5-Aminolevulinic Acid Tumor Paint and Photodynamic Therapy for Chordoma: An In Vitro Study

Abstract

Abstract Background: Chordomas are malignant tumors originating from embryonic notochord remnants. Wide resection is challenging due to lack of intraoperative visualization, leading to unacceptably high recurrence rates. Known techniques using five-aminolevulinic acid (5-ALA) for tumor paint and photodynamic therapy (PDT) may improve outcomes but have not been well described for chordomas. The purpose of this study was to analyze chordoma cell response to 5-ALA tumor paint and PDT in vitro. Methods: Tumor Paint: Experiment 1: Human chordoma cells (MUG-Chor1) co-cultured with green fluorescence protein (GFP) rat adipose derived stromal cells (ADS) were observed 5 hours after 5-ALA exposure using emission filters showing GFP cells as green and 5-ALA positive cells as red. Experiment 2: GFP-ADS and MUG-Chor1 micromass pellets were co-cultured and observed 5 hours after 5-ALA exposure. Experiment 3: co-cultured human ADS and MUG-Chor1 cells were observed 5 hours after 5-ALA exposure. Photodynamic Therapy: MUG-Chor1 cells that had been exposed to 3 hours of 5-ALA were visualized at 10× with 405-nm excitation and emission at 603-738-nm. Time-lapse images of live cells were captured every second for 15 minutes and then visualized sequentially at 50× speed. Results: Tumor Paint: In the first experiment, chordoma cells displayed minimal red fluorescence while GFP-ADS cells appeared to fluoresce expectedly green. In the second experiment, chordoma cells fluoresced brightly red. As a micromass, the distinction between chordoma cells and GFP-ADS cells was clearly seen. The third experiment showed strong red fluorescence in high cellular density areas, yet minimal fluorescence in low cellular density areas. PDT: Initially, chordoma cells fluoresced red in response to the 405-nm excitation laser exposure. As time progressed, increased intracellular swelling occurred with concomitant rapid formation of multiple vesicles exiting from the cellular membrane. Conclusions: The results of these experiments demonstrate the effective in vitro application of 5-ALA tumor paint and PDT on chordoma cells. This will be the first time this phenomenon is documented visually using time-lapse photography for this tumor subtype, further validating prior research in this field. 5-ALA, with its double-edged applications of selective tumor identification and kill may lead to transformative change in the management of soft tissue sarcomas.