Low dose amiodarone reduces tumor growth and angiogenesis.

Amiodarone is an anti-arrhythmic drug that was approved by the US Food and Drug Administration (FDA) in 1985. Pre-clinical studies suggest that Amiodarone induces cytotoxicity in several types of cancer cells, thus making it a potential candidate for use as an anti-cancer treatment. However, it is also known to cause a variety of severe side effects. We hypothesized that in addition to the cytotoxic effects observed in cancer cells Amiodarone also has an indirect effect on angiogensis, a key factor in the tumor microenvironment. In this study, we examined Amiodarone's effects on a murine tumor model comprised of U-87 MG glioblastoma multiforme (GBM) cells, known to form highly vascularized tumors. We performed several in vitro assays using tumor and endothelial cells, along with in vivo assays utilizing three murine models. Low dose Amiodarone markedly reduced the size of GBM xenograft tumors and displayed a strong anti-angiogenic effect, suggesting dual cancer fighting properties. Our findings lay the ground for further research of Amiodarone as a possible clinical agent that, used in safe doses, maintains its dual properties while averting the drug's harmful side effects.

Necrosis in the Tumor Microenvironment and Its Role in Cancer Recurrence.

Cancer recurrence is one of the most imminent problems in the current world of medicine, and it is responsible for most of the cancer-related death rates worldwide. Long-term administration of anticancer cytotoxic drugs may act as a double-edged sword, as necrosis may lead to renewed cancer progression and treatment resistance. The lack of nutrients, coupled with the induced hypoxia, triggers cell death and secretion of signals that affect the tumor niche. Many efforts have been made to better understand the contribution of hypoxia and metabolic stress to cancer progression and resistance, but mostly with respect to inflammation. Here we provide an overview of the direct anticancer effects of necrotic signals, which are not necessarily mediated by inflammation and the role of DAMPs (damage-associated molecular patterns) on the formation of a pro-cancerous environment.

Encapsulated Carbenoxolone Reduces Lung Metastases.

Carbenoxolone is an anti-inflammatory compound and a derivate of a natural substance from the licorice plant. We previously showed that carbenoxolone reduces the metastatic burden in the lungs of mice through its antagonistic effect on high mobility group box 1 (HMGB1). To further enhance carbenoxolone's activity and localization in the lungs, thereby reducing the potential adverse side effects resulting from systemic exposure, we developed a poly(lactic-co-glycolic acid) (PLGA) slow-release system for pulmonary delivery which maintains drug activity in-vitro, as demonstrated in the anoikis assay. Both systemic and intranasal administrations of carbenoxolone effectively minimize metastatic formation in a lung colonization model in mice. Our results show a decrease in the metastatic burden in the lung tissue. Notably, the therapeutic effect of a single intranasal administration of 25 mg/kg carbenoxolone, in the form of drug-loaded particles, had a similar effect in reducing metastatic lesions in the lungs to that of a 10-fold dose of the free drug via intraperitoneal injections, three times per week over the course of four weeks. These data offer new means to potentiate the anti-cancer activity of carbenoxolone and simultaneously reduce the requirement for high dosage administration; the upshot substantially improves therapeutic effect and avoidance of side effects.

Tissue necrosis and its role in cancer progression.

Great efforts have been made in revealing the mechanisms governing cancer resistance and recurrence. The in-situ effects of cell death, caused by hypoxia and metabolic stress, were largely studied in association with inflammation. However, in this work, we focused on the direct effects of necrosis on cancer promotion and on the tumor microenvironment. The conditions leading to cell necrosis, upon nutrient and oxygen deprivation, were recapitulated in-vitro and were used to generate samples for computational proteomic analysis. Under these conditions, we identified clusters of enriched pathways that may be involved in tumor resistance, leading to cancer recurrence. We show that the content of necrotic cells enhances angiogenesis and proliferation of endothelial cells, induces vasculature, as well as increases migration, invasion, and cell-cell interactions. In-vivo studies, where MDA-MB-231 xenografts were exposed to necrotic lysates, resulted in an increase in both proliferation and angiogenesis. Histological analysis of tumor tissues revealed high expression levels of key mediators that were identified by proteomic analysis. Moreover, when cells were injected systemically, coupled with necrotic lysates, a higher number of large lesions was detected in the lung. Finally, using xenografts, we demonstrated that combining an antagonist of a necrotic signal with an anticancer treatment potentiates the prolonged therapeutic effect. This approach suggests a paradigm shift in which targeting late necrotic-secreted factors may increase survival and enhance the efficacy of anticancer therapy.

Matrix metalloproteinase 12 promotes tumor propagation in the lung.

OBJECTIVE: Past studies are inconsistent with regard to the role of matrix metalloproteinase 12 in lung tumorigenesis. This is due, in part, to differential tumorigenesis based on tumor-derived versus immune-derived matrix metalloproteinase 12 expression. Our study aims to thoroughly dissect the role of matrix metalloproteinase 12 in lung tumorigenesis. METHODS: We tested matrix metalloproteinase 12 expression and the association with prognosis using a tissue array and a published non-small cell lung cancer gene expression database. In addition, we characterized the contribution of matrix metalloproteinase 12 to tumor propagation in the lung using a series of in vitro and in vivo studies. RESULTS: Tumor cells of a diverse set of human lung cancers stained positive for matrix metalloproteinase 12, and high matrix metalloproteinase 12 mRNA levels in the tumor were associated with reduced survival. The lung microenvironment stimulated endogenous production of matrix metalloproteinase 12 in lung cancer cells (human 460 lung cancer cell line, Lewis lung carcinoma). In vitro, matrix metalloproteinase 12 knockout Lewis lung carcinoma and Lewis lung carcinoma cells had the same proliferation rate, but Lewis lung carcinoma showed increased invasiveness. In vivo, deficiency of matrix metalloproteinase 12 in Lewis lung carcinoma cells, but not in the host, reduced tumor growth and invasiveness. CONCLUSIONS: We suggest that tumor cell-derived matrix metalloproteinase 12 promotes tumor propagation in the lung and that in the context of pulmonary malignancies matrix metalloproteinase 12 should further be tested as a potential novel therapeutic target.

Tumor cells and their crosstalk with endothelial cells in 3D spheroids.

Recapitulating the tumor microenvironment is a central challenge in the development of experimental model for cancer. To provide a reliable tool for drug development and for personalized cancer therapy, it is critical to maintain key features that  exist in the original tumor. Along with this effort, 3-dimentional (3D) cellular models are being extensively studied. Spheroids are self-assembled cell aggregates that possess many important components of the physiological spatial growth and cell-cell interactions. In this study we aimed to investigate the interconnection between tumor and endothelial cells (EC) in hybrid spheroids containing either tumor cell (TC) lines or patient derived cancer cells. Preparation protocols of hybrid spheroids were optimized and their morphology and tissue-like features were analyzed. Our finding show that capillary-like structures are formed upon assembly and growth of TC:EC spheroids and that spheroids' shape and surface texture may be an indication of spatial invasiveness of cells in the extra-cellular matrix (ECM). Establishing a model of hybrid tumor/stroma spheroids has a crucial importance in the experimental approach for personalized medicine, and may offer a reliable and low-cost method for the goal of predicting drug effects.

High mobility group box 1 antagonist limits metastatic seeding in the lungs via reduction of cell-cell adhesion.

Metastatic spread is the leading cause for cancer-related mortality, with the lungs being a major site for metastatic seeding. Available therapies for patients with metastatic disease are extremely limited. Therefore, there is a desperate need for new strategies to prevent or limit metastatic dissemination and treat existing metastases. The metastatic cascade is highly complex and is affected by multiple factors related to both tumor cells themselves and the microenvironment in the future site of metastasis. We hypothesized that modifying the lung microenvironment by blocking central ubiquitous signals may affect metastatic seeding in the lungs. Given the high basal levels of the Receptor for Advanced Glycation End products (RAGE) in the pulmonary tissue, and its pro-inflammatory properties, we investigated the consequences of interfering with its ligand; High Mobility Group Box 1 (HMGB1). To this end, we tested the effect of Carbenoxolone, an HMGB1 antagonist, on primary tumor growth and metastatic progression in several murine tumor models. We show that antagonizing HMGB1 prevents the adhesion and colonization of cancer cells in the lungs through the reduction of their adhesion and cell-cell interaction both in vitro and in vivo. We demonstrated that these activities are mediated by downregulation of the adhesion molecule Intercellular Adhesion Molecule 1 (ICAM1) and ultimately result in reduced metastatic burden. Carbenoxolone decreases significantly lung metastases formation and can be used potentially as prophylactic therapy for metastatic diseases.