The peritoneal tumour microenvironment of high-grade serous ovarian cancer

High-grade serous ovarian cancer (HGSC) disseminates early and extensively throughout the peritoneal space, causing multiple lesions that are a major clinical problem. The aim of this study was to investigate the cellular composition of peritoneal tumour deposits in patient biopsies and their evolution in mouse models using immunohistochemistry, intravital microscopy, confocal microscopy, and 3D modelling. Tumour deposits from the omentum of HGSC patients contained a prominent leukocyte infiltrate of CD3(+) T cells and CD68(+) macrophages, with occasional neutrophils. Alpha-smooth muscle actin(+) (α-SMA(+) ) pericytes and/or fibroblasts surrounded these well-vascularized tumour deposits. Using the murine bowel mesentery as an accessible mouse peritoneal tissue that could be easily imaged, and two different transplantable models, we found multiple microscopic tumour deposits after i.p. injection of malignant cells. Attachment to the peritoneal surface was rapid (6-48 h) with an extensive CD45(+) leukocyte infiltrate visible by 48 h. This infiltrate persisted until end point and in the syngeneic murine ID8 model, it primarily consisted of CD3(+) T lymphocytes and CD68(+) macrophages with α-SMA(+) cells also involved from the earliest stages. A majority of tumour deposits developed above existing mesenteric blood vessels, but in avascular spaces new blood vessels tracked towards the tumour deposits by 2-3 weeks in the IGROV-1 xenografts and 6 weeks in the ID8 syngeneic model; a vigorous convoluted blood supply was established by end point. Inhibition of tumour cell cytokine production by stable expression of shRNA to CXCR4 in IGROV-1 cells did not influence the attachment of cells to the mesentery but delayed neovascularization and reduced tumour deposit size. We conclude that the multiple peritoneal tumour deposits found in HGSC patients can be modelled in the mouse. The techniques described here may be useful for assessing treatments that target the disseminated stage of this disease

Oncolytic adenoviral gene therapy in ovarian cancer: why we are not wasting our time

Preclinical gene-therapy studies in the past 15 years have repeatedly raised hopes that we were about to enter a brave new era. However, many clinical trials have disappointed. For tumor types with poor response rates to first-line conventional cytotoxic chemotherapy and/or high rates of chemorefractory disease, there remain very few treatment options. In this article we review gene therapy within the context of ovarian cancer. We examine why clinical data have been discouraging and discuss how the lessons learned from earlier trials are being applied to current research

Pharmacological inhibition of 3 integrin reduces the inflammatory toxicities caused by oncolytic adenovirus without compromising anticancer activity

Adenoviruses have been clinically tested as anticancer therapies but their utility has been severely limited by rapid, systemic cytokine release and consequent inflammatory toxicity. Here, we describe a new approach to tackling these dangerous side effects. Using human ovarian cancer cell lines as well as malignant epithelial cells harvested from the ascites of women with ovarian cancer, we show that tumor cells do not produce cytokines in the first 24 hours following in vitro infection with the oncolytic adenovirus dl922-947. In contrast, dl922-947 does induce inflammatory cytokines at early time points following intraperitoneal delivery in mice with human ovarian cancer intraperitoneal xenografts. In these animals, cytokines originate predominantly in murine tissues, especially in macrophage-rich organs such as the spleen. We use a nonreplicating adenovirus to confirm that early cytokine production is independent of adenoviral replication. Using β3 integrin knockout mice injected intraperitoneally with dl922-947 and β3 null murine peritoneal macrophages, we confirm a role for macrophage cell surface β3 integrin in this dl922-947–induced inflammation. We present new evidence that co-administration of a cyclic RGD-mimetic–specific inhibitor of β3 integrin significantly attenuates the cytokine release and inflammatory hepatic toxicity induced by dl922-947 in an intraperitoneal murine model of ovarian cancer. Importantly, we find no evidence that β3 inhibition compromises viral infectivity and oncolysis in vitro or anticancer efficacy in vivo. By enabling safe, systemic delivery of replicating adenoviruses, this novel approach could have a major impact on the future development of these effective anticancer agents

Assessment of the Na/I symporter as a reporter gene to visualize oncolytic adenovirus propagation in peritoneal tumours

<p>Purpose: In vivo imaging of the spread of oncolytic viruses using the Na/I symporter (NIS) has been proposed. Here, we assessed whether the presence of NIS in the viral genome affects the therapeutic efficacy of the oncolytic adenovirus dl922-947 following intraperitoneal administration, in a mouse model of peritoneal ovarian carcinoma.</p> <p>Methods: We generated AdAM7, a dl922-947 oncolytic adenovirus encoding the NIS coding sequence. Iodide uptake, NIS expression, infectivity and cell-killing activity of AdAM7, as well as that of relevant controls, were determined in vitro. In vivo, the propagation of this virus in the peritoneal cavity of tumour-bearing mice was determined using SPECT/CT imaging and its therapeutic efficacy was evaluated.</p> <p>Results: In vitro infection of ovarian carcinoma IGROV-1 cells with ADAM7 led to functional expression of NIS. However, the insertion of NIS into the viral genome resulted in a loss of efficacy of the virus in terms of replication and cytotoxicity. In vivo, on SPECT/CT imaging AdAM7 was only detectable in the peritoneal cavity of animals bearing peritoneal ovarian tumours for up to 5 days after intraperitoneal administration. Therapeutic experiments in vivo demonstrated that AdAM7 is as potent as its NIS-negative counterpart.</p> <p>Conclusion: This study demonstrated that despite the detrimental effect observed in vitro, insertion of the reporter gene NIS in an oncolytic adenovirus did not affect its therapeutic efficacy in vivo. We conclude that NIS is a highly relevant reporter gene to monitor the fate of oncolytic adenovectors in live subjects.</p&gt

Proinflammatory characteristics of SMAC/DIABLO-induced cell death in antitumor therapy

Molecular mimetics of the caspase activator second mitochondria-derived activator of caspase (SMAC) are being investigated for use in cancer therapy, but an understanding of in vivo effects remains incomplete. In this study, we offer evidence that SMAC mimetics elicit a proinflammatory cell death in cancer cells that engages an adaptive antitumor immune response. Cancer cells of different histologic origin underwent apoptosis when transduced with lentiviral vectors encoding a cytosolic form of the SMAC mimetic LV-tSMAC. Strikingly, treatment of tumor-bearing mice with LV-tSMAC resulted in the induction of apoptosis, activation of antitumor immunity, and enhanced survival. Antitumor immunity was accompanied by an increase of tumor-infiltrating lymphocytes displaying low PD-1 expression, high lytic capacity, and high levels of IFN-γ when stimulated. We also noted in vivo a decrease in regulatory T cells along with in vitro activation of tumor-specific CD8+ T cells by dendritic cells (DC) isolated from tumor draining lymph nodes. Last, tumor-specific cytotoxic T cells were also found to be activated in vivo. Mechanistic analyses showed that transduction of cancer cells with LV-tSMAC resulted in exposure of calreticulin but not release of HMGB1 or ATP. Nevertheless, DCs were activated upon engulfment of dying cancer cells. Further validation of these findings was obtained by their extension in a model of human melanoma using transcriptionally targeted LV-tSMAC. Together, our findings suggest that SMAC mimetics can elicit a proinflammatory cell death that is sufficient to activate adaptive antitumor immune responses in cancer

The intracellular uptake of CD95 modified paclitaxel-loaded poly(lactic-co-glycolic acid) microparticles

The CD95/CD95L receptor-ligand system is mainly recognised in the induction of apoptosis. However, it has also been shown that CD95L is over-expressed in many cancer types where it modulates immune-evasion and together with its receptor CD95 promotes tumour growth. Here, we show that CD95 surface modification of relatively large microparticles >0.5 μm in diameter, including those made from biodegradable polylactic-co-glycolic acid (PLGA), enhances intracellular uptake by a range of CD95L expressing cells in a process akin to phagocytosis. Using this approach we describe the intracellular uptake of microparticles and agent delivery in neurons, medulloblastoma, breast and ovarian cancer cells in vitro. CD95 modified paclitaxel-loaded PLGA microparticles are shown to be significantly more effective compared to conventional paclitaxel therapy (Taxol) at the same dose in subcutaneous medulloblastoma (∗∗∗P < 0.0001) and orthotopic ovarian cancer xenograft models where a >65-fold reduction in tumour bioluminescence was measured after treatment (∗P = 0.012). This drug delivery platform represents a new way of manipulating the normally advantageous tumour CD95L over-expression towards a therapeutic strategy. CD95 functionalised drug carriers could contribute to the improved function of cytotoxics in cancer, potentially increasing drug targeting and efficacy whilst reducing toxicity

Interleukin-6 as a therapeutic target in human ovarian cancer

<p>Purpose: We investigated whether inhibition of interleukin 6 (IL-6) has therapeutic activity in ovarian cancer via abrogation of a tumor-promoting cytokine network.</p> <p>Experimental Design: We combined preclinical and in silico experiments with a phase 2 clinical trial of the anti-IL-6 antibody siltuximab in patients with platinum-resistant ovarian cancer.</p> <p>Results: Automated immunohistochemistry on tissue microarrays from 221 ovarian cancer cases showed that intensity of IL-6 staining in malignant cells significantly associated with poor prognosis. Treatment of ovarian cancer cells with siltuximab reduced constitutive cytokine and chemokine production and also inhibited IL-6 signaling, tumor growth, the tumor-associated macrophage infiltrate and angiogenesis in IL-6–producing intraperitoneal ovarian cancer xenografts. In the clinical trial, the primary endpoint was response rate as assessed by combined RECIST and CA125 criteria. One patient of eighteen evaluable had a partial response, while seven others had periods of disease stabilization. In patients treated for 6 months, there was a significant decline in plasma levels of IL-6–regulated CCL2, CXCL12, and VEGF. Gene expression levels of factors that were reduced by siltuximab treatment in the patients significantly correlated with high IL-6 pathway gene expression and macrophage markers in microarray analyses of ovarian cancer biopsies.</p> <p>Conclusion: IL-6 stimulates inflammatory cytokine production, tumor angiogenesis, and the tumor macrophage infiltrate in ovarian cancer and these actions can be inhibited by a neutralizing anti-IL-6 antibody in preclinical and clinical studies.</p&gt