Immunogenic Treatment for Metastatic Breast Cancer Using Targeted Carbon Nanotube Mediated Photothermal Therapy in Combination with Anti-PD-1

The high prevalence of breast cancer is a global health concern, but there are no safe or effective treatments for it at its advanced stages. These facts urge the development of novel treatment strategies. Annexin A5 (ANXA5) is a natural human protein that binds with high specificity to phosphatidylserine, a phospholipid tightly maintained in the inner leaflet of the cell membrane on most healthy cells but externalized in tumor cells and the tumor vasculature. Here, we have developed a targeted photosensitizer for photothermal therapy (PTT) of solid tumors through the functionalization of single walled carbon nanotubes (SWCNTs) to ANXA5-the SWCNT-ANXA5 conjugate. The ablation of tumors through the SWCNT-ANXA5-mediated PTT synergizes with checkpoint inhibition, creating a systemic anti-cancer immune response. In vitro ablation of cells incubated with the conjugate promoted cell death in a dose-dependent and targeted manner. This treatment strategy was tested in vivo with the orthotopic EMT6 breast tumor model in female balb/cJ mice. Enhanced therapeutic effects were achieved by using intratumoral injection of the conjugate and treating tumors at a lower PTT temperature (45oC). Intratumoral injection prevented the accumulation of the SWCNTs in major clearance organs. When combined with checkpoint inhibition of anti-PD-1, SWCNT-ANXA5-mediated PTT increased survival and 80% of the mice survived for 100 days. Evidence of immune system activation by flow cytometry of splenic cells strengthens the hypothesis of an abscopal effect as a mechanism of prolonged survival. Significance Statement This study demonstrated a relatively high survival rate (80% at 100 days) of mice with aggressive breast cancer when treated with photothermal therapy using the SWCNT-ANXA5 conjugate injected intratumorally and combined with immune stimulation using the anti-PD-1 checkpoint inhibitor. Photothermal therapy was accomplished by maintaining the tumor temperature at a relatively low level of 45oC and avoiding accumulation of the nanotubes in the clearance organs by using intratumoral administration.