GIT1-IN-1

GIT1-IN-1 is an inhibitor of ARF GTPase-activating protein 1 (GIT1), with a KD value of 6.2 μM. It induces apoptosis (cell death) in liver cancer and colon cancer cells, halts the cell cycle in the G2/M phase, and suppresses cell proliferation, colony formation, and migration. Furthermore, GIT1-IN-1 can inhibit the activity of MEK and ERK in liver and colon cancer cells, reduce cyclin D1 expression, and stabilize cyclin B1 protein. This compound is applicable for research in liver and colon cancer.

GIT1-IN-1 (Compound C3) at concentrations of 10-100 μM over 24 hours inhibits the viability and proliferation of liver and colon cancer cells, specifically HepG2, Hep3B, MzChA-1, HT-29, and RKO, with an IC50 of approximately 20 μM, but shows no effect on non-malignant AML12, HEK293 cells, or primary mouse and human hepatocytes up to 100 μM. At 1-5 μM for 24 hours, GIT1-IN-1 induces dose-dependent G2/M phase arrest in HepG2 and RKO liver and colon cancer cells without affecting the cell cycle progression of non-malignant AML12 or HEK293 cells. Treatment with 5 μM for 24-48 hours prompts time-dependent apoptosis in RKO colon cancer cells and, after 48 hours, increases apoptosis and necrosis in HepG2 liver cancer cells without inducing cell death in non-malignant AML12 or HEK293 cells or primary mouse/human hepatocytes. Additionally, GIT1-IN-1 at 1-10 μM over 24 hours inhibits colony formation in HepG2 liver cancer cells and RKO colon cancer cells in a dose-dependent manner, while not impacting non-malignant AML12 cells. A concentration of 1-2 μM for 24 hours shows dose-dependent inhibition of migration in RKO colon cancer cells and MzChA-1 liver cancer cells. At doses of 5-10 μM for 24 hours, GIT1-IN-1 disrupts the interaction of GIT1-MAT2B and MEK1/2-cRAF/BRAF/ERK1/2/GIT1 at 10 μM, and at 5 μM disrupts interactions within cells of GIT1-CDC20, GIT1-APC3, cyclin B1-CDC20, and cyclin B1-APC3, while enhancing GIT1-cyclin B1 interaction. GIT1-IN-1 also suppresses the activity of the MEK and ERK pathways in HepG2, RKO, and MC38 cancer cells by decreasing levels of pMEK1/2, pERK1/2, and cyclin D1, with no impact on these pathways in non-malignant AML12 cells at 2-10 μM for 24 hours. The compound, at 2-5 μM for 24 hours, enhances cyclin B1 protein stability in HepG2 and RKO cancer cells, promotes cyclin B1-CDK1 complex formation, and increases CDK1 activity, evidenced by the extension of cyclin B1 half-life, reduced CDK1 inhibitory phosphorylation, and elevated cyclin B1 activating phosphorylation.

GIT1-IN-1 (Compound C3) administered at 100 μM via intratumoral injection every two days for six days in C57BL/6 mice effectively inhibits subcutaneous colorectal cancer xenograft growth by suppressing MEK activity, reducing tumor cell proliferation, and promoting apoptosis. Additionally, GIT1-IN-1 at 25 mg/kg, delivered by intraperitoneal injection once daily for five consecutive days, significantly reduces the growth of human colorectal cancer intrahepatic xenografts in nude mice. Moreover, GIT1-IN-1 at doses of 15-20 mg/kg administered intermittently via intraperitoneal injection dramatically inhibits colorectal cancer liver metastasis in immunocompetent C57BL/6 mice without causing detectable hepatotoxicity.