Cellular models with extreme metabophenotypes to study tumor metabolic reprogramming: Development of antitumor nanodevices focused on metabolic targets

Abstract

Tumor cells need to adapt and evolve efficiently in a hostile cellular niche, frequently subjected to a hypoxic microenvironment and with fluctuations in availability of nutrients. These metabolic adaptations of cancer cells have not arisen as a consequence of an accidental adaptation, but due to a very precise regulated process for cell transformation and oncogenesis. Therefore, we intend to study many of the metabolic targets that are still to be deciphered, particularly in some cancer-specific subtypes such as respiratory deficient tumoral cells that can play a key role in tumor evolution. In addition, the discovery of these key metabolic targets can improve the development and optimization of new therapeutic strategies through the use of nanotechnology based approaches. The main aims of this project are: (a) to elucidate the most important adaptations and metabolic targets in tumor cells, mainly in models of carcinogenic cells deficient in respiration, such as the extreme metabolic phenotype proposed in this work; (b) to take advantage of the metabolic knowledge to identify therapeutic targets susceptible of nanotechnological applications, that allow a selective and specific impact in the cancerous cells using our cellular models to develop a new approach for the study of cell proliferation through the use of nanoparticles.; (c) To develop an efficient nanotechnology fluorescence based method to track cell proliferation as the uncontrolled cell proliferation is a hallmark of cancer cells, being crucial cell proliferation assays to study the influence of metabolic changes in cancer progression. A deep study of the map of metabolic status in tumor cell models according to the following study parameters to corroborate the proposed metabofotypes has been carried out (Aim 1). Results has been achieved that include (i) analysis of the degree of tumor "addiction" to certain metabolites that play a central role in cellular bioenergetics, (ii) study of the dependence of these metabolites establishing profiles of resistance / susceptibility to inhibitors of key enzymes of their metabolism. (iii) study the expression and activity profiles of certain oncogenes to determine their relevance in the metabolic reprogramming process of the cellular models. Additionally in this section a metabolomic approximation by mass spectrometry to identify the most important metabolic routes and targets is described. To implement these nanotechnology approaches based on tumor metabolism (Aim 2), the results achieved are: (i) evaluation of nanodevices efficiency in cancer cell model based on respiration deficient cells with an extreme metabolic phenotype, (ii) development and evaluation of a controlled drug delivery system for combined metabolic-based therapy and (iii) development and validation of multifunctionalized nanodevices for targeted drug delivery system to tumor cells with selective metabolic profiles. To develop a novel tracking method based on nanoparticles (Aim 3), the method has been validated in several adherent cells and also in suspension cells including hard-to-transfect cells. Even more interesting is the fact that monitoring of cell proliferation of lymphocytes has been successfully achieved (so far the only efficient method to do this lymphocytes monitoring is CFSE staining). The method was also validated in a cell-based assay which determined the induced cellular arrest through MMC effect, showing the power of the method for long-term cellular assays. Furthermore, this new method does not alter the cell cycle hence presenting no cytotoxic effects.