Fig. 1.

DHP-B suppresses colorectal cancer cell growth and induces apoptosis in vitro. (A) Chemical structure of DHP-B, highlighting its α-methylene-γ-butyrolactone moiety in red; (B) Dose-response curves of DHP-B against five colorectal cancer cell lines; (C) Comparison of IC₅₀ values of DHP-B and positive drug oxaliplatin (OXA) against colorectal cancer cells; (D) Comparison of DHP-B and peperomin B (PB) in inhibiting colorectal cancer cell proliferation; (E) Representative images of 3D spheroids of normal colon epithelial cells NCM460 and colorectal cancer cells treated with DHP-B (scale bar: 100 μm) and statistical graphs of the changes in 3D spheroid area at different concentrations on 0, 3, 6, 9, and 12 days (lines represent mean area, different colored bands represent SD values, n = 5); (F) Immunofluorescence staining and quantification of Ki-67 (green) in SW620 and LS180 tumor spheroids treated with 5 μM DHP-B or DMSO (Ctrl) for 6 days (scale bar: 20 μm). The number of Ki-67 positive cells were quantified using Image J 1.5.3 software; (G) Flow cytometry analysis and quantification of apoptosis in colorectal cancer cells treated with 5 μM DHP-B for 24 h; (H) Flow cytometry analysis and quantification of apoptosis in SW620 cells treated with different concentrations of DHP-B for 24 h. All of the studies above were examined using at least five biological replicates, and the results were expressed as mean ± SD; NS, not significant, *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, upaired t-test. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)