The major therapeutic challenge in Ph + leukemia is to efficiently treat patients with BCR/ABL harboring the T315I mutation. The T315I mutation is the most resistant to inhibition because of a combination of several factors, including steric hindrance of drug binding, loss of a key hydrogen-bonding interaction with the T315 side-chain hydroxyl group exploited by Imatinib, Nilotinib and Dasatinib and potentially through increasing aberrant intrinsic kinase activity accompanied by aberrant substrate phosphorylation [13
]. Unfortunately, T315I confers resistance not only against ABL kinase inhibitors but also against the allosteric inhibition by GNF-2 [11
]. Allosteric inhibition is a novel approach for targeting BCR/ABL, which overcomes the resistance mediated by the T315I in combination with inhibition of oligomerization [4
]. The fact that the competitive peptides for oligomerization inhibition are still far from clinical application led us to explore whether the allosteric inhibition could also improve the response of BCR/ABL-T315I to competitive ATP analogues.
GNF-2 and its analogues are non-ATP competitive ABL kinase inhibitors, which bind to the MBP in the kinase domain. It seems that the binding of GNF-2 to the MBP stabilizes the protein in an inhibited conformation resulting in a structural reorganization of ABL that disrupts the catalytic machinery located in the ATP-binding region [7
]. Thus, one can speculate that GNF-2 introduces changes in the overall conformations of BCR/ABL-T315I, which renders the ATP-binding site more accessible to Dasatinib. This result is confirmed by recent biophysical studies showing that Dasatinib induces conformational changes in unmutated BCR/ABL but not in BCR/ABL-T315I. In contrast, GNF-5 leads to the same changes in both unmutated BCR/ABL and BCR/ABL-T315I [15
An additive but not synergistic effect was shown for the combination of Nilotinib with GNF-2 or GNF-5 on BCR/ABL-T315I-related resistance. The stronger effects may be attributed to the fact that Dasatinib, originally developed as a SRC-kinase inhibitor, not only inhibits the BCR/ABL kinase but also targets a broader range of kinases compared to Nilotinib, the spectrum of which is mainly limited to ABL, c-KIT and PDGFR [16
]. An additional effect of GNF-2 itself on SRC family kinases is unlikely. c-SRC is also myristoylated and harbors a putative MBP, which is involved in the regulation of c-SRC kinase activity, but in a manner very different from that for c-ABL [17
Our data further establish allosteric inhibition as alternative or additional molecular therapy approach for the treatment of Ph+ leukemia. In fact, it not only overcomes the resistance mediated by the “gatekeeper” mutation T315I but also increases the response of unmutated BCR/ABL to AKI. In the clinical setting, this feature could contribute to a more efficient use of AKI at a lower dosage in “normally” responsive patients and the possibility to further increase dosage in patients early in the progression of disease, in the absence of BCR/ABL mutations, for whom dosage escalation is still a therapeutic option.
The results presented here contribute to the further development of allosteric inhibition for the molecular targeting of both unmutated BCR/ABL and BCR/ABL harboring the multi-resistance mutation T315I.