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Updated: May 12, 2026
Implementation of In Vitro Drug Resistance Assays: Maximizing the Potential for Uncovering Clinically Relevant Resistance Mechanisms
Published on: December 9, 2015
Sequential inhibitor therapy in CML: in vitro simulation elucidates the pattern of resistance mutations after second-
Robert C Bauer1, Jana Sänger, Christian Peschel
1III. Medizinische Klinik und Poliklinik, Klinikum rechts der Isar, Technische Universität München, München, Germany.
Purpose:
Dasatinib and nilotinib are active in imatinib-resistant chronic myelocytic leukemia (CML) and many patients undergo sequential treatment. We aimed at modeling sequential tyrosine kinase inhibitor (TKI) resistance in vitro to compare the sequences imatinib-nilotinib-dasatinib and imatinib-dasatinib-nilotinib.
Experimental Design:
We designed an in vitro model for sequential TKI resistance in CML. Replicates of imatinib-resistant cell lines were treated with dasatinib or nilotinib. Second-line resistant replicates were exposed to third-line treatment.
Results:
Growth of all replicates in all three lines of treatment was associated with T315I. However, T315I occurred with low abundance and did not increase during sequential treatment. Nilotinib second-line more often gave rise to sequential resistance compared with dasatinib due to pre-existing P-loop mutations, especially at suboptimal drug concentration. In contrast, mutations predisposing to dasatinib resistance such as F317C/V and V299L did not occur before dasatinib exposure. Nilotinib third-line did not overcome imatinib-dasatinib resistance due to pre-existing T315I or P-loop/V299L or P-loop/F317 exchanges. Dasatinib third-line suppressed imatinib-nilotinib-resistant replicates with residual sensitivity.
Conclusions:
Sequential acquisition of BCR-ABL drug resistance mutations in CML might be underestimated. Resistance to sequential TKI monotherapy in vitro more often was associated with stepwise acquisition of drug-specific compound mutations compared with T315I. Pre-existing mutations strongly limited the activity of both third-line treatments, and the activity of nilotinib second-line in vitro critically depended on drug concentration.
Insights
Sequential tyrosine kinase inhibitor (TKI) resistance in chronic myelocytic leukemia (CML) can be underestimated. Pre-existing mutations significantly limit third-line TKI efficacy, with nilotinib
Area of Science:
- Oncology
- Hematology
- Molecular Biology
Background:
- Sequential tyrosine kinase inhibitor (TKI) therapy is common in chronic myelocytic leukemia (CML).
- Understanding resistance mechanisms to sequential TKIs is crucial for optimizing treatment strategies.
- Dasatinib and nilotinib are effective second-line options after imatinib failure.
Purpose of the Study:
- To model and compare sequential TKI resistance in vitro.
- To evaluate the efficacy of imatinib-nilotinib-dasatinib versus imatinib-dasatinib-nilotinib sequences.
- To investigate the role of specific mutations in acquired TKI resistance.
Main Methods:
- Development of an in vitro model for sequential TKI resistance in CML cell lines.
- Exposure of imatinib-resistant cells to sequential treatment with dasatinib and nilotinib.
- Analysis of resistance mutations, including T315I and P-loop mutations, after sequential drug exposure.
Main Results:
- Sequential TKI resistance was associated with T315I mutation, though often at low abundance.
- Nilotinib as a second-line treatment more frequently induced sequential resistance than dasatinib, particularly at suboptimal concentrations.
- Pre-existing mutations (T315I, P-loop, F317, V299L) significantly impaired the efficacy of third-line TKIs.
- Dasatinib demonstrated residual sensitivity against imatinib-nilotinib-resistant cells.
Conclusions:
- The sequential acquisition of BCR-ABL drug resistance mutations in CML may be underestimated.
- Stepwise acquisition of drug-specific compound mutations, rather than solely T315I, drives resistance to sequential TKI monotherapy.
- Pre-existing mutations are critical determinants of third-line treatment outcomes.
- Nilotinib's efficacy in the second-line setting is highly dependent on drug concentration.
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