Chapter 3 │ Page 87 generated short-lived RONS (•OH, •NO, O/O3) as main drivers for ICD induction in cancer cells [26]. Therefore, we hypothesized that the ICD-inducing capacity could potentially enhance the immunogenicity of cancer cell killing by CDDP, and assessed this by evaluating a set of key DAMPs according to their dynamic expression pattern over a course of 72 hours. We first investigated the ICD-associated expression of ecto-CRT, and HSP70 and 90 on the cell surface using IF staining. As a direct test of whether our novel NTPCDDP combination enhanced DAMP expression, we demonstrated that the addition of NTP promoted the translocation and surface expression of all three markers across di erent HNSCC spheroids 24 hours post treatment (Figure 6, and quantified in Figure 7). Figure 6: Microscopic imaging of membrane-associated DAMP expression in HNSCC spheroids. Surface-presented DAMPs ecto-CRT (left section), HSP70 (middle section), and HSP90 (right section) were visualized with IF 24 hours after treatment application. Representative images show marker expression in untreated samples vs samples exposed to the COMBOhigh regimen in the Cal27 cell line. Scale bar is 400µm.
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