Lung cancers represent the first cause of cancer-related death worldwide. The most common subtype (i.e. lung adenocarcinoma) is characterized by high genomic instability, drug resistance and the insurgence of metastasis in distal organs. Unfortunately, the designing of in vitro systems that fully resemble the in microenvironment, that cooperates with tumor cells during cancer progression to create a pervasive matrix. For this reason, over the years, animal models have been used to test drug efficacy and mechanisms that lead to therapy resistance. Nonetheless, the cost, the differences with the human system and the ethical concerns lead to the development of new and more efficient in vitro models which could replace the in vivo ones. For this reason, the aim of this project is to design a 3D in vitro system of lung adenocarcinoma to be used for the study of tumor cellular and molecular features and drug responses. In the first part of the work, the development of a matrix which could support the growth of the tumor model has been described. In particular, methacrylated gelatin (GelMA) has been selected for its biocompatibility, the presence of cell-adhesive motifs and printability. Two different models of lung adenocarcinoma have been identified according to the tumor cells used, i.e. A549 and H1299 with the fibroblasts (i.e. MRC-5 cells) and their response to the new matrices has been analyzed. Further, the feasibility of promote spheroids formation inside 3D printed GelMA scaffold has been demonstrated. As reported in the literature, vasculature plays a crucial role in nutrient supply and in tumor progression. An effective approach to introduce this aspect in the models could be the extrusion-based 3D-bioprinting, by using sacrificial inks, which allow to obtain empty channels inside the scaffold. In this view, printability of Carbopol, a copolymer of acrylic acid usually used in topic drug formulation and noteworthy for its thixotropic properties, has been investigated. Carbopol can be easy remove, thanks to buffer dilution and the endothelial component could be seeded onto the empty channels. In the last part of the work, the design of a microfluidic platform has been described. Recent studies demonstrated that dynamic culture increases the reliability of the response of the cells to drug treatment. In this work, digital light processing (DLP) based printing has been selected to test the printability of several acrylate resins, thanks to its fast printing speed and high resolution. PEGDA and acrylate PDMS resulted to be the best candidates for the development of a microfluidic device, for the growth and development of lung adenocarcinoma 3D in vitro models and drug testing.
3D-printed cell culture system as an in vitro platform for non-small cell lung cancer (NSCLC) modeling
baruffaldi, desiree
2023-01-01
Abstract
Lung cancers represent the first cause of cancer-related death worldwide. The most common subtype (i.e. lung adenocarcinoma) is characterized by high genomic instability, drug resistance and the insurgence of metastasis in distal organs. Unfortunately, the designing of in vitro systems that fully resemble the in microenvironment, that cooperates with tumor cells during cancer progression to create a pervasive matrix. For this reason, over the years, animal models have been used to test drug efficacy and mechanisms that lead to therapy resistance. Nonetheless, the cost, the differences with the human system and the ethical concerns lead to the development of new and more efficient in vitro models which could replace the in vivo ones. For this reason, the aim of this project is to design a 3D in vitro system of lung adenocarcinoma to be used for the study of tumor cellular and molecular features and drug responses. In the first part of the work, the development of a matrix which could support the growth of the tumor model has been described. In particular, methacrylated gelatin (GelMA) has been selected for its biocompatibility, the presence of cell-adhesive motifs and printability. Two different models of lung adenocarcinoma have been identified according to the tumor cells used, i.e. A549 and H1299 with the fibroblasts (i.e. MRC-5 cells) and their response to the new matrices has been analyzed. Further, the feasibility of promote spheroids formation inside 3D printed GelMA scaffold has been demonstrated. As reported in the literature, vasculature plays a crucial role in nutrient supply and in tumor progression. An effective approach to introduce this aspect in the models could be the extrusion-based 3D-bioprinting, by using sacrificial inks, which allow to obtain empty channels inside the scaffold. In this view, printability of Carbopol, a copolymer of acrylic acid usually used in topic drug formulation and noteworthy for its thixotropic properties, has been investigated. Carbopol can be easy remove, thanks to buffer dilution and the endothelial component could be seeded onto the empty channels. In the last part of the work, the design of a microfluidic platform has been described. Recent studies demonstrated that dynamic culture increases the reliability of the response of the cells to drug treatment. In this work, digital light processing (DLP) based printing has been selected to test the printability of several acrylate resins, thanks to its fast printing speed and high resolution. PEGDA and acrylate PDMS resulted to be the best candidates for the development of a microfluidic device, for the growth and development of lung adenocarcinoma 3D in vitro models and drug testing.
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