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1.
Ann Hematol ; 102(1): 107-115, 2023 Jan.
Article in English | MEDLINE | ID: mdl-36369497

ABSTRACT

Ibrutinib revolutionized therapy for relapsed/refractory (R/R) mantle cell lymphoma (MCL). Real-world data on the outcome of unselected patients are still limited. We analyzed 77 R/R MCL patients receiving ibrutinib with at least one prior systemic anti-lymphoma therapy. After a median follow-up of 14.0 months, 56 patients relapsed/progressed, and 45 died. The overall response rate was 66%, with 31% of complete metabolic remissions on PET/CT. The median progression-free and overall survival (OS) rates were 10.3 and 23.1 months, respectively. The median OS from ibrutinib failure was 3.7 months. High proliferation rate by Ki67 (≥ 30%) and two or more previous therapy lines both negatively correlated with outcome (HR = 2.2, p = 0.04, and HR = 2.06, p = 0.08, respectively). Female gender borderline correlated with better outcome (HR = 0.53, p = 0.08). In multivariate analysis, Ki67 and response to ibrutinib both correlated with OS (p < 0.05). Importantly, ibrutinib appeared to better control nodal and extranodal lymphoma than bone marrow (BM) involvement. From 20 patients with detectable BM infiltration (before ibrutinib initiation) achieving complete (n = 13) or partial (n = 7) metabolic remission, none achieved remission in BM. We confirmed good efficacy of ibrutinib in unselected heavily pre-treated MCL patients. Our findings support the use of a combination of ibrutinib and rituximab in patients with BM involvement.


Subject(s)
Lymphoma, Mantle-Cell , Adult , Humans , Female , Lymphoma, Mantle-Cell/pathology , Ki-67 Antigen , Czech Republic , Positron Emission Tomography Computed Tomography
2.
Lab Invest ; 102(9): 957-965, 2022 09.
Article in English | MEDLINE | ID: mdl-35488033

ABSTRACT

Non-Hodgkin lymphomas (NHL) represent the most common hematologic malignancies. Patient-derived xenografts (PDXs) are used for various aspects of translational research including preclinical in vivo validation of experimental treatment approaches. While it was repeatedly demonstrated that PDXs keep majority of somatic mutations with the primary lymphoma samples, from which they were derived, the composition of PDX tumor microenvironment (TME) has not been extensively studied. We carried out a comparative genetic and histopathological study of 15 PDX models derived from patients with various types of NHL including diffuse large B-cell lymphoma (DLBCL; n = 7), Burkitt lymphoma (BL; n = 1), mantle cell lymphoma (MCL; n = 2), and peripheral T-cell lymphomas (PTCL; n = 5). Whole exome sequencing (WES) of the PDXs and primary lymphoma cells was implemented in 13 out of 15 cases with available DNA samples. Standard immunohistochemistry (IHC) was used to analyze the composition of PDX TME. WES data confirmed that PDXs maintained the genetic heterogeneity with the original primary lymphoma cells. In contrast, IHC analysis revealed the following recurrently observed alterations in the composition of PDX tumors: more blastoid lymphoma cell morphology, increased proliferation rate, lack of non-malignant cellular components including T cells and (human or murine) macrophages, and significantly lower intratumoral microvessel density and microvessel area composed of murine vessels. In addition, PDX tumors derived from T-NHL displayed additional differences compared to the primary lymphoma samples including markedly lower desmoplasia (i.e., the extent of both reticular and collagen fibrosis), loss of expression of cytotoxic granules (i.e., perforin, TIA, granzyme B), or loss of expression of T-cell specific antigens (i.e., CD3, CD4, CD8). Our data suggest that despite keeping the same genetic profiles, PDX models of aggressive NHL do not recapitulate the microenvironmental heterogeneity of the original lymphomas. These findings have implications on the relevance of PDX models in the context of preclinical research.


Subject(s)
Antineoplastic Agents , Lymphoma, Large B-Cell, Diffuse , Adult , Animals , Disease Models, Animal , Heterografts , Humans , Mice , Tumor Microenvironment
3.
Blood ; 138(10): 871-884, 2021 09 09.
Article in English | MEDLINE | ID: mdl-33876201

ABSTRACT

Despite the development of novel targeted drugs, the molecular heterogeneity of diffuse large B-cell lymphoma (DLBCL) still poses a substantial therapeutic challenge. DLBCL can be classified into at least 2 major subtypes (germinal center B cell [GCB]-like and activated B cell [ABC]-like DLBCL), each characterized by specific gene expression profiles and mutation patterns. Here we demonstrate a broad antitumor effect of dimethyl fumarate (DMF) on both DLBCL subtypes, which is mediated by the induction of ferroptosis, a form of cell death driven by the peroxidation of phospholipids. As a result of the high expression of arachidonate 5-lipoxygenase in concert with low glutathione and glutathione peroxidase 4 levels, DMF induces lipid peroxidation and thus ferroptosis, particularly in GCB DLBCL. In ABC DLBCL cells, which are addicted to NF-κB and STAT3 survival signaling, DMF treatment efficiently inhibits the activity of the IKK complex and Janus kinases. Interestingly, the BCL-2-specific BH3 mimetic ABT-199 and an inhibitor of ferroptosis suppressor protein 1 synergize with DMF in inducing cell death in DLBCL. Collectively, our findings identify the clinically approved drug DMF as a promising novel therapeutic option in the treatment of both GCB and ABC DLBCLs.


Subject(s)
Dimethyl Fumarate/pharmacology , Ferroptosis/drug effects , Lymphoma, Large B-Cell, Diffuse/metabolism , NF-kappa B/metabolism , Neoplasm Proteins/metabolism , STAT3 Transcription Factor/metabolism , Signal Transduction/drug effects , Animals , Gene Expression Regulation, Neoplastic/drug effects , Humans , Lipid Peroxidation/drug effects , Lipid Peroxidation/genetics , Lymphoma, Large B-Cell, Diffuse/drug therapy , Lymphoma, Large B-Cell, Diffuse/genetics , Lymphoma, Large B-Cell, Diffuse/pathology , Mice , NF-kappa B/genetics , Neoplasm Proteins/genetics , STAT3 Transcription Factor/genetics , Signal Transduction/genetics , Xenograft Model Antitumor Assays , Zebrafish
4.
Ultrasound Med Biol ; 47(4): 1099-1107, 2021 04.
Article in English | MEDLINE | ID: mdl-33455807

ABSTRACT

Tumor oxygenation and vascularization are important parameters that determine the aggressiveness of the tumor and its resistance to cancer therapies. We introduce dual-modality ultrasound and photoacoustic imaging (US-PAI) for the direct, non-invasive real-time in vivo evaluation of oxygenation and vascularization of patient-derived xenografts (PDXs) of B-cell mantle cell lymphomas. The different optical properties of oxyhemoglobin and deoxyhemoglobin make it possible to determine oxygen saturation (sO2) in tissues using PAI. High-frequency color Doppler imaging enables the visualization of blood flow with high resolution. Tumor oxygenation and vascularization were studied in vivo during the growth of three different subcutaneously implanted patient-derived xenograft (PDX) lymphomas (VFN-M1, VFN-M2 and VFN-M5 R1). Similar values of sO2 (sO2 Vital), determined from US-PAI volumetric analysis, were obtained in small and large VFN-M1 tumors ranging from 37.9 ± 2.2 to 40.5 ± 6.0 sO2 Vital (%) and 37.5 ± 4.0 to 35.7 ± 4.6 sO2 Vital (%) for small and large VFN-M2 PDXs. In contrast, the higher sO2 Vital values ranging from 57.1 ± 4.8 to 40.8 ± 5.7 sO2 Vital (%) (small to large) of VFN-M5 R1 tumors corresponds with the higher aggressiveness of that PDX model. The different tumor percentage vascularization (assessed as micro-vessel areas) of VFN-M1, VFN-M2 and VFN-M5 R1 obtained by color Doppler (2.8 ± 0.1%, 3.8 ± 0.8% and 10.3 ± 2.7%) in large-stage tumors clearly corresponds with their diverse growth and aggressiveness. The data obtained by color Doppler were validated by histology. In conclusion, US-PAI rapidly and accurately provided relevant and reproducible information on tissue oxygenation in PDX tumors in real time without the need for a contrast agent.


Subject(s)
Lymphoma, Mantle-Cell/diagnostic imaging , Lymphoma, Mantle-Cell/physiopathology , Neovascularization, Pathologic/diagnostic imaging , Oxygen/metabolism , Photoacoustic Techniques , Ultrasonography, Doppler, Color , Animals , Cell Hypoxia , Female , Hemoglobins/metabolism , Humans , Lymphoma, Mantle-Cell/pathology , Mice , Microvascular Density , Microvessels/diagnostic imaging , Multimodal Imaging , Neoplasm Transplantation , Oxyhemoglobins/metabolism , Tumor Burden
5.
Leuk Lymphoma ; 62(4): 861-867, 2021 04.
Article in English | MEDLINE | ID: mdl-33238780

ABSTRACT

Platelet/endothelial cell adhesion molecule 1 (PECAM-1, CD31) is an immunoglobulin superfamily member expressed on the surface of platelets, leukocytes and endothelial cells. The role of CD31 in biology of lymphomas has not yet been systemically studied. Expression of cell surface CD31 was analyzed by flow cytometry on primary MCL cells isolated from peripheral blood, bone marrow or malignant effusions obtained from 29 newly diagnosed MCL patients. CD31 was significantly more expressed in patients with documented extranodal involvement. Knock-down of CD31 expression in JEKO1 and MINO MCL cell lines hampered their subcutaneous engraftment in immunodeficient mice and prolonged overall survival of intravenously-xenografted animals. In contrast, transgenic overexpression of CD31 accelerated growth of subcutaneous JEKO1 and MINO tumors, shortened overall survival of intravenously-xenografted mice, and resulted in significantly increased frequency of extramedullary murine tissue infiltration Our observations suggest that CD31 facilitate survival and regulate extranodal spread of MCL cells.


Subject(s)
Lymphoma, Mantle-Cell , Adult , Animals , Blood Platelets , Bone Marrow , Endothelial Cells , Humans , Lymphoma, Mantle-Cell/genetics , Mice , Platelet Endothelial Cell Adhesion Molecule-1/genetics
6.
Acta Biomater ; 119: 349-359, 2021 01 01.
Article in English | MEDLINE | ID: mdl-33186784

ABSTRACT

Mantle cell lymphoma (MCL) is a rare subtype of B-cell non-Hodgkin lymphoma (B-NHL) with chronically relapsing clinical course. Implementation of cytarabine (araC) into induction and salvage regimen became standard of care for majority of MCL patients. In this study, tailored N-(2-hydroxypropyl)methacrylamide (HPMA)-based polymer nanotherapeutics containing covalently bound araC (araC co-polymers) were designed, synthesized and evaluated for their anti-lymphoma efficacy in vivo using a panel of six patient-derived lymphoma xenografts (PDX) derived from newly diagnosed and relapsed / refractory (R/R) MCL. While free araC led to temporary inhibition of growth of MCL tumors, araC co-polymers induced long-term disappearance of the engrafted lymphomas with no observed toxicity even in the case of PDX models derived from patients, who relapsed after high-dose araC-based treatments. The results provide sound preclinical rationale for the use of HPMA-based araC co-polymers in induction, salvage or palliative therapy of MCL patients.


Subject(s)
Lymphoma, Mantle-Cell , Adult , Antineoplastic Combined Chemotherapy Protocols , Cytarabine/pharmacology , Humans , Lymphoma, Mantle-Cell/drug therapy , Neoplasm Recurrence, Local , Rituximab/therapeutic use , Treatment Outcome
7.
J Control Release ; 328: 160-170, 2020 12 10.
Article in English | MEDLINE | ID: mdl-32860930

ABSTRACT

B-cell non-Hodgkin lymphomas (B-NHL) represent the most common type of hematologic malignancies in the Western hemisphere. The therapy of all B-NHL is based on the combination of different genotoxic cytostatics and anti-CD20 monoclonal antibody (mAb) rituximab. Unfortunately, many patients relapse after the mentioned front-line treatment approaches. The therapy of patients with relapsed/refractory (R/R) B-NHL represents an unmet medical need. We designed, developed and tested novel actively targeted hybrid mAb-polymer-drug conjugate (APDC) containing anti-CD20, anti-CD38 or anti-CD19 mAbs. Biocompatible copolymers based on N-(2-hydroxypropyl)methacrylamide (HPMA) with cytostatic agent doxorubicin attached via stimuli-sensitive hydrazone bond were employed for the mAb grafting. Anti-lymphoma efficacy of the APDC nanotherapeutics was evaluated in vivo on a panel of three patient-derived lymphoma xenografts derived from two patients with R/R B-NHL and one patient with so far untreated B-NHL. In both PDX models derived from patients with R/R B-NHL, the targeting with anti-CD38 antibody daratumumab demonstrated highly improved anti-lymphoma efficacy compared to the targeting with anti-CD20 rituximab, two experimental anti-CD19 antibodies and non-targeted controls. The results represent a proof-of-concept of a new algorithm of personalized anti-tumor therapy based on highly innovative APDC biomaterials.


Subject(s)
Antineoplastic Agents , Lymphoma, Non-Hodgkin , Pharmaceutical Preparations , Antibodies, Monoclonal/therapeutic use , Antineoplastic Agents/therapeutic use , Humans , Lymphoma, Non-Hodgkin/drug therapy , Polymers/therapeutic use , Rituximab
8.
Clin Cancer Res ; 25(14): 4455-4465, 2019 07 15.
Article in English | MEDLINE | ID: mdl-31004002

ABSTRACT

PURPOSE: Mantle cell lymphoma (MCL) is an aggressive subtype of B-cell non-Hodgkin lymphomas characterized by (over)expression of BCL2. A BCL2-targeting drug, venetoclax, has promising anticancer activity in MCL. We analyzed molecular mechanisms of venetoclax resistance in MCL cells and tested strategies to overcome it. EXPERIMENTAL DESIGN: We confirmed key roles of proapoptotic proteins BIM and NOXA in mediating venetoclax-induced cell death in MCL. Both BIM and NOXA are, however, differentially expressed in cell lines compared with primary cells. First, NOXA protein is significantly overexpressed in most MCL cell lines. Second, deletions of BIM gene harbored by three commonly used MCL cell lines (JEKO-1, MINO, and Z138) were not found by array comparative genomic hybridization using a validation set of 24 primary MCL samples. RESULTS: We demonstrated that MCL1 and NOXA play important roles in mediating resistance to venetoclax. Consequently, we tested an experimental treatment strategy based on cotargeting BCL2 with venetoclax and MCL1 with a highly specific small-molecule MCL1 inhibitor S63845. The combination of venetoclax and S63845 demonstrated synthetic lethality in vivo on a panel of five patient-derived xenografts established from patients with relapsed MCL with adverse cytogenetics. CONCLUSIONS: Our data strongly support investigation of venetoclax in combination with S63845 as an innovative treatment strategy for chemoresistant MCL patients with adverse cytogenetics in the clinical grounds.


Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Drug Synergism , Lymphoma, Mantle-Cell/drug therapy , Myeloid Cell Leukemia Sequence 1 Protein/antagonists & inhibitors , Neoplasm Recurrence, Local/drug therapy , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Pyrimidines/pharmacology , Sulfonamides/pharmacology , Thiophenes/pharmacology , Animals , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Drug Resistance, Neoplasm , Female , Humans , Lymphoma, Mantle-Cell/metabolism , Lymphoma, Mantle-Cell/pathology , Mice , Mice, Inbred NOD , Myeloid Cell Leukemia Sequence 1 Protein/metabolism , Neoplasm Recurrence, Local/metabolism , Neoplasm Recurrence, Local/pathology , Proto-Oncogene Proteins c-bcl-2/metabolism , Xenograft Model Antitumor Assays
9.
Blood ; 130(3): 310-322, 2017 07 20.
Article in English | MEDLINE | ID: mdl-28202458

ABSTRACT

Activated B-cell-like (ABC) and germinal center B-cell-like diffuse large B-cell lymphoma (DLBCL) represent the 2 major molecular DLBCL subtypes. They are characterized by differences in clinical course and by divergent addiction to oncogenic pathways. To determine activity of novel compounds in these 2 subtypes, we conducted an unbiased pharmacologic in vitro screen. The phosphatidylinositol-3-kinase (PI3K) α/δ (PI3Kα/δ) inhibitor AZD8835 showed marked potency in ABC DLBCL models, whereas the protein kinase B (AKT) inhibitor AZD5363 induced apoptosis in PTEN-deficient DLBCLs irrespective of their molecular subtype. These in vitro results were confirmed in various cell line xenograft and patient-derived xenograft mouse models in vivo. Treatment with AZD8835 induced inhibition of nuclear factor κB signaling, prompting us to combine AZD8835 with the Bruton's tyrosine kinase inhibitor ibrutinib. This combination was synergistic and effective both in vitro and in vivo. In contrast, the AKT inhibitor AZD5363 was effective in PTEN-deficient DLBCLs through downregulation of the oncogenic transcription factor MYC. Collectively, our data suggest that patients should be stratified according to their oncogenic dependencies when treated with PI3K and AKT inhibitors.


Subject(s)
Antineoplastic Agents/pharmacology , Gene Expression Regulation, Neoplastic , Lymphoma, Large B-Cell, Diffuse/drug therapy , Oxadiazoles/pharmacology , Piperidines/pharmacology , Protein Kinase Inhibitors/pharmacology , Pyrazoles/pharmacology , Pyrimidines/pharmacology , Pyrroles/pharmacology , Adenine/analogs & derivatives , Agammaglobulinaemia Tyrosine Kinase , Animals , Apoptosis/drug effects , Drug Combinations , Drug Screening Assays, Antitumor , Drug Synergism , Humans , Lymphoma, Large B-Cell, Diffuse/classification , Lymphoma, Large B-Cell, Diffuse/genetics , Lymphoma, Large B-Cell, Diffuse/pathology , Mice , NF-kappa B/antagonists & inhibitors , NF-kappa B/genetics , NF-kappa B/metabolism , Organ Specificity , PTEN Phosphohydrolase/deficiency , PTEN Phosphohydrolase/genetics , Phosphatidylinositol 3-Kinases/genetics , Phosphatidylinositol 3-Kinases/metabolism , Phosphoinositide-3 Kinase Inhibitors , Protein-Tyrosine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/genetics , Protein-Tyrosine Kinases/metabolism , Proto-Oncogene Proteins c-akt/antagonists & inhibitors , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins c-myc/antagonists & inhibitors , Proto-Oncogene Proteins c-myc/genetics , Proto-Oncogene Proteins c-myc/metabolism , Signal Transduction , Xenograft Model Antitumor Assays
11.
Oncoimmunology ; 5(4): e1115940, 2016 Apr.
Article in English | MEDLINE | ID: mdl-27141398

ABSTRACT

Tumor immunotherapy based on the use of chimeric antigen receptor modified T cells (CAR T cells) is a promising approach for the treatment of refractory hematological malignancies. However, a robust response mediated by CAR T cells is observed only in a minority of patients and the expansion and persistence of CAR T cells in vivo is mostly unpredictable.Lenalidomide (LEN) is an immunomodulatory drug currently approved for the treatment of multiple myeloma (MM) and mantle cell lymphoma, while it is clinically tested in the therapy of diffuse large B-cell lymphoma of activated B cell immunophenotype. LEN was shown to increase antitumor immune responses at least partially by modulating the activity of E3 ubiquitin ligase Cereblon, which leads to increased ubiquitinylation of Ikaros and Aiolos transcription factors, which in turn results in changed expression of various receptors on the surface of tumor cells. In order to enhance the effectiveness of CAR-based immunotherapy, we assessed the anti-lymphoma efficacy of LEN in combination with CAR19 T cells or CAR20 T cells in vitro and in vivo using various murine models of aggressive B-cell non-Hodgkin lymphomas (B-NHL).Immunodeficient NSG mice were transplanted with various human B-NHL cells followed by treatment with CAR19 or CAR20 T cells with or without LEN. Next, CAR19 T cells were subjected to series of tests in vitro to evaluate their response and signaling capacity following recognition of B cell in the presence or absence of LEN.Our data shows that LEN significantly enhances antitumor functions of CAR19 and CAR20 T cells in vivo. Additionally, it enhances production of interferon gamma by CAR19 T cells and augments cell signaling via CAR19 protein in T cells in vitro. Our data further suggests that LEN works through direct effects on T cells but not on B-NHL cells. The biochemical events underlying this costimulatory effect of LEN are currently being investigated. In summary, our data supports the use of LEN for augmentation of CAR-based immunotherapy in the clinical grounds.

12.
Clin Cancer Res ; 22(5): 1138-49, 2016 Mar 01.
Article in English | MEDLINE | ID: mdl-26467384

ABSTRACT

PURPOSE: To investigate the roles of BCL2, MCL1, and BCL-XL in the survival of diffuse large B-cell lymphoma (DLBCL). EXPERIMENTAL DESIGNS: Immunohistochemical analysis of 105 primary DLBCL samples, and Western blot analysis of 18 DLBCL cell lines for the expression of BCL2, MCL1, and BCL-XL. Pharmacologic targeting of BCL2, MCL1, and BCL-XL with ABT-199, homoharringtonine (HHT), and ABT-737. Analysis of DLBCL clones with manipulated expressions of BCL2, MCL1, and BCL-XL. Immunoprecipitation of MCL1 complexes in selected DLBCL cell lines. Experimental therapy aimed at inhibition of BCL2 and MCL1 using ABT-199 and HHT, single agent, or in combination, in vitro and in vivo on primary cell-based murine xenograft models of DLBCL. RESULTS: By the pharmacologic targeting of BCL2, MCL1, and BCL-XL, we demonstrated that DLBCL can be divided into BCL2-dependent and MCL1-dependent subgroups with a less pronounced role left for BCL-XL. Derived DLBCL clones with manipulated expressions of BCL2, MCL1, and BCL-XL, as well as the immunoprecipitation experiments, which analyzed MCL1 protein complexes, confirmed these findings at the molecular level. We demonstrated that concurrent inhibition of BCL2 and MCL1 with ABT-199 and HHT induced significant synthetic lethality in most BCL2-expressing DLBCL cell lines. The marked cytotoxic synergy between ABT-199 and HHT was also confirmed in vivo using primary cell-based murine xenograft models of DLBCL. CONCLUSIONS: As homoharringtonine is a clinically approved antileukemia drug, and ABT-199 is in advanced phases of diverse clinical trials, our data might have direct implications for novel concepts of early clinical trials in patients with aggressive DLBCL.


Subject(s)
Lymphoma, Large B-Cell, Diffuse/genetics , Myeloid Cell Leukemia Sequence 1 Protein/biosynthesis , Proto-Oncogene Proteins c-bcl-2/biosynthesis , bcl-X Protein/biosynthesis , Animals , Apoptosis/drug effects , Biphenyl Compounds/administration & dosage , Bridged Bicyclo Compounds, Heterocyclic/administration & dosage , Cell Line, Tumor , Cell Proliferation/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Harringtonines/administration & dosage , Homoharringtonine , Humans , Lymphoma, Large B-Cell, Diffuse/classification , Lymphoma, Large B-Cell, Diffuse/drug therapy , Lymphoma, Large B-Cell, Diffuse/pathology , Mice , Myeloid Cell Leukemia Sequence 1 Protein/genetics , Nitrophenols/administration & dosage , Piperazines/administration & dosage , Proto-Oncogene Proteins c-bcl-2/genetics , Sulfonamides/administration & dosage , Xenograft Model Antitumor Assays
13.
Blood ; 127(9): 1128-37, 2016 Mar 03.
Article in English | MEDLINE | ID: mdl-26675347

ABSTRACT

Mantle cell lymphoma (MCL) is characterized by an aggressive clinical course and inevitable development of refractory disease, stressing the need to develop alternative therapeutic strategies. To this end, we evaluated pevonedistat (MLN4924), a novel potent and selective NEDD8-activating enzyme inhibitor in a panel of MCL cell lines, primary MCL tumor cells, and 2 distinct murine models of human MCL. Pevonedistat exposure resulted in a dose-, time-, and caspase-dependent cell death in the majority of the MCL cell lines and primary tumor cells tested. Of interest, in the MCL cell lines with lower half-maximal inhibitory concentration (0.1-0.5 µM), pevonedistat induced G1-phase cell cycle arrest, downregulation of Bcl-xL levels, decreased nuclear factor (NF)-κB activity, and apoptosis. In addition, pevonedistat exhibited additive/synergistic effects when combined with cytarabine, bendamustine, or rituximab. In vivo, as a single agent, pevonedistat prolonged the survival of 2 MCL-bearing mouse models when compared with controls. Pevonedistat in combination with rituximab led to improved survival compared with rituximab or pevonedistat monotherapy. Our data suggest that pevonedistat has significant activity in MCL preclinical models, possibly related to effects on NF-κB activity, Bcl-xL downregulation, and G1 cell cycle arrest. Our findings support further investigation of pevonedistat with or without rituximab in the treatment of MCL.


Subject(s)
Cyclopentanes/therapeutic use , Enzyme Inhibitors/therapeutic use , Lymphoma, Mantle-Cell/drug therapy , Pyrimidines/therapeutic use , Rituximab/therapeutic use , Ubiquitins/antagonists & inhibitors , Animals , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Apoptosis/drug effects , Caspases/metabolism , Cell Cycle Checkpoints/drug effects , Cell Line, Tumor , Cell Separation , Cell Survival/drug effects , Cyclopentanes/pharmacology , Enzyme Inhibitors/pharmacology , Gene Expression Profiling , Gene Expression Regulation, Neoplastic/drug effects , Humans , Lymphoma, Mantle-Cell/genetics , Lymphoma, Mantle-Cell/pathology , Mice, SCID , NEDD8 Protein , NF-kappa B/metabolism , Proto-Oncogene Proteins c-bcl-2/metabolism , Pyrimidines/pharmacology , Rituximab/pharmacology , Small Molecule Libraries/pharmacology , Small Molecule Libraries/therapeutic use , Ubiquitins/metabolism
15.
PLoS One ; 10(8): e0135314, 2015.
Article in English | MEDLINE | ID: mdl-26285204

ABSTRACT

Mantle cell lymphoma (MCL) is a chronically relapsing aggressive type of B-cell non-Hodgkin lymphoma considered incurable by currently used treatment approaches. Fludarabine is a purine analog clinically still widely used in the therapy of relapsed MCL. Molecular mechanisms of fludarabine resistance have not, however, been studied in the setting of MCL so far. We therefore derived fludarabine-resistant MCL cells (Mino/FR) and performed their detailed functional and proteomic characterization compared to the original fludarabine sensitive cells (Mino). We demonstrated that Mino/FR were highly cross-resistant to other antinucleosides (cytarabine, cladribine, gemcitabine) and to an inhibitor of Bruton tyrosine kinase (BTK) ibrutinib. Sensitivity to other types of anti-lymphoma agents was altered only mildly (methotrexate, doxorubicin, bortezomib) or remained unaffacted (cisplatin, bendamustine). The detailed proteomic analysis of Mino/FR compared to Mino cells unveiled over 300 differentially expressed proteins. Mino/FR were characterized by the marked downregulation of deoxycytidine kinase (dCK) and BTK (thus explaining the observed crossresistance to antinucleosides and ibrutinib), but also by the upregulation of several enzymes of de novo nucleotide synthesis, as well as the up-regulation of the numerous proteins of DNA repair and replication. The significant upregulation of the key antiapoptotic protein Bcl-2 in Mino/FR cells was associated with the markedly increased sensitivity of the fludarabine-resistant MCL cells to Bcl-2-specific inhibitor ABT199 compared to fludarabine-sensitive cells. Our data thus demonstrate that a detailed molecular analysis of drug-resistant tumor cells can indeed open a way to personalized therapy of resistant malignancies.


Subject(s)
Antineoplastic Agents/pharmacology , Biomarkers, Tumor/metabolism , Drug Resistance, Neoplasm , Lymphoma, Mantle-Cell/metabolism , Proteomics/methods , Vidarabine/analogs & derivatives , Chromatography, Liquid/methods , Humans , Isotope Labeling/methods , Lymphoma, Mantle-Cell/drug therapy , Male , Tandem Mass Spectrometry/methods , Tumor Cells, Cultured , Vidarabine/pharmacology
16.
Mol Cancer ; 13: 159, 2014 Jun 27.
Article in English | MEDLINE | ID: mdl-24972933

ABSTRACT

BACKGROUND: Mantle cell lymphoma (MCL) is an aggressive type of B-cell non-Hodgkin lymphoma associated with poor prognosis. Implementation of high-dose cytarabine (araC) into induction therapy became standard-of-care for all newly diagnosed younger MCL patients. However, many patients relapse even after araC-based regimen. Molecular mechanisms responsible for araC resistance in MCL are unknown and optimal treatment strategy for relapsed/refractory MCL patients remains elusive. METHODS: Five araC-resistant (R) clones were derived by long-term culture of five MCL cell lines (CTRL) with increasing doses of araC up to 50 microM. Illumina BeadChip and 2-DE proteomic analysis were used to identify gene and protein expression changes associated with araC resistance in MCL. In vitro cytotoxicity assays and experimental therapy of MCL xenografts in immunodeficient mice were used to analyze their relative responsiveness to a set of clinically used anti-MCL drugs. Primary MCL samples were obtained from patients at diagnosis and after failure of araC-based therapies. RESULTS: Marked downregulation of deoxycytidine-kinase (DCK) mRNA and protein expression was identified as the single most important molecular event associated with araC-resistance in all tested MCL cell lines and in 50% primary MCL samples. All R clones were highly (20-1000x) cross-resistant to all tested nucleoside analogs including gemcitabine, fludarabine and cladribine. In vitro sensitivity of R clones to other classes of clinically used anti-MCL agents including genotoxic drugs (cisplatin, doxorubicin, bendamustine) and targeted agents (bortezomib, temsirolimus, rituximab) remained unaffected, or was even increased (ibrutinib). Experimental therapy of immunodeficient mice confirmed the anticipated loss of anti-tumor activity (as determined by overall survival) of the nucleoside analogs gemcitabine and fludarabine in mice transplanted with R clone compared to mice transplanted with CTRL cells, while the anti-tumor activity of cisplatin, temsirolimus, bortezomib, bendamustine, cyclophosphamide and rituximab remained comparable between the two cohorts. CONCLUSIONS: Acquired resistance of MCL cells to araC is associated with downregulation of DCK, enzyme of the nucleotide salvage pathway responsible for the first phosphorylation (=activation) of most nucleoside analogs used in anti-cancer therapy. The data suggest that nucleoside analogs should not be used in the therapy of MCL patients, who relapse after failure of araC-based therapies.


Subject(s)
Cladribine/pharmacology , Cytarabine/pharmacology , Deoxycytidine Kinase/metabolism , Deoxycytidine/analogs & derivatives , Down-Regulation/drug effects , Drug Resistance, Neoplasm/drug effects , Lymphoma, Mantle-Cell/enzymology , Vidarabine/analogs & derivatives , Animals , Antibodies, Monoclonal, Murine-Derived/pharmacology , Antibodies, Monoclonal, Murine-Derived/therapeutic use , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Blotting, Western , Cell Line, Tumor , Clone Cells , Deoxycytidine/pharmacology , Electrophoresis, Gel, Two-Dimensional , Gene Expression Profiling , Gene Expression Regulation, Neoplastic/drug effects , Humans , Lymphoma, Mantle-Cell/drug therapy , Lymphoma, Mantle-Cell/genetics , Mass Spectrometry , Mice , Proteomics , Rituximab , Vidarabine/pharmacology , Xenograft Model Antitumor Assays , Gemcitabine
17.
Lab Invest ; 94(7): 806-17, 2014 Jul.
Article in English | MEDLINE | ID: mdl-24862967

ABSTRACT

Mantle cell lymphoma (MCL) is an aggressive type of B-cell non-Hodgkin lymphoma (NHL) associated with poor prognosis. Animal models of MCL are scarce. We established and characterized various in vivo models of metastatic human MCL by tail vein injection of either primary cells isolated from patients with MCL or established MCL cell lines (Jeko-1, Mino, Rec-1, Hbl-2, and Granta-519) into immunodeficient NOD.Cg-Prkdc(scid) Il2rg(tm1Wjl)/SzJ mice. MCL infiltration was assessed with immunohistochemistry (tissues) and flow cytometry (peripheral blood). Engraftment of primary MCL cells was observed in 7 out of 12 patient samples. The pattern of engraftment of primary MCL cells varied from isolated involvement of the spleen to multiorgan infiltration. On the other hand, tumor engraftment was achieved in all five MCL cell lines used and lymphoma involvement of murine bone marrow, spleen, liver, and brain was observed. Overall survival of xenografted mice ranged from 22 ± 1 to 54 ± 3 days depending on the cell line used. Subsequently, we compared the gene expression profile (GEP) and phenotype of the engrafted MCL cells compared with the original in vitro growing cell lines (controls). We demonstrated that engrafted MCL cells displayed complex changes of GEP, protein expression, and sensitivity to cytotoxic agents when compared with controls. We further demonstrated that our MCL mouse models could be used to test the therapeutic activity of systemic chemotherapy, monoclonal antibodies, or angiogenesis inhibitors. The characterization of MCL murine models is likely to aid in improving our knowledge in the disease biology and to assist scientists in the preclinical and clinical development of novel agents in relapsed/refractory MCL patients.


Subject(s)
Disease Models, Animal , Gene Expression Regulation, Neoplastic , Lymphoma, Mantle-Cell/genetics , Transcriptome/genetics , Aged , Animals , Bone Marrow/metabolism , Brain/metabolism , Cell Line, Tumor , Female , Humans , Immunohistochemistry , Immunophenotyping , Interleukin Receptor Common gamma Subunit/deficiency , Interleukin Receptor Common gamma Subunit/genetics , Kaplan-Meier Estimate , Liver/metabolism , Lymphoma, Mantle-Cell/drug therapy , Lymphoma, Mantle-Cell/metabolism , Male , Mice , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Middle Aged , Spleen/metabolism , Transplantation, Heterologous , Tumor Cells, Cultured
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