Exploiting iron-binding proteins for drug delivery.

Currently, many therapies fail due to an insufficient drug dose reaching the target site and high systemic toxicity. Protein-based drug delivery systems that allow an increase in drug concentration at a specific location in the body or predominantly target malignant cells are promising technologies. Due to the high need for iron in many disorders including various types of cancer, iron-binding proteins: transferrin, ferritin and hemoglobin, are a promising tool as drug carriers. In this review we summarize the characteristics of human iron-binding proteins and present their use in targeted drug delivery strategies.

Wnt signaling pathway in development and cancer.

Wnt signaling pathway is one of the most important signaling pathways. The complexity of Wnt signals and their functional role is crucial in development and growth. It is the most active during embryogenesis facilitating new organism formation by cell differentiation, polarization and migration. Its activation is also common during development of many tumors and others diseases. In this review we shortly describe a role of Wnt pathway in development in order to better understand its role in cancer progression. We also describe current anti-cancer therapies targeting Wnt pathway.

Interferon lambda 2 promotes mammary tumor metastasis via angiogenesis extension and stimulation of cancer cell migration.

Myeloid-derived suppressor cells (MDSCs) support tumor development by stimulation of angiogenesis and immune response inhibition. In our previous study, we showed that interferon lambda 2 (IFN-λ2), secreted by MDSCs, enhances production of pro-angiogenic factors by cancer cells via phosphorylation of STAT3 and therefore promotes blood vessels formation. In the present study IFN-λ2 level was evaluated by ELISA in serum of tumor-bearing mice, whereas its expression in MDSCs isolated from the lungs with metastatic tumors and normal lungs was assessed by qPCR. The effect of IFN-λ2 on mouse mammary cancer cells motility was tested in Boyden chamber migration assay. In order to evaluate its pro-angiogenic function we performed in vitro tubule formation assay and in ovo angiogenesis assay on chicken embryo chorioallantoic membrane (CAM). Moreover, in order to design small molecule inhibitors of IFN-λ2 and its receptor we performed molecular modeling followed by the identification of potential natural inhibitors. Then, we examined their ability to inhibit angiogenesis in vitro. Our results showed that IFN-λ2 predisposed mouse mammary cancer cells to migration in vitro. It also enhanced angiogenesis induced by mouse mammary cancer cells in vitro and in ovo. For the first time we selected potential IFN-λ2 inhibitors and we validated that they were capable to abolish pro-angiogenic effect of IFN-λ2, similarly to blocking antibodies. Therefore, IFN-λ2 and its receptor may become targets of anti-cancer therapy, but their mechanism of action requires further investigation.

Drug delivery systems improving chemical and physical properties of anticancer drugs currently investigated for treatment of solid tumors.

Cancer is the second leading cause of death worldwide. Conventional cancer treatment like chemotherapy do not fulfil the expectations of both patients and physicians and there is a pressing need for a new kind of therapies that will increase drug delivery to the tumor mass. Standard chemotherapy does not show either specific tumor-targeting, or selective mode of action for cancer cells. Moreover, tumor microenvironments additionally disturb drug perfusion and diffusion. Currently approved anticancer drugs have many limitations and therefore special delivery systems improving their chemical and physical properties are beneficial. In the present review paper we discuss various drug delivery systems for solid tumors that are actually at various stages of pre-clinical tests or approved for therapy.

Tumour-associated macrophages influence canine mammary cancer stem-like cells enhancing their pro-angiogenic properties.

Cancer stem-like cells as cells with ability to self-renewal and potential to differentiate into various types of cells are known to be responsible for tumour initiation, recurrence and drug resistance. Hence a comprehensive research is concentrated on discovering cancer stem-like cells biology and interdependence between them and other cells. The aim of our study was to evaluate the impact of macrophages on cancer stem-like cells in canine mammary carcinomas. As recent studies indicated presence of macrophages in cancer environment stimulates cancer cells into more motile and invasive cells by acquisition of macrophage phenotypes. From two canine mammary tumour cell lines, CMT-U27 and P114 cancer stem-like cells were stained with Sca1, CD44 and EpCAM monoclonal antibodies and isolated. Those cells were next co-cultured with macrophages for 5 days and used for further experiments. Canine Gene Expression Microarray revealed 29 different expressed transcripts in cancer stem-like cells co-cultured with macrophages compared to those in mono-culture. Up-regulation of C-C motif chemokine 2 was considered as the most interesting for further investigation. Additionally, those cells showed overexpression of genes involved in non-canonical Wnt pathway. The results of 3D tubule formation in endothelial cells induced by cancer stem-like cells co-cultured with macrophages compared to cancer stem-like cells from mono-cultures and with addition of Recombinant Canine CCL2/MCP-1 revealed the same stimulating effect. Based on those results we can conclude that macrophages have an impact on cancer stem-like cells increasing secretion of pro-angiogenic factors.

Analysis of microRNA expression in canine mammary cancer stem-like cells indicates epigenetic regulation of transforming growth factor-beta signaling.

Cancer stem cells (CSCs) display both unique self-renewal ability as well as the ability to differentiate into many kinds of cancer cells. They are supposed to be responsible for cancer initiation, recurrence and drug resistance. Despite the fact that a variety of methods are currently employed in order to target CSCs, little is known about the regulation of their phenotype and biology by miRNAs. The aim of our study was to assess miRNA expression in canine mammary cancer stem-like cells (expressing stem cell antigen 1, Sca-1; CD44 and EpCAM) sorted from canine mammary tumour cell lines (CMT-U27, CMT-309 and P114). In order to prove their stem-like phenotype, we conducted a colony formation assay that confirmed their ability to form colonies from a single cell. Profiles of miRNA expression were investigated using Agilent custom-designed microarrays. The results were further validated by real-time rt-PCR analysis of expression of randomly selected miRNAs. Target genes were indicated and analysed using Kioto Encyclopedia of Genes and Genomes (KEGG) and BioCarta databases. The results revealed 24 down-regulated and nine up-regulated miRNAs in cancer stem-like cells compared to differentiated tumour cells. According to KEGG and BioCarta databases, target genes (n=240) of significantly down-regulated miRNAs were involved in transforming growth factor-beta signaling, mitogen-activated protein kinases (MAPK) signaling pathway, anaplastic lymphoma receptor tyrosine kinase (ALK) and peroxisome proliferator-activated receptor gamma, coactivator 1 alpha (PGC1A) pathways. The analysis of single-gene overlapping with different pathways showed that the most important genes were: TGFBR1, TGFBR2, SOS1, CHUK, PDGFRA, SMAD2, MEF2A, MEF2C and MEF2D. All of them are involved in tumor necrosis factor-beta signaling and may indicate its important role in cancer stem cell biology. Increased expression of TGFBR2, SMAD2, MEF2A and MEF2D in canine mammary cancer stem-like cells was further confirmed by real-time-qPCR. The results of our study point at epigenetic differences between cancer stem-like cells and differentiated tumour cells, which may be important not only for veterinary medicine but also for comparative oncology.