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1.
Sci Transl Med ; 15(695): eabp9229, 2023 05 10.
Article in English | MEDLINE | ID: mdl-37163618

ABSTRACT

This first-in-human study evaluated RO7122290, a bispecific fusion protein carrying a split trimeric 4-1BB (CD137) ligand and a fibroblast activation protein α (FAP) binding site that costimulates T cells for improved tumor cell killing in FAP-expressing tumors. Patients with advanced or metastatic solid tumors received escalating weekly intravenous doses of RO7122290 as a single agent (n = 65) or in combination with a 1200-milligram fixed dose of the anti-programmed death-ligand 1 (anti-PD-L1) antibody atezolizumab given every 3 weeks (n = 50), across a tested RO7122290 dose range of 5 to 2000 milligrams and 45 to 2000 milligrams, respectively. Three dose-limiting toxicities were reported, two at different RO7122290 single-agent doses (grade 3 febrile neutropenia and grade 3 cytokine release syndrome) and one for the combination (grade 3 pneumonitis). No maximum tolerated dose was identified. The pharmacokinetic profile of RO7122290 suggested nonlinearity in elimination. The observed changes in peripheral and tissue pharmacodynamic (PD) biomarkers were consistent with the postulated mechanism of action. Treatment-induced PD changes included an increase in proliferating and activated T cells in peripheral blood both in the single-agent and combination arms. Increased infiltration of intratumoral CD8+ and Ki67+CD8+ T cells was observed for both treatment regimens, accompanied by the up-regulation of T cell activation genes and gene signatures. Eleven patients experienced a complete or partial response, six of whom were confirmed to be immune checkpoint inhibitor naive. These results support further evaluation of RO7122290 in combination with atezolizumab or other immune-oncology agents for the treatment of solid tumors.


Subject(s)
CD8-Positive T-Lymphocytes , Neoplasms , Humans , CD8-Positive T-Lymphocytes/metabolism , Neoplasms/pathology , Fibroblasts/pathology
2.
Blood Adv ; 6(3): 1025-1037, 2022 02 08.
Article in English | MEDLINE | ID: mdl-34941996

ABSTRACT

Glofitamab, a novel CD20xCD3, T-cell-engaging bispecific antibody, exhibited single-agent activity in Study NP30179, a first-in-human, phase 1 trial in relapsed/refractory B-cell non-Hodgkin lymphoma. Preclinical studies showed that glofitamab leads to T-cell activation, proliferation, and tumor cell killing upon binding to CD20 on malignant cells. Here, we provide evidence of glofitamab's clinical activity, including pharmacodynamic profile, mode of action, and factors associated with clinical response, by evaluating biomarkers in patient samples from the dose-escalation part of this trial. Patients enrolled in Study NP30179 received single-dose obinutuzumab pretreatment (1000 mg) 7 days before IV glofitamab (5 µg-25 mg). Glofitamab treatment lasted ≤12 cycles once every 2 or 3 weeks. Blood samples were collected at predefined time points per the clinical protocol; T-cell populations were evaluated centrally by flow cytometry, and cytokine profiles were analyzed. Immunohistochemical and genomic biomarker analyses were performed on tumor biopsy samples. Pharmacodynamic modulation was observed with glofitamab treatment, including dose-dependent induction of cytokines, and T-cell margination, proliferation, and activation in peripheral blood. Gene expression analysis of pretreatment tumor biopsy samples indicated that tumor cell intrinsic factors such as TP53 signaling are associated with resistance to glofitamab, but they may also be interlinked with a diminished effector T-cell profile in resistant tumors and thus represent a poor prognostic factor per se. This integrative biomarker data analysis provides clinical evidence regarding glofitamab's mode of action, supports optimal biological dose selection, and will further guide clinical development. This trial was registered at www.clinicaltrials.gov as #NCT03075696.


Subject(s)
Antibodies, Bispecific , Lymphoma, B-Cell , Lymphoma, Non-Hodgkin , Antibodies, Bispecific/pharmacology , Antibodies, Bispecific/therapeutic use , Antigens, CD20/therapeutic use , Humans , Lymphoma, Non-Hodgkin/drug therapy
3.
Transl Oncol ; 14(2): 100984, 2021 Feb.
Article in English | MEDLINE | ID: mdl-33338877

ABSTRACT

Vanucizumab is a novel bispecific antibody inhibiting vascular endothelial growth factor (VEGF-A) and angiopoietin-2 (Ang-2) that demonstrated safety and anti-tumor activity in part I of a phase I study of 42 patients with advanced solid tumors. Part II evaluated the pharmacodynamic effects of vanucizumab 30 or 15 mg/kg every 2 weeks in 32 patients. Serial plasma samples, paired tumor, and skin-wound-healing biopsies were taken over 29 days to evaluate angiogenic markers. Vanucizumab was associated with marked post-infusion reductions in circulating unbound VEGF-A and Ang-2. By day 29, tumor samples revealed mean reductions in density of microvessels (-32.2%), proliferating vessels (-47.9%) and Ang-2 positive vessels (-62.5%). Skin biopsies showed a mean reduction in density of microvessels (-49.0%) and proliferating vessels (-25.7%). Gene expression profiling of tumor samples implied recruitment and potential activation of lymphocytes. Biopsies were safely conducted. Vanucizumab demonstrated a consistent biological effect on vascular-related biomarkers, confirming proof of concept. Skin-wound-healing biopsies were a valuable surrogate for studying angiogenesis-related mechanisms.

4.
Front Immunol ; 11: 550250, 2020.
Article in English | MEDLINE | ID: mdl-33193316

ABSTRACT

The development and progression of solid tumors such as colorectal cancer (CRC) are known to be affected by the immune system and cell types such as T cells, natural killer (NK) cells, and natural killer T (NKT) cells are emerging as interesting targets for immunotherapy and clinical biomarker research. In addition, CD3+ and CD8+ T cell distribution in tumors has shown positive prognostic value in stage I-III CRC. Recent developments in digital computational pathology support not only classical cell density based tumor characterization, but also a more comprehensive analysis of the spatial cell organization in the tumor immune microenvironment (TiME). Leveraging that methodology in the current study, we tried to address the question of how the distribution of myeloid derived suppressor cells in TiME of primary CRC affects the function and location of cytotoxic T cells. We applied multicolored immunohistochemistry to identify monocytic (CD11b+CD14+) and granulocytic (CD11b+CD15+) myeloid cell populations together with proliferating and non-proliferating cytotoxic T cells (CD8+Ki67+/-). Through automated object detection and image registration using HALO software (IndicaLabs), we applied dedicated spatial statistics to measure the extent of overlap between the areas occupied by myeloid and T cells. With this approach, we observed distinct spatial organizational patterns of immune cells in tumors obtained from 74 treatment-naive CRC patients. Detailed analysis of inter-cell distances and myeloid-T cell spatial overlap combined with integrated gene expression data allowed to stratify patients irrespective of their mismatch repair (MMR) status or consensus molecular subgroups (CMS) classification. In addition, generation of cell distance-derived gene signatures and their mapping to the TCGA data set revealed associations between spatial immune cell distribution in TiME and certain subsets of CD8+ and CD4+ T cells. The presented study sheds a new light on myeloid and T cell interactions in TiME in CRC patients. Our results show that CRC tumors present distinct distribution patterns of not only T effector cells but also tumor resident myeloid cells, thus stressing the necessity of more comprehensive characterization of TiME in order to better predict cancer prognosis. This research emphasizes the importance of a multimodal approach by combining computational pathology with its detailed spatial statistics and gene expression profiling. Finally, our study presents a novel approach to cancer patients' characterization that can potentially be used to develop new immunotherapy strategies, not based on classical biomarkers related to CRC biology.


Subject(s)
Colorectal Neoplasms/etiology , Colorectal Neoplasms/metabolism , Myeloid-Derived Suppressor Cells/immunology , Myeloid-Derived Suppressor Cells/metabolism , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Adult , Aged , Aged, 80 and over , Biomarkers , Colorectal Neoplasms/pathology , Colorectal Neoplasms/surgery , Computational Biology/methods , Disease Susceptibility , Female , Gene Expression Profiling/methods , Humans , Immunohistochemistry , Immunomodulation , Intestinal Mucosa/metabolism , Intestinal Mucosa/pathology , Lymphocytes, Tumor-Infiltrating/immunology , Lymphocytes, Tumor-Infiltrating/metabolism , Lymphocytes, Tumor-Infiltrating/pathology , Male , Middle Aged , Neoplasm Grading , Neoplasm Metastasis , Neoplasm Staging , Prognosis , Tumor Microenvironment/genetics , Tumor Microenvironment/immunology
5.
Proc Natl Acad Sci U S A ; 117(1): 541-551, 2020 01 07.
Article in English | MEDLINE | ID: mdl-31889004

ABSTRACT

Cancer immunotherapies are increasingly combined with targeted therapies to improve therapeutic outcomes. We show that combination of agonistic anti-CD40 with antiangiogenic antibodies targeting 2 proangiogenic factors, vascular endothelial growth factor A (VEGFA) and angiopoietin 2 (Ang2/ANGPT2), induces pleiotropic immune mechanisms that facilitate tumor rejection in several tumor models. On the one hand, VEGFA/Ang2 blockade induced regression of the tumor microvasculature while decreasing the proportion of nonperfused vessels and reducing leakiness of the remaining vessels. On the other hand, both anti-VEGFA/Ang2 and anti-CD40 independently promoted proinflammatory macrophage skewing and increased dendritic cell activation in the tumor microenvironment, which were further amplified upon combination of the 2 treatments. Finally, combined therapy provoked brisk infiltration and intratumoral redistribution of cytotoxic CD8+ T cells in the tumors, which was mainly driven by Ang2 blockade. Overall, these nonredundant synergistic mechanisms endowed T cells with improved effector functions that were conducive to more efficient tumor control, underscoring the therapeutic potential of antiangiogenic immunotherapy in cancer.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/pharmacology , CD40 Antigens/agonists , Neoplasms/drug therapy , Neovascularization, Pathologic/drug therapy , Tumor Microenvironment/drug effects , Angiogenesis Inhibitors/pharmacology , Angiogenesis Inhibitors/therapeutic use , Angiopoietin-2/antagonists & inhibitors , Angiopoietin-2/metabolism , Animals , Antineoplastic Agents, Immunological/pharmacology , Antineoplastic Agents, Immunological/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , CD40 Antigens/immunology , Cell Line, Tumor/transplantation , Disease Models, Animal , Drug Synergism , Female , Humans , Lymphocytes, Tumor-Infiltrating/drug effects , Lymphocytes, Tumor-Infiltrating/immunology , Mice , Neoplasms/blood supply , Neoplasms/immunology , Neoplasms/pathology , T-Lymphocytes, Cytotoxic/drug effects , T-Lymphocytes, Cytotoxic/immunology , Tumor Microenvironment/immunology , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor A/metabolism
6.
Nat Cell Biol ; 21(4): 511-521, 2019 04.
Article in English | MEDLINE | ID: mdl-30886344

ABSTRACT

Recent studies have revealed a role for macrophages and neutrophils in limiting chemotherapy efficacy; however, the mechanisms underlying the therapeutic benefit of myeloid-targeting agents in combination with chemotherapy are incompletely understood. Here, we show that targeting tumour-associated macrophages by colony-stimulating factor-1 receptor (CSF-1R) blockade in the K14cre;Cdh1F/F;Trp53F/F transgenic mouse model for breast cancer stimulates intratumoural type I interferon (IFN) signalling, which enhances the anticancer efficacy of platinum-based chemotherapeutics. Notably, anti-CSF-1R treatment also increased intratumoural expression of type I IFN-stimulated genes in patients with cancer, confirming that CSF-1R blockade is a powerful strategy to trigger an intratumoural type I IFN response. By inducing an inflamed, type I IFN-enriched tumour microenvironment and by further targeting immunosuppressive neutrophils during cisplatin therapy, antitumour immunity was activated in this poorly immunogenic breast cancer mouse model. These data illustrate the importance of breaching multiple layers of immunosuppression during cytotoxic therapy to successfully engage antitumour immunity in breast cancer.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Interferon Type I/physiology , Mammary Neoplasms, Experimental/drug therapy , Receptor, Macrophage Colony-Stimulating Factor/antagonists & inhibitors , Animals , Antibodies, Monoclonal/therapeutic use , Antibodies, Monoclonal, Humanized , Cell Line, Tumor , Cisplatin/therapeutic use , Female , Humans , Immunity, Innate/drug effects , Macrophages/drug effects , Mammary Neoplasms, Experimental/immunology , Mammary Neoplasms, Experimental/pathology , Mammary Neoplasms, Experimental/secondary , Mice , Mice, Knockout , Mice, Transgenic
7.
Cancer Discov ; 8(4): 395-402, 2018 04.
Article in English | MEDLINE | ID: mdl-29545369

ABSTRACT

Checkpoint inhibitor therapy has been a breakthrough in cancer research, but only some patients with cancer derive substantial benefit. Although mechanisms underlying sensitivity and resistance to checkpoint inhibitors are being elucidated, the importance of organ-specific regulation of immunity is currently underappreciated. Here, we call for a greater understanding of tissue-specific immunoregulation, namely, "tissue-specific immunostats," to make advances in treatments for cancer. A better understanding of how individual organs at baseline regulate the immune system could enable an improved precision medicine approach to cancer immunotherapy. Cancer Discov; 8(4); 395-402. ©2018 AACR.


Subject(s)
Immune System , Neoplasms/immunology , Neoplasms/therapy , Animals , Humans , Immunotherapy , Mice , Organ Specificity , Precision Medicine
8.
J Exp Med ; 215(3): 859-876, 2018 03 05.
Article in English | MEDLINE | ID: mdl-29436396

ABSTRACT

Depletion of immunosuppressive tumor-associated macrophages (TAMs) or reprogramming toward a proinflammatory activation state represent different strategies to therapeutically target this abundant myeloid population. In this study, we report that inhibition of colony-stimulating factor-1 receptor (CSF-1R) signaling sensitizes TAMs to profound and rapid reprogramming in the presence of a CD40 agonist before their depletion. Despite the short-lived nature of macrophage hyperactivation, combined CSF-1R+CD40 stimulation of macrophages is sufficient to create a proinflammatory tumor milieu that reinvigorates an effective T cell response in transplanted tumors that are either responsive or insensitive to immune checkpoint blockade. The central role of macrophages in regulating preexisting immunity is substantiated by depletion experiments, transcriptome analysis of ex vivo sorted TAMs, and gene expression profiling of whole tumor lysates at an early treatment time point. This approach enabled the identification of specific combination-induced changes among the pleiotropic activation spectrum of the CD40 agonist. In patients, CD40 expression on human TAMs was detected in mesothelioma and colorectal adenocarcinoma.


Subject(s)
Immunity , Macrophages/immunology , Neoplasms/immunology , Neoplasms/pathology , Animals , CD40 Antigens/agonists , CD40 Antigens/metabolism , CD8-Positive T-Lymphocytes/immunology , Female , Humans , Inflammation/pathology , Mice, Inbred BALB C , Mice, Inbred C57BL , Models, Biological , Phenotype , Receptor, Macrophage Colony-Stimulating Factor/antagonists & inhibitors , Receptor, Macrophage Colony-Stimulating Factor/metabolism
9.
Sci Transl Med ; 9(385)2017 04 12.
Article in English | MEDLINE | ID: mdl-28404865

ABSTRACT

Pathological angiogenesis is a hallmark of cancer and a therapeutic target. Vascular endothelial growth factor A (VEGFA) and angiopoietin-2 (ANGPT2; also known as ANG2) are proangiogenic cytokines that sustain tumor angiogenesis and limit antitumor immunity. We show that combined ANGPT2 and VEGFA blockade by a bispecific antibody (A2V) provided superior therapeutic benefits, as compared to the single agents, in both genetically engineered and transplant tumor models, including metastatic breast cancer (MMTV-PyMT), pancreatic neuroendocrine tumor (RIP1-Tag2), and melanoma. Mechanistically, A2V promoted vascular regression, tumor necrosis, and antigen presentation by intratumoral phagocytes. A2V also normalized the remaining blood vessels and facilitated the extravasation and perivascular accumulation of activated, interferon-γ (IFNγ)-expressing CD8+ cytotoxic T lymphocytes (CTLs). Whereas the antitumoral activity of A2V was, at least partly, CTL-dependent, perivascular T cells concurrently up-regulated the expression of the immune checkpoint ligand programmed cell death ligand 1 (PD-L1) in tumor endothelial cells. IFNγ neutralization blunted this adaptive response, and PD-1 blockade improved tumor control by A2V in different cancer models. These findings position immune cells as key effectors of antiangiogenic therapy and support the rationale for cotargeting angiogenesis and immune checkpoints in cancer therapy.


Subject(s)
Angiopoietin-2/metabolism , Programmed Cell Death 1 Receptor/metabolism , Vascular Endothelial Growth Factor A/metabolism , Angiopoietin-2/genetics , Animals , B7-H1 Antigen/metabolism , Blood Vessels/metabolism , CD8-Positive T-Lymphocytes/metabolism , Cell Line, Tumor , Mice , Mice, Inbred C57BL , Mice, Transgenic , Programmed Cell Death 1 Receptor/genetics , Vascular Endothelial Growth Factor A/genetics
10.
Mol Cancer Ther ; 15(12): 3077-3086, 2016 12.
Article in English | MEDLINE | ID: mdl-27582524

ABSTRACT

Blockade of colony-stimulating factor-1 receptor (CSF-1R) enables the therapeutic targeting of tumor-associated macrophages (TAM) in cancer patients. Various CSF-1R inhibitors, mAbs, and tyrosine kinase inhibitors are currently evaluated in early clinical trials. Presence of an alternative survival signal, such as GM-CSF, rescues human monocyte-derived macrophages from CSF-1R inhibitor-induced apoptosis. In this study, we sought to identify additional factors that mediate resistance to CSF-1R-blocking antibody RG7155 (emactuzumab). We investigated the impact of hypoxia, macrophage-polarizing cytokines IL4 and IL10, and genetic alterations within the CSF1R locus and mitochondrial DNA. Among all investigated factors, only IL4 completely rescued viability of RG7155-treated macrophages in vitro This RG7155-resistant population was characterized by a substantially increased mannose receptor-1 (CD206) expression. Analysis of CD206 and the hemoglobin scavenger receptor CD163 expression on normal tissue allowed for discrimination of distinct macrophage populations according to localization and frequency. In emactuzumab-treated cancer patients, we found a significant reduction of CSF-1R, CD204, and CD163 mRNA levels in contrast to a less pronounced decrease of CD206 expression by transcriptome analysis of tumor biopsies. However, we detected in normal skin tissue, which shows lower IL4 mRNA expression compared with melanoma tissue, significant reduction of CD206+ dermal macrophages in RG7155-treated skin biopsies. These results suggest that in cancers where the cytokines IL4 and GM-CSF are sufficiently expressed to induce very high CD206 expression on macrophages, CSF-1R inhibition may not deplete CD206hi TAM. This observation can help to identify those patients most likely to benefit from CSF-1R-targeting agents. Mol Cancer Ther; 15(12); 3077-86. ©2016 AACR.


Subject(s)
Antibodies, Monoclonal/pharmacology , Antineoplastic Agents/pharmacology , Macrophages/drug effects , Macrophages/metabolism , Antibodies, Monoclonal, Humanized , Biomarkers , Biopsy , Cell Line, Tumor , Cell Survival/drug effects , Cytokines/metabolism , Drug Resistance , Humans , Immunophenotyping , Monocytes/metabolism , Neoplasms/metabolism , Neoplasms/pathology , Phenotype , STAT1 Transcription Factor/metabolism , Signal Transduction/drug effects , Skin/cytology
11.
Br J Cancer ; 115(6): 691-702, 2016 09 06.
Article in English | MEDLINE | ID: mdl-27529514

ABSTRACT

BACKGROUND: To assess antivascular effects, and evaluate clinically translatable magnetic resonance imaging (MRI) biomarkers of tumour response in vivo, following treatment with vanucizumab, a bispecific human antibody against angiopoietin-2 (Ang-2) and vascular endothelial growth factor-A (VEGF-A). METHODS: Colo205 colon cancer xenografts were imaged before and 5 days after treatment with a single 10 mg kg(-1) dose of either vanucizumab, bevacizumab (anti-human VEGF-A), LC06 (anti-murine/human Ang-2) or omalizumab (anti-human IgE control). Volumetric response was assessed using T2-weighted MRI, and diffusion-weighted, dynamic contrast-enhanced (DCE) and susceptibility contrast MRI used to quantify tumour water diffusivity (apparent diffusion coefficient (ADC), × 10(6) mm(2) s(-1)), vascular perfusion/permeability (K(trans), min(-1)) and fractional blood volume (fBV, %) respectively. Pathological correlates were sought, and preliminary gene expression profiling performed. RESULTS: Treatment with vanucizumab, bevacizumab or LC06 induced a significant (P<0.01) cytolentic response compared with control. There was no significant change in tumour ADC in any treatment group. Uptake of Gd-DTPA was restricted to the tumour periphery in all post-treatment groups. A significant reduction in tumour K(trans) (P<0.05) and fBV (P<0.01) was determined 5 days after treatment with vanucizumab only. This was associated with a significant (P<0.05) reduction in Hoechst 33342 uptake compared with control. Gene expression profiling identified 20 human genes exclusively regulated by vanucizumab, 6 of which are known to be involved in vasculogenesis and angiogenesis. CONCLUSIONS: Vanucizumab is a promising antitumour and antiangiogenic treatment, whose antivascular activity can be monitored using DCE and susceptibility contrast MRI. Differential gene expression in vanucizumab-treated tumours is regulated by the combined effect of Ang-2 and VEGF-A inhibition.


Subject(s)
Adenocarcinoma/drug therapy , Angiogenesis Inhibitors/therapeutic use , Antibodies, Bispecific/therapeutic use , Antibodies, Monoclonal/therapeutic use , Colonic Neoplasms/drug therapy , Gene Expression Profiling , Magnetic Resonance Imaging/methods , Molecular Targeted Therapy , Neovascularization, Pathologic/drug therapy , Adenocarcinoma/blood supply , Adenocarcinoma/diagnostic imaging , Adenocarcinoma/pathology , Angiogenesis Inhibitors/immunology , Angiopoietin-2/antagonists & inhibitors , Angiopoietin-2/immunology , Animals , Antibodies, Monoclonal/immunology , Antibodies, Monoclonal, Humanized , Bevacizumab/therapeutic use , Cell Line, Tumor , Colonic Neoplasms/blood supply , Colonic Neoplasms/diagnostic imaging , Colonic Neoplasms/pathology , DNA Replication/drug effects , Gene Expression Regulation, Neoplastic/drug effects , Humans , Immunoglobulin E/immunology , Mice , Neovascularization, Pathologic/diagnostic imaging , Neovascularization, Pathologic/pathology , Omalizumab/therapeutic use , Tumor Burden , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor A/immunology , Xenograft Model Antitumor Assays
12.
Nat Cell Biol ; 18(7): 790-802, 2016 07.
Article in English | MEDLINE | ID: mdl-27295554

ABSTRACT

Tumour-associated macrophages (TAMs) largely express an alternatively activated (or M2) phenotype, which entails immunosuppressive and tumour-promoting capabilities. Reprogramming TAMs towards a classically activated (M1) phenotype may thwart tumour-associated immunosuppression and unleash anti-tumour immunity. Here we show that conditional deletion of the microRNA (miRNA)-processing enzyme DICER in macrophages prompts M1-like TAM programming, characterized by hyperactive IFN-γ/STAT1 signalling. This rewiring abated the immunosuppressive capacity of TAMs and fostered the recruitment of activated cytotoxic T lymphocytes (CTLs) to the tumours. CTL-derived IFN-γ exacerbated M1 polarization of Dicer1-deficient TAMs and inhibited tumour growth. Remarkably, DICER deficiency in TAMs negated the anti-tumoral effects of macrophage depletion by anti-CSF1R antibodies, and enabled complete tumour eradication by PD1 checkpoint blockade or CD40 agonistic antibodies. Finally, genetic rescue of Let-7 miRNA activity in Dicer1-deficient TAMs partly restored their M2-like phenotype and decreased tumour-infiltrating CTLs. These findings suggest that DICER/Let-7 activity opposes IFN-γ-induced, immunostimulatory M1-like TAM activation, with potential therapeutic implications.


Subject(s)
DEAD-box RNA Helicases/metabolism , Interferon-gamma/metabolism , Macrophage Activation/immunology , Macrophages/immunology , MicroRNAs/genetics , Ribonuclease III/metabolism , Tumor Microenvironment/genetics , Animals , Cells, Cultured , DEAD-box RNA Helicases/deficiency , Humans , Mice , Neoplasms/genetics , Neoplasms/immunology , Ribonuclease III/deficiency
13.
Haematologica ; 99(4): 697-705, 2014 Apr.
Article in English | MEDLINE | ID: mdl-24162792

ABSTRACT

C/EPBα proteins, encoded by the CCAAT-enhancer-binding protein α gene, play a crucial role in granulocytic development, and defects in this transcription factor have been reported in acute myeloid leukemia. Here, we defined the C/EBPα signature characterized by a set of genes up-regulated upon C/EBPα activation. We analyzed expression of the C/EBPα signature in a cohort of 525 patients with acute myeloid leukemia and identified a subset characterized by low expression of this signature. We referred to this group of patients as the C/EBPα dysfunctional subset. Remarkably, a large percentage of samples harboring C/EBPα biallelic mutations clustered within this subset. We hypothesize that re-activation of the C/EBPα signature in the C/EBPα dysfunctional subset could have therapeutic potential. In search for small molecules able to reverse the low expression of the C/EBPα signature we applied the connectivity map. This analysis predicted positive connectivity between the C/EBPα activation signature and histone deacetylase inhibitors. We showed that these inhibitors reactivate expression of the C/EBPα signature and promote granulocytic differentiation of primary samples from the C/EBPα dysfunctional subset harboring biallelic C/EBPα mutations. Altogether, our study identifies histone deacetylase inhibitors as potential candidates for the treatment of certain leukemias characterized by down-regulation of the C/EBPα signature.


Subject(s)
Antineoplastic Agents/pharmacology , CCAAT-Enhancer-Binding Protein-alpha/metabolism , Gene Expression Regulation, Leukemic/drug effects , Histone Deacetylase Inhibitors/pharmacology , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/metabolism , Transcriptome , CCAAT-Enhancer-Binding Protein-alpha/genetics , Cell Differentiation , Cell Line, Tumor , Cluster Analysis , Gene Expression Profiling , Humans , Mutation/drug effects , Mutation/genetics , Transcriptional Activation
14.
EJNMMI Res ; 4(1): 34, 2014 Dec.
Article in English | MEDLINE | ID: mdl-26116108

ABSTRACT

BACKGROUND: Inhibition of mitogen-activated protein kinase (MEK, also known as MAPK2, MAPKK), a key molecule of the Ras/MAPK (mitogen-activated protein kinase) pathway, has shown promising effects on B-raf-mutated and some RAS (rat sarcoma)-activated tumors in clinical trials. The objective of this study is to examine the efficacy of a novel allosteric MEK inhibitor RO4987655 in K-ras-mutated human tumor xenograft models using [(18)F] FDG-PET imaging and proteomics technology. METHODS: [(18)F] FDG uptake was studied in human lung carcinoma xenografts from day 0 to day 9 of RO4987655 therapy using microPET Focus 120 (CTI Concorde Microsystems, Knoxville, TN, USA). The expression levels of GLUT1 and hexokinase 1 were examined using semi-quantitative fluorescent immunohistochemistry (fIHC). The in vivo effects of RO4987655 on MAPK/PI3K pathway components were assessed by reverse phase protein arrays (RPPA). RESULTS: We have observed modest metabolic decreases in tumor [(18)F] FDG uptake after MEK inhibition by RO4987655 as early as 2 h post-treatment. The greatest [(18)F] FDG decreases were found on day 1, followed by a rebound in [(18)F] FDG uptake on day 3 in parallel with decreasing tumor volumes. Molecular analysis of the tumors by fIHC did not reveal statistically significant correlations of GLUT1 and hexokinase 1 expressions with the [(18)F] FDG changes. RPPA signaling response profiling revealed not only down-regulation of pERK1/2, pMKK4, and pmTOR on day 1 after RO4987655 treatment but also significant up-regulation of pMEK1/2, pMEK2, pC-RAF, and pAKT on day 3. The up-regulation of these markers is interpreted to be indicative of a reactivation of the MAPK and activation of the compensatory PI3K pathway, which can also explain the rebound in [(18)F] FDG uptake following MEK inhibition with RO4987655 in the K-ras-mutated human tumor xenografts. CONCLUSIONS: We have performed the first preclinical evaluation of a new MEK inhibitor, RO4987655, using a combination of [(18)F] FDG-PET imaging and molecular proteomics. These results provide support for using preclinical [(18)F] FDG-PET imaging in early, non-invasive monitoring of the effects of MEK and perhaps other Ras/MAPK signaling pathway inhibitors, which should facilitate a wider implementation of clinical [(18)F] FDG-PET to optimize their clinical use.

15.
Clin Cancer Res ; 17(9): 2657-67, 2011 May 01.
Article in English | MEDLINE | ID: mdl-21415212

ABSTRACT

PURPOSE: MicroRNAs (miRNA) play pivotal oncogenic and tumor-suppressor roles in several human cancers. We sought to discover novel tumor-suppressor miRNAs in gastric cancer (GC). EXPERIMENTAL DESIGN: Using Agilent miRNA microarrays, we compared miRNA expression profiles of 40 primary gastric tumors and 40 gastric normal tissues, identifying miRNAs significantly downregulated in gastric tumors. RESULTS: Among the top 80 miRNAs differentially expressed between gastric tumors and normals (false discovery rate < 0.01), we identified hsa-miR-486 (miR-486) as a significantly downregulated miRNA in primary GCs and GC cell lines. Restoration of miR-486 expression in GC cell lines (YCC3, SCH and AGS) caused suppression of several pro-oncogenic traits, whereas conversely inhibiting miR-486 expression in YCC6 GC cells enhanced cellular proliferation. Array-CGH analysis of 106 primary GCs revealed genomic loss of the miR-486 locus in approximately 25% to 30% of GCs, including two tumors with focal genomic losses specifically deleting miR-486, consistent with miR-486 playing a tumor-suppressive role. Bioinformatic analysis identified the secreted antiapoptotic glycoprotein OLFM4 as a potential miR-486 target. Restoring miR-486 expression in GC cells decreased endogenous OLFM4 transcript and protein levels, and also inhibited expression of luciferase reporters containing an OLFM4 3' untranslated region with predicted miR-486 binding sites. Supporting the biological relevance of OLFM4 as a miR-486 target, proliferation in GC cells was also significantly reduced by OLFM4 silencing. CONCLUSIONS: miR-486 may function as a novel tumor-suppressor miRNA in GC. Its antioncogenic activity may involve the direct targeting and inhibition of OLFM4.


Subject(s)
Carcinoma/genetics , Gene Deletion , Granulocyte Colony-Stimulating Factor/genetics , MicroRNAs/physiology , Stomach Neoplasms/genetics , Apoptosis/genetics , Apoptosis Regulatory Proteins/genetics , Apoptosis Regulatory Proteins/metabolism , Carcinoma/pathology , Cell Line, Tumor , Cell Proliferation , Disease Progression , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Genes, Tumor Suppressor , Genomic Instability/genetics , Genomic Instability/physiology , Granulocyte Colony-Stimulating Factor/metabolism , Humans , MicroRNAs/genetics , Microarray Analysis , Neoplasms/genetics , Neoplasms/pathology , Stomach Neoplasms/pathology
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