Annihilation of metastatic 4T1 breast tumor micrometastases by photodynamic therapy with gemcitabine

Background: STL-6014, is an RGD-coupled bacteriochlorophyll photosensitizer that targets the tumor vasculature, cells and extracellular matrix. Gemcitabine (Gemzar) is an approved chemotherapeutic agent that attenuates immune suppressor cells. We hypothesized that STL-6014 PDT preceded by administration of Gemzar, will improve primary tumor control and anti-tumor immunity.

Methods: 4T1-Luc tumor grafts in the mouse mammary pad were subjected to PDT by intravenous administration of STL-6014 and 753 nm laser illumination (120 mW/cm², 10 min) 6h thereafter. Mice were intraperitoneally injected with Gemzar (75 mg/kg) at 5 days intervals from day -2 to day 25. Metastases were identified by luciferin bioluminescence. Single-cell RNA sequencing of splenocytes isolated on days 7 and 26 from control, Gemzar-, PDT- and PDT/Gemzar-treated mice was performed using the 10X Genomics platform.

Results: FACS analyses and immunohistochemistry showed STL-6014 accumulation at 6h in cancer, endothelial, and pro-tumor immune cells. Primary tumor eradication was observed in 62% of the PDT-treated animals with 90% of the animals showing metastatic dissemination, by week 3-4. PDT combined with Gemzar, resulted in 53% disease-free survival. Cancer cells were rejected in 70% of these animals at three months re-challenge test. Single-cell 10X Genomics indicated dramatic increase of mature B cells, CD8, CD4 T cells, natural killer cells and dendritic cells, alongside a dramatic decrease in myeloid-derived suppressor cell counts in the treated animals.

Conclusion: Vascular-targeted PDT synchronized with a metronomic Gemzar regimen presents a new means for disseminated disease control through evolution of antitumor immunity.