Adaptation of cancer cells to changes in the microenvironment in an expanding tumor mass is a crucial aspect of malignant progression. To proliferate, cells depend on nutrient and waste transport from surrounding vasculature that becomes less orderly as tumor volume increases, creating chemical gradients in extracellular pH (pHe) and oxygen (O2). Hypoxia inducible factor 1? (HIF-1?), a monomer of a heterodimeric transcription factor regulated by a complex network, mediates cellular responses to physiologic and pathologic processes that allow changes in angiogenesis, cell proliferation and survival, chemosensitivity, and metabolism. The activation of the glycolytic pathway even in the presence of oxygen is a characteristic of many cancer cells, leading to an increase in metabolic consumption rates that can be directly correlated to pHe and O2. HIF-1? regulates energy metabolism by activating the gene expression of glycolytic enzymes, regulatory enzymes, and glucose transporters. HIF-1? is itself post-transcriptionally regulated through blockage of proteosomal degradation as it depends on oxygen concentration. We hypothesize that pHe also plays a role in the regulation of HIF-1? by influencing the half-life of the transcription factor potentially explaining clinical observations of increased chemosensitivity even under hypoxic conditions traditionally thought to cause chemoresistance. Currently, there are no experimental platforms that provide the tools to test such a hypothesis. The fellowship-training plan, through three specific aims, will develop a new in vitro 3D cell culture perfusion system with integrated fluorescence nanosensing capabilities for in situ detection of pHe and O2 gradients. The instrument is designed to enable spatially correlated investigation of cellular responses to the measured pHe and O2 gradients by determination of metabolic consumption rates using a simple transport model. Recovering cells and supernatants from defined regions within the device allows for biochemical and molecular analysis of HIF-1?, proteins regulated by HIF-1?, cell proliferation and survival, and VEGF as adapted by cells in different microenvironments. The three aims will ultimately yield a new integrated model system that will produce advances in: 1) quantitative, manipulable in vitro 3D models of the tumor microenvironment; 2) direct spatial correlation of pHe and O2 gradients with cellular adaptations; and 3) understanding tumor cell response to gradients of pHe and O2 as a function HIF-1?.
Rapid expansion of cancer cells in solid tumors and the impact on interactions in the tumor microenvironment is a dynamic and multiscale process. The integration of improved tools and hypothesis driven experimentation will lead to a better understanding of these interactions and allow progress in the tumor microenvironment field. The goal of this fellowship-training proposal is to utilize this integrated approach to address the interacting dynamics of HIF-1? mediated responses to pH and oxygen microenvironments.
