultivation, 10μl of vaccines against influenza were introduced into the culture of Immature DCs (i-DCs): a non-adjuvant split vaccine (Vaxigripp, Sanofi Pasteur) and an immunoadjuvant subunit vaccine (Grippol plus, Petrovax), as well as immunomodulator Polyoxidonium. Results: Insertion of influenza vaccines into i-DC culture induced the acquisition by DCs typical morphological signs of maturation. DCs became large with eccentrically located of irregular shape nucleus, densified cytoplasm, numerous processes. By immunophenotypic examination decrease in monocyte/macrophage pool, cells with expression of CD34 immaturity marker, increase in expressing CD11c/CD86 costimulatory molecules and CD83 terminal differentiation molecules were observed. Although Polyoxidonium caused a decrease in number of CD11c/CD14 cells (18, 5%), but compared to vaccines, its activity was lower (p<0, 05). Grippol plus more actively induced differentiation of TLR2 and TLR8 expressing cells, whereas Vaxigripp-expression of TLR4 and TLR8 on DCs. Conclusion: The possibility of using in vitro model of DCs obtained from human bone marrow cells by cytokine stimulation for examination of the ability of influenza vaccines to induce DC maturation processes has been demonstrated.

CREB Plays a Critical Role in the Regulation of Normal and Malignant Hematopoiesis.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1224-1224
Author(s):  
Jerry C. Cheng ◽  
Dejah Judelson ◽  
Kentaro Kinjo ◽  
Jenny Chang ◽  
Elliot Landaw ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Cell Proliferation ◽  
Progenitor Cells ◽  
Myeloid Leukemia ◽  
Bone Marrow Cells ◽  
Leukemia Cells ◽  
Mouse Bone Marrow ◽  
T315i Mutation

Abstract The cAMP Response Element Binding Protein, CREB, is a transcription factor that regulates cell proliferation, memory, and glucose metabolism. We previously demonstrated that CREB overexpression is associated with an increased risk of relapse in a small cohort of adult acute myeloid leukemia (AML) patients. Transgenic mice that overexpress CREB in myeloid cells develop myeloproliferative/myelodysplastic syndrome after one year. Bone marrow cells from these mice have increased self-renewal and proliferation. To study the expression of CREB in normal hematopoiesis, we performed quantitative real-time PCR in both mouse and human hematopoietic stem cells (HSCs). CREB expression was highest in the lineage negative population and was expressed in mouse HSCs, common myeloid progenitors, granulocyte/monocyte progenitors, megakaryocyte/erythroid progenitors, and in human CD34+38- cells. To understand the requirement of CREB in normal HSCs and myeloid leukemia cells, we inhibited CREB expression using RNA interference in vitro and in vivo. Bone marrow progenitor cells infected with CREB shRNA lentivirus demonstrated a 5-fold decrease in CFU-GM but increased Gr-1/Mac-1+ cells compared to vector control infected cells (p<0.05). There were fewer terminally differentiated Mac-1+ cells in the CREB shRNA transduced cells (30%) compared to vector control (50%), suggesting that CREB is critical for both myeloid cell proliferation and differentiation. CREB downregulation also resulted in increased apoptosis of mouse bone marrow progenitor cells. Given our in vitro results, we transplanted sublethally irradiated mice with mouse bone marrow cells transduced with CREB or scrambled shRNA. At 5 weeks post-transplant, we observed increased Gr-1+/Mac-1+ cells in mice infused with CREB shRNA transduced bone marrow compared to controls. After 12 weeks post-transplant, there was no difference in hematopoietic reconstitution or in the percentage of cells expressing Gr-1+, Mac-1+, Gr-1/Mac-1+, B22-+, CD3+, Ter119+, or HSCs markers, suggesting that CREB is not required for HSC engraftment. To study the effects of CREB knockdown in myeloid leukemia cells, K562 and TF-1 cells were infected with CREB shRNA lentivirus, sorted for GFP expression, and analyzed for CREB expression and proliferation. Within 72 hours, cells transduced with CREB shRNA demonstrated decreased proliferation and survival with increased apoptosis. In cell cycle experiments, we observed increased numbers of cells in G1 and G2/M with CREB downregulation. Expression of cyclins A1 and D, which are known target genes of CREB, was statistically significantly decreased in TF-1 and K562 cells transduced with CREB shRNA lentivirus compared to controls. To study the in vivo effects of CREB knockdown on leukemic progression, we injected SCID mice with Ba/F3 cells expressing bcr-abl or bcr-abl with the T315I mutation and the luciferase reporter gene. Cells were transduced with either CREB or scrambled shRNA. Disease progression was monitored using bioluminescence imaging. The median survival of mice injected with CREB shRNA transduced Ba/F3 bcr-abl or bcr-abl with the T315I mutation was increased with CREB downregulation compared to controls (p<0.05). Our results demonstrate that CREB is a critical regulator of normal and neoplastic hematopoiesis both in vitro and in vivo.

ARC Is Regulated by MAPK and PI3K and Confers Drug Resistance and Survival Advantage to AML in Vitro and in Vivo

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 893-893
Author(s):  
Po Yee Mak ◽  
Duncan H Mak ◽  
Yuexi Shi ◽  
Vivian Ruvolo ◽  
Rodrigo Jacamo ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Leukemia Cell ◽  
Leukemia Cells ◽  
Apoptosis Resistance ◽  
Control Mscs ◽  
Extrinsic Apoptosis ◽  
Induced Apoptosis ◽  
And Control

Abstract Abstract 893 ARC (Apoptosis repressor with caspase recruitment domain) is a unique antiapoptotic protein that has been shown to suppress the activation of both intrinsic and extrinsic apoptosis. We previously reported that ARC is one of the most potent adverse prognostic factors in AML and that high ARC protein expression predicted shorter survival and poor clinical outcome in patients with AML (Carter BZ et al., Blood 2011). Here we report how ARC is regulated and its role in inhibition of AML apoptosis and in cell survival. We provide evidence that ARC expression is regulated by MAPK and PI3K signaling. Inhibition of MAPK and PI3K pathways decreased ARC mRNA and protein levels in AML cells. ARC expression in AML cells is upregulated in co-cultures with bone marrow-derived mesenchymal stromal cells (MSCs) and the upregulation is suppressed in the presence of MAPK or PI3K inhibitors. To investigate the role of ARC in apoptosis resistance in AML, we generated stable ARC overexpressing (O/E) KG-1 and stable ARC knock down (K/D) OCI-AML3 and Molm13 cells and treated them with Ara-C and agents selectively inducing intrinsic (ABT-737) or extrinsic (TRAIL) apoptosis. We found that ARC O/E cells are more resistant and ARC K/D cells more sensitive to Ara-C, ABT-737, and TRAIL-induced apoptosis: EC50s of Ara-C, ABT-737, or TRAIL treatment at 48 hours for ARC O/E KG-1 and control cells were 1.5 ± 0.1 μM vs. 83.5 ± 4.6 nM, 2.2 ± 0.2 μM vs. 60.2 ± 3.1 nM, or 0.97 ± 0.03 μg/mL vs. 0.17 ± 0.08 μg/mL, respectively and for ARC K/D OCI-AML3 and control cells were 0.33 ± 0.02 μM vs. 3.4 ± 0.2 μM, 0.24 ± 0.01 μM vs. 1.3 ± 0.1 μM, or 0.13 ± 0.09 μg/mL vs. 0.36 ± 0.03 μg/mL, respectively. Bone marrow microenvironment is known to play critical roles in AML disease progression and in protecting leukemia cells from various therapeutic agent-induced apoptosis. Leukemia cells were co-cultured with MSCs in vitro study to mimic the in vivo condition. ARC was found to be highly expressed in MSCs and stable ARC K/D MSCs were generated. AML cell lines and primary patient samples were co-cultured with ARC K/D or control MSCs and treated with Ara-C, ABT-737, or TRAIL. Interestingly, ARC K/D MSCs lost their protective activity for leukemia cells treated with these agents. EC50s for OCI-AML3 cells co-cultured with ARC K/D or control MSCs for 48 hours treated with Ara-C, ABT-737, or TRAIL were 1.0 ± 0.04 μM vs. 4.5 ± 0.2 μM, 0.15 ± 0.06 μM vs. 0.53 ± 0.02 μM, or 1.4 ± 0.8 μg/mL vs. 8.1 ± 0.3 μg/mL, respectively. In addition, ARC O/E KG-1 cells grew faster and ARC K/D OCI-AML3 and Molm13 cells and ARC K/D MSCs grew slower than their respective controls. We then injected KG-1 cells into mice and found that NOD-SCID mice harboring ARC O/E KG-1 had significantly shorter survival than mice injected with the vector control KG-1 (median 84 vs. 111 days) as shown in the figure. Collectively, results demonstrate that ARC plays critical roles in AML. ARC is regulated by MSCs through various signaling pathways in AML cells, protects leukemia cells from apoptosis induced by chemotherapy and by agents selectively inducing intrinsic and extrinsic apoptosis. ARC regulates leukemia cell growth in vitro and in vivo. The results suggest that ARC is a potential target for AML therapy. In addition, targeting ARC in MSCs suppresses microenvironmental protection of AML cells. Disclosures: No relevant conflicts of interest to declare.

Stathmin 1 Is Involved In The Highly Proliferative Phenotype Of High-Risk Myelodysplastic Syndromes and Acute Leukemia Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 861-861
Author(s):  
João Agostinho Machado-Neto ◽  
Paula de Melo Campos ◽  
Patricia Favaro ◽  
Mariana Lazarini ◽  
Irene Lorand-Metze ◽  
...  
Keyword(s):  
Bone Marrow ◽  
High Risk ◽  
Bone Marrow Cells ◽  
Hematopoietic Cells ◽  
Leukemia Cell ◽  
Low Risk ◽  
Leukemia Cells ◽  
Healthy Donors ◽  
Stathmin 1 ◽  
Marrow Cells

Abstract Introduction : Stathmin 1, also known as Oncoprotein 18 (OP18) or Leukemia-associated phosphoprotein p18 (LAP18), is an important cytoplasmic microtubule-destabilizing protein that plays a critical role in the process of mitosis, proliferation and accurate chromosome segregation through regulation of microtubule dynamics. High levels of Stathmin 1 have been reported in solid tumors and have been associated with poor prognosis in various types of cancers. The identification of overactive proteins in leukemia cells, compared to normal hematopoietic cells, as well as understanding the molecular and cellular basis of the disease may provide new therapeutic opportunities. Aims: To evaluate Stathmin 1 expression in proliferating and non-proliferating hematopoietic cells, in bone marrow cells from healthy donors and from patients with myelodysplastic syndromes (MDS), acute myeloid leukemia (AML) and acute lymphoid leukemia (ALL). In addition, we evaluated the effect of Stathmin 1 silencing on proliferation and apoptosis in the U937 acute myeloid leukemia cell line. Materials and Methods: A panel of human leukemia cell lines that included myeloid (K562, KU812, NB4, HL60, P39, HEL, U937, KG1 and THP1) and lymphoid cells (Jurkat, MOLT4, Daudi, Raji, Namalwa and Karpas 422) in exponential growth was used. Peripheral blood lymphocytes (PBL) were induced, or not, to proliferate upon PHA stimulation for 72 hours. A total of 30 healthy donors and 117 patients at diagnosis (MDS=52 [low-risk=36, high-risk=16], AML=49, and ALL=16) were included in the study. Stathmin 1 gene and protein expression was evaluated by qPCR and Western blot. Stathmin 1 was stably knocked down with specific shRNA-expressing lentiviral vector and cell growth was examined by MTT assay, clonogenicity by colony formation and apoptosis by AnnexinV/PI. Appropriate statistical analyses were performed; results are expressed as median (minimum- maximum). Results: A higher expression of Stathmin 1 was observed in all leukemia cell lines, when compared with normal non-proliferating hematopoietic cells. We also observed a marked increase in Stathmin 1 expression in PBL induced to proliferate with PHA after 72 hours. Stathmin 1 transcripts were significantly increased in total bone marrow cells from patients with AML (2.01 [0.35-8.88]; p=.0009) and ALL (2.94 [1.16-10.82]; p=.0004), compared with healthy donors (1.01 [0.38-4.08]). No difference in Stathmin 1 expression was observed between healthy donors and MDS patients. When the MDS group was stratified by the WHO classification into low and high-risk MDS, Stathmin 1 expression was significantly higher in the high-risk, when compared with low-risk MDS (1.62 [0.42–3.28] vs. 1.13 [0.36–2.61], p=.03). Similar results were found in isolated CD34+ bone marrow cells, Stathmin 1 transcripts were significantly increased in CD34+ AML cells compared with CD34+ normal cells, and in high-risk compared with low-risk MDS (all p≤.02). Interestingly, 3 out of 5 MDS patients showed a significant increase in Stathmin 1 transcripts after disease progression. Also, a significant positive correlation was observed between percentage of bone marrow blasts and Stathmin 1 expression in MDS patients (p=.03; r=.31). In U937 leukemia cells, Stathmin 1 silencing significantly reduced cell proliferation (p=.02) and clonal growth (p<.0001), but did not modulate apoptosis. Conclusions: Stathmin 1 is overexpressed in high-risk MDS and acute leukemia cells, and is upregulated during MDS progression, suggesting that Stathmin 1 plays a role in the highly proliferative phenotype. Our study adds new insights to the role of Stathmin 1 in leukemogenesis. Future studies are necessary to validate whether Stathmin 1 is a predictive marker for MDS progression, and to determinate whether Stathmin 1 is a “driver” or a “passenger” during malignant transformation. Disclosures: No relevant conflicts of interest to declare.

Type I interferons produced by dendritic cells promote their phenotypic and functional activation

Blood ◽  
2002 ◽  
Vol 99 (9) ◽  
pp. 3263-3271 ◽  
Author(s):  
Maria Montoya ◽  
Giovanna Schiavoni ◽  
Fabrizio Mattei ◽  
Ion Gresser ◽  
Filippo Belardelli ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
T Cell ◽  
Bone Marrow Cells ◽  
Type I Interferons ◽  
Type I ◽  
Type I Ifn ◽  
Type I Ifns

Abstract Resting dendritic cells (DCs) are resident in most tissues and can be activated by environmental stimuli to mature into potent antigen-presenting cells. One important stimulus for DC activation is infection; DCs can be triggered through receptors that recognize microbial components directly or by contact with infection-induced cytokines. We show here that murine DCs undergo phenotypic maturation upon exposure to type I interferons (type I IFNs) in vivo or in vitro. Moreover, DCs either derived from bone marrow cells in vitro or isolated from the spleens of normal animals express IFN-α and IFN-β, suggesting that type I IFNs can act in an autocrine manner to activate DCs. Consistent with this idea, the ability to respond to type I IFN was required for the generation of fully activated DCs from bone marrow precursors, as DCs derived from the bone marrow of mice lacking a functional receptor for type I IFN had reduced expression of costimulatory and adhesion molecules and a diminished ability to stimulate naive T-cell proliferation compared with DCs derived from control bone marrow. Furthermore, the addition of neutralizing anti–IFN-α/β antibody to purified splenic DCs in vitro partially blocked the “spontaneous” activation of these cells, inhibiting the up-regulation of costimulatory molecules, secretion of IFN-γ, and T-cell stimulatory activity. These results show that DCs both secrete and respond to type I IFN, identifying type I interferons as autocrine DC activators.

Adipocytes Secrete Factors That Cause Leukemia Drug Resistance.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1346-1346
Author(s):  
James W Behan ◽  
Jason P Yun ◽  
Marina P Proektor ◽  
Ehsan A Ehsanipour ◽  
Anna Butturini ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Chemotherapy Resistance ◽  
Leukemia Cell ◽  
Conditioned Media ◽  
Leukemia Cells ◽  
Cell Contact ◽  
Human Leukemia ◽  
Bone Marrow Niche

Abstract We have previously shown that obesity is an independent predictor of leukemia relapse in children. We have also shown that obese mice transplanted with syngeneic leukemia cells have poorer survival after chemotherapy, even when they are dosed proportional to body weight. Since interactions between leukemia cells and cells of the bone marrow niche are considered important for chemotherapy resistance and relapse, and adipocytes can comprise ~50% of the bone marrow niche, we developed in vivo and in vitro models to investigate the role of adipocytes in the leukemia microenvironment. Obese C57Bl/6J mice were transplanted with GFP+ murine preB cell ALL (“8093”) cells and then treated with vincristine (0.5 mg/kg/week × 3 weeks). At the time of relapse, we found that GFP+ leukemia cells persisted in the fat pads of the mice. We then developed an in vitro co-culture system in which human or murine leukemia cells were cultured together with adipocytes (differentiated 3T3-L1s). Undifferentiated 3T3-L1 cells, which are fibroblastic in nature, were used as a control. In this model, adipocytes severely diminished the anti-leukemic effect of all chemotherapeutics tested against murine 8093 cells, including vincristine, dexamethasone, nilotinib, daunorubicin, and L-asparaginase. Adipocytes also protected murine T-cell ALL and human SD-1, RCH-ACV, and BV173 cells from vincristine and daunorubicin. Adipocyte protection of leukemia cells occurred independent of cell contact. Further experiments demonstrated that media conditioned by adipocytes was able to protect 8093 cells from a 3-day exposure to 25 nM dexamethasone (viable cells were at 40±12% of their plated value in regular media, 66±17% in fibroblast-conditioned media, and 109±24% in adipocyte-conditioned media, p&lt;0.05). Surprisingly, adipocyte-conditioned media did not protect leukemia cells from daunorubicin. However, media conditioned by the presence of both adipocytes and leukemia cells simultaneously conferred a high degree of resistance to the leukemia cells (n=3, p&lt;0.05 compared to all other media types). In summary, adipose tissue is a reservoir for relapsed leukemia cells in vivo. Adipocytes engender protection from multiple chemotherapies in murine and human leukemia cell lines. Adipocytes secrete factor(s) that confer dexamethasone and daunorubicin resistance to leukemia cells, though for the latter drug it appears that a two-way communication between leukemia and adipocytes may be necessary for this protection. Figure Figure

Eptifibatide Reduced Cell Adhesion and Recruitment Of The Myeloid Sarcoma-Inducing Leukemia Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4220-4220
Author(s):  
Jen-Fen Fu ◽  
Lee-Yung Shih
Keyword(s):  
Bone Marrow ◽  
Cell Adhesion ◽  
Leukemia Cell ◽  
Myeloid Sarcoma ◽  
Leukemia Cells ◽  
Antiplatelet Drug ◽  
Ras Gene ◽  
Cell Cell

AML patients with myeloid sarcoma (MS) usually had a poor outcome. Our clinical data revealed that AML patients harboring MLL/AF10 and RAS gene mutations were associated with MS formation. By using retroviral transduction/transplantation assay, we demonstrated that the mice transplanted with bone marrow (BM) cells carrying cooperating MLL/AF10(OM-LZ) and KRAS-G12C mutations induced MPD-like myeloid leukemia and MS. Gene expression analyses identified Gpr125, an adhesion G protein-coupled receptor, was up-regulated in the cells carrying cooperating mutations than the cells carrying MLL/AF10(OM-LZ) alone. Knockdown of Gpr125 by RNA interference reduced the number and the size of MS, suggesting that Gpr125 was involved in the MS formation. As Gpr125 contains a HormR domain with Lysine-Glycine-Aspartic acid (KGD) motif which is known to involve in the cell-extracellular matrix (ECM) and cell-cell adhesion, we investigated whether a cyclic RGD peptide drug, eptifibatide (Ep), could interfere MS formation. An in vitro cell-ECM binding assay showed that Gpr125 interacted with fibronectin. Ep reduced leukemia cell-fibronectin binding. Ep also reduced homotypic leukemia cell adhesion and leukemia cell-adipocyte adhesion. In vivo assay demonstrated that Ep reduced leukemia cells recruitment to the adipose tissues, spleen and bone marrow. Our results suggested that blocking Gpr125-mediated cell-ECM and cell-cell adhesion might be helpful to interfere MS formation and BM/spleen recruitment of leukemia cells. Disclosures: Off Label Use: Eptifibatide (Integrilin, Millennium Pharmaceuticals, also co-promoted by Schering-Plough/Essex), is an antiplatelet drug of the glycoprotein IIb/IIIa inhibitor class.

Disulfiram, an clinically used anti-alcoholism drug has the potential to target acute myeloid leukemia cells in a copper-dependent manner in vivo

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5040-5040
Author(s):  
Bing Xu ◽  
Rongwei Li ◽  
Huijuan Dong ◽  
Feili Chen ◽  
Yuejian Liu ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Specific Antigen ◽  
Leukemia Cell ◽  
Leukemia Cells ◽  
Control Group ◽  
Acute Myeloid

Abstract Background Disulfiram(DS), an old drug clinically used for alcoholism, was reported to have antitumor effects, recent studies have found that Copper(Cu) can significantly enhance the DS-induced cell death in vitro in a variety of tumor cells. Our previous studies also demonstrated that disulfiram/copper (DS/Cu) couldtarget human leukemia cell lines(like KG1α,Molt4) through the activation of JNK, in vitro. However, there is few report about the ability of DS/Cu in killing cancer cells in vivo. Aims This study aims to explore the effect of DS/Cu on acute myeloid leukemia cell line KG1αin vivo and clarify the underlining mechanism. Methods 6-8 week old female NOD/SCID mice were sublethally irradiated with 2Gy X-ray the day before transplantation, followed by intravenous injection of KG1α cells (1×107 cells) suspended in 0.2 mL of PBS. 5 weeks after transplantation mice were randomly divided into three treatment groups: vehicle (0.9% saline), a combination of DS and Cu daily for 2 weeks, Ara-C alone twice before killing. Mice were sacrificed after 2 weeks treatment with tissues of spleen, liver, bone marrow being observed using histopathology method to detect the invasion of leukemia. The DS/Cu-induced p-c-jun activation was also examined by western blot using tissues of spleen, liver, bone marrow. Statistical analysis was carried out with one-way ANOVA to assess statistical significance (*p < 0.05). Results 4 weeks after transplantation, mice were dispirited with low appetite, down-bent gait, wrinkled fur, slow move, just like suffered from leukemia. What’s more, immature blasts like morphology similar to KG1α were found in the peripheral blood of the mice(11%±3.41). All the mice were sacrificed after 2 weeks treatment, mice in control group were observed with slightly larger spleen and liver with the morphology of invasion of leukemia such as a granular appearance than the other two groups. Histopathology examination showed that leukemia cells infiltrate liver, spleen and bone marrow, and the immunohistochemistry examination found that the leukemia cells in spleen, liver and bone marrow expressed human specific antigen CD45 with the highest expression level in the control group. Moreover, solid tumor could be observed in the peritoneal cavity of two mice in the control group with expression of human specific antigen CD45detected by immunohistochemistry examination. Western blot in this study showed DS/Cu complex induced phosphorylation of c-Jun expression in the spleen, liver and bone marrow. Conclusion DS/Cu complex could effectively target the acute myeloid leukemia cells in the acute leukemia NOD/SCID mice while inhibiting the invasion of leukemia to some extent, and the activation of JNK might play a functional role in DS/Cu mediated antileukemic effects. Disclosures: No relevant conflicts of interest to declare.

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