ABSTRACT Purpose It has been suggested that heavy-ion radiation therapy may contribute to the control of distal metastases. These distant responses may include immune cell activation. Immunostimulation resulting from radiation-induced immunogenic cell death (ICD) of cancer cells, leads to the recruitment of anti-tumor T cells. Specific markers of ICD include translocation of calreticulin (CRT) and extracellular release of high mobility group box 1 protein (HMGB1), and ATP. However, the LET dependence of these effects remains unknown. Materials and Methods Expression of the molecular indicators described above were tested in a panel of human cancer cell lines, that included pancreatic cancer (Panc1 and Paca2), glioblastoma (U87 and LN18) and melanoma (HTB129 and SK-Mel5). Cells were irradiated with 5 Gy of particles spanning a range of LETs, from 10 KeV/μm to 150 KeV/μm and assayed for relocalization of calreticulin and release of HMGB1 and ATP were assayed 24 hours later. Results In the pancreatic cancer cell lines (Panc1 and Paca2) there was a continued increase in the membrane relocalization of calreticulin as a function of increasing LET up to 150 KeV/μm. The melanoma cell lines, HTB129 and Sk-Mel5 showed similar patterns. In contrast, calreticulin levels were higher, but not LET-dependent, in irradiated U87 and LN18 (glioblastoma) lines. With the exception of the response in Paca2, increases in LET correlated with increases in HMGB1 that seemed to peak at 100 KeV/μm and then either remain unchanged or decrease at 150 KeV/μm. while the ATP levels were elevated in media from some of the irradiated groups, there were no clear patterns either by cell type or LET. Conclusions Our results indicate that at equal doses, although there is an overall trend of increases in the responses to increasing LETs, there are significant cell line-specific differences in the patterns of expression of these key ICD markers.

LET-dependence of radiation-induced makers of Immunogenic Cell Death in human cancer cell lines