HK2-IN-5

HK2-IN-5 (compound 12) (0-12.5 μM; 10 min preincubation) inhibits HK2 enzyme activity with an IC₅₀ of 1.17 μM; HK2-IN-5 shows weak inhibition of HK3 and inhibits only approximately 20% of GCK enzyme activity at 100 μM, demonstrating subtype selectivity for HK2^[1].

HK2-IN-5 (48 h) inhibits the proliferation/cell viability of MDA-MB-231, HCT116, SW480, HT1080, and A375 tumor cells with IC₅₀ values of 4.04, 12.85, 32.00, 6.64, and 5.92 μM, respectively; its IC₅₀ values against Vero and NCM460 cells are 137.21 and 35.12 μM, respectively, corresponding to selectivity indices of 34 and 8.7 relative to MDA-MB-231 cells^[1].
The cytotoxicity of HK2-IN-5 is attenuated in HK2-knockdown MDA-MB-231 cells; its IC₅₀ values in the siNC, siHK2-1, siHK2-2, and siHK2-3 groups are 6.44, 12.31, 6.34, and 11.52 μM, respectively, among which siHK2-1 and siHK2-3, which effectively reduce HK2 expression, simultaneously decrease cellular sensitivity to HK2-IN-5^[1].
HK2-IN-5 (0.5-8 μM; 48 h) inhibits colony formation of cultured cells in a concentration-dependent manner, gradually reducing both the number and size of colonies, and progressively decreases the wound healing efficiency^[1].

HK2-IN-5 (4 μM; 48 h) increases the protein retention levels of HK2 after heat treatment at 40, 43, 46, and 49°C in the CETSA assay in MDA-MB-231 cells, supporting intracellular target engagement of HK2-IN-5 with HK2^[1].

HK2-IN-5 (2-8 μM; 12 h) concentration-dependently increases intracellular ROS levels in MDA-MB-231 cells^[1].

HK2-IN-5 (2-8 μM; 24 h) decreases the mitochondrial membrane potential (ΔΨm) of MDA-MB-231 cells in a concentration-dependent manner, with ΔΨm reduced by 42% at 8 μM compared with the control group^[1].