TCIPS preceded aggregation. (A-B) WPs (300 × 10³ platelets per μL) were stimulated with increasing concentration (0.1-100 × 10³ cells per mL) of Caco-2 (A, n = 6), PC3M-luc (B, n = 6), or PC3 cells (B, n = 3) and representative dose-response profile for luminescence arbitrary absorbance unit (AAU) deriving from platelet ATP/ADP release are shown. The lines in gray display the amount of ATP/ADP released by cancer cells in the absence of any platelets. TRAP (10 μM)–stimulated platelets were used as positive control (data are expressed as mean ± SEM). (C-D) Progress time curves of TCIPS and aggregation (TCIPA; n = 6). The curves precisely depict the temporal relationship between platelet secretion and aggregation induced by max concentration (100 × 10³ cells per mL) of Caco-2 (C) or PC3M-luc (D) cells. Values (mean ± SEM) are expressed as the percentage of the maximum platelet response, identifiable with the percentage of the highest platelet secretion or aggregation measured for each donor at the indicated time point. (E) Platelet α-granule release in response to maximum concentration of cancer cells (100 × 10³ cells per mL) was measured by quantifying changes in CD62P surface expression in flow cytometry (***P < .0001; **P < .01; mean ± SEM). Each experiment was repeated 4 times in presence of PC3M and Caco-2 and 3 times in presence of PC3 cells. The level of platelets expressing P-selectin was defined as a fraction of the 10 000 platelets that exhibit specific CD62P binding. The percentage of CD62P binding was determined as the percentage gated and the fold increase in cancer cell–stimulated platelets compared with resting platelets was recorded. AAU, arbitrary absorbance unit.