Hitchhiking Nanoparticles: Mesenchymal Stem Cell-Mediated Delivery of Theranostic Nanoparticles.

Nanotechnology has emerged as a promising solution to permanent elimination of cancer. However, nanoparticles themselves lack specificity to tumors. Due to enhanced migration to tumors, mesenchymal stem cells (MSCs) were suggested as cell-mediated delivery vehicles of nanoparticles. In this study, we have constructed a complex composed of photoluminescent quantum dots (QDs) and a photosensitizer chlorin e6 (Ce6) to obtain multifunctional nanoparticles, combining cancer diagnostic and therapeutic properties. QDs serve as energy donors-excited QDs transfer energy to the attached Ce6 via Förster resonance energy transfer, which in turn generates reactive oxygen species. Here, the physicochemical properties of the QD-Ce6 complex and singlet oxygen generation were measured, and the stability in protein-rich media was evaluated, showing that the complex remains the most stable in protein-free medium. In vitro studies on MSC and cancer cell response to the QD-Ce6 complex revealed the complex-loaded MSCs' potential to transport theranostic nanoparticles and induce cancer cell death. In vivo studies proved the therapeutic efficacy, as the survival of tumor-bearing mice was statistically significantly increased, while tumor progression and metastases were slowed down.

Skin-derived mesenchymal stem cells as quantum dot vehicles to tumors.

PURPOSE: Cell-mediated delivery of nanoparticles is emerging as a new method of cancer diagnostics and treatment. Due to their inherent regenerative properties, adult mesenchymal stem cells (MSCs) are naturally attracted to wounds and sites of inflammation, as well as tumors. Such characteristics enable MSCs to be used in cellular hitchhiking of nanoparticles. In this study, MSCs extracted from the skin connective tissue were investigated as transporters of semiconductor nanocrystals quantum dots (QDs). MATERIALS AND METHODS: Cytotoxicity of carboxylated CdSe/ZnS QDs was assessed by lactate dehydrogenase cell viability assay. Quantitative uptake of QDs was determined by flow cytometry; their intracellular localization was evaluated by confocal microscopy. In vitro tumor-tropic migration of skin-derived MSCs was verified by Transwell migration assay. For in vivo migration studies of QD-loaded MSCs, human breast tumor-bearing immunodeficient mice were used. RESULTS: QDs were found to be nontoxic to MSCs in concentrations no more than 16 nM. The uptake studies showed a rapid QD endocytosis followed by saturating effects after 6 h of incubation and intracellular localization in the perinuclear region. In vitro migration of MSCs toward MDA-MB-231 breast cancer cells and their conditioned medium was up to nine times greater than the migration toward noncancerous breast epithelial cells MCF-10A. In vivo, systemically administered QD-labeled MSCs were mainly located in the tumor and metastatic tissues, evading most healthy organs with the exception being blood clearance organs (spleen, kidneys, liver). CONCLUSION: Skin-derived MSCs demonstrate applicability in cell-mediated delivery of nanoparticles. The findings presented in this study promise further development of a cell therapy and nanotechnology-based tool for early cancer diagnostics and therapy.

Gene and miRNA expression profiles of mouse Lewis lung carcinoma LLC1 cells following single or fractionated dose irradiation.

In clinical practice ionizing radiation (IR) is primarily applied to cancer treatment in the form of fractionated dose (FD) irradiation. Despite this fact, a substantially higher amount of current knowledge in the field of radiobiology comes from in vitro studies based on the cellular response to single dose (SD) irradiation. In addition, intrinsic and acquired resistance to IR remains an issue in clinical practice, leading to radiotherapy treatment failure. Numerous previous studies suggest that an improved understanding of the molecular processes involved in the radiation-induced DNA damage response to FD irradiation could improve the effectiveness of radiotherapy. Therefore, the present study examined the differential expression of genes and microRNA (miRNA) in murine Lewis lung cancer (LLC)1 cells exposed to SD or FD irradiation. The results of the present study indicated that the gene and miRNA expression profiles of LLC1 cells exposed to irradiation were dose delivery type-dependent. Data analysis also revealed that mRNAs may be regulated by miRNAs in a radiation-dependent manner, suggesting that these mRNAs and miRNAs are the potential targets in the cellular response to SD or FD irradiation. However, LLC1 tumors after FD irradiation exhibited no significant changes in the expression of selected genes and miRNAs observed in the irradiated cells in vitro, suggesting that experimental in vitro conditions, particularly the tumor microenvironment, should be considered in detail to promote the development of efficient radiotherapy approaches. Nevertheless, the present study highlights the primary signaling pathways involved in the response of murine cancer cells to irradiation. Data presented in the present study can be applied to improve the outcome and development of radiotherapy in preclinical animal model settings.

Gene and miRNA expression signature of Lewis lung carcinoma LLC1 cells in extracellular matrix enriched microenvironment.

BACKGROUND: The extracellular matrix (ECM), one of the key components of tumor microenvironment, has a tremendous impact on cancer development and highly influences tumor cell features. ECM affects vital cellular functions such as cell differentiation, migration, survival and proliferation. Gene and protein expression levels are regulated in cell-ECM interaction dependent manner as well. The rate of unsuccessful clinical trials, based on cell culture research models lacking the ECM microenvironment, indicates the need for alternative models and determines the shift to three-dimensional (3D) laminin rich ECM models, better simulating tissue organization. Recognized advantages of 3D models suggest the development of new anticancer treatment strategies. This is among the most promising directions of 3D cell cultures application. However, detailed analysis at the molecular level of 2D/3D cell cultures and tumors in vivo is still needed to elucidate cellular pathways most promising for the development of targeted therapies. In order to elucidate which biological pathways are altered during microenvironmental shift we have analyzed whole genome mRNA and miRNA expression differences in LLC1 cells cultured in 2D or 3D culture conditions. METHODS: In our study we used DNA microarrays for whole genome analysis of mRNA and miRNA expression differences in LLC1 cells cultivated in 2D or 3D culture conditions. Next, we indicated the most common enriched functional categories using KEGG pathway enrichment analysis. Finally, we validated the microarray data by quantitative PCR in LLC1 cells cultured under 2D or 3D conditions or LLC1 tumors implanted in experimental animals. RESULTS: Microarray gene expression analysis revealed that 1884 genes and 77 miRNAs were significantly altered in LLC1 cells after 48 h cell growth under 2D and ECM based 3D cell growth conditions. Pathway enrichment results indicated metabolic pathway, MAP kinase, cell adhesion and immune response as the most significantly altered functional categories in LLC1 cells due to the microenvironmental shift from 2D to 3D. Comparison of the expression levels of selected genes and miRNA between LLC1 cells grown in 3D cell culture and LLC1 tumors implanted in the mouse model indicated correspondence between both model systems. CONCLUSIONS: Global gene and miRNA expression analysis in LLC1 cells under ECM microenvironment indicated altered immune response, adhesion and MAP kinase pathways. All these processes are related to tumor development, progression and treatment response, suggesting the most promising directions for the development of targeted therapies using the 3D cell culture models.