EFFECT OF SUB-SATURATION OXYGEN LEVEL IN CANCER CELL GROWTH AND METASTATIC MARKER EXPRESSION

Abstract

Cancer remains a leading cause of death worldwide, with incident rates projected to increase. Although mortality rates have improved, a cancer diagnosis remains a significant burden for patients and their families. This burden extends beyond psychological and physical hardships, but also economic strain. For example, the economic cost of cancer treatment is estimated to reach approximately 25.2 trillion international dollars. Contributing to these rising costs is the limited clinical success of modern therapeutics, which restricts their introduction and drives the cost of successful therapeutics. One of the responses includes the field of tissue engineering has expanded into cancer research. The current in vitro cancer culture models generally rely on traditional 2D monolayer cell culture. This method lacks the native cell conformation, function, and physiological accuracy. Additionally, many therapeutic screenings appear promising in the pre-clinical research stage but ultimately fail to translate into clinical trials due to these oversimplifications. Furthermore, the nude mouse models lack critical immune response components. As the newer therapeutics are shifting from cytotoxic drugs to immunotherapeutics, nude mouse models are losing effectiveness. Thus, there is a demand for novel, more accurate in vitro cancer research models. Tissue engineering allowed the development of 3D cellular structures that satisfy many of the criteria required in in vitro cancer research models but necessitate special culturing conditions. Bioreactors are becoming increasingly popular in the field of tissue engineering and are great candidates to fill this void. Flow-perfusion bioreactors in particular are uniquely suited to create 3D macro-scale cell culture environments with the additional benefit of mechanical stimulation when needed. Culture conditions can be monitored through modular sensors in the liquid phase and gas phase, which provides a method for real time monitoring. The cellular physiological limitations of traditional 2D culture models led to the guiding questions of this thesis: 1) “In which ways do sub-saturation of oxygen levels affect the proliferation, oxygen uptake, and phenotypic modification of cancer cells in 3D in vitro culture with flow-perfusion bioreactor models?” and 2) “How does sub-saturation oxygen levels drive metastatic markers in 3D cancer models?”. These questions will be addressed by utilizing a previously modified flow-perfusion bioreactor system with the incorporation of 3D collagen hydrogel scaffolds to mimic more physiologically representative in vitro solid tumors. The sub-saturation of oxygen was created by directing the flow of non-reoxygenated media through subsequent chambers in series. Thus, the oxygen saturation was lowered in a stepwise manner. The verification of sub-saturation oxygen levels was confirmed with optical sensors and the verification of cell culture was determined using cell growth and viability. Further the genetic expression was analyzed using quantitative real time polymerase chain reaction.