Microelectrode measurements of photochemical oxygen depletion in multicell tumor spheroids during photodynamic therapy

We have previously reported results of experiments in which EMT6/Ro spheroids were subjected to Photofrin-PDT consisting of a fixed incident fluence (60 J-cm-2) delivered at 200, 50, and 25 mW-cm-2. Surviving fractions from treated spheroids decreased as the incident fluence rate was lowered over this range. We have interpreted these data using a model wherein the cells compromising the surviving fractions are assumed to originate from within a therapy-induced anoxic volume resulting from Type-II photochemistry. In this paper, we demonstrate direct measurements of the phenomenon in individual photosensitized EMT6 spheroids. Steady-state measurements of 3O2 gradients in and around metabolizing spheroids allow determination of the 3O2 diffusion constant and the rate of metabolic 3O2 consumption within a spheroid. Time-dependent measurements obtained at a single spatial location during laser irradiation are fit to numerical solutions of a pair of time-dependent diffusion with consumption equations. Fits allow a determination of the rate of PDT-induced 3O2 consumption. Based on these fits, it is possible to calculate the spatial and temporal distributions of oxygen within a spheroid undergoing PDT.