Abstract
Asciminib (previously ABL001), which binds the myristate-binding pocket of the Bcr-Abl kinase domain, is in phase I clinical trials as monotherapy and in combination with imatinib, nilotinib and dasatinib for the treatment of patients with refractory CML or Ph+ ALL. Asciminib sensitivity was evaluated in asciminib naïve BCR-ABL1+ cell lines K562 (negligible ABCB1/ABCG2 expression), K562-Dox (ABCB1-overexpressing through doxorubicin exposure) and K562-ABCG2 (ABCG2 overexpression via transduction) with results demonstrating asciminib efflux by both ABCB1 and ABCG2 transporters. K562-Dox and K562-ABCG2 cells demonstrated increased LD50asciminib vs K562 control cells: 256 and 299 nM respectively vs 24 nM, p < 0.001. Sensitivity was completely restored with specific inhibitors cyclosporine (ABCB1) and Ko143 (ABCG2): K562-Dox LD50asciminib+cyclosporine = 13 nM, K562-ABCG2 LD50asciminib+Ko143 = 15 nM (p < 0.001). When asciminib resistance was modelled in vitro, ABCB1 and ABCG2 overexpression was integral in the development of asciminib resistance. In K562 asciminib-resistant cells, ABCG2 expression increased prior to BCR-ABL1 overexpression and remained high (up to 7.6-fold greater levels in resistant vs control cells, p < 0.001). K562-Dox asciminib-resistant cells had increased ABCB1 expression (2.1-fold vs control cells p = 0.0033). KU812 asciminib-resistant cells overexpressed ABCB1 (5.4-fold increase, p < 0.001) and ABCG2 (6-fold increase, p < 0.001) before emergence of a novel myristate-binding pocket mutation (F497L). In all three cell lines, asciminib resistance was reversible upon chemical inhibition of ABCB1, ABCG2 or both (p < 0.001). When K562 asciminib-resistant cells were treated with asciminib in combination with clinically achievable doses of either imatinib or nilotinib, reversal of the resistance phenotype was also observed (p < 0.01). Overexpression of efflux transporters will likely be an important pathway for asciminib resistance in the clinical setting. Given the lack of evidence for ABCG2-mediated transport of nilotinib or imatinib at clinically relevant concentrations, our data provide an additional rationale for using asciminib in combination with either TKI.
Highlights
The first generation ATP-competitive tyrosine kinase inhibitor (TKI) imatinib was designed to bind the ATP-pocket of Bcr-Abl [1]
Asciminib-mediated cell death was evaluated in K562Dox (ABCB1 overexpressing) and K562-ABCG2 overexpressing cells compared with parental K562 cells
While difficult to determine the exact sensitivity of KU812 10 μM asciminib cells to imatinib, nilotinib and dasatinib due to the multiple overlapping resistance mechanisms present in this cell line (F497L, ABCB1, ABCG2, BCR-ABL1 overexpression; Figure 1D) we have evaluated TKI-sensitivity in the absence of transporter overexpression, which is responsible for the majority of asciminib resistance (Figure 3C)
Summary
The first generation ATP-competitive tyrosine kinase inhibitor (TKI) imatinib was designed to bind the ATP-pocket of Bcr-Abl [1]. The second generation inhibitors nilotinib and dasatinib, have resulted in excellent overall and event free survival rates in chronic myeloid leukemia (CML) patients [2,3,4]. T315I demonstrates resistance to all first and second generation inhibitors [12, 13], and the frequency of development increases with disease progression and exposure to multiple TKIs. While the third generation inhibitor ponatinib demonstrates activity against cells harboring the T315I mutation in vitro [14] and is successful at reducing disease burden in vivo [15, 16], it is associated with significant safety concerns [17]
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