Comparison of Imatinib, Dasatinib, Nilotinib and INNO-406 in Imatinib-Resistant Cell Lines.

Second-generation ABL tyrosine kinase inhibitors (TKIs) such as dasatinib, nilotinib and INNO-406 (formerly NS-187) have been developed to override imatinib-resistance mechanisms. We directly compared the growth-inhibitory effects in imatinib-sensitive and -resistant CML cell lines and the inhibitory profile for SRC family kinases (SFKs) among ABL TKIs. Dasatinib was the most potent inhibitor in all six CML cell lines evaluated (K562, BV173, KU812, MEG01, KT-1, MYL). Despite both nilotinib and INNO-406 being 2-phenylaminopyrimidine derivatives, INNO-406 demonstrated 2.5 times more activity than nilotinib against BV173 and KU812, although in the other four cell lines, there was no significant difference between these two TKIs. In three imatinib-resistant cell lines including K562/D1-9 (P-glycoprotein overexpressing), K562-IMR (BCR-ABL overexpressing) and MYL-R (BCR-ABL and LYN overexpressing), dasatinib also showed the greatest potency. INNO-406 was 3.1 times more effective than nilotinib against MYL-R, confirming a possible effect of INNO-406 against LYN. Nilotinib showed more potency than INNO-406 against K562/D1-9, suggesting less affinity to P-gp. None of the ABL TKIs inhibited the growth of Ba/F3/T315I. Dasatinib showed at least six-fold greater potency than nilotinib and INNO-406 against most Ba/F3 harboring BCR-ABL mutants. However, dasatinib was not effective against T315A, F317L and F317A, which have been detected in dasatinib-resistant CML patients. Interestingly, nilotinib and INNO-406 inhibited T315A, F317L and F317V (Table 1). To determine why dasatinib was ineffective against the T315A, F317L and F317V, the X-ray crystal structures of the dasatinib/ABL (PDB ID: 2GQG) and INNO-406/ABL (PDB ID: 2E2B) complexes were closely explored. While the P-loop of ABL closely locates INNO-406 and tightly grips INNO-406, it locates remotely from dasatinib and does not directly interact with dasatinib. The T315A, F317L and F317A mutations cause decreased steric and hydrogen-bonding interactions. Accordingly, the P-loop is likely to compensate these decreased interactions for INNO-406 but not for dasatinib. Next we compared the inhibitory effect of dasatinib and INNO-406 against SFKs. Dasatinib inhibited all eight SFKs at very low concentrations, while INNO-406 inhibited only LCK and LYN. In conclusion, dasatinib showed the strongest potency against BCR-ABL with less selectivity over SFKs. Nilotinib showed weaker affinity for SFKs compared to the other compounds, but was highly specific for ABL and may be useful for P-glycoprotein overexpressing leukemic cells. INNO-406 had intermediate affinity between dasatinib and nilotinib and inhibited LCK and LYN, in addition to ABL. Both nilotinib and INNO-406 were potent inhibitors of the dasatinib-resistant T315A, F317L and F317V BCR-ABL mutations. These findings should be useful for treating imatinib-resistant patients with second-generation ABL TKIs.