GROWTH DELAY STUDIES OF THE RESPONSE OF V‐79 MULTICELL SPHEROIDS EXPOSED TO DHE and RED LIGHT

Abstract

Multicell tumour spheroids (MTS) of V‐79 Chinese hamster cells have been used to study the role of a number of treatment and microenvironmental parameters in the modification of tumour response to Photodynamic Therapy (PDT) using visible light in combination with the photosensitizing compound dihematoporphyrin ether (DHE). The kinetics of DHE uptake into MTS, determined by fluorimetry of extracted porphyrins, indicate that after extended incubation (i.e. 24 h) the mean cellular DHE content in larger (˜300 μ.m and 400 u.m) MTS is significantly less than that for smaller (˜200 μm) MTS, consistent with a hypothesis that DHE uptake into the internal regions of spheroids is diffusion‐limited. The response of spheroids to PDT, as assessed by the endpoint of growth delay, indicates that the kinetics of spheroid volume alteration and cell loss, as well as the potential for regfrrwth, are markedly dependent on both the drug and light exposure levels used. The oxygen dependence of this response has been investigated after light irradiation of spheroid cultures equilibrated with either 21% O2 (i.e. air) or 0% 02 (i.e. N2). While treatment in air results in significant growth delay, the growth kinetics of DHE‐treated spheroids irradiated under N2 were essentially unchanged from those of untreated spheroids. These observations clearly demonstrate an important role for oxygen, at the time of irradiation, in determining the response of spheroids to PDT.

Monitoring of cell and tissue responses to photodynamic therapy by electrical impedance spectroscopy

“…Subsequent growth delay was observed for both PDT doses (figure 10(a)), with an initial transient swelling in the high-dose group. Similar responses have been described by Jeeves et al (1987). Light and electron microscopy revealed that the depth of damage below the surface increased with light dose (figure 9).…”

“…Spheroids thus represent a 3D tumour model to investigate purely cellular changes without extraneous in vivo factors. Spheroids have been used to study PDT response (Jeeves et al 1987, West 1988) and the effects of the microenvironment (O 2 , pH) on this (Chan et al 1991) and to quantify the effects of photobleaching and photochemical depletion of oxygen during PDT (Mitra and Foster 2000). The goal here was to determine whether EIS is sensitive to purely cellular PDT damage and whether it can discriminate between necrosis and apoptosis, both of which are believed to occur with PDT, depending on the exact treatment conditions (Oleinick and Evans 1998).…”

Multicell tumor spheroids in photodynamic therapy

“…This, and other early studies, focused mainly on evaluating hematoporphyrin (Hpd) distributions and the effects of Hpd-PDT in a variety of MCS models [15][16][17][18][19]. In particular, fundamental studies by West [16] and West et al [19] demonstrated that human colon carcinoma spheroids were more resistant to Hpd-PDT compared to monolayers and that the sensitivity to PDT decreased with increasing spheroid size.…”

The biology of photodynamic therapy

“…Three-dimensional multicellular spheroids, tumour models intermediate between cell monolayers and organized tumours, are typically more resistant to PDT than exponentially growing monolayers (Christensen et al 1984, Jeeves et al 1985 and this resistance is spheroid size dependent (West 1989). A decrease in photosensitizer uptake with increasing spheroid size has been demonstrated (Jeeves et al 1987, West 1989, West and Moore 1992) and an increase in heterogeneity of intracellular drug levels with increasing spheroid size (leading to the loss of the shoulder of the survival curve) for large spheroids (West and Moore 1989b). Cellular sub-populations resistant to PDT, that have been sorted from spheroids, have been shown to have smaller median cell volumes, to be enhanced in G 1 -type cells and to show reduced LDL uptake (West and Moore 1992).…”