Abstract B15: The pan-BCR-ABL inhibitor ponatinib inhibits viability of gatekeeper mutant BCR-ABLT315I cells with greater potency than a nilotinib/MEK inhibitor combination

Abstract

Abstract Introduction: The second generation BCR-ABL inhibitor nilotinib has recently been shown to have weak off-target activity against RAF and drive the paradoxical activation of BRAF and CRAF in cells with activated RAS. Importantly, this effect has been seen in cells expressing T315I mutant BCR-ABL, against which nilotinib is ineffective (Packer, Cancer Cell 20:715). Interestingly, nilotinib was shown to synergize with an inhibitor of MEK, a downstream effector of RAF, to inhibit the viability of cells expressing BCR-ABLT315I. Ponatinib is a pan-BCR-ABL inhibitor that potently kills cells expressing BCR-ABLT315I and is in a pivotal phase 2 trial in patients with CML. Here we examined the activity of ponatinib against RAF and compared the cellular potency of ponatinib against BCR-ABLT315I to that of a nilotinib/MEK inhibitor combination. Results: Ponatinib potently inhibited the in vitro kinase activity of ARAF, BRAF, and CRAF with IC50s of 71, 33, and 17 nM, respectively, compared to IC50s of 4390, 235, and 188 for nilotinib. In 2 RAS mutant cell lines tested, nilotinib (1 to 10 μM) induced feedback activation of RAF/MEK/ERK signaling as reported previously; however ponatinib (0.01 to 10 μM) did not. Both ponatinib and nilotinib potently inhibited viability of Ba/F3 cells expressing native BCR-ABL, with IC50s of 1.5 and 10 nM, respectively. While nilotinib has no activity against cells expressing BCR-ABLT315I, when combined with a concentration of the MEK inhibitor AZD6244 that inhibits MEK signaling (2 μM), nilotinib inhibited viability with an IC50 of 550 nM. In contrast, ponatinib potently inhibited viability of cells expressing BCR-ABLT315I with an IC50 of 6 nM. Similarly, single agent ponatinib induced apoptosis in BCR-ABLT315I cells with 100-fold greater potency than the combination of nilotinib and AZD6244. Conclusion: In contrast to nilotinib, ponatinib does not appear to induce the paradoxical activation of RAF/MEK/ERK signaling in RAS mutant cells, possibly due to its inhibition of all 3 RAF family members. Although nilotinib synergizes with a MEK inhibitor to kill BCR-ABLT315I mutant cells, this effect requires a concentration of nilotinib approximately 50-fold higher than that required to kill native BCR-ABL cells. Substantially greater potency, at clinically achievable drug concentrations, is achieved via the direct inhibition of BCR-ABLT315I by ponatinib.