Resumen en inglés

ABSTRACT: according to the Global Cancer Observatory, colorectal cancer (CRC) is the third most recurrent cancer worldwide and in Colombia. For 2022, the American Cancer Society previewed that it will be the cause of death of 52,580 persons. Current conventional treatments can generate adverse effects and there is a need for new alternative methods to fight this disease. In order to test these new alternative methods, there is a need for a model that correctly represents the human tissue microenvironment to determine the possible effect of these new treatments. 3D models have the ability to form cell-to-cell interactions, mimic in vivo inflammatory response, develop drug resistance and allows the possibility for evaluating the treatment targeting capabilities. In this work the use of 3D cancer cell cultures is proposed as a possible model for the evaluation of novel alternative methods to fight colorectal cancer. The main objective of this project is to evaluate the ability of cancer cell line Caco-2 to form a 3D cell culture for possible applications in alternative colorectal cancer treatment. For the project execution, the culture conditions for 2D cell cultures were established based on a literature review in order to determine the 3D cell culture conditions. For the formation of the 2D culture and subcultures different experimental setups were tested to identify the best conditions for the 2D cell culture. The 3D cell culture was formed using the hanging drops method in droplets of 25 µL to obtain Caco-2 derived spheroids. The spheroids growth formed with different cell concentrations was observed to determine which conditions provided the best results for the formation of the spheroids. After the spheroids were formed the viability was evaluated by measuring the metabolic activity of the cells in the spheroids and the response to fluorescent dyes. To evaluate the proliferation a measure of the diameter of a single cell inside the droplet was recorded in day 1 and compared to the diameter of the spheroids formed. Morphology was observed under Scanning Electron Microscopy and all the results were evaluated using statistical analysis to determine the significance. As a result, Caco-2 derived spheroids were formed in 6 days under different cell concentrations with average diameters ranged between 70 and 80 µm in complete culture media supplemented with 20% FBS and 0.1% gentamycin with no additional supplements.