Ng, Jane;
Henriquez, Nico;
MacRobert, Alexander;
Kitchen, Neil;
Williams, Norman;
Bown, Stephen;
(2022)
Bioluminescence-Activated Photodynamic Therapy for Luciferase Transfected, Grade 4 Astrocytoma cells in vitro.
Photodiagnosis and Photodynamic Therapy
, 38
, Article 102856. 10.1016/j.pdpdt.2022.102856.
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Abstract
BACKGROUND: Grade 4 astrocytoma is incurable due to the diffusely infiltrative nature of the disease. Photodynamic therapy (PDT) is a promising therapeutic option, but external light delivery is not feasible when cancer cells infiltrate unknown areas of normal brain. Hence the search for endogenous sources such as bioluminescence that can generate light at cancer cells. This requires a substrate (a luciferin) and an enabling enzyme (a luciferase), neither seen in mammalian cells. METHODS: Preliminary studies confirmed that U87 cells (derived from a human grade 4 astrocytoma) could be killed by conventional PDT using the photosensitisers hypericin or mTHPC. U87 cells were then transfected with firefly and other luciferases and light generating cell lines (U87-luc, U87-hRluc, U87-CBG68luc) identified using the appropriate substrate. Reagent doses and conditions were optimized and U87-luc cells incubated with hypericin or mTHPC with d-luciferin added to initiate bioluminescence activated PDT (bPDT). Cell survival was assessed by MTT assay, haemocytometry and growth assay. Control groups included U87-luc cells with no added active reagents, substrate only, photosensitiser only and non-transfected U87 cells. Results were expressed as a percentage of surviving cells compared with untreated U87-luc controls. RESULTS: There was no bPDT effect on non-transfected cells. The mean survival of treated transfected cells was 36%, (P<0.001) using hypericin and 35% (P<0.001) using mTHPC, compared with untreated U87-luc cells. bPDT effects were suppressed by the anti-oxidant, lycopene. CONCLUSIONS: bPDT can kill Grade 4 astrocytoma cells transfected with luciferase in vitro. This justifies progression to in vivo studies.
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