Inhibition of T315I Bcr-Abl and Other Imatinib-Resistant Bcr-Abl Mutants by the Selective Abl Kinase Inhibitor SGX70393.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1373-1373 ◽  
Author(s):  
Thomas O’Hare ◽  
Christopher A. Eide ◽  
Jeffrey W. Tyner ◽  
Matthew J. Wong ◽  
Caitlyn A. Smith ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Imatinib Resistance ◽  
Abl Kinase ◽  
Imatinib Resistant ◽  
Escape Mutants ◽  
A Cell ◽  
Almost All ◽  
Proliferation Assays

Abstract Imatinib effectively inhibits the tyrosine kinase activity of Bcr-Abl, the molecular driver of CML. Emergence of imatinib resistance due to mutations within the Bcr-Abl kinase domain (KD) has prompted the development of new Abl kinase inhibitors. A particularly important target is Bcr-Abl(T315I), which accounts for 15–20% of patients with resistance. To address this unresolved need, we profiled the novel Abl kinase inhibitor SGX70393 against native and mutant Bcr-Abl. Methods: We assessed the efficacy of SGX70393 in cellular and biochemical assays against a panel of KD mutants. Cell proliferation assays and Bcr-Abl tyrosine phosphorylation immunoblot analyses were performed for parental Ba/F3 cells, Ba/F3 cells expressing unmutated Bcr-Abl, or Ba/F3 cells expressing a single Bcr-Abl KD mutation (M244V, G250E, Q252H, Y253F, Y253H, E255K, E255V, F311L, T315I, F317L, M351T, F359V, V379I, L387M, H396P, or H396R). The resistance profile of SGX70393 was also evaluated using a recently developed accelerated, cell-based mutagenesis assay (Bradeen, et al. Blood, June 2006; doi:10.1182). Results: SGX70393 inhibited growth of cells expressing Bcr-Abl(T315I) (IC50: 7.3 nM) or unmutated Bcr-Abl (IC50: 12 nM). Sensitivity of Bcr-Abl mutants to SGX70393 partitioned into three categories: high (IC50<25 nM: M244V, T315I, F359V, V379I, L387M, H396P, and H396R), medium (IC50<300 nM: Q252H, Y253H, E255K, and F311L), and low (IC50>500 nM: G250E, Y253F, E255V, and F317L). A cell-based mutagenesis screen for Bcr-Abl kinase domain escape mutants emerging in the presence of SGX70393 revealed a concentration-dependent reduction in surviving clones, with five previously reported Bcr-Abl mutations (L248M; G250E; Y253F; E255V; F317V) accounting for almost all resistance. Conclusions: (a) SGX70393 is a potent inhibitor of native and T315I mutant Bcr-Abl. (b) SGX70393 coverage extends to most clinically relevant mutants except mutations of the p-loop and F317.

Optimal Treatment for Patients after Dasatinib or Nilotinib: Comparison of Preclinical Activities of Approved and Promising Investigational Kinase Inhibitors Against BCR-ABL Kinase Domain Mutations That Confer Clinical Resistance to Dasatinib or Nilotinib.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4589-4589
Author(s):  
Corynn Kasap ◽  
Christopher Weier ◽  
Neil P. Shah
Keyword(s):  
Second Generation ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Inhibitor Therapy ◽  
Drug Resistant ◽  
Imatinib Resistance ◽  
Abl Kinase ◽  
Clinical Resistance ◽  
Kinase Domain Mutations

Abstract The optimal management of patients with chronic myeloid leukemia (CML) is increasingly reliant upon molecular studies. Loss of response to imatinib in CML is most commonly associated with selection for a limited number of BCR-ABL kinase domain mutations that impair the ability of imatinib to effectively bind to BCR-ABL Molecular understanding of imatinib resistance mechanisms has led to the development of effective “second generation” BCR-ABL kinase inhibitors, such as dasatinib and nilotinib, which have clinical activity against most, but not all, drug-resistant mutations. Analysis of the BCR-ABL kinase domain in patients who develop resistance to second-generation inhibitors has implicated further selection of drug-resistant BCR-ABL kinase domain mutants in nearly all cases reported to date. Encouragingly, the number of resistant mutations capable of conferring clinical resistance to the most clinically-advanced second-generation agents, dasatinib (approved by the US FDA and EMEA) and nilotinib (approved in Mexico and Switzerland), appears to be restricted to a relatively small number of amino acid substitutions. As clinical experience with dasatinib and nilotinib grows, an understanding of the relative sensitivities of dasatinib- and nilotinib-resistant BCR-ABL mutants to other kinase inhibitors, both approved and investigational, is critical to optimize clinical outcomes in patients with resistance to dasatinib or nilotinib. At the present time, kinase inhibitor therapy options for patients with resistance to one of these agents include the investigational options bosutinib and MK-0457 (VX-680), as well as dasatinib and nilotinib (for patients not yet exposed to one of these agents) and re-exposure imatinib. It is likely that the success of therapeutic intervention in these cases can be predicted based upon the preclinical sensitivity of the mutation(s) involved with the agent chosen. We have therefore conducted a thorough biochemical and biological cross-analysis of the activities of each of these clinically-useful kinase inhibitors against mutations that confer clinical resistance to dasatinib or nilotinib. These studies provide clinicians with a useful reference for choosing an appropriate kinase inhibitor based upon the identity of the resistant BCR-ABL kinase domain mutation(s) detected at the time of relapse when faced with a patient who has lost response to dasatinib or nilotinib. It is hoped that the application of such “personalized medicine” strategies to the clinical management of CML cases will further improve outcomes in patients treated with kinase inhibitor therapy.

Comparison of Imatinib, AMN107 and Dasatinib in an Accelerated Cell-Based Mutagenesis Screen.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 691-691 ◽  
Author(s):  
Michael W.N. Deininger ◽  
Heather Bradeen ◽  
Taiping Jia ◽  
Thomas O’Hare ◽  
Stephanie G. Willis ◽  
...  
Keyword(s):  
Structural Changes ◽  
Kinase Inhibitors ◽  
Kinase Domain ◽  
Abl Kinase ◽  
Drug Concentrations ◽  
A Cell ◽  
Vitro Model ◽  
Proliferation Assays ◽  
Abl Kinase Inhibitors

Abstract Background. Mutations in the Bcr-Abl kinase domain (KD) are the leading cause of acquired imatinib (IM) resistance. Dasatinib (BMS354825) and AMN107 are potent alternate Abl inhibitors with activity at nanomolar levels against wild type Bcr-Abl and most KD mutants, with the exception of T315I. In a cell-line based mutagenesis assay we compared incidence and type of Bcr-Abl mutants emerging in the presence of IM, AMN107 and dasatinib. Methods. BaF3-p210Bcr-Abl cells were mutagenized by 24 hours exposure to 0.42 μM N-ethyl-N-nitrosourea (ENU), a dose with minimal cytotoxicity. After ENU washout cells were seeded at 5 x 105/well in 96-well plates and observed for growth for up to 4 weeks. Cells from wells with growth were expanded and subjected to BCR-ABL KD sequencing. Results: The frequency of wells with growth decreased with higher doses of all 3 inhibitors (table 1) and growth tended to occur later. Only isolated wells had growth without ENU exposure. At ≥2 μM IM (2-fold the IC90 in cell proliferation assays) 18 different mutations were seen, with highly resistant mutants prevailing at higher concentrations (table 2). At 50 nM AMN107 (2-fold the IC90) Y253H, G250E, F359C, E255K, L384M, L387F, E292V and T315I were detected, at 500 nM Y253H, E255V and T315I were recovered and only T315I at 2000 nM. At 5 nM dasatinib (2-fold the IC90), E255K, L284V, F317V were detected in addition to T315I, at 10 nM T315I, F317V/I and V299L were found and at 25 nM only T315I. All resistant clones growing out at ≥4 μM IM, 500 nM AMN107 or 10 nM dasatinib were KD mutant, suggesting that KD mutations were the sole cause of the observed resistance. Conclusions: (i) At drug concentrations corresponding to 2-fold IC9018 different mutations were recovered with IM, 9 with AMN107 and 6 with dasatinib, suggesting that the conformational requirements for dasatinib binding to Abl may be least stringent. If free plasma trough levels ≥25 nM dasatinib or ≥2000 nM AMN107 are achievable, the only mutant predicted to emerge clinically is T315I. (ii) No additional mutations were observed with AMN107 compared to IM, suggesting the structural changes in AMN107 compared to IM did not generate novel vulnerable sites. (iii) At least in this in vitro model, resistance to Abl kinase inhibitors is entirely dependent on Bcr-Abl, despite the fact that ENU treatment is expected to induce multiple additional mutations. Thus a T315I inhibitor combined with AMN107 or dasatinib may be effective at preventing the emergence of resistance to Abl kinase inhibitors. Table 1 Recovery of resistant clones (representative experiment) Imatinib (microM) Wells with mutations/wells sequenced/wells with growth Dasatinib (nM) Wells with mutations/wells sequenced/wells with growth AMN107 (nM) Wells with mutations/wells sequenced/wells with growth 2 62/62/82 5 8/24/96 10 0/24/96 4 74/74/74 10 38/38/56 50 20/24/96 8 27/27/42 25 22/22/24 500 45/45/46 16 12/12/12 100 20/20/21 2000 23/23/24 Table 2 Percentage of resistant clones with T315I mutations Imatinib (μM) number of different mutations/% T315I Dasatinib (nM) number of different mutations/% T315I AMN107 (nM) number of different mutations/% T315I 2 16/27.8 5 4/16.7 10 0/0.0 4 7/43.2 10 3/63.2 50 8/20.8 8 4/37.4 25 1/100 500 3/62.0 16 4/50.0 100 1/100.0 2000 1/100.0

Philadelphia-positive patients who already harbor imatinib-resistant Bcr-Abl kinase domain mutations have a higher likelihood of developing additional mutations associated with resistance to second- or third-line tyrosine kinase inhibitors

Blood ◽  
2009 ◽  
Vol 114 (10) ◽  
pp. 2168-2171 ◽  
Author(s):  
Simona Soverini ◽  
Alessandra Gnani ◽  
Sabrina Colarossi ◽  
Fausto Castagnetti ◽  
Elisabetta Abruzzese ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Tyrosine Kinase Inhibitors ◽  
Kinase Inhibitors ◽  
Sequential Therapy ◽  
Initial Response ◽  
Kinase Domain ◽  
Imatinib Resistance ◽  
Mutation Status ◽  
Abl Kinase ◽  
Imatinib Resistant

Abstract Dasatinib and nilotinib are tyrosine kinase inhibitors (TKIs) developed to overcome imatinib resistance in Philadelphia-positive leukemias. To assess how Bcr-Abl kinase domain mutation status evolves during sequential therapy with these TKIs and which mutations may further develop and impair their efficacy, we monitored the mutation status of 95 imatinib-resistant patients before and during treatment with dasatinib and/or nilotinib as second or third TKI. We found that 83% of cases of relapse after an initial response are associated with emergence of newly acquired mutations. However, the spectra of mutants conferring resistance to dasatinib or nilotinib are small and nonoverlapping, except for T315I. Patients already harboring mutations had higher likelihood of relapse associated with development of further mutations compared with patients who did not harbor mutations (23 of 51 vs 8 of 44, respectively, for patients who relapsed on second TKI; 13 of 20 vs 1 of 6, respectively, for patients who relapsed on third TKI).

A Cell-Based Screening Method for Resistance of Bcr-Abl Positive Leukemia Identifies the Mutation Pattern for an Alternative Abl Kinase Inhibitor.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 558-558
Author(s):  
Nikolas von Bubnoff ◽  
Darren R. Veach ◽  
Heiko van der Kuip ◽  
Walter E. Aulitzky ◽  
Jana Saenger ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Screening Method ◽  
Point Mutations ◽  
Kinase Domain ◽  
Screening Strategy ◽  
Culture Conditions ◽  
Cross Resistance ◽  
Imatinib Resistance ◽  
Abl Kinase ◽  
A Cell

Abstract The increasing impact of targeted cancer treatment demands strategies to identify and evaluate resistance mechanisms toward kinase inhibitors prior to their therapeutic application. Point mutations within the Bcr-Abl kinase domain constitute the major mechanism of resistance toward imatinib mesylate in Philadelphia-positive (Ph+) leukemia. Using Bcr-Abl-transformed Ba/F3 cells, we established a cell-based screening strategy for the prediction of specific kinase mutations that cause resistance toward kinase inhibitors. With imatinib at clinically relevant concentrations, we generated 368 resistant Ba/F3 sublines that were derived from resistant colonies. Thirty-two different single point mutations within the kinase domain of Bcr-Abl were identified in twenty-five per cent (liquid culture conditions) and seventy-two per cent (solid culture conditions) of these lines at known and novel positions. Using imatinib, the pattern and relative frequency of mutations reflected matters observed in patients with imatinib resistance. We then applied this screen to the pyrido-pyrimidine PD166326 (PD16), an investigational Abl kinase inihibitor. Compared to imatinib, we observed a five to seven times lower frequency of resistant colonies with equipotent concentrations of PD16. In addition, PD16 produced a distinct pattern of Bcr-Abl mutations. P-loop, A-loop and the known imatinib contact site T315 were affected with both inhibitors, whereas C-helix and SH2 contact sites were affected in imatinib resistant colonies exclusively. In contrast to imatinib, where kinase domain mutations were still widely distributed over the kinase domain even at at 4μM, mutations observed with PD16 at a concentration of 100nM narrowed to the exchange at position T315 to iseulicine. We did not detect mutations outside the kinase domain. Some resistant sublines displayed increased Bcr-Abl activity. Mutations that were derived from the screen were cloned and examined for the extent of cross-resistance to both inhibitors. The majority of mutations were effectively suppressed by PD16 at 50–500nM. In contrast, only few mutations were inhibited by imatinib at 5–10μM. However, exchanges at position F317 mediated resistance toward PD16, but were inhibited by standard concentrations of imatinib. Since this cell-based system produced results that are clinically significant, it may be used to predict resistance mutations in Bcr-Abl and other oncogenic kinases like cKit, EGFR, FIP1L1-PDGFRalpha or FLT3 towards clinically applicated and investigational drugs. Thus, this robust and simple screening strategy provides a rational basis for combinatorial and sequential treatment strategies.

Complete Suppression of in Vitro Resistance by AP24534, a Pan-BCR-ABL Inhibitor

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 726-726 ◽  
Author(s):  
Thomas O’Hare ◽  
Christopher A. Eide ◽  
Lauren T. Adrian ◽  
Victor M. Rivera ◽  
William C. Shakespeare ◽  
...  
Keyword(s):  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Complete Suppression ◽  
In Vitro Mutagenesis ◽  
Mutagenesis Screen ◽  
Abl Kinase ◽  
Imatinib Resistant ◽  
Escape Mutants ◽  
Selection Of

Abstract The BCR-ABL inhibitor imatinib is front-line therapy for chronic myeloid leukemia (CML). The second-line inhibitors dasatinib and nilotinib provide treatment options for controlling imatinib-resistant CML associated with BCR-ABL kinase domain mutations. However, the T315I mutant of BCR-ABL is resistant to all 3 clinical inhibitors, and is a frequent cause of salvage therapy failure. AP24534 is an oral, multi-targeted kinase inhibitor with activity against native and kinase domain-mutant BCR-ABL, including T315I. We have previously utilized an in vitro mutagenesis-screening assay to successfully predict the profile of mutations that confer resistance to imatinib, dasatinib, and nilotinib in patients. Here we use the in vitro mutagenesis screen to test whether BCR-ABL mutants can emerge in the presence of AP24534. Methods: To determine a resistance profile for AP24534, Ba/F3 cells expressing native BCR-ABL were mutagenized with ENU, washed, and plated in the presence of graded concentrations of AP24534 (5–80 nM). For each condition, 4.8×107 mutagenized cells were distributed into 480 wells and observed for growth for 4 weeks. Resistant clones were expanded in the continued presence of AP24534 and sequenced for mutations in the BCR-ABL kinase domain. Results: We first established IC50 values for inhibition of proliferation of Ba/F3 cells expressing native BCR-ABL (IC50: 0.5 nM) and an extensive panel of imatinib-resistant BCR-ABL mutants (IC50 range: 0.5 nM to 35.7 nM) including T315I (IC50: 11.4 nM) and E255V (IC50: 35.7 nM). Parental Ba/F3 cells were not inhibited up to a concentration of 1713 nM AP24534. Corresponding immunoblot analyses confirmed the same rank order for effective inhibition of CrkL phosphorylation in cells expressing native BCR-ABL, the T315I mutant, or the E255V mutant. Inhibition of CrkL phosphorylation was also demonstrated with primary hematopoetic cells from CML patients harboring native BCR-ABL or the T315I mutant. In the mutagenesis screen starting with Ba/F3 cells expressing native BCR-ABL, resistant clones recovered in 10 nM AP24534 expressed native BCR-ABL or one of several imatinib-resistant BCR-ABL mutants (168/1440 wells in 3 independent experiments). By contrast, when the screen was conducted in the presence of 20 nM AP24534, the frequency of outgrowth of escape mutants was extremely low and limited to cells expressing the T315I mutant (2/1440 wells) or the E255V mutant (1/1440 wells). Remarkably, outgrowth was completely suppressed by 40 nM AP24534. Conclusions: AP24534 is a potent inhibitor of native BCR-ABL and all tested BCR-ABL mutants, including T315I. Mutagenesis screening reveals that single-agent AP24534 (40 nM) completely suppressed outgrowth of escape mutants. This is in marked contrast to any of the BCR-ABL inhibitors previously profiled in this assay, where outgrowth was evident at the highest tested drug concentrations and complete suppression was observed only when dasatinib or nilotinib was combined with an investigational T315I inhibitor (PNAS2008; 105: 5507). As sequential BCR-ABL kinase inhibitor therapy has been linked to selection of rare subclones in which 2 mutations occur in the same BCR-ABL molecule, compound mutations are potentially capable of thwarting any of the current clinical BCR-ABL inhibitors, even in combination. Front-line therapy with a pan-BCR-ABL inhibitor could improve the depth and durability of responses by preventing selection of drug-resistant kinase domain point mutants. Our pre-clinical profiling indicates that AP24534 is an important new option in controlling resistance in CML. A phase 1 clinical trial designed to evaluate AP24534 treatment in patients with refractory CML and other hematologic malignancies has recently commenced.

BCR-ABL P-Loop Deletion Detected in Imatinib-Resistant CML Patients Corresponds to Splice Variant Associated with L248V Point Mutation, Retains BCR-ABL Kinase Activity, and Responds to Dasatinib Treatment.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2168-2168
Author(s):  
Nikolas von Bubnoff ◽  
Philipp Erben ◽  
Martin Müller ◽  
Tanja Lahaye ◽  
Susanne Schnittger ◽  
...  
Keyword(s):  
Splice Variant ◽  
Kinase Activity ◽  
Kinase Inhibitors ◽  
Point Mutations ◽  
Kinase Domain ◽  
Imatinib Resistance ◽  
Abl Kinase ◽  
Imatinib Resistant ◽  
Exon 4

Abstract Clonal selection of cells harboring point mutations of the BCR-ABL kinase domain are considered a major cause of resistance to imatinib. More than 40 different point mutations have been described that cause a variable degree of imatinib resistance, and display a differential response to alternative kinase inhibitors, like dasatinib or nilotinib. Here, we describe three cases (2 m, 1 f) with imatinib resistant chronic myelogenous leukemia (CML) associated with a specific deletion of 81 bp of ABL exon 4. Patients were diagnosed with chronic phase (CP) CML at the age of 52, 54, and 68 years. After initial interferon alpha based therapies for 32, 60, and 71 mo, imatinib therapy was initiated at dosages between 400–800 mg per day. After 18, 24, and 29 mo patients lost hematologic response in CP CML (n=2) or progressed to lymphoid blast crisis (BC, n=1). Molecular analysis of the ABL kinase domain revealed a deletion of a 81 bp fragment associated with a loss of amino acids 248–274 in all cases. In one patient, an additional M351T mutation was found. In the two cases with CP CML, dasatinib was commenced for imatinib resistance, resulting in a partial hematologic and minor cytogenetic response (60 and 70% Ph+ metaphases, respectively) after 14 mo of therapy. The patient with lymphoid BC was treated with vincristine and prednisone and died 24 mo after appearance of imatinib resistance. In two cases, sequencing of genomic DNA revealed an underlying CTG/GTG mutation associated with a L248V amino acid switch. The point mutation activated a cryptic splice site within ABL exon 4 leading to an in-frame splice variant characterized by the loss of a 81 bp 3′ portion of exon 4. We sought to evaluate the BCR-ABL kinase activity of the splice variant and the response to tyrosine kinase inhibitors in vitro. The 81 bp deletion of p210 BCR-ABL was cloned using cDNA from one of the patients. Using this construct, retrovirally transduced Ba/F3 cells were transformed upon growth factor withdrawal. These cells expressed BCR-ABL at the transcript and protein levels. Presence of the 81 bp deletion was confirmed by sequencing. Despite the presence of the corresponding 27 amino acid P-loop deletion (Δ248–274), Western blot indicated strong autophosphorylation of BCR-ABL, which decreased in the presence of imatinib to non-detectable levels at concentrations of 1.25μM and above. In the presence of imatinib/nilotinib/dasatinib, the growth of BCR-ABL expressing Ba/F3 cells was shifted from an IC50 of 125/30/0.5nM for wild-type BCR-ABL to 470/185/1.9nM for Δ248–274 cells. Thus, in vitro data demonstrate that deletion of almost the entire P-loop does not abrogate BCR-ABL kinase activity and results in only marginal resistance towards ABL kinase inhibitors. We conclude that deletions of BCR-ABL may be the result of alternative splicing generated by point mutations associated with resistance to imatinib. The Δ248–274 splice variant retains BCR-ABL kinase activity and sensitivity to imatinib, nilotinib, and dasatinib.

Sequential Kinase Inhibitor Therapy in CML Patients Can Select for Cells Harboring Compound BCR-ABL Kinase Domain Mutations with Increased Oncogenic Potency: Rationale for Early Combination Therapy of ABL Kinase Inhibitors.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 751-751 ◽  
Author(s):  
Neil P. Shah ◽  
Brian Skaggs ◽  
Susan Branford ◽  
Timothy P. Hughes ◽  
John M. Nicoll ◽  
...  
Keyword(s):  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Inhibitor Therapy ◽  
Significant Finding ◽  
Abl Kinase ◽  
Imatinib Resistant ◽  
Clinical Resistance ◽  
T315i Mutation ◽  
Kinase Domain Mutations

Abstract A critical question in the targeted therapy era relates to whether treatment outcomes will be optimized by sequential or combinatorial use of targeted agents. Selection for CML cells with BCR-ABL kinase domain mutations is the main mechanism responsible for loss of response to imatinib. Dasatinib is an ABL tyrosine kinase inhibitor that has activity against nearly all imatinib-resistant mutations and is approved for the treatment of imatinib-resistant and -intolerant BCR-ABL-associated leukemias. Acquired clinical resistance to sequential use of dasatinib following imatinib failure has been observed. We analyzed the BCR-ABL kinase domain at the time of relapse in 15 patients who lost an initial response to dasatinib, and found evolution of a total of three new mutations at the time of relapse in all cases. The highly resistant BCR-ABL/T315I mutation was detected in 11 cases. The four remaining cases were associated with the evolution of novel mutations (V299L, 3 cases; T315A, 1 case). V299L was also detected in a fourth case that had also evolved T315I. These three dasatinib-resistant mutations were part of a small number of amino acid substitutions previously isolated in a preclinical mutagenesis screen for dasatinib resistance-conferring BCR-ABL mutations. While the T315I mutation is highly resistant to imatinib, V299L and T315A retain sensitivity to imatinib in vitro and have not been previously described in imatinib-resistant cases, raising the potential utility of imatinib rechallenge in select dasaitinib-resistant cases. A significant finding of our studies is the evolution of five unique “compound” mutations (i.e. greater than one mutation on a DNA strand) in the BCR-ABL kinase domain of patients treated sequentially with imatinib and dasatinib. It is noteworthy that although the imatinib-sensitive V299L and T315A mutations evolved in five cases, they were detected in the context of a pre-existing imatinib-resistant mutation in three of these cases, and these cases are therefore unlikely to respond to rechallenge with IM. The T315A mutation was detected in the context of 2 pre-existing IM-resistant mutations (M244V/L364I). Interestingly, in bone marrow transformation assays, the clinically-identified dasatinib-resistant M244V/L364I/T315A mutation was more potently oncogenic than non-mutated BCR-ABL, in contrast to the baseline imatinib resistant M244V/L364I, which like T315A in isolation, was less potent than native BCR-ABL Our studies of CML cases resistant to sequential kinase inhibitor therapy reinforce BCR-ABL kinase domain mutation as the predominant mechanism of resistance to kinase inhibitor therapy, and provide evidence that compound mutations acquired as a result of sequential therapy can not only limit further therapeutic options, but also create more biologically aggressive isoforms of BCR-ABL. Together, these findings provide a strong rationale for early treatment of CML with combinations of kinase inhibitors that have the capacity to collectively prevent selection of resistant kinase domain mutations.

Loss of Response to Imatinib: Mechanisms and Management

Hematology ◽  
2005 ◽  
Vol 2005 (1) ◽  
pp. 183-187 ◽  
Author(s):  
Neil P. Shah
Keyword(s):  
Kinase Inhibitor ◽  
Kinase Domain ◽  
Imatinib Resistance ◽  
Early Clinical Trial ◽  
Abl Kinase ◽  
Imatinib Resistant ◽  
Trial Experience ◽  
Highly Effective ◽  
Mutant Forms

AbstractThe treatment of chronic myeloid leukemia (CML) has been revolutionized by the small molecule BCR-ABL-selective kinase inhibitor imatinib. Although imatinib is highly effective initially and generally well-tolerated, relapse is increasingly encountered clinically. Until recently, for the majority of CML patients with disease no longer responsive to imatinib, as well as for patients with imatinib intolerance, few effective therapeutic options existed. Our understanding of the major mechanisms of imatinib resistance has led to the clinical development of two novel BCR-ABL inhibitors that harbor significant therapeutic promise in early clinical trial experience. These agents, dasatinib (BMS-354825) and AMN107, are more potent inhibitors of BCR-ABL than imatinib, and moreover, harbor activity against nearly all imatinib-resistant BCR-ABL kinase domain mutant forms tested in vitro. Notably, neither of these compounds is effective against the imatinib-resistant BCR-ABL/T315I mutation. The potential availability of highly effective medications for the treatment of imatinib-resistant and intolerant cases of CML is expected to further complicate the timing of other effective therapies, such as allogeneic stem cell transplantation. Additionally, periodic genotyping of the BCR-ABL kinase domain to screen for drug-resistant mutations may play an increasingly important role in the future management of CML cases.

Bcr-Abl kinase domain mutations, drug resistance, and the road to a cure for chronic myeloid leukemia

Blood ◽  
2007 ◽  
Vol 110 (7) ◽  
pp. 2242-2249 ◽  
Author(s):  
Thomas O'Hare ◽  
Christopher A. Eide ◽  
Michael W. N. Deininger
Keyword(s):  
Chronic Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Kinase Inhibitor ◽  
Therapeutic Potential ◽  
Kinase Domain ◽  
Imatinib Resistance ◽  
The Road ◽  
Abl Kinase ◽  
Imatinib Resistant ◽  
Kinase Domain Mutations

Mutations in the kinase domain (KD) of BCR-ABL are the most prevalent mechanism of acquired imatinib resistance in patients with chronic myeloid leukemia (CML). Here we examine predisposing factors underlying acquisition of KD mutations, evidence for acquisition of mutations before and during therapy, and whether the detection of a KD mutation universally implies resistance. We also provide a perspective on how the second-line Abl inhibitors dasatinib and nilotinib are faring in the treatment of imatinib-resistant CML, especially in relation to specific KD mutations. We discuss the growing importance of the multi-inhibitor–resistant 315T>I mutant and the therapeutic potential that a 315T>I inhibitor would have. Last, we assess the potential of Abl kinase inhibitor combinations to induce stable responses even in advanced CML and interpret the emerging data in the context of CML pathogenesis.

Several Bcr-Abl kinase domain mutants associated with imatinib mesylate resistance remain sensitive to imatinib

Blood ◽  
2003 ◽  
Vol 101 (11) ◽  
pp. 4611-4614 ◽  
Author(s):  
Amie S. Corbin ◽  
Paul La Rosée ◽  
Eric P. Stoffregen ◽  
Brian J. Druker ◽  
Michael W. Deininger
Keyword(s):  
Imatinib Mesylate ◽  
Kinase Inhibitor ◽  
Blast Crisis ◽  
Chronic Phase ◽  
Kinase Domain ◽  
Myelogenous Leukemia ◽  
Imatinib Resistance ◽  
Abl Kinase ◽  
Cellular Assays ◽  
Kinase Domain Mutations

Abstract Imatinib mesylate is a selective Bcr-Abl kinase inhibitor, effective in the treatment of chronic myelogenous leukemia. Most patients in chronic phase maintain durable responses; however, many in blast crisis fail to respond, or relapse quickly. Kinase domain mutations are the most commonly identified mechanism associated with relapse. Many of these mutations decrease the sensitivity of the Abl kinase to imatinib, thus accounting for resistance to imatinib. The role of other mutations in the emergence of resistance has not been established. Using biochemical and cellular assays, we analyzed the sensitivity of several mutants (Met244Val, Phe311Leu, Phe317Leu, Glu355Gly, Phe359Val, Val379Ile, Leu387Met, and His396Pro/Arg) to imatinib mesylate to better understand their role in mediating resistance.While some Abl mutations lead to imatinib resistance, many others are significantly, and some fully, inhibited. This study highlights the need for biochemical and biologic characterization, before a resistant phenotype can be ascribed to a mutant.

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