Search

Patients with B-cell non-Hodgkin’s lymphoma (B-NHL) who fail to respond to first-line treatment regimens or develop resistance, exhibit poor prognosis. This signifies the need to develop alternative treatment strategies. CD19-chimeric antigen receptor (CAR) T cell-redirected immunotherapy is an attractive and novel option, which has shown encouraging outcomes in phase I clinical trials of relapsed/refractory NHL. However, the underlying mechanisms of, and approaches to overcome, acquired anti-CD19CAR CD8⁺ T cells (CTL)-resistance in NHL remain elusive. CD19CAR transduced primary human CTLs kill CD19⁺ human NHLs in a CD19- and caspase-dependent manner, mainly via the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) apoptotic pathway. To understand the dynamics of the development of resistance, we analyzed several anti-CD19CAR CTL-resistant NHL sublines (R-NHL) derived by serial exposure of sensitive parental lines to excessive numbers of anti-CD19CAR CTLs followed by a limiting dilution analysis. The R-NHLs retained surface CD19 expression and were efficiently recognized by CD19CAR CTLs. However, R-NHLs developed cross-resistance to CD19CAR transduced human primary CTLs and the Jurkat human T cell line, activated Jurkat, and lymphokine activated killer (LAK) cells, suggesting the acquisition of resistance is independent of CD19-loss and might be due to aberrant apoptotic machinery. We hypothesize that the R-NHL refractoriness to CD19CAR CTL killing could be partially rescued by small molecule sensitizers with apoptotic-gene regulatory effects. Chromatin modifiers and Celecoxib partially reversed the resistance of R-NHL cells to the cytotoxic effects of anti-CD19CAR CTLs and rhTRAIL. These in vitro results, though they require further examination, may provide a rational biological basis for combination treatment in the management of CD19CAR CTL-based therapy of NHL.

Malignant melanoma is the most aggressive form of skin cancer and has a very low survival rate. Over 50% of melanomas harbor various BRAF mutations with the most common being the V600E. BRAF V600E mutation that causes constitutive activation of the MAPK pathway leading to drug-, immune-resistance, apoptosis evasion, proliferation, survival, and metastasis of melanomas. The ATP competitive BRAF V600E selective inhibitor, vemurafenib, has shown dramatic success in clinical trials; promoting tumor regression and an increase in overall survival of patients with metastatic melanoma. Regrettably, vemurafenib-resistance develops over an average of six months, which renders melanomas resistant to other therapeutic strategies. Elucidation of the underlying mechanism(s) of acquisition of vemurafenib-resistance and design of novel approaches to override resistance is the subject of intense clinical and basic research. In this review, we summarize recent developments in therapeutic approaches and clinical investigations on melanomas with BRAF V600E mutation to establish a new platform for the treatment of melanoma.

Autocrine VEGF is necessary for endothelial survival, although the cellular mechanisms supporting this function are unknown. Here, we show that--even after full differentiation and maturation--continuous expression of VEGF by endothelial cells is needed to sustain vascular integrity and cellular viability. Depletion of VEGF from the endothelium results in mitochondria fragmentation and suppression of glucose metabolism, leading to increased autophagy that contributes to cell death. Gene-expression profiling showed that endothelial VEGF contributes to the regulation of cell cycle and mitochondrial gene clusters, as well as several--but not all--targets of the transcription factor FOXO1. Indeed, VEGF-deficient endothelium in vitro and in vivo showed increased levels of FOXO1 protein in the nucleus and cytoplasm. Silencing of FOXO1 in VEGF-depleted cells reversed expression profiles of several of the gene clusters that were de-regulated in VEGF knockdown, and rescued both cell death and autophagy phenotypes. Our data suggest that endothelial VEGF maintains vascular homeostasis through regulation of FOXO1 levels, thereby ensuring physiological metabolism and endothelial cell survival., (© 2015. Published by The Company of Biologists Ltd.)

The chondroitin sulfate-bearing proteoglycans, also known as lecticans, are a major component of the extracellular matrix (ECM) in the central nervous system and regulate neural plasticity. Growing evidence indicates that endogenous, extracellular metalloproteinases that cleave lecticans mediate neural plasticity by altering the structure of ECM aggregates. The bulk of this in vivo data examined the matrix metalloproteinases, but another metalloproteinase family that cleaves lecticans, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS), modulates structural plasticity in vitro, although few in vivo studies have tested this concept. Thus, the purpose of this study was to examine the neurological phenotype of a mouse deficient in ADAMTS1. Adamts1 mRNA was absent in the ADAMTS1 null mouse frontal cortex, but there was no change in the abundance or proteolytic processing of the prominent lecticans brevican and versican V2. However, there was a marked increase in the perinatal lectican neurocan in juvenile ADAMTS1 null female frontal cortex. More prominently, there were declines in synaptic protein levels in the ADAMTS1 null female, but not male, frontal cortex beginning at postnatal day 28. These synaptic marker declines did not affect learning or memory in the adult female ADAMTS1 null mice when tested with the radial-arm water maze. These results indicate that in vivo Adamts1 knockout leads to sexual dimorphism in frontal cortex synaptic protein levels. Since changes in lectican abundance and proteolytic processing did not accompany the synaptic protein declines, ADAMTS1 may play a nonproteolytic role in regulating neural plasticity.

Cancer stem cells have been hypothesized to explain tumor plasticity, including the capability to adopt distinct differentiation commitments. Among the mechanisms of tumor neovascularization, the ability of some malignant cells to mimic an endothelial phenotype has been recognized by a capacity to form matrix-enriched pseudovascular structures. In addition to the expression of genes associated with an endothelial nature, the molecular dynamism of specific microenvironments may also be critical. Here, we report the identification of the extracellular protease ADAMTS1 as a critical molecule for tumor cells to acquire endothelial-like properties. In a fibrosarcoma model, ADAMTS1 increased tumor growth rate in an angiogenesis-independent manner, influencing the tumor cells to display an exclusive endothelial-like gene signature. We documented the relevant expression of ADAMTS1 in aggressive and highly plastic melanoma and Ewing sarcoma cells. Notably, inhibiting ADAMTS1 action compromised the endothelial mimetic attributes observed in this setting. Our findings provide insights into how the tumor microenvironment can elicit endothelial mimicry by tumor cells., (Copyright 2010 AACR.)

Tissue injury initiates a complex series of events that act to restore structure and physiological homeostasis. Infiltration of inflammatory cells and vascular remodeling are both keystones of this process. However, the role of inflammation and angiogenesis in general and, more specifically, the significance of inflammatory cell-derived VEGF in this context are unclear. To determine the role of inflammatory cell-derived VEGF in a clinically relevant and chronically inflamed injury, pulmonary fibrosis, we deleted the VEGF-A gene in myeloid cells. In a model of pulmonary fibrosis in mice, deletion of VEGF in myeloid cells resulted in significantly reduced formation of blood vessels; however, it causes aggravated fibrotic tissue damage. This was accompanied by a pronounced decrease in epithelial cell survival and a striking increase in myofibroblast invasion. The drastic increase in fibrosis following loss of myeloid VEGF in the damaged lungs was also marked by increased levels of hypoxia-inducible factor (HIF) expression and Wnt/beta-catenin signaling. This demonstrates that the process of angiogenesis, driven by myeloid cell-derived VEGF, is essential for the prevention of fibrotic damage.

Tumor-infiltrating myeloid cells (TIMs) support tumor growth by promoting angiogenesis and suppressing antitumor immune responses. CSF-1 receptor (CSF1R) signaling is important for the recruitment of CD11b(+)F4/80(+) tumor-associated macrophages (TAMs) and contributes to myeloid cell-mediated angiogenesis. However, the impact of the CSF1R signaling pathway on other TIM subsets, including CD11b(+)Gr-1(+) myeloid-derived suppressor cells (MDSCs), is unknown. Tumor-infiltrating MDSCs have also been shown to contribute to tumor angiogenesis and have recently been implicated in tumor resistance to antiangiogenic therapy, yet their precise involvement in these processes is not well understood. Here, we use the selective pharmacologic inhibitor of CSF1R signaling, GW2580, to demonstrate that CSF-1 regulates the tumor recruitment of CD11b(+)Gr-1(lo)Ly6C(hi) mononuclear MDSCs. Targeting these TIM subsets inhibits tumor angiogenesis associated with reduced expression of proangiogenic and immunosuppressive genes. Combination therapy using GW2580 with an anti-VEGFR-2 antibody synergistically suppresses tumor growth and severely impairs tumor angiogenesis along with reverting at least one TIM-mediated antiangiogenic compensatory mechanism involving MMP-9. These data highlight the importance of CSF1R signaling in the recruitment and function of distinct TIM subsets, including MDSCs, and validate the benefits of targeting CSF1R signaling in combination with antiangiogenic drugs for the treatment of solid cancers.

Metastasis relies on angiogenesis for tumor expansion. Tumor angiogenesis is restrained by a variety of endogenous inhibitors, including thrombospondin 1 (TSP1). The principal antiangiogenic activity of TSP1 resides in a domain containing three TSP1 repeats (3TSR), and TSP1 cleavage is regulated, in part, by the metalloproteinase ADAMTS1. In this study, we examined the role of TSP1 and ADAMTS1 in controlling metastatic disease in the liver and lung. TSP1 overexpression inhibited metastatic growth of colon or renal carcinoma cells in liver but not lung. Metastatic melanoma in liver grew more rapidly in Tsp1-null mice compared with controls, whereas in lung grew similarly in Tsp1-null mice or controls. Recombinant TSP1 was cleaved more efficiently in lysates from liver than lung. ADAMTS1 inhibition by neutralizing antibody, small interfering RNA, or genetic deletion abrogated cleavage activity. To confirm that lack of cleavage of TSP1 ablated its antiangiogenic function in the lung, we generated colon cancer cells stably secreting only the 3TSR domain and found that they inhibited formation of both liver and lung metastases. Collectively, our results indicate that the antiangiogenic activity of TSP1 is differentially regulated by ADAMTS1 in the liver and lung, emphasizing the concept that regulation of angiogenesis is varied in different tissue environments.

ADAMTS1 is an extracellular metalloproteinase known to participate in a variety of biological processes that includes inflammation, angiogenesis, and development of the urogenital system. Many of its functions rely on its catalytic activity, which thus far has been limited to the cleavage of the matrix proteoglycans aggrecan and versican. However, it is likely that other substrates exist. Using a yeast two-hybrid screen, we identified the Kunitz-type inhibitor, tissue factor pathway inhibitor-2 (TFPI-2), as a binding partner of ADAMTS1. The interaction was confirmed by several biochemical and cell-based assays. In addition, our studies revealed alterations in the pattern of TFPI-2-secreted isoforms and in its extracellular location caused by the specific action of ADAMTS1. Interestingly, we found that TFPI-2 is a novel substrate of ADAMTS1. The cleavage removes a protease-sensitive C-terminal region in TFPI-2, altering its binding properties. The proposed role of TFPI-2 as a maintenance factor of extracellular remodeling suggests the indirect function of ADAMTS1 as an additional homeostatic player by its ability to alter the extracellular location of TFPI-2 and, therefore, to disrupt the remodeling machinery, a phenomenon directly associated to pathologies such as atherosclerosis and tumor progression.