Chronic myelogenous leukaemia (CML) and Philadelphia chromosome positive (Ph+) acute lymphoblastic leukaemia (ALL) are caused by the BCR-ABL oncogene. Imatinib inhibits the tyrosine kinase activity of the BCR-ABL protein and is an effective, frontline therapy for chronic-phase CML. However, accelerated or blast-crisis phase CML patients and Ph+ ALL patients often relapse due to drug resistance resulting from the emergence of imatinib-resistant point mutations within the BCR-ABL tyrosine kinase domain. This has stimulated the development of new kinase inhibitors that are able to over-ride resistance to imatinib. The novel, selective BCR-ABL inhibitor, AMN107, was designed to fit into the ATP-binding site of the BCR-ABL protein with higher affinity than imatinib. In addition to being more potent than imatinib (IC50<30 nM) against wild-type BCR-ABL, AMN107 is also significantly active against 32/33 imatinib-resistant BCR-ABL mutants. In preclinical studies, AMN107 demonstrated activity in vitro and in vivo against wild-type and imatinib-resistant BCR-ABL-expressing cells. In phase I/II clinical trials, AMN107 has produced haematological and cytogenetic responses in CML patients, who either did not initially respond to imatinib or developed imatinib resistance. Dasatinib (BMS-354825), which inhibits Abl and Src family kinases, is another promising new clinical candidate for CML that has shown good efficacy in CML patients. In this review, the early characterisation and development of AMN107 is discussed, as is the current status of AMN107 in clinical trials for imatinib-resistant CML and Ph+ ALL. Future trends investigating prediction of mechanisms of resistance to AMN107, and how and where AMN107 is expected to fit into the overall picture for treatment of early-phase CML and imatinib-refractory and late-stage disease are discussed.

Background: Methods: Results: Conclusion:

The tyrosine kinase activity of the Bcr/Abl oncogene is required for transformation of hematopoietic cells. The tyrosine kinase inhibitor STI571 (formerly called CGP57148B, Novartis Pharmaceuticals) inhibits BCR/ABL, TEL/ABL, and v-ABL kinase activity and inhibits growth and viability of cells transformed by any of these ABL oncogenes. Here we report the generation of 2 BCR/ABL–positive cell lines that have developed partial resistance to STI571. BCR/ABL–transformed Ba/F3 hematopoietic cells and Philadelphia-positive human K562 cells were cultured in gradually increasing concentrations of STI571 over a period of several months to generate resistant lines. Resistant Ba/F3.p210 cells were found to have an increase in Bcr/Abl messenger RNA, amplification of the Bcr/Abl transgene, and a greater than tenfold increase in the level of BCR/ABL protein. In contrast to Ba/F3.p210 cells, drug-resistant K562 cells did not undergo detectable amplification of the BCR/ABL gene, although they displayed a 2-fold to 3-fold increase in p210BCR/ABL protein. The addition of STI571 to both resistant Ba/F3.p210 and K562 cells resulted in a rapid reduction of tyrosine phosphorylation of cellular proteins, similar to that observed for nonresistant cells. However, the inhibition of kinase activity was transient and partial and was not accompanied by apoptosis. The results suggest that resistance to STI571 may be multifactorial. Increased expression of the target protein BCR/ABL was observed in both lines, and resulted from oncogene amplification in one line. However, altered drug metabolism, transport, or other related mechanisms may also contribute to drug resistance.

The molecular mechanisms by which Notch receptors induce diverse biological responses are not fully understood. We recently cloned a mammalian homologue of the Mastermind gene of Drosophila melanogaster, MAML1 (Mastermind-like-1 molecule) and determined that it functions as a transcriptional coactivator for Notch receptors. In this report, we characterize two additional genes in this Mastermind-like gene family: MAML2 and MAML3. The three MAML genes are widely expressed in adult tissues but exhibit distinct expression patterns in mouse early spinal cord development. All MAML proteins localize to nuclear bodies, share a conserved basic domain in their N termini that binds to the ankyrin repeat domain of Notch, and contain a transcriptional activation domain in their C termini. Moreover, as determined by using coimmunoprecipitation assays, each MAML protein was found to be capable of forming a multiprotein complex with the intracellular domain of each Notch receptor (ICN1 to -4) and CSL in vivo. However, MAML3 bound less efficiently to the ankyrin repeat domain of Notch1. Also, in U20S cells, whereas MAML1 and MAML2 functioned efficiently as coactivators with each of the Notch receptors to transactivate a Notch target HES1 promoter construct, MAML3 functioned more efficiently with ICN4 than with other forms of ICN. Similarly, MAML1 and MAML2 amplified Notch ligand (both Jagged2 and Delta1)-induced transcription of the HES-1 gene, whereas MAML3 displayed little effect. Thus, MAML proteins may modify Notch signaling in different cell types based on their own expression levels and differential activities and thereby contribute to the diversity of the biological effects resulting from Notch activation.