Iron is an essential nutrient that plays a complex role in cancer biology. Iron metabolism must be tightly controlled within cells. Whilst fundamental to many cellular processes and required for cell survival, excess labile iron is toxic to cells. Increased iron metabolism is associated with malignant transformation, cancer progression, drug resistance and immune evasion. Depleting intracellular iron stores, either with the use of iron chelating agents or mimicking endogenous regulation mechanisms, such as microRNAs, present attractive therapeutic opportunities, some of which are currently under clinical investigation. Alternatively, iron overload can result in a form of regulated cell death, ferroptosis, which can be activated in cancer cells presenting an alternative anti-cancer strategy. This review focuses on alterations in iron metabolism that enable cancer cells to meet metabolic demands required during different stages of tumorigenesis in relation to metastasis and immune response. The strength of current evidence is considered, gaps in knowledge are highlighted and controversies relating to the role of iron and therapeutic targeting potential are discussed. The key question we address within this review is whether iron modulation represents a useful approach for treating metastatic disease and whether it could be employed in combination with existing targeted drugs and immune-based therapies to enhance their efficacy.

Sorafenib remains the only approved drug for treating patients with advanced hepatocellular carcinoma (HCC). However, the therapeutic effect of sorafenib is transient, and patients invariably develop sorafenib resistance (SR). Recently, TYRO3, a member of the TYRO3-AXL-MER family of receptor tyrosine kinases, was identified as being aberrantly expressed in a significant proportion of HCC; however, its role in SR is unknown. In this study, we generated two functionally distinct sorafenib-resistant human Huh-7 HCC cell lines in order to identify new mechanisms to abrogate acquired SR as well as new potential therapeutic targets in HCC. Initially, we investigated the effects of a microRNA (miR), miR-7-5p (miR-7), in both in vitro and in vivo preclinical models of human HCC and identified miR-7 as a potent tumor suppressor of human HCC. We identified TYRO3 as a new functional target of miR-7, which regulates proliferation, migration, and invasion of Huh-7 cells through the phosphoinositide 3-kinase/protein kinase B pathway and is markedly elevated with acquisition of SR. Furthermore, miR-7 effectively silenced TYRO3 expression in both sorafenib-sensitive and sorafenib-resistant Huh-7 cells, inhibiting TYRO3/growth arrest specific 6-mediated cancer cell migration and invasion. Conclusion: We identified a mechanism for acquiring SR in HCC that is through the aberrant expression of the TYRO3/phosphoinositide 3-kinase/protein kinase B signal transduction pathway, and that can be overcome by miR-7 overexpression. Taken together, these data suggest a potential role for miR-7 as an RNA-based therapeutic to treat refractory and drug-resistant HCC. (HEPATOLOGY 2018;67:216-231) L iver cancer ranks sixth among all malignancies in the world and is the second leading cause of cancer-related mortality. (1) The rising incidence of hepatocellular carcinoma (HCC), the most common primary malignant neoplasm of the liver, has been linked to hepatitis B or C infection and increasing worldwide obesity associated with fatty liver disease. (2) In 2017, it is estimated that 40,710 patients will be diagnosed with HCC in the United States, and more than 70% of them will eventually die from this

Cancer-associated fibroblasts (CAF) remain a poorly characterized, heterogeneous cell population. Here we characterized two previously described tumor-promoting CAF sub-types, smooth muscle actin (SMA)-positive myofibroblasts and senescent fibroblasts, identifying a novel link between the two. Analysis of CAF cultured ex vivo, showed that senescent CAF are predominantly SMA-positive; this was confirmed by immunochemistry in head & neck (HNSCC) and esophageal (EAC) cancers. In vitro, we found that fibroblasts induced to senesce develop molecular, ultrastructural and contractile features typical of myofibroblasts and this is dependent on canonical TGF-β signaling. Similar to TGF-β1-generated myofibroblasts, these cells secrete soluble factors that promote tumor cell motility. However, RNA-sequencing revealed significant transcriptomic differences between the two SMA-positive CAF groups, particularly in genes associated with extracellular matrix (ECM) deposition and organization, which differentially promote tumor cell invasion. Notably, second harmonic generation imaging and bioinformatic analysis of SMA-positive human HNSCC and EAC showed that collagen fiber organization correlates with poor prognosis, indicating that heterogeneity within the SMA-positive CAF population differentially impacts on survival. These results show that non-fibrogenic, SMA-positive myofibroblasts can be directly generated through induction of fibroblast senescence and suggest that senescence and myofibroblast differentiation are closely linked processes.

BackgroundmicroRNAs (miRNAs) are short non-coding RNAs that fine-tune gene expression. The aberrant expression of miRNAs is associated with many diseases and they have both therapeutic and biomarker potential. However, our understanding of their usefulness is dependent on the tools we have to study them. Previous studies have identified the need to optimise and standardise RNA extraction methods in order to avoid biased results. Herein, we extracted RNA from murine lung, liver and brain tissues using five commercially available total RNA extraction methods. These included either: phenol: chloroform extraction followed by alcohol precipitation (TRIzol), phenol:chloroform followed by solid-phase extraction (column-based; miRVana and miRNeasy) and solid-phase separation with/without affinity resin (Norgen total and Isolate II). We then evaluated each extraction method for the quality and quantity of RNA recovered, and the expression of miRNAs and target genes.ResultsWe identified differences between each of the RNA extraction methods in the quantity and quality of RNA samples, and in the analysis of miRNA and target gene expression. For the purposes of consistency in quantity, quality and high recovery of miRNAs from tissues, we identified that Phenol:chloroform phase separation combined with silica column-based solid extraction method was preferable (miRVana microRNA isolation). We also identified a method that is not appropriate for miRNA analysis from tissue samples (Bioline Isolate II). For target gene expression any of the kits could be used to analyse mRNA, but if interested in analysing mRNA and miRNA from the same RNA samples some methods should be avoided.ConclusionsDifferent methods used to isolate miRNAs will yield different results and therefore a robust RNA isolation method is required for reproducibility. Researchers should optimise these methods for their specific application and keep in mind that “total RNA” extraction methods do not isolate all types of RNA equally.Electronic supplementary materialThe online version of this article (10.1186/s12896-018-0421-6) contains supplementary material, which is available to authorized users.

Senescent cancer-associated fibroblasts (CAF) develop a senescence-associated secretory phenotype (SASP) that is believed to contribute to cancer progression. The mechanisms underlying SASP development are, however, poorly understood. Here we examined the functional role of microRNA in the development of the SASP in normal fibroblasts and CAF. We identified a microRNA, miR-335, up-regulated in the senescent normal fibroblasts and CAF and able to modulate the secretion of SASP factors and induce cancer cell motility in co-cultures, at least in part by suppressing the expression of phosphatase and tensin homologue (PTEN). Additionally, elevated levels of cyclo-oxygenase 2 (PTGS2; COX-2) and prostaglandin E2 (PGE2) secretion were observed in senescent fibroblasts, and inhibition of COX-2 by celecoxib reduced the expression of miR-335, restored PTEN expression and decreased the pro-tumourigenic effects of the SASP. Collectively these data demonstrate the existence of a novel miRNA/PTEN-regulated pathway modulating the inflammasome in senescent fibroblasts.

BackgroundInflammation of the aortic wall is recognised as a key pathogenesis of abdominal aortic aneurysm (AAA). This study was undertaken to determine whether inflammatory cytokines could be used as biomarkers for the presence of AAA.Methods and ResultsTissue profiles of 27 inflammatory cytokine were examined in AAA (n=14) and nonaneurysmal (n=14) aortic tissues. Three cytokines, regulated upon activation normally T‐cell expressed and secreted (RANTES), eotaxin, and macrophage inflammatory protein 1 beta (MIP‐1b), had increased expression in AAA, particularly within the adventitial layer of the aortic wall. Basic fibroblast growth factor (bFGF) had reduced expression in all layers of the AAA wall. Examination of the circulating plasma profiles of AAA (n=442) and AAA‐free controls (n=970) suggested a (risk factor adjusted) AAA‐association with eotaxin, RANTES, and high sensitivity C‐reactive protein (hsCRP). A plasma inflammatory cytokine score, calculated using these three markers, suggested a strong risk association with AAA (odds ratio, 4.8; 95% CI, 3.5–6.7; P<0.0001), independent of age, sex, history of ischemic heart disease, and smoking.ConclusionsContrary to reports suggesting a distinct T helper 2–associated inflammatory profile in AAA, this current study suggests a more‐generalized pattern of inflammation, albeit with some potentially distinct features, including elevated plasma eotaxin and decreased plasma RANTES. In combination with hsCRP, these markers may have potential utility as AAA biomarkers.

Different strategies, based on CVD risk, resulted in AAA prevalence rates that were significantly greater than that in CVD-free individuals. This may provide opportunities for a targeted approach to community AAA screening in parts of the world where more sophisticated national screening programmes do not exist.

These findings suggest that tobacco constituents influence the expression of miRNA within oral fibroblasts promoting a phenotype that increases oral cancer migration and sheds new light on the mechanisms underlying oral cancer pathogenesis.