Oral Administration of the Opium Alkaloid Noscapine (NOS) Exhibits Potent Anti-Tumor Activity Against Non-Hodgkin’s Lymphomas and Myeloma Cell Lines In Vitro and In Vivo in SCID-beige Murine Xenograft Models.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2427-2427
Author(s):  
Owen O’Connor ◽  
J. Mastrella ◽  
E. Smith ◽  
L. Toner ◽  
A. Auerbach ◽  
...  
Keyword(s):  
Oral Administration ◽  
Cell Lines ◽  
Drug Exposure ◽  
Structural Similarity ◽  
Myeloma Cell ◽  
Oral Gavage ◽  
Trypan Blue Exclusion ◽  
Hodgkin's Lymphomas

Abstract Noscapine is an oral opiate alkaloid with structural similarity to colchicine that induces cell cycle arrest and apoptosis. Its mechanism of action appears similar to vincristine (VCN). In a trypan blue exclusion assay, noscapine exhibits increasing potency with increasing duration of exposure in B- and T-cell lymphomas, and myelomas (MM). For example, following 24 hours of exposure to noscapine, the OPM2 (MM), H9 (TCL) and RL (BCL) lines exhibited an IC50 of approximately 10 to 15 mM. Exposure for up to 96 hours effected IC50 of 30 nM, 700 nM, and 500 nM for the OPM2, H9 and RL lines respectively. Across all cell lines studied, increasing the duration of exposure by only a couple of days resulted in multiple log decreases in IC50. Combining noscapine with doxorubicin and 4-hydroxycyclophosphamide in vitro resulted in at least additive or better cytotoxicity. In vivo, both intraperitoneal and oral doses of noscapine were evaluated on a daily schedule for 14 days (200, 300, 400 and 600 mg/kg). These data demonstrated that the toxicity was much greater when administered by the i.p. route, where 60% of animals died as a result of toxicity in the 600 mg/kg i.p. cohort, while none died from toxicity when given by oral gavage. Preliminary efficacy experiments demonstrated that by day 15, animals treated with 600 and 400 mg/kg noscapine by oral gavage exhibited tumor growth delays that were 30 to 50% of the control tumor volume. Activation of caspase 3 by noscapine demonstrated a linear dose response relationship which was strongly related to duration of drug exposure. Noscapine may represent a new well tolerated oral therapy for the control of lymphoma and myeloma. Antimitotic agents with effects like VCN may be improved with higher area under of the curve of exposure, and thus are well suited for oral administration. Ongoing phase I studies are determining the MTD and DLT associated with noscapine. Phase 2 studies are warranted.

CYTOTOXICITY, ANTITUMOUR AND ANTICARCINOGENIC ACTIVITY OF CURCUMA LONGA ESSENTIAL OIL

INDIAN DRUGS ◽  
2014 ◽  
Vol 51 (03) ◽  
pp. 28-34
Author(s):  
V.B Liju ◽  
◽  
K Jeena ◽  
R. Kuttan
Keyword(s):  
Essential Oil ◽  
Oral Administration ◽  
Life Span ◽  
Cell Lines ◽  
Curcuma Longa ◽  
Cytochrome P450 Enzyme ◽  
Dalton’S Lymphoma ◽  
Dalton's Lymphoma

In the present study, we have evaluated the antitumour and anticarcinogenic activity of turmeric essential oil in vivo. Turmeric essential oil was found to have significant in vitro cytotoxic activity against Dalton’s lymphoma ascites cells (DLA) and Ehrlich ascites carcinoma (EAC) cancer cell lines. Concentration needed for 50% cytotoxicity (IC50) was 8 μg for DLA cells and 18 μg to EAC cell lines. Oral administration of turmeric essential oil was found to significantly increase the life span (56.25%) of Dalton’s Lymphoma Ascites (DLA) induced ascites tumour bearing mice as well as significantly reduced (P<0.001) the solid tumours. 3-Methyl cholanthrene induced sarcoma development was also delayed and there was significant increase in the life span of mice after oral administration of turmeric essential oil. Moreover, turmeric essential oil significantly (P<0.001) inhibited phenobarbitone induced cytochrome p450 enzyme activity in rats.

Constitutive and Immunoproteasome Inhibitors Increase IκB and Decrease NF-κB Expression In Dendritic Cell

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3493-3493
Author(s):  
Ahmad-Samer Samer Al-Homsi ◽  
Zhongbin Lai ◽  
Tara Sabrina Roy ◽  
Niholas Kouttab
Keyword(s):  
Multiple Myeloma ◽  
T Cell ◽  
Cell Lines ◽  
Mechanism Of Action ◽  
Peripheral Blood ◽  
Research Funding ◽  
Myeloma Cell ◽  
Multiple Myeloma Cell

Abstract Introduction Constitutive and immunoproteasome inhibitors (C&IPI) were thought to suppress nuclear factor-κB (NF-κB) pathway by preventing IκB degradation, which prevents NF-κB translocation into the nucleus. This mechanism of action has since been questioned by a number of studies. First, bortezomib promoted constitutive NF-κB activity in endothelial cell carcinoma. Second, NF-κB constitutive activity was resistant to bortezomib in multiple myeloma cell lines. Third, bortezomib increased IκB mRNA but post-transcriptionally downregulated IκB in normal cells and in multiple myeloma cell lines resulting in induced canonical NF-κB activation. Lastly, bortezomib increased nuclear levels of IκB as opposed to lowering cytoplasmic levels in cutaneous T cell lymphoma cell line suggesting that nuclear translocation of IκB was possibly responsible for NF-κB inhibition. The inhibitory activity of C&IPI on dendritic cells (DC) is of interest in the prevention of graft versus host disease (GvHD). It has been shown that different C&IPI impede DC maturation and T cell priming both in vitro and in vivo. Herein we sought to understand the mechanism of action of proteasome and immunoproteasome inhibitors on DC and to test their effect on IκB and NF-IκB expression. Materials and Methods We first performed RT PCR on lysates of DC obtained from the peripheral blood of 7 patients who received post-transplant cyclophosphamide and bortezomib as prevention of GvHD on a phase I clinical trial. Patients received allogeneic transplantation from matched-related or unrelated donors. Patients received no other immunosuppressive therapy except for rabbit anti-thymocyte globulin for those receiving graft from unrelated donor. Steroids were not allowed on the study. Samples were obtained on days +1, +4, and +7. The results were analyzed in comparison to samples obtained on day 0 before stem cell infusion. We then performed the same experiment on lysates of DC obtained from the peripheral blood of healthy volunteer donors. DC were untreated or incubated with bortezomib (10 nM for 4 h), carfilzomib (30 nM for 1 h), oprozomib (100 nM and 300 nM for 4 h), ONX 0914 (200 nM for 1 h), PR-825 (125 nM for 1 h), or PR-924 (1000 nM for 1 h). The drug concentration and duration of exposure were chosen based on the IC50 on proteasome activity and to reproduce in vivo conditions. We also performed IκB western blot on DC isolated from peripheral blood of healthy volunteers, untreated or incubated with bortezomib (10 nM for 4 h) or oprozomib (300 nM for 4 h). Each experiment was performed at least in triplicate. Results We found that the combination of cyclophosphamide and bortezomib significantly and progressively increased IκB mRNA while decreasing NF-κB mRNA in DC studied ex vivo. We also found that all studied C&IPI increased IκB mRNA to a variable degree while only oprozomib (300 nM) decreased NF-κB mRNA in DC in vitro. Finally, both bortezomib and oprozomib increased IκB protein level in DC in vitro (figure). Conclusion Our data suggest that C&IPI increase IκB expression in DC. As opposed to the previously reported data in other cell types, the effect is not associated with post-transcriptional downregulation. Cyclophosphamide and bortezomib also decrease NF-κB expression in DC in vivo while only oprozomib had the same effect in vitro. The effect of C&IPI on IκB and NF-κB expression may represent a new mechanism of action and suggests their effect may be cell-type dependent. Disclosures: Al-Homsi: Millennium Pharmaceuticals: Research Funding. Off Label Use: The use of cyclophosphamide and bortezomib for GvHD prevention. Lai:Millennium Pharmaceuticals: Research Funding.

siRNA for the Ig Light Chain Constant Region Reduces Light Chain Production and Secretion By Human Plasma Cells and in a Murine Xenograft Model

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5722-5722
Author(s):  
Xun Ma ◽  
Ping Zhou ◽  
Monika Pilichowska ◽  
Chakra P Chaulagain ◽  
Sandy Wong ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Human Plasma ◽  
Cell Lines ◽  
Light Chain ◽  
Plasma Cells ◽  
Xenograft Model ◽  
Myeloma Cell ◽  
Constant Region

Abstract Background Ig light chain (LC) diseases such as AL amyloidosis and monoclonal light-chain deposition disease are caused by pathologic free LC. Treatment is aimed at eliminating LC production but success is limited. RNA interference (RNAi) can stop LC production but the diversity of LC variable region sequences poses a challenge that targeting consensus sequences in the constant region (CR) of LC mRNA may overcome (Blood 2014;123:3440). We have developed siRNA pools designed to target the κ or λ LC CR mRNA in human plasma cells and impair LC production and secretion, and have shown that the pool targeting the λ LC CR can do so, and can also trigger a terminal unfolded protein response in clones producing intact Ig due to intracellular accumulation of unpaired heavy chains (ibid). Here we report the results of continued in vitro and in vivo testing of these pools in patient specimens and in a murine xenograft model. Methods Pools of siRNA for the κ or λ LC CR (si[IGLCκCR], si[IGLCλCR]) were custom produced with a non-target control (si[-]). They were introduced in vitro into human plasma cells by an optimized streptolysin O-based method (SLO) and in a NOD.SCID xenograft flank plasmacytoma model by in vivo electroporation as per Gene Therapy 2011;18:1150. In vitro we evaluated LC gene expression, production and secretion at 24 hours in human myeloma cell lines and CD138-selected specimens from patients with plasma cell neoplasms, using real-time PCR (qPCR) for LC mRNA, flow cytometry for intracellular LC mean fluorescence intensity (MFI) and ELISA (Bethyl Laboratories) for LC secretion in 24-hour suspension cultures (106 cells/ml). In vivo we inoculated each of the flanks of NOD.SCID mice with 107 human myeloma cells (ALMC-1 or ALMC-2). When plasmacytomas were 0.5cm3 we injected si[IGLCλCR] or si[-] one time to each flank plasmacytoma respectively, allowing each mouse to serve as its own control. Two days later, the mice were sacrificed and the plasmacytomas excised for qPCR for λ LC mRNA and serum was obtained to measure human λ LC levels by ELISA. Results We have previously described results with siRNA targeting the λ LC CR in human cell lines that make λ LC (ALMC-1, ALMC-2, EJM, OPM2, MM.1S, and MM.1R) and in 16 AL λ patient specimens. We demonstrated significant decreases in LC mRNA, intracellular LC MFI, and λ LC secretion by cell lines (Blood 2014;123:3220); moreover, transcriptional profiling indicated minimal off-target effects (ibid; Supplement). We now report that in vitro secretion of λ LC by CD138-selected plasma cells from AL patients (n=3, newly diagnosed λ) treated with si[IGLCλCR] was reduced by 65% from a mean of 3.1 to 1.0µg/ml and that the residual λ LC mRNA was 49% of control. Similarly we treated κ LC secreting human myeloma cell lines with si[IGLCκCR] and si[-] (IM9, H929, JJN-3, and ARH77). By qPCR the residual κ LC mRNA was 13%, by flow cytometry the MFI was reduced by a median of 67.3% (22.5-90.8), and by ELISA mean κ LC secretion was reduced from 3.7 to 0.8µg/ml (P = 0.055, paired t test). We treated CD138-selected κ patient samples (AL 3, LCDD 1, MM 6) in the same way. By qPCR the residual κ LC mRNA was 57% control, by flow cytometry the MFI was reduced by a median of 37.5% (14-69.8), and by ELISA secretion was reduced from 9.4 to 6.5µg/ml (P = 0.02, paired t test). In the murine dual-flank xenograft model employing λ secreting cells, by qPCR there was a reduction in λ LC mRNA with si[IGLCλCR] treatment in 13 of 16 mice (ALMC-1 11/114, ALMC-2 2/2). In these 14, the median λ LC expression was 66% of control (range, 17-97). In 6/13 the average reduction in λ LC expression was 59%. Of note, measurable levels of human λ LC were found in the blood of all mice at sacrifice. Conclusion With one pool of siRNA targeting the constant region of the κ or λ LC we can significantly reduce production and secretion of LC by clonal human plasma cells, including patient cells, and also reduce the expression of LC in xenograft plasmacytomas in vivo. Two methods of siRNA delivery have been employed in this work thus far, SLO and in vivo electroporation, neither of which require endosomal escape. The specificity of the siRNA pools for plasma cell LC genes and the possible receptivity of plasma cells to RNAi are important positive aspects of this work. Further pre-clinical development of Ig LC CR RNAi employing lipid-based nanoparticle platforms is warranted in order to optimize cell-specific delivery, delivery efficiency and siRNA targeting. Disclosures No relevant conflicts of interest to declare.

Effect of a novel agent, SL-401, targeting interleukin-3 (IL-3)-receptor on plasmacytoid dendritic cell (pDC)-induced myeloma cell growth and drug resistance.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. 8582-8582
Author(s):  
Dharminder Chauhan ◽  
Arghya Ray ◽  
Christopher Brooks ◽  
Eric K. Rowinsky ◽  
Kenneth Carl Anderson
Keyword(s):  
Drug Resistance ◽  
Cell Growth ◽  
Cell Lines ◽  
Plasmacytoid Dendritic Cell ◽  
Myeloma Cell ◽  
Healthy Donors ◽  
Signaling Axis ◽  
Novel Therapeutic Strategies

8582 Background: Multiple myeloma (MM) remains incurable despite novel therapies, highlighting the need for further identification of factors mediating disease progression and drug resistance. The bone marrow (BM) microenvironment confers growth, survival, and drug resistance in MM cells. Our recent study utilized in vitro and in vivo MM xenograft models to show that plasmacytoid dendritic cells (pDCs) were significantly increased in MM BM and promote MM growth (Chauhan et al., Cancer Cell 2009, 16:309). Importantly, we found increased IL-3 levels upon pDC-MM interaction, which in turn, trigger MM cell growth and pDCs survival. IL-3R is highly expressed on pDCs. We utilized SL-401, a novel biologic conjugate that targets IL-3R, to examine whether abrogation of IL-3–IL-3R signaling axis affects pDC-MM interaction and its tumor promoting sequelae. Methods: MM cell lines, patient MM cells, and pDCs from healthy donors or MM patients were utilized to study the anti-MM activity of SL-401. MM cells and pDCs were cultured alone or together in the presence or absence of SL-401, followed by analysis of cell growth or viability. Results: SL-401 significantly decreased the viability of pDCs at low concentrations (IC50: 0.83 ng/ml; P < 0.005, n = 3). SL-401 also decreased the viability of MM cells at clinically achievable doses. Co-culture of pDCs with MM cells induced growth of MM cell lines; and importantly, low doses (0.8 ng/ml) of SL-401 blocked MM cell growth-promoting activity of pDCs. MM patient-derived pDCs induced growth of MM cell lines and primary MM cells as well; conversely, SL-401 inhibited pDC-triggered MM cell growth (P < 0.005, n= 5). Tumor cells from 3 of the 5 patients were from patients whose disease was progressing while on bortezomib, dexamethasone, and lenalidomide therapies. In agreement with these results, SL-401 blocked pDC-induced growth of dexamethasone-resistant MM cell lines. Conclusions: Our study therefore provides the basis for directly targeting pDCs or blocking the pDC-MM interaction, as well as targeting MM, in novel therapeutic strategies with SL-401 to enhance MM cytotoxicity, overcome drug-resistance, and improve patient outcome.

Cytotoxic effect in vivo and in vitro of CHS 828 on human myeloma cell lines

2004 ◽  
Vol 15 (1) ◽  
pp. 63-70 ◽  
Author(s):  
Peter Hovstadius ◽  
Elin Lindhagen ◽  
Sadia Hassan ◽  
Kenneth Nilsson ◽  
Helena Jernberg-Wiklund ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cytotoxic Effect ◽  
Myeloma Cell ◽  
Chs 828 ◽  
Human Myeloma Cell

scholarly journals The Combination of CUDC-907 and Gilteritinib Shows Promising Antileukemic Activity in Vitro and In Vivo in Preclinical Models of FLT3-ITD AML

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 1262-1262
Author(s):  
Tristan Knight ◽  
Xinan Qiao ◽  
Jun Ma ◽  
Holly Edwards ◽  
Lisa Polin ◽  
...  
Keyword(s):  
Combination Therapy ◽  
Mouse Model ◽  
Cell Lines ◽  
Vehicle Control ◽  
Antileukemic Activity ◽  
Oral Gavage ◽  
In Vivo Efficacy ◽  
Flt3 Expression

Introduction FMS-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) mutations are found in approximately one quarter of acute myeloid leukemia (AML) cases. Its presence results in constitutive activation of the FLT3 receptor tyrosine kinase and its downstream growth/pro-survival pathways including MAPK/ERK, PI3K/AKT, and JAK/STAT, and confers a poor prognosis. Gilteritinib is a selective inhibitor of FLT3 recently approved by the Food and Drug Administration for treatment of patients with relapsed/refractory AML and a FLT3 mutation. However, gilteritinib exposure induces upregulation of FLT3 - a mechanism of resistance. Previously, we showed that CUDC-907, a dual PI3K/histone deacetylase inhibitor, downregulates FLT3 expression (Li X, et al. Haematologica. 2019; epub ahead of print). We therefore hypothesized that combining CUDC-907 with gilteritinib would abrogate FLT3 upregulation and expression, resulting in synergistic antileukemic activities against FLT3-mutated AML. Methods FLT3-ITD AML cell lines and primary patient samples were treated with gilteritinib or CUDC-907, alone or in combination at clinically achievable concentrations, and subjected to annexin V/propidium iodide staining and flow cytometry analysis to quantify apoptosis. Protein levels of FLT3, Bcl-2 family proteins, and key components of the MAPK/ERK, PI3K/AKT, and JAK/STAT pathways were examined using western blotting. The impact of the observed alterations upon apoptosis were confirmed via overexpression, knockdown, and targeted inhibitor experiments. Real-time RT-PCR was used to determine FLT3 transcript levels. The FLT3-ITD AML cell line MV4-11 was used to generate a xenograft mouse model to assess in vivo efficacy of the two agents. Results CUDC-907 and gilteritinib demonstrated potent synergistic antileukemic effects in FLT3-ITD AML cell lines in vitro and patient samples ex vivo, with combined therapy. CUDC-907 abolished gilteritinib-induced expression of FLT3 in both cell lines and primary patient samples. Gilteritinib treatment reduced p-AKT, p-S6, and p-STAT5 and increased p-ERK, while CUDC-907 reduced p-AKT and p-ERK, and upregulated p-STAT5. The combination of gilteritinib and CUDC-907 decreased not only p-AKT and p-S6, but also p-ERK and p-STAT5. Targeted inhibition of ERK and JAK2/STAT5 signaling by SCH772984 and AZD1480, respectively, confirmed their roles in resistance to gilteritinib and CUDC-907 monotherapies, respectively. Combined gilteritinib and CUDC-907 treatment reduced expression of the anti-apoptotic BCL-2 family member Mcl-1 and increased expression of the pro-apoptotic protein Bim. MCL-1 overexpression and BIM knockdown partially rescued FLT3-ITD AML cells upon drug treatment, confirming their role in the antileukemic activity of combined gilteritinib and CUDC-907. To determine in vivo efficacy of the two agents, NSGS mice were injected with MV4-11 cells. Three days later, the mice were randomized into vehicle control (n=5), 40 mg/kg gilteritinib (oral gavage; n=5), 100 mg/kg CUDC-907 (oral gavage; n=5) or combination (40 mg/kg gilteritinib + 100 mg/kg CUDC-907; n=6) groups. CUDC-907 was given daily for 5 days on, 2 days off, for a total of 4 cycles. Gilteritinib was administered daily for 28 days. Both agents were well tolerated; maximal weight loss was 5.5%, 0.9%, and 6.7% in the CUDC-907, gilteritinib, and combination groups, respectively. Median survival of mice in the vehicle control group was 43 days. Median survival in the CUDC-907 monotherapy and gilteritinib monotherapy arm was 40.5 days and 104 days, respectively. One mouse in the combination therapy arm died on day 138, while the remaining 5 mice in the combination therapy arm continue to survive, as of time of writing (day 168), and are asymptomatic (Figure 1). Conclusion We confirmed that the combination of CUDC-907 plus gilteritinib synergistically induces apoptosis in both FLT3-ITD AML cell lines and primary patient samples, and that gilteritinib-induced FLT3 expression is abolished by CUDC-907. Cooperative inhibition of the PI3K-AKT, JAK-STAT, and RAS-RAF pathways, as well as upregulation of Bim/downregulation of Mcl-1 all appear to contribute to this observed antileukemic synergy. Our cell line-derived xenograft mouse model provides strong evidence of in vivo efficacy and robust grounds for clinical translation of this therapeutic combination. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Effects of PI3K-δ/γ Inhibitor, Duvelisib, in Mantle Cell Lymphoma in Vitro and in Patient-Derived Xenograft Studies

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 3016-3016 ◽  
Author(s):  
Jack Wang ◽  
Victoria Zhang ◽  
Taylor Bell ◽  
Yang Liu ◽  
Hui Guo ◽  
...  
Keyword(s):  
Growth Inhibition ◽  
Mantle Cell Lymphoma ◽  
Cell Lines ◽  
Cell Lymphoma ◽  
Oral Gavage ◽  
Patient Derived Xenograft ◽  
Pdx Model ◽  
Mantle Cell

Abstract Background: Mantle cell lymphoma (MCL) is an incurable subtype of B-cell lymphoma. Ibrutinib, a first-in-class, once-daily, oral covalent inhibitor of Bruton's tyrosine kinase (BTK) was approved by the FDA for the treatment of MCL in patients previously treated. In our prior multicenter Phase 2 clinical trial, the overall response rate in relapsed/refractory MCL was 68%, with a median progression-free survival (PFS) of 13.9 months. However, the majority of MCL patients treated with ibrutinib relapsed; in these relapsed patients, the one-year survival rate was only 22%. Therefore, there exists an urgent need for additional novel targeted therapies to improve the mortality rate in these patients. In this study, we assessed the in vitro and in vivo effects of duvelisib, a PI3K-δ,-γ inhibitor, in MCL. Methods: The PI3K/AKT/mTOR and other cell survival signaling pathways were investigated by RNASeq and reverse phase protein array (RPPA) in ibrutinib-sensitive and -resistant MCL samples. The expression of PI3K isoforms, α, β, γ, and δ was tested in 11 MCL cell lines, patient and patient-derived xenograft (PDX) MCL cells by western blot analysis. We then investigated the growth inhibition and apoptosis of duvelisib (IPI-145, Infinity Pharmaceuticals, Inc.) in MCL cells by CellTiter-Glo® Luminescent Cell Viability Assay (Promega) and Annexin V-binding assay (BD Biosciences). We established a primary MCL-bearing PDX model and passaged the primary MCL tumor to next generations. Mice were administrated with 50 mg/kg duvelisib daily by oral gavage. Tumor burden and survival time were investigated in the MCL-PDX model. Results: We found that the PI3K/AKT/mTOR signaling pathway was activated in both primary and acquired ibrutinib-resistant MCL cell lines and PDX MCL cells. We immunoblotted PI3K isoforms, α, β, γ, and δ in 11 MCL cell lines and the result demonstrated that both ibrutinib-sensitive and ibrutinib-resistant MCL cells dominantly expressed PI3K-δ and -γ. Next, we tested the effects of duvelisib on these MCL cells. Duvelisib had effects on the growth inhibition and apoptosis in both ibrutinib-sensitive and ibrutinib-resistant MCL cells as good as the PI3K-δ inhibitor, idelalisib (Cal-101, GS-1101). The PI3K-δ isoform could play a very important role in PI3K-mediated signals in MCL. We then investigated the effects of duvelisib in vivo through our established MCL-bearing PDX mouse models. These models are created by inoculating the primary tumor cells from MCL patients into a human fetal bone chip implanted into NSG mice to provide a microenvironment that reconstitutes the human environment. MCL tumor mass was then passaged to next generations for therapeutic investigation of duvelisib. Mice were treated with 50 mg/kg duvelisib daily by oral gavage. Our data demonstrated that duvelisib significantly inhibited tumor growth and prolonged survival of MCL-PDX mice. Conclusion: Duvelisib, an oral dual inhibitor of PI3K-δ,-γ, inhibits MCL growth both in vitro and in PDX mice. These preclinical results suggests duvelisib may be effective in the treatment of patients with relapsed/refractory MCL. Disclosures No relevant conflicts of interest to declare.

Mcl-1 Is Overexpressed in Multiple Myeloma and Correlates with Disease Severity.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1419-1419
Author(s):  
Soraya Wuilleme-Toumi ◽  
Nelly Robillard ◽  
Patricia Gomez-Bougie ◽  
Philippe Moreau ◽  
Steven Le Gouill ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
Disease Severity ◽  
Cell Lines ◽  
Plasma Cells ◽  
Myeloma Cell ◽  
Lymphocytic Leukemia ◽  
Myeloma Cells

Abstract Multiple Myeloma (MM) is a fatal malignancy of B-cell origin characterized by the accumulation of plasma cells within the bone marrow. The expression of the pro-survival members of the Bcl-2 family has been shown to be a key process in the survival of myeloma cells. More particularly, Mcl-1 expression turned out to be critical for their survival. Indeed, knockdown of Mcl-1 by antisenses induces apoptosis in myeloma cells. Finally, Mcl-1 was found to be the only anti-apoptotic Bcl-2 family member which level of expression was modified by cytokine treatment of myeloma cells. For these reasons, we have evaluated the expression of Mcl-1 in vivo in normal, reactive and malignant plasma cells (PC) i.e., myeloma cells from 55 patients with MM and 20 human myeloma cell lines using flow cytometry. We show that Mcl-1 is overexpressed in MM in comparison with normal bone marrow PC. Forty-seven percent of patients with MM at diagnosis (p=.017) and 80% at relapse (p=.014 for comparison with diagnosis) overexpress Mcl-1. Of note, only myeloma cell lines but not reactive plasmocytoses have abnormal Mcl-1 expression, although both plasmocyte expansion entities share similar high proliferation rates (&gt;20%). Of interest, Bcl-2 as opposed to Mcl-1, does not discriminate malignant from normal PC. This shows that the overexpression of Mcl-1 is clearly related to malignancy rather than to proliferation. It will be important to know whether the overexpression of Mcl-1 is related to an abnormal response to cytokines like Interleukin-6 or to mutations of the promoter of the Mcl-1 gene as already described in B chronic lymphocytic leukemia. Finally, level of Mcl-1 expression is related to disease severity, the highest values being correlated with the shortest event-free survival (p=.01). In conclusion, Mcl-1 which has been shown to be essential for the survival of human myeloma cells in vitro is overexpressed in vivo in MM and correlates with disease severity. Mcl-1 represents a major therapeutical target in MM.

Targeting FGFR3 in t(4;14) Mutiple Myeloma: Pre-Clinical Studies of PRO-001, a Novel Anti-FGFR3 Neutralizing Antibody.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2479-2479
Author(s):  
Suzanne Trudel ◽  
Ellen Wei ◽  
Zhi Hua Li ◽  
Eran Rom ◽  
Ira Chumakov ◽  
...  
Keyword(s):  
Growth Factor ◽  
Cell Lines ◽  
Neutralizing Antibody ◽  
Myeloma Cell ◽  
Chromosomal Translocations ◽  
Minimal Growth ◽  
Erk Phosphorylation ◽  
Human Myeloma Cell

Abstract As with other B-cell malignancies, chromosomal translocations to the immunoglobulin heavy-chain (IgH) locus on chromosome 14q32 are believed to be a hallmark of multiple myeloma (MM), occurring in approximately 50% of patients. Identification of these chromosomal translocations has resulted in the discovery of powerful prognostic tools and novel molecular targets that promise to revolutionize the treatment of this malignancy. Five recurrent translocation partners have been defined, resulting in the dysregulation of the genes encoding cyclin D1 and D3, c-maf, mafB and Fibroblast Growth Factor Receptor 3 (FGFR3) together with MMSET. Genetic analysis of 14q32 translocations in MM has identified distinct groups of patients with separate clinical outcomes supporting a biological correlation of these genes in MM. In particular, the t(4;14) translocation portends a particularly bad prognosis. The association of FGFR3 expression with t(4;14) myeloma and the demonstration of the transforming potential of this receptor tyrosine kinase (RTK), make this a particularly attractive target for drug development for this poor prognosis group. We report here the development of a novel and highly specific anti-FGFR3 neutralizing antibody (PRO-001) isolated from a phage display human combinatorial antibody library. PRO-001 binds with high affinity (Kd=1.3 nM) to FGFR3 in in vitro binding assays and blocks ligand-dependent and independent FGFR3 phosphorylation and signal transduction in cell-based assays. Furthermore, PRO-001 potently inhibits FGFR3-dependent solid tumor growth in mouse xenograft models. We found that PRO-001 bound to, and competed with FGF binding to the surface of FGFR3 on human myeloma cell lines. PRO-001 inhibited FGF-induced phosphorylation of wild-type FGFR3 and downstream ERK phosphorylation in stable B9 cell transfectants (B9-WT) and FGFR3 expressing human myeloma cell lines. The antibody inhibited FGF-mediated growth of B9-WT with an IC50 of 3 μg/ml as determined by MTT proliferation assay. Growth of these cells could be rescued by IL-6 demonstrating selectivity of PRO-001 for FGFR3. PRO-001 inhibited the viability of the FGFR3 expressing, human myeloma cell line, UTMC2. Inhibition of viability was still observed when cells were co-cultured with stroma or in the presence of IL-6, a potent growth factor for MM cells. Several myeloma cell lines lacking FGFR3, showed minimal growth inhibition demonstrating selectivity and lack of non-specific toxic at effective dose concentrations. Finally, PRO-001 bound to FGFR3 on the cell surface, inhibited ERK phosphorylation, and induced cytotoxic responses in primary MM samples derived from t(4;14) positive patients. A xenograft mouse model has been established and studies assessing in vivo activity of PRO-001 are planned and will be reported. Taken together, the data demonstrate that PRO-001 is a specific and potent inhibitor of FGFR3 and that it deserves further study for targeted therapy in MM.

Ascorbic Acid Overcomes Drug Resistance in Myeloma and Significantly Increases the Anti-Myeloma Effects of both Arsenic Trioxide and Melphalan in Vitro and in Vivo.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2470-2470 ◽  
Author(s):  
Richard A. Campbell ◽  
Haiming Chen ◽  
Daocheng Zhu ◽  
Janice C. Santos ◽  
Benjamin Bonavida ◽  
...  
Keyword(s):  
Ascorbic Acid ◽  
Arsenic Trioxide ◽  
Cell Lines ◽  
Synergistic Effects ◽  
Myeloma Cell ◽  
Tumor Burden ◽  
Early Results ◽  
Mtt Assays

Abstract Glutathione levels have previously been shown to be associated with the development of resistance to a variety of anti-myeloma therapies. Ascorbic acid (AA) depletes intracellular glutathione levels which, in turn, should increase the sensitivity of tumor cells to anti-myeloma agents such as arsenic trioxide (ATO) and melphalan. To determine the synergistic effects of combining AA, with ATO and/or melphalan, we evaluated the effects of these combinations with MTT assays on myeloma cell lines in vitro and using our severe combined immunodeficient (SCID)-hu murine myeloma models. We determined the synergistic effects of combining AA with ATO and/or melphalan on the myeloma cell lines RPMI8226, 8226/dox, U266, and U266/dox in vitro. MTT assays demonstrated marked synergistic anti-proliferative effects of AA at 10 mM when added to these cell lines in the presence of ATO concentrations ranging from 5x10−5 M – 5x10−9 M, and melphalan concentrations ranging from 3x10−5 M – 3x10−9 M. In order to provide further evidence for the clinical relevance of these synergistic effects of AA, we investigated the potential of AA to increase the efficacy of current anti-myeloma therapies in our SCID-hu murine model of human myeloma LAGλ–1 (Yang H et al. Blood 2002). Each SCID mouse was implanted with a 0.5 cm3 LAGλ–1 tumor fragment into the left hind limb muscle. Twenty-eight days following implantation, mice then received treatment intraperitoneally (IP) with either AA (300 mg/kg) daily x5/week, ATO (1.25 mg/kg) daily x5/week, or melphalan (3.0 mg/kg) x1/week, or the combination of these agents. AA, ATO, and melphalan alone have no anti-myeloma effects at these doses, whereas AA+melphalan results in significantly decreased tumor burden and paraprotein levels. The most profound anti-myeloma effects were observed in animals treated with all three drugs together. These data show not only the additional synergistic anti-myeloma effects of AA on both ATO and melphalan in vitro but for the first time suggest that these effects are also present in vivo. This provides the rationale for combining AA with these agents in myeloma patients with resistant disease. In support of this, early results of clinical trials using the combination of AA, ATO and low doses of oral melphalan are promising.

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