FAK-IN-26 is a blood-brain barrier-penetrating inhibitor of Focal Adhesion Kinase (FAK) with an IC50 of 0.87 nM. It significantly reduces tumor cell viability, cancer stem cell activity, and cell migration in A549 and SKOV-3 cell lines. FAK-IN-26 exhibits potent anticancer activity, achieving tumor inhibition rates of 59.15% and 57.9% in A549 and SKOV-3 tumor mouse models, respectively.

FAK-IN-26 (Compound A8) exhibits excellent binding affinity for FAK with a Kd value of 15 μM. At 1 μmol/L, it broadly inhibits various kinases, achieving inhibition rates over 95% for both FAK and FYNα. FAK-IN-26 reduces the viability of A549 and SKOV-3 cells significantly at lower concentrations compared to Defactinib (VS6063) at 0.5-10 μM. It also decreases the cancer stem cell population in A549 and SKOV-3 cells in a dose-dependent manner at 0.2-1.6 μM. Moreover, FAK-IN-26 induces significant G2/M phase arrest in these cell lines at concentrations of 62.5-1000 nM over a period of 72 hours. The compound reduces cell migration and distance traveled by A549 and SKOV-3 cells in a dose-dependent manner within 0.1-10 nM over 6-48 hours, with significant inhibition observed at 1 nM and 10 nM. It effectively inhibits FAK autophosphorylation in a dose-dependent manner in the cell lines at 31.25-62.5 nM over 48 hours, with stronger inhibition noted in A549 cells. Additionally, FAK-IN-26 demonstrates good metabolic stability in human liver microsomes, with CL int of 31.8 μL/min/mg and a half-life (T 1/2) of 43.6 minutes.

In the A549 and SKOV-3 tumor mouse models, FAK-IN-26 (Compound A8) [(25-50 mg/kg, oral administration, once or twice daily for 5 days, followed by a 2-day break, over 28 days)] exhibits superior antitumor efficacy compared to Defactinib (VS6063), Erlotinib, and Paclitaxel, with enhanced effects when combined with Paclitaxel in the SKOV-3 model. FAK-IN-26 shows good tolerability in mice up to the highest tested dose of 2000 mg/kg without acute toxicity. Additionally, FAK-IN-26 demonstrates effective tumor uptake and retention, with peak uptake of 4.16 ID/g at 30 minutes in S180 tumor mice and penetrates the BBB, registering brain uptake of 2.63% ID/g at 15 minutes and 1.62% ID/g at 120 minutes.