Abstract 4769: Mapping cellular metabolic activity within U87MR glioblastoma cells in response to chemical and environmental insult using label-free quantitative phase imaging

Abstract

Abstract Introduction: One of our drug development target diseases is Glioblastoma multiforme (GBM), a serious neurological disease with a very poor prognosis for the patients due, in part, to the large number of metabolic pathways it can follow. Materials and Methods: We are using U87-MR cells as a model. We employ time-lapse quantitative phase imaging technology that quantifies the refractive index (RI) of cells via light interference. There is no need for labeling of the cells. The RI is an intrinsic physical property of the cells which lends itself to highly precise and accurate quantitative analysis. Our basic protocol involves seeding cells, providing atmosphere and temperature control. Time-lapse images are automatically obtained at intervals ranging from 10 seconds up to 5 minutes and over time periods from minutes to days. Cells are unlabeled, and can be untreated, or treated with drugs or components of multi-drug formulations. We used drugs that have known effects on the cell cycle, mitochondria and induce apoptosis. We also tested drugs such as temozolomide (TMZ) the first line regimen in GBM therapy protocols. Results: Our first notable finding relates to U87-MR motility. The cells self-assemble into a pattern of well separated cell clusters with tubular connections. These connections are initiated by mitochondrial tunneling and are stable over time. The tunnels are reinforced with actin, and mitochondrial fusion occurs at substrate contact foci. These connections between cells also are conduits for intercellular communication and the transport of objects such as mitochondria (MT). Many drugs that have nuclear targets, including demecolcine, topoisomerase inhibitors, present effects in the cytoplasm of the cells, including vacuoles and possible the remnants of DNA repair. MT contain their DNA. The necessary genes are needed to control their own replication to replace damaged MT and to initiate alternative energy sources, such as mitophagy where cellular protein digestion is utilized as a source of energy. There are numerous factors that cause a wide range of textures within cells, including MT fission, lipid droplets, and autophagy. In many treatments, we see a definite region forming with very active textural changes. Due to its proximity to the nuclear membrane as well as the endoplasmic reticulum, we hypothesize that this may be where mTOR1 and mTOR2C activities originate. In the TMZ studies, we found a strong correlation between the dosage, and the morphology of the cells with a transformation to mesenchymal morphologies at the higher dosages. Discussion: GBM cells adapt to and control a number of different metabolic pathways which are primarily characterized by biochemical analysis. We have vastly improved-time lapse imaging capabilities. We produced first evidence of successful characterization of the metabolic changes based on RI quantifications and visual effects. Citation Format: Ed Luther, Livia P. Mendes, Vladimir P. Torchilin. Mapping cellular metabolic activity within U87MR glioblastoma cells in response to chemical and environmental insult using label-free quantitative phase imaging [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4769.