Activity of ABL Kinase Inhibitors in Two Distinct Models of Imatinib Resistance.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 4819-4819
Author(s):  
Nicholas J. Donato ◽  
Ji Wu ◽  
William Bornmann ◽  
Moshe Talpaz
Keyword(s):  
Cell Lines ◽  
Inhibitory Activity ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
K562 Cells ◽  
Mutant Form ◽  
Imatinib Resistance ◽  
Resistant Variant ◽  
Lyn Kinase ◽  
Kinase Inhibitory Activity

Abstract The tyrosine kinase inhibitor imatinib mesylate (Gleevec) is effective in controlling BCR-ABL expressing leukemias but resistance occurs in a small subset of early stage patients and is very common in advanced stage patients. Resistance is associated with overexpression and/or mutations in the BCR-ABL gene but patients are also increasingly reported to fail imatinib therapy while retaining wild-type BCR-ABL expression. To circumvent or overcome resistance novel kinase inhibitors have been synthesized and tested clinically. However, while investigators have designed models to measure and predict activity of novel compounds in mutation-mediated imatinib resistance, other mechanisms of resistance have not been modeled or shown clinical relevance. In this report, the activity of four novel kinase inhibitors (norlotinib, dasatinib, SKI-606, ON012380) was compared in two distinct models of imatinib resistance. These models include cell lines established from natural resistant variant clones selected from imatinib sensitive cell lines and associated with a T315I BCR-ABL mutation (BV-173R) or overexpressed Lyn kinase (K562R). Kinase inhibitory activity in these models was compared to transfectant-based resistance models including BaF3 cells expressing the T315I mutant form of BCR-ABL and overexpression of Lyn kinase in K562 cells. K562R cells were completely resistant to imatinib and norlotinib but highly sensitive to Src/Abl inhibitors (dasatinib, SKI-606) but only partially sensitive to ON012380. Overexpression of Lyn in K562 cells reduced imatinib and norlotinib sensitivity (3-fold) but did not affect sensitivity to the other kinase inhibitors. BV-173R cells expressing the T315I mutant form of BCR-ABL were completely resistant to Abl-selective kinase inhibitors (imatinib, norlotinib) and ~100-fold less sensitive (IC50 ~ 2 microM) to Src/Abl-directed inhibitors (dasatinib, SKI-606) while the non-ATP competitive kinase inhibitor, ON012380, was equally effective against both BV-173 and BV-173R cells. Expression of the T315I mutant form of BCR-ABL in BaF3 cells completely blocked kinase inhibitory activity of all inhibitors except ON012380. IL-3 dependent BaF3 cells were not inhibited by imatinib, norlotinib or SKI-606 but were equally sensitive to ON012380 when compared to IL-3 independent BCR-ABL transfectants. Cellular sensitivity was associated with reduced phosphorylation of BCR-ABL, CrkL and Lyn kinase with all inhibitors except ON012380, which mediated apoptosis in the absence of alterations in tyrosine phosphorylation. Together, our results suggest that imatinib resistant cell models are useful in evaluating the activity of novel kinase inhibitors but need to be carefully interpreted and mechanistically tested. Additional mediators of imatinib resistance need to be defined and modeled so that an appropriate individualized therapy can be applied to most effectively overcome resistant disease.

Dasatinib (BMS-354825) Overcomes Multiple Mechanisms of Imatinib Resistance in Chronic Myeloid Leukemia (CML).

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1994-1994 ◽  
Author(s):  
Francis Y. Lee ◽  
Mei-Li Wen ◽  
Rajeev Bhide ◽  
Amy Camuso ◽  
Stephen Castenada ◽  
...  
Keyword(s):  
Cell Lines ◽  
Kinase Inhibitor ◽  
Leukemic Cell ◽  
K562 Cells ◽  
Src Family Kinases ◽  
Imatinib Resistance ◽  
Over Expression ◽  
Imatinib Resistant

Abstract Resistance to imatinib is a growing concern in CML, particularly in advanced disease. The most common cause of resistance is mutations in BCR-ABL, but other mechanisms have also been identified, including over-expression of BCR-ABL, activation of SRC family kinases and the P-glycoprotein (PGP) efflux pump (via MDR1 over-expression). Dasatinib (BMS-354825) is a novel, oral, multi-targeted tyrosine kinase inhibitor that targets BCR-ABL and SRC kinases. Dasatinib has 325-fold greater potency versus imatinib in cell lines transduced with wild-type BCR-ABL and is active against 18 out of 19 BCR-ABL mutations tested that confer imatinib resistance (Shah et al, Science305:399, 2004; O’Hare et al, Cancer Res65:4500–5, 2005), and preliminary results from a Phase I study show that it is well tolerated and has significant activity in imatinib-resistant patients in all phases of CML (Sawyers et al, J Clin Oncol23:565s, 2005; Talpaz et al, J Clin Oncol23:564s, 2005). We assessed the ability of dasatinib to overcome a variety of mechanisms of imatinib resistance. First, the leukemic-cell killing activity of dasatinib was tested in vitro in three human imatinib-resistant CML cell lines (K562/IM, MEG-01/IM and SUP-B15/IM). Based on IC50 values, dasatinib had >1000-fold more potent leukemic-cell killing activity compared with imatinib versus all three cell lines. Furthermore, in mice bearing K562/IM xenografts, dasatinib was curative at doses >5 mg/kg, while imatinib had little or no impact at doses as high as 150 mg/kg, its maximum tolerated dose. We determined that the MEG-01/IM and SUP-B15/IM cell lines carried BCR-ABL mutations known to confer imatinib resistance to imatinib clinically (Q252H and F359V, respectively). In K562/IM cells, BCR-ABL mutations or BCR-ABL over-expression were not detected, but the SRC family member FYN was over-expressed. PP2, a known inhibitor of SRC family kinases but not BCR-ABL, could reverse the imatinib resistance in these cells. Together, these data suggest that activation of FYN may be a cause of imatinib resistance in K562/IM. Based on cell proliferation IC50, we found that the anti-leukemic activity of dasatinib in K562/IM cells was 29-fold more potent compared with AMN107 (a tyrosine kinase inhibitor that inhibits BCR-ABL but not SRC family kinases). Given that the human serum protein binding of dasatinib, imatinib and AMN107 were 93, 92 and >99% respectively, the difference in potency between dasatinib and AMN107 in vivo may be far greater than the simple fold-difference in the in vitro IC50 values. Finally, in K562 cells over-expressing PGP (K562/ADM), we found that dasatinib was only 6-fold less active than in parental K562 cells. Because of the extreme potency of dasatinib in K562 cells, this reduced potency still afforded an IC50 of 3 nM, which is readily achievable in vivo. Indeed, in mice bearing K562/ADM xenografts, dasatinib was curative at 30 mg/kg, with significant anti-leukemic activity at 15 mg/kg. In conclusion, the rational design of dasatinib as a multi-targeted kinase inhibitor allows this agent to overcome a variety of mechanisms of resistance to imatinib in CML, including mechanisms that are not overcome by agents with a narrower spectrum of inhibition, such as AMN107. Dasatinib is currently in Phase II evaluation in imatinib-resistant/-intolerant patients in the ‘START’ program, and in Phase I evaluation in solid tumors.

A New Abl Kinase Inhibitor (AMN107) Has In Vitro Activity on Chronic Myeloid Leukaemia (CML) Ph+ Cells Resistant to Imatinib.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2004-2004 ◽  
Author(s):  
Giovanni Martinelli ◽  
Alberto M. Martelli ◽  
Tiziana Grafone ◽  
Irina Mantovani ◽  
Alessandra Cappellini ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Lines ◽  
Cell Cycle Progression ◽  
Kinase Inhibitor ◽  
K562 Cells ◽  
Sh2 Domain ◽  
Regulatory Subunit ◽  
Imatinib Resistance ◽  
First Line Therapy

Abstract Imatinib mesylate (Novartis Pharma), an inhibitor of the bcr/abl tyrosine kinase, has rapidly become the first-line therapy for CML. Imatinib has proved remarkably effective at reducing the number of leukaemia cells in individual CML patients and promises to prolong life substantially in comparison with earlier treatments. However, in patients in advanced phases of the disease, the development of resistance to this drug is a frequent setback. Therefore, new inhibitors of bcr/abl are needed. Very recently, a new bcr/abl inhibitor, AMN107 (Novartis Pharma), has been developed. We have tested AMN107 on human leukaemia cell lines and on blasts isolated from imatinib-resistant CML patients. After a 24 h incubation, AMN107 (10 nM) blocked K562 cells in the G1 phase of the cell cycle. To obtain the same effect with imatinib, a 200 nM concentration was required. AMN107 had no affect on cell cycle progression of bcr/abl-negative cell lines such as HL60 and NB4, even if the concentration was raised to 500 nM. After 48 h incubation, AMN107 (10 nM) was capable of inducing a massive apoptosis of K562 cells whereas, once again, 200 nM imatinib was required to obtain the same effect. Western blot analysis with phosphospecific antibodies revealed that in K562 cells AMN107 (50 nM) markedly down-regulated autophosphorylation of bcr/abl Tyr177 and Tyr412, whereas autophosphorylation of Thr735 was unaffected. In contrast, imatinib even if used at 200 nM, did not diminish phosphorylation of either bcr/abl Tyr177 or Tyr412. This finding seems particularly important because recent evidence has demonstrated that the signalling pathway emanating from Tyr177 plays a major role in the pathogenesis of CML. Indeed, phosphorylated Tyr177 forms a high-affinity binding site for the SH2 domain of the adapter Grb2. The main effectors of Grb2 are Sos and Ras, however Grb2 also recruits the scaffolding adapter protein Gab2 to bcr/abl via a Grb2-Gab2 complex, which results in activation of phosphoinositide 3-kinase (PI3K)/Akt and Erk signalling networks. Consistently, we found by immunoprecipitation decreased levels of bcr/abl-associated Gab2, Grab2, and p85 regulatory subunit of PI3K in AMN107-treated cells. AMN107 treatment of K562 cells also caused a reduction of STAT5, cCBL, CRKL, and Akt phosphorylation levels, as well as Bcl-XL expression. AMN107 (5 μM for 24h) significantly increased the apoptosis rate of CML blasts isolated from patients resistant to imatinib. Therefore, AMN107 might represent a new bcr/abl selective inhibitor useful for overcoming imatinib resistance.

Lyn Kinase Alters Gab2 Phosphorylation and c-Cbl Protein Levels To Regulate Imatinib Sensitivity and Survival of Chronic Myelogenous Leukemia Cells.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2132-2132
Author(s):  
Ji Wu ◽  
Feng Meng ◽  
Moshe Talpaz ◽  
Nicholas J. Donato
Keyword(s):  
Tyrosine Phosphorylation ◽  
Kinase Inhibitor ◽  
K562 Cells ◽  
Imatinib Resistance ◽  
Kinase Inhibition ◽  
Protein Levels ◽  
Abl Kinase ◽  
Lyn Kinase ◽  
Imatinib Resistant ◽  
Cbl Protein

Abstract The tyrosine kinase inhibitor imatinib mesylate (Gleevec) is effective in controlling BCR-ABL expressing leukemias but resistance occurs in some early phase patients while it is more common in advanced disease. Resistance has been generally associated with mutations in the BCR-ABL kinase that effect drug affinity. However patients are also increasingly reported to fail imatinib therapy while retaining wild-type BCR-ABL expression. Our previous studies suggested a role for Lyn, a Src-related kinase, in imatinib resistance. K562 cells selected for imatinib resistance (K562R) overexpress Lyn kinase and its targeted silencing overcomes imatinib resistance and engages apoptosis. Overexpression of Lyn in K562 cells reduces imatinib sensitivity (3-fold) and patients that fail imatinib therapy in the absence of BCR-ABL mutations express a highly activated Lyn kinase that is not suppressed by imatinib. Silencing Lyn expression in patient specimens induces changes in cell survival that are proportional to the level of Lyn protein reduction. To understand the role of Lyn kinase in imatinib resistance and apoptosis we examined proteins associated with this kinase in imatinib resistant cell lines, leukemic cells overexpressing Lyn and specimens derived from imatinib resistant patients. Lyn overexpression blocked complete suppression of BCR-ABL tyrosine phosphorylation by imatinib and affected BCR-ABL signaling adaptors. Although BCR-ABL forms a stable complex with the leukemogenic-critical adaptor protein Gab2 in imatinib sensitive cells, Lyn overexpression resulted in the formation of Lyn:Gab2 complexed in resistant cells. BCR-ABL kinase inhibition failed to reduce tyrosine phosphorylation of Gab2 in these cells while Lyn silencing or kinase inhibition (with dasatinib) completely suppressed Gab2 tyrosine phosphorylation and correlated with the induction of apoptosis. Lyn silencing in K562R cells also lead to a reciprocal increase in the tyrosine phosphorylation and association with a protein of ~120kDa, identified as the E3 ligase, c-Cbl. Lyn overexpression in K562 cells reduced their imatinib sensitivity and reduced c-Cbl protein levels. Kinase inhibitor and co-transfection studies demonstrated that tyrosine phosphorylation of c-Cbl at a critical signaling site (Y774) is primarily controlled by BCR-ABL and deletion or mutation of the c-Cbl RING domain altered its BCR-ABL phosphorylation. These results suggest that c-Cbl complexes are regulated at both the protein and phosphorylation level by Lyn and BCR-ABL kinase activities, respectively. Overexpression and/or activation of Lyn may disrupt the balance and regulation of critical regulators of leukemogenic signaling (Gab2) or protein trafficking and stability (c-Cbl), resulting in increased cell survival and reduced responsiveness to BCR-ABL kinase inhibition. We conclude that Lyn alters the level and function of critical signaling adaptor proteins in CML cells.

A New Abl Kinase Inhibitor (AMN107) Has In Vitro Activity on CML Ph+Blast Cells Resistant to Imatinib.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4687-4687 ◽  
Author(s):  
Giovanni Martinelli ◽  
Alberto M. Martelli ◽  
Tiziana Grafone ◽  
Gianantonio Rosti ◽  
Irina Mantovani ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Lines ◽  
Cell Cycle Progression ◽  
Kinase Inhibitor ◽  
Leukemic Cell ◽  
K562 Cells ◽  
Sh2 Domain ◽  
Adapter Protein ◽  
Imatinib Resistance

Abstract Imatinib mesylate (STI571), an inhibitor of the bcr/abl tyrosine kinase, is rapidly becoming the first-line therapy for chronic myeloid leukemia (CML). Imatinib has proved remarkably effective at reducing the number of leukemia cells in individual CML patients and promises to prolong life substantially in comparison with earlier treatments. However, the development of resistance to this drug is a frequent setback, particularly in patients in advanced phases of the disease. Therefore, new inhibitors of bcr/abl are needed. Very recently, a new bcr/abl inhibitor, AMN107, has been synthesized. We have tested AMN107 on leukemic cell lines and on blasts isolated from imatinib-resistant CML patients. Western blot analysis with phosphospecific antibodies revealed that in K562 cells AMN107 (10 nM) down-regulated phosphorylation of bcr/abl Tyr177, while the phosphorylation levels of Tyr412 were unaffected. This finding seems particularly important because recent evidence has demonstrated that the signaling pathway emanating from Tyr177 plays a major role in the pathogenesis of CML. Indeed, phosphorylated Tyr 177 forms a high-affinity binding site for the SH2 domain of the adapter protein Grb2. The main effectors of Brb2 are Sos and Ras, however Grb2 also recruits the scaffolding adapter protein Gab2 to bcr/abl via a Grb2,Gab2 complex. Which results in activation of the PI3K/Akt and Erk signalling networks. In contrast, STI571, even if used at 200 nM, did not diminish phosphorylation of bcr/abl Tyr177. At 10 nM AMN107 blocked K562 cells in the G1 phase of the cell cycle. To obtain the same effect with imatinib, a 200 nM concentration was required. AMN107 did not affect cell cycle progression of bcr/abl-negative cell lines such as HL60 and NB4, even if the concentration was raised to 200 nM. AMN107 (5 μM for 24 h) significantly increased apoptosis rate in CML blasts isolated from patients resistant to the same concentration of imatinib. Therefore, AMN107 might represent a new bcr/abl selective inhibitor useful for overcoming imatinib resistance.

Imatinib-Resistant BCR-ABL Mutant Cellular Screening Paradigm To Characterize Promising Small-Molecule Inhibitors: T315I-BCR-ABL Targeted Drug Discovery.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 2181-2181
Author(s):  
Mohammad Azam ◽  
William C. Shakespeare ◽  
Chester Metcalf ◽  
Yihan Wang ◽  
Raji Sunderamoorthi ◽  
...  
Keyword(s):  
Drug Discovery ◽  
Structural Analysis ◽  
Cell Lines ◽  
Small Molecule ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Drug Binding ◽  
Imatinib Resistance ◽  
Inhibition Of Proliferation ◽  
Imatinib Resistant

Abstract In patients with chronic myeloid leukemia (CML), kinase domain mutations account for imatinib resistance in the majority of cases. Mutations cause either a direct steric hindrance to drug binding or a conformational change that favors kinase activation, which therefore precludes imatinib binding. We have previously characterized the dual Src-Abl kinase inhibitor AP23464 and found it to effectively suppress the growth of cells expressing native and essentially all imatinib-resistant variants of BCR-ABL, with the notable exception of the gatekeeper T315I mutant (Azam et al., Proc. Natl. Acad. Sci. USA, 103: 9244, 2006). Following this work, we have used mutant panel screening and integrated structural analysis to further characterize key analogs designed to overcome T315I resistance, as exemplified by AP23846 and AP24163. Both molecules effectively inhibit the tyrosine kinase activity of wild type (WT) and T315I variants of BCR-ABL, and inhibit the proliferation of BaF3-derived cell lines expressing these enzymes (see Table below). AP24163 was further characterized against a broader panel of imatinib-resistant BCR-ABL-expressing cell lines and showed a promising profile of proliferation inhibition. Comparison of these data with structural models of the mutants provides insights into the basis for the ability of AP24163 to overcome imatinib resistance. Refinement of small-molecule kinase inhibitors by the integration of sequential screening of panels of mutants coupled with structural analysis is a powerful drug discovery paradigm that is applicable to an increasing number of targeted therapeutic agents. INHIBITION OF PROLIFERATION OF BAF3 CELLS EXPRESSING BCR-ABL AND ITS VARIANTS (IC50 in nM) IMATINIB AP23464 AP23846 AP24163 WT 600 14 500 7 T315I >20000 >1000 500 480 L248R >20000 92 ND 64 G250E 5000 25 ND 63 Q252H 3000 40 ND 42 Y253H 18000 32 ND 44 E255K 12000 74 ND 24 BAF3+IL3 >20000 >1000 500 >10000 Figure Figure

scholarly journals Identification of Hsp90 inhibitors as potential drugs for the treatment of TSC1/TSC2 deficient cancer

2021 ◽  
Author(s):  
Evelyn M. Mrozek ◽  
Vineeta Bajaj ◽  
Yanan Guo ◽  
Izabela Malinowska ◽  
Jianming Zhang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Xenograft Model ◽  
Growth Delay ◽  
Hsp90 Inhibitors ◽  
Null Cell ◽  
Standard Dosage

Inactivating mutations in either TSC1 or TSC2 cause Tuberous Sclerosis Complex, an autosomal dominant disorder, characterized by multi-system tumor and hamartoma development. Mutation and loss of function of TSC1 and/or TSC2 also occur in a variety of sporadic cancers, and rapamycin and related drugs show highly variable treatment benefit in patients with such cancers. The TSC1 and TSC2 proteins function in a complex that inhibits mTORC1, a key regulator of cell growth, which acts to enhance anabolic biosynthetic pathways. In this study, we identified and validated five cancer cell lines with TSC1 or TSC2 mutations and performed a kinase inhibitor drug screen with 197 compounds. The five cell lines were sensitive to several mTOR inhibitors, and cell cycle kinase and HSP90 kinase inhibitors. The IC50 for Torin1 and INK128, both mTOR kinase inhibitors, was significantly increased in three TSC2 null cell lines in which TSC2 expression was restored.  Rapamycin was significantly more effective than either INK128 or ganetespib (an HSP90 inhibitor) in reducing the growth of TSC2 null SNU-398 cells in a xenograft model. Combination ganetespib-rapamycin showed no significant enhancement of growth suppression over rapamycin. Hence, although HSP90 inhibitors show strong inhibition of TSC1/TSC2 null cell line growth in vitro, ganetespib showed little benefit at standard dosage in vivo. In contrast, rapamycin which showed very modest growth inhibition in vitro was the best agent for in vivo treatment, but did not cause tumor regression, only growth delay.

Acquired Extramedullary Resistance to Dasatinib Due to Selection of Philadelphia-Positive Lymphoblast Clone Harboring a T315I BCR-ABL Gene Mutation: Reversal by Dose Escalation and Hydroxyurea.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4579-4579
Author(s):  
Tuija Lundan ◽  
Franz Gruber ◽  
Martin Hoglund ◽  
Bengt Simonsson ◽  
Sakari Knuutila ◽  
...  
Keyword(s):  
Dose Escalation ◽  
Low Dose ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Mutant Cell ◽  
Skin Lesions ◽  
Fish Analysis ◽  
Imatinib Resistance

Abstract Most patients with advanced Philadelphia-positive (Ph+) hematologic malignancies develop resistance to imatinib. Acquired resistance to imatinib is commonly a result of selection for subclones bearing point-mutations in the catalytic kinase domain of BCR-ABL. Dasatinib (BMS-354825), a dual-specific SRC/ABL kinase inhibitor, has shown activity in imatinib-resistant Ph+ diseases both in vitro and in vivo. Preliminary data also indicate efficacy in patients. Based on laboratory evidence, dasatinib appears to inhibit all known BCR-ABL mutant clones, with the exception of T315I, a gatekeeper mutation conferring resistance to several kinase inhibitors. Here we describe a Ph+ ALL patient, who initially developed imatinib resistance (hematologic) possibly due to BCR-ABL amplification (FISH). His disease relapsed as extensive extramedullary tumors bearing wild-type BCR-ABL. He received dasatinib 70 mg BID as part of the BMS CA180–015 study and achieved a very good partial remission. After 5 months of therapy, the disease relapsed as a solitary axillary tumor and several small palmar skin lesions. He also had blasts in the CSF indicative of neuroleukemia. Bone marrow remained in cytogenetic remission. FISH analysis of the tumor revealed 2–3 copies of BCR-ABL as previously. A highly sensitive, quantitative, mutation-specific PCR (Gruber F, ASH 2004) showed the presence of the T315I mutation, which was confirmed by sequencing. A very low level of T315I transcript was also detected in the blood. Dasatinib dose was escalated to 100 mg BID, and low-dose hydroxyurea 500 mg BID was initiated to putatively enhance the access of dasatinib in the CSF sanctuary. He also received two doses of i.t. therapy (methotrexate, cytarabine). Patient’s symptoms (confusion, headache) related to neuroleukemia resolved rapidly, skin lesions disappeared and axillary tumor decreased in size. He is currently symptom-free and has no signs of active ALL. The favorable response to dasatinib dose escalation and low-dose hydroxyurea was unexpected. Preclinical data on T315I mutant cell lines would argue against a significant concentration dependence in kinase inhibition by dasatinib. Putatively, targets other than BCR-ABL may be of importance in particular in Ph+ ALL (e.g. Src, Lyn), and this effect may account for the response. Similar off-target activity of hydroxyurea is utilized in clinical trials to overcome resistance to multidrug HIV therapy - a setting resembling current treatment of Ph+ malignancies with kinase inhibitors.

Frequency of NUP214-ABL1 Oncogene in Patients with T-Cell Acute Lymphoblastic Leukemia (T-ALL) and Analysis of the Activity of Imatinib and Nilotinib in NUP214-ABL1-Expressing T-ALL Cell Lines.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 710-710
Author(s):  
Alfonso Quintas-Cardama ◽  
Weigang Tong ◽  
Taghi Manshouri ◽  
Jan Cools ◽  
D. Gary Gilliland ◽  
...  
Keyword(s):  
Acute Lymphoblastic Leukemia ◽  
T Cell ◽  
Cell Lines ◽  
Inhibitory Activity ◽  
Lymphoblastic Leukemia ◽  
Fusion Transcript ◽  
Lymphoblastic Lymphoma ◽  
Abl Kinase ◽  
Cell Acute Lymphoblastic Leukemia ◽  
Kinase Inhibitory Activity

Abstract The fusion of ABL1 with BCR results in the hybrid BCR-ABL1 oncogene that encodes the constitutively active Bcr-Abl tyrosine kinase encountered in the majority of patients with chronic myeloid leukemia (CML) and in approximately 30% of pts with B-cell acute lymphoblastic leukemia (B-ALL). Recently, the episomal amplification of ABL1 has been described in 6% of pts with T-ALL (Nat Genet2004;36:1084–9). Molecular analysis demonstrated the oncogenic fusion of ABL1 with the nuclear pore complex protein NUP214 (NUP214-ABL1). We screened 29 pts with T-cell lymphoblastic lymphoma (T-LBL) and T-ALL for the presence of the NUP214-ABL1 fusion transcript by RT-PCR using specific primers for the 5 different transcripts thus far described. Three (10%) pts were found to express this fusion transcript, including 2 with T lymphoblastic lymphoma (NUP214 exon 31) and 1 with T-ALL (NUP214 exon 29). This was confirmed by direct sequencing in all cases. All pts received therapy with hyperCVAD and achieved a complete remission (CR). However, 2 of them died 6 and 9 months into therapy, respectively. One other pt remains in CR (19+ months) by morphologic and flow cytometry criteria. However, NUP214-ABL1 is still detectable in peripheral blood by nested PCR, thus suggesting minimal residual disease (MRD). We then studied the activity of the tyrosine kinase inhibitors imatinib and nilotinib in the NUP214-ABL1-expressing cell lines PEER and BE-13. Although PEER and BE-13 cell viability was reduced with both agents, the IC50 was almost 10-fold higher for imatinib (643 nM) than for nilotinib (68 nM) (F test, p<0.001), which parallels the 10− to 30− fold higher Abl kinase inhibitory activity of nilotinib compared to imatinib in BCR-ABL-expressing cells. Nilotinib also potently inhibited the cell proliferation of BE-13 cells (IC50 131 nM). In contrast, Jurkat cells, a T-ALL cell line which does not carry NUP214-ABL1, were remarkably resistant to both imatinib and nilotinib with an IC50 values greater than 5 μM indicating that the cytotoxicity mediated by both TKIs is not related to a general toxic effect on T-ALL cell lines. The inhibition of cellular proliferation by imatinib and nilotinib was associated with a dose- and time-dependent induction of apoptosis in both PEER and BE-13 cells. In Western blotting, higher inhibition of phospho-Abl and phospho-CRKL (a surrogate of Bcr-Abl kinase status) was observed in PEER cells upon exposure to nilotinib as compared with imatinib at their respective IC50 concentrations for cell growth inhibition. We conclude that NUP214-ABL1 can be detected in 10% of pts with T-cell malignancies and its detection can be used as a sensitive marker of MRD. Imatinib and nilotinib potently inhibits the growth of NUP214-ABL1-expressing cells. Given the higher Abl kinase inhibitory activity of nilotinib with respect to imatinib, this agent must be further investigated in clinical studies targeting patients with T-ALL and T-LBL expressing the NUP214-ABL1 fusion kinase.

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.

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