Fig. 6: P-LPK bound to SLC1A5 and entered CRC cells through endocytosis.
From: Anti-tumor effects of P-LPK-CPT, a peptide-camptothecin conjugate, in colorectal cancer
a, b Western blotting analysis of SLC1A5 in NCM460, HCT116, LoVo, SW480, HT29 and Colo320HSR cells. After knocking down SLC1A5 in HCT116 and HT29, the binding intensity of FITC-P-LPK to cells was observed by confocal microscopy. Bar, 10 μm. c, d Effects of different endocytosis inhibitors on the internalization of FITC-P-LPK. HCT116 cells were pre-treated with 10 µM chlorpromazine, 50 μM methyl-β-cyclodextrin, 20 µM Amiloride hydrochloride for 30 min at 37 °C. Subsequently, the cells were incubated with FITC-P-LPK for 2 h. Then the fluorescence intensity was observed by confocal microscopy and detected using a multi-well plate reader (n = 5, means ± SD, NC vs CPZ, p = 0.0001; NC vs MβCD, p = 0.9069; NC vs EIPA, p = 0.9812) (****p < 0.0001). Bar, 10 μm. e The competition experiment with SLC1A5 substrates glutamine. FITC-P-LPK was incubated with increasing concentrations (0–32 mM) of glutamine in HCT116 cells at 37 °C for 2 h. The fluorescence intensity was measured in a multi-well plate reader (n = 5, means ± SD).
