5-Aminolevulinic acid tumor paint and photodynamic therapy for myxofibrosarcoma: an in vitro study

Abstract

Background: 5-Aminolevulinic acid (5-ALA), a fluorescent contrast agent, has been used for tumor paint and photodynamic therapy (PDT) for various tumors, but its use with soft tissue sarcomas is not well documented. Myxofibrosarcoma, a subtype of soft tissue sarcoma with a high local recurrence rate, may benefit from similar types of treatment. The purpose of this study was to analyze the effects of 5-ALA tumor paint and PDT on a myxofibrosarcoma cell line. Methods: Tumor paint was assessed by exposing micromass pellets of human adipose-derived stromal (ADS) cells or myxofibrosarcoma (MUG-Myx1) cells to 5-ALA. Cell pellets were then visualized using a microscope at established excitation and emission wavelengths. Corrected total cell fluorescence was calculated per accepted protocols. Photodynamic therapy was similarly assessed by exposing ADS and MUG-Myx1 cells to 5-ALA, with subsequent analysis via flow cytometry and real-time confocal microscopy. Results: The use of 5-ALA tumor paint led to a selective fluorescence in MUG-Myx1 cells. Findings were confirmed by flow cytometry. Interestingly, flow cytometry results showed progressive selective cell death with increasing 5-ALA exposure as a result of the PDT effect. PDT was further confirmed using confocal microscopy, which revealed progressive cellular bubble formation consistent with advancing stages of cell death-a finding that was not seen in control ADS cells. Conclusions: 5-ALA tumor paint and PDT were successfully used on a human myxofibrosarcoma cell line (MUG-Myx1). Results from this study showed both selective fluorescent tagging and selective cytotoxicity of 5-ALA toward malignant myxofibrosarcoma cells, while sparing benign adipose control cells. This finding was further confirmed in a dramatic time-lapse video, visually confirming active, targeted cell death. 5-ALA's two-pronged application of selective tumor identification and cytotoxicity may transform surgical and medical approaches for treating soft tissue sarcomas.

Consistent localization of SARS-CoV-2 spike glycoprotein and ACE2 over TMPRSS2 predominance in placental villi of 15 COVID-19 positive maternal-fetal dyads

“…Images were measured by a total of three blinded reviewers (authors EST, YB, and KR). These values were then used to calculate a corrected total cell fluorescence (CTCF) per published protocols using the following calculation: CTCF = Integrated density – (Area of selected image x average mean background fluorescence) [ [40] , [41] , [42] ]. An average CTCF was also calculated from secondary only negative control slides (n = 3 per staining assay) and used to calculate a final Fluorescence Ratio: target antigen CTCF/secondary only control CTCF.…”

Decidual immune response following COVID-19 during pregnancy varies by timing of maternal SARS-CoV-2 infection

“…Image area and integrated density were measured via ImageJ software (imagej.net) for each immunofluorescent 200x image (n=5/slide) along with mean fluorescence values from 5 randomly selected background readings per image cohort to calculate a corrected total cell fluorescence (CTCF) per published protocols ( Kenan et al, 2020 , Taglauer et al, 2020 , Benarroch et al, 2021 ). An average CTCF was calculated from secondary only negative control slides (n=3 per staining assay) and used to calculate a final Fluorescence Ratio: target antigen CTCF/secondary only control CTCF.…”

5-ALA-mediated fluorescence of musculoskeletal tumors in a chick chorio-allantoic membrane model: preclinical in vivo qualification analysis as a fluorescence-guided surgery agent in Orthopedic Oncology

“…Kusuzaki et al reported that tumor fluorescence could be detected in lesions measuring as small as 1 mm using acridine orange (AO) in a mouse osteosarcoma model [ 6 ]. The qualification of 5-ALA in photodynamic detection (PDD) and photodynamic therapy (PDT) of musculoskeletal sarcoma has also been investigated with promising results in in-vitro studies [ 7 , 8 ].…”