Cancer Drug Imatinib Research Articles

Structural mechanism of a drug-binding process involving a large conformational change of the protein target

Proteins often undergo large conformational changes when binding small molecules, but atomic-level descriptions of such events have been elusive. Here, we report unguided molecular dynamics simulations of Abl kinase binding to the cancer drug imatinib. In the simulations, imatinib first selectively engages Abl kinase in its autoinhibitory conformation. Consistent with inferences drawn from previous experimental studies, imatinib then induces a large conformational change of the protein to reach a bound complex that closely resembles published crystal structures. Moreover, the simulations reveal a surprising local structural instability in the C-terminal lobe of Abl kinase during binding. The unstable region includes a number of residues that, when mutated, confer imatinib resistance by an unknown mechanism. Based on the simulations, NMR spectra, hydrogen-deuterium exchange measurements, and thermostability measurements and estimates, we suggest that these mutations confer imatinib resistance by exacerbating structural instability in the C-terminal lobe, rendering the imatinib-bound state energetically unfavorable.

High-resolution AP-SMALDI MSI as a tool for drug imaging in Schistosoma mansoni

Schistosoma mansoni is a parasitic flatworm causing schistosomiasis, an infectious disease affecting several hundred million people worldwide. Schistosomes live dioeciously, and upon pairing with the male, the female starts massive egg production, which causes pathology. Praziquantel (PZQ) is the only drug used, but it has an inherent risk of resistance development. Therefore, alternatives are needed. In the context of drug repurposing, the cancer drug imatinib was tested, showing high efficacy against S. mansoni in vitro. Besides the gonads, imatinib mainly affected the integrity of the intestine in males and females. In this study, we investigated the potential uptake and distribution of imatinib in adult schistosomes including its distribution kinetics. To this end, we applied for the first time atmospheric-pressure scanning microprobe matrix-assisted laser desorption/ionization mass spectrometry imaging (AP-SMALDI MSI) for drug imaging in paired S. mansoni. Our results indicate that imatinib was present in the esophagus and intestine of the male as early as 20 min after in vitro exposure, suggesting an oral uptake route. After one hour, the drug was also found inside the paired female. The detection of the main metabolite, N-desmethyl imatinib, indicated metabolization of the drug. Additionally, a marker signal for the female ovary was successfully applied to facilitate further conclusions regarding organ tropism of imatinib. Our results demonstrate that AP-SMALDI MSI is a useful method to study the uptake, tissue distribution, and metabolization of imatinib in S. mansoni. The results suggest using AP-SMALDI MSI also for investigating other antiparasitic compounds and their metabolites in schistosomes and other parasites.Graphical abstract

Multiwalled Carbon Nanotubes/Gold Nanoparticles Hybrid Electrodes for Enzyme-Free Electrochemical Glucose Sensor.

We followed a facile strategy to fabricate glucose sensors using mildly oxidized multiwalled carbon nanotubes (MWCNTs), gold nanoparticles (AuNPs) and thiol acids including mercaptoacetic acid (MAA), mercaptopropionic acid (MPA), and mercaptosuccinic acid (MSA). The thiol acids separately bonded to MWCNTs anchored AuNPs of average diameter 14 nm, and yielded three different nanohybrids; MWCNTs-MAA-AuNPs, MWCNTs-MPA-AuNPs and MWCNTs-MSA-AuNPs. The nanohybrids after coating onto glassy carbon (GC) electrode resulted into enzyme free glucose sensors (GC-MWCNTs-MAA-AuNPs, GC-MWCNTs-MPA-AuNPs and GC-MWCNTs-MSA-AuNPs). Their electrocatalytic glucose sensing ability was examined through cyclic voltammetry and amperometry. GC-MWCNTs-MSA-AuNPs electrode showed high stability and activity in the electrocatalytic oxidation of glucose compared to other two sensors. It also showed a wide range of response for glucose concentration from 0.12 to 4.0 μM, and low detection limit of 0.036 μM (S/N = 3). The optimum rate of applied potential was 0.8 V/s, which proves the effective sensing of glucose. The selective sensing of glucose in the presence of H₂O₂, uric acid and blood cancer drug (imatinib mesylate) was verified through amperometry. The electrode can be a new addition to glucose sensors and bioanalytical techniques.

Cytotoxic activity of fucoxanthin, alone and in combination with the cancer drugs imatinib and doxorubicin, in CML cell lines

In the present study, we evaluate the in vitro cytotoxicity of fucoxanthin (Fx) on two human leukemia cell lines, K562 and TK6, alone and in combination with the conventional anticancer drugs imatinib (Imat) and doxorubicin (Dox). For the purpose, we assessed the cytotoxic and proliferation effects by cell count, induction of DNA damage by comet assay, and cell death by nuclear condensation, annexin V staining, coupled with propidium iodide uptake, and protein expression of Bax, caspase-3, and Bcl-2 (western blot). Our results show that Imat increased cytotoxicity in TK6 cells and inhibited proliferation in K562 cells, while Dox decreased cell viability and proliferation in both cell lines. Fx per se increased cytotoxicity against K562 cells and decreased cell proliferation of K562 and TK6 cells. The effects were confirmed by phase contrast microscopy. However, the antiproliferative effects are not explained by induction of DNA damage or cell death. In co-incubation, Fx increased antiproliferative effects of both drugs in the cell lines tested, however no differences where observed relative to Fx alone. This study unveiled in vitro cytotoxicity of Fx by inhibition of cell proliferation in both cell lines. Further studies are needed to elucidate the signal transduction pathways and molecular targets involved in that effect.

Structural Mechanisms Determining Inhibition of the Collagen Receptor DDR1 by Selective and Multi-Targeted Type II Kinase Inhibitors

The discoidin domain receptors (DDRs), DDR1 and DDR2, form a unique subfamily of receptor tyrosine kinases that are activated by the binding of triple-helical collagen. Excessive signaling by DDR1 and DDR2 has been linked to the progression of various human diseases, including fibrosis, atherosclerosis and cancer. We report the inhibition of these unusual receptor tyrosine kinases by the multi-targeted cancer drugs imatinib and ponatinib, as well as the selective type II inhibitor DDR1-IN-1. Ponatinib is identified as the more potent molecule, which inhibits DDR1 and DDR2 with an IC50 of 9nM. Co-crystal structures of human DDR1 reveal a DFG-out conformation (DFG, Asp-Phe-Gly) of the kinase domain that is stabilized by an unusual salt bridge between the activation loop and αD helix. Differences to Abelson kinase (ABL) are observed in the DDR1 P-loop, where a β-hairpin replaces the cage-like structure of ABL. P-loop residues in DDR1 that confer drug resistance in ABL are therefore accommodated outside the ATP pocket. Whereas imatinib and ponatinib bind potently to both the DDR and ABL kinases, the hydrophobic interactions of the ABL P-loop appear poorly satisfied by DDR1-IN-1 suggesting a structural basis for its DDR1 selectivity. Such inhibitors may have applications in clinical indications of DDR1 and DDR2 overexpression or mutation, including lung cancer.

Probing an Ancient Protein's Dynamics with NMR

The tyrosine kinases Abl and Src are very similar in function and fold. However, the cancer drug imatinib exhibits highly selective binding for Abl compared to Src. For a long time it was assumed that these differences arise through a conformational selection mechanism of a DFG-loop, which allows binding of imatinib only if it is in the correct conformation. Recently, an additional induced fit step in the process of imatinib binding has been shown to account for the differences in binding of the drug to Abl in comparison with Src. To investigate the binding mechanism of imatinib to Abl we used resurrected enzymes along the nodes in a phylogenetic tree between the common ancestor of Abl and Src and the contemporary Abl kinase. We show that the binding affinities of each of the four resurrected ancestor enzymes are gradually increasing from the common ancestor towards the contemporary Abl and that the mechanism of binding is comprised of a conformational selection step followed by an induced fit step. To probe the conformational changes the enzyme undergoes upon imatinib binding, we use CPMG relaxation dispersion experiments. We show that most of the exchanging residues are in an intermediate or fast time regime. Combining the CPMG data with molecular dynamics will allows us to get a more detailed look at which residues play an import role for the selectivity of imatinib towards Abl kinase.

Transnational Mobilisation on Access to Medicines: The Global Movement Around the Imatinib Mesylate Case and Its Roots in the AIDS Movement

Starting in 2006, a patent dispute emerged in India with significant implications for global access to medicine. An application by Novartis for a ‘new formulation’ patent on the beta crystalline form of the cancer drug imatinib mesylate was rejected. Novartis responded by challenging the legality of provisions designed to protect access to medicines in India’s patent legislation. A global activist movement emerged in defence of these aspects of Indian patent legislation mainly because the challenge threatened access to medicine at a systemic level, given India’s critical role as a supplier of generic medicines to developing countries. The mobilisation described in this paper exploited existing activist networks and broader ‘networks of influence’ built during prior campaigns conducted around access to antiretroviral drugs. AIDS activists consequently played a key role in the global mobilisation around the imatinib mesylate dispute, aimed at generating public outrage and political pressure against Novartis’s challenge to Indian patent law. A critical ‘broker’ in these transnational networks is identified as the medical charity Medicins sans Frontieres (MSF), and specifically its dedicated medicines access campaign, which linked disparate activist groups together, disseminated information and mobilised AIDS activists across the world in support of the Indian patent flexibilities. In 2013 the Indian Supreme Court upheld the relevant sections of the law, representing a significant victory for the international medicines access movement. This movement illustrates the power of transnational mobilisation to help bring about ‘moral consensus’ and to deploy influence and social power at the global level sufficient to overcome substantial corporate power. But we also argue that both the emergence of the ‘global AIDS movement’ and the emergence of a transnational social movement around the imatinib mesylate dispute depended on a set of historical circumstances that are unlikely to persist and demonstrate that the broader medicines access movement faces significant challenges to its sustainability and future success.

International comparison on the factors influencing reimbursement of expensive cancer drugs.

6614 Background: Reimbursement policy for expensive cancer drugs is heterogeneous and inconsistent among countries. We compared the pattern of publically funded drug programs of ten countries. Methods: We investigated reimbursement policies of 19 indications with expensive cancer drugs (nilotinib, dasatinib, imatinib, pemetrexed, bevacizumab, sunitinib, temsirolimus, lapatinib, crizotinib, erlotinib, cetuximab, sorafenib and lenalidomide) in ten countries (Australia, Canada, England, France, Germany, Japan, Korea, Sweden, Taiwan, and the United States). Microarray analysis methodology is adopted to cluster the countries according to the pattern of reimbursement, based on incremental cost effectiveness ratio (ICER) and other variables of 19 indications. The earlier a country starts reimbursement for the indication, the higher reimbursement score (RS) is assigned to the country. Fairness index (FI) was determined for each country, depending on whether reimbursement decision follows ICER sequence. Results: Except for US, Japan and France were more likely to reimburse the indications (16/19), whereas Sweden and UK were less likely to reimburse them (5/19 and 6/19, respectively). From array analysis, we observed three separate groups: 1) France and Germany, 2) Korea and Japan, 3) UK, Sweden, Canada, Taiwan and Austrailia. The details of analysis will be presented. Regarding FI, Sweden (0.75), France (0.73) and UK (0.71) were the countries with high FI, in which health system was financed by general taxation and health technology assessment (HTA) was performed by government agency. Korea (0.34), Taiwan (0.40) and Germany (0.46) were the countries with low FI, in which social health insurance was financed by payroll tax or premium rather than general taxation. HTA was performed by non-government agency (Germany) or recently established government agency (2007 in Taiwan and 2009 in Korea) in these countries. Conclusions: Reimbursement policies for expensive cancer drugs are variable among the nations. Countries where health system is funded by general taxation and HTA is performed by government agency showed consistency of reimbursement policy based on ICER.

International variability in the reimbursement of cancer drugs by publically funded drug programs.

Evaluate inter-country variability in the reimbursement of publically funded cancer drugs, and identify factors such as cost containment measures that may contribute to variability. As of February 28, 2010, licensed indications for 10 cancer drugs (bevacizumab, bortezomib, cetuximab, erlotinib, imatinib, pemetrexed, rituximab, sorafenib, sunitinib, and trastuzumab) were obtained from the drug registries of 6 licensing authorities corresponding to 13 countries or regions: Australia, Canada (Ontario), England, Finland, France, Italy, Germany, Japan, New Zealand, the Netherlands, Scotland, Sweden, and the United States (Medicare Parts B and D). Number of licensed indications reimbursed by public payers and the use of cost containment measures were obtained by survey of health authorities involved in reimbursement and through public documents. The 48 identified licensed indications varied between agencies (range: 36-44 indications). Finland, France, Germany, Sweden, and the United States reimbursed the highest percentage of indications (range: 90%-100%). Canada (54%), Australia (46%), Scotland (40%), England (38%), and New Zealand (25%) reimbursed the least. All 5 countries with the lowest rate of reimbursement incorporated a cost-effectiveness analysis into reimbursement decisions and rejected submissions for reimbursement mainly because of lack of cost effectiveness; in New Zealand, lack of cost effectiveness was the second leading cause of rejection after excessive cost. In 9 countries, risk-sharing agreements were used to contain costs. Indications initially not recommended for reimbursement (9 in Australia, 5 in Canada, and 3 in England, New Zealand, and Scotland) were subsequently approved with risk-sharing agreements or special pricing arrangements. Reimbursement of publically funded cancer drugs varies globally. The cause is multifactorial.

Abstract B65: MicroRNAs in cancer drug resistance and sensitivity

Abstract microRNAs are small noncoding RNAs of 18–25 nucleotides that have been shown to regulate gene expression posttranscriptionally and play fundamental roles in diverse biological and pathological processes. MiRNApedia is an in silico discovery tool designed to integrate all microRNA-related information including genetic variation, gene expression, epigenetic regulation, pathways, diseases, clinical outcomes and particularly drug actions. The system documented more than 1,000 microRNAs and 600 therapeutic and experimental compounds including well-known cancer drugs cisplatin, doxorubicin, 5-flouracil, gemicitabine, paclitaxel, docetaxal, imatinib, vorinostat, sorafenib and gefitinib etc. For each compound, microRNAs and genes related to its sensitivity and function were compiled from literature and used to construct comprehensive drug-gene-microRNA networks. MicroRNAs played important roles in determining resistance and sensitivity of cancer drugs. For example, miR-21 reduced sensitivities to daunorubicin, topotecan, 5-FU, taxol, doxorubicin, docetaxel, ara-C, arsenic trioxide, gemcitabine and dexamethasome, while miR-146a increased sensitivity to bleomycin and over-expression of miR-221 conferred resistance to tamoxifen. Multiple microRNAs might be involved in the efficacy of a single drug. MiRs -98, -214, -106a, -125b and -630 contributed to cisplatin resistance, while -451, -181a, -148a, -138 and let-7i enhanced its sensitivity. Meanwhile, cisplatin was shown to down-regulate miRs -150 and -106 while up-regulated miRs -34, -16, -17-5p and -125. Drug combination would be the choice to overcome resistance in mono-therapies. Prior to experimental testing, the combination effects of multiple drugs could be assessed computationally by combining annotated information in microRNA targets, signaling pathways and transcriptional regulations etc. For example, trichostatin A (TSA), a HDAC inhibitor, might enhance sensitivity of cisplatin by up-regulating p53 and its transcriptional target miR-34 which subsequently down-regulated BCL2, a well-known drug resistance factor. However, if miR-125b, another transcriptional target of p53, was up-regulated, the combination might turn out to be antagonistic. Mechanistically, miR-125b targeted BAK1 which was required for cisplatin sensitivity. Such in silico modeling exercises not only predicted synergism or antagonism, but also identified underlined drivers which could serve as biomarkers for desired therapeutic effects. The potential biomarkers identified by the network modeling approach could be used to guide selection of experimental system (e.g., cell line models) and monitor the combination effectiveness. The present study devised a mechanism to systematically capture microRNA information in regards to cancer drug resistance and established a computational framework to identify potential drug combinations to overcome resistance in cancer therapies.

Crystal Structure of the Kinase Domain of ABL in Complex with a Potent Rationally-Designed Derivative of the PP1 Inhibitor, AB129

As part of our effort to understand the determinants of drug binding to the Abl tyrosine kinase, we determined the crystal structure of the Abl kinase domain bound to a derivative of the PP1 nucleoside analog. Just as the cancer drug imatinib selectively inhibits the tyrosine kinase activity of c-Abl, c-Kit, and the PDGF receptor, but is a poor inhibitor of the closely related Src kinases (1), PP1 is a selective inhibitor of the Src family (2), c-Kit (3), and PDGFR kinases but not of Abl tyrosine kinase activity (3, 4). In fact, PP1 is 1000X less effective at inhibiting Abl and Bcr-Abl kinases than at inhibiting the Src kinases (3, 4). We rationally designed a derivative of the PP1 compound to introduce hydrogen bonding between it and a strictly conserved lysine residue that coordinates the alpha and beta phosphate groups of ATP in protein kinases (5). By expanding the interactions between the inhibitor and Abl kinase, we sought to make the PP1 derivative, AB129, less selective. Our crystal structure of the Abl kinase domain bound to the AB129 inhibitor shows three important features. First, the compound binds to the kinase domain in the active conformation: alpha C helix is rotated inward facilitating a salt bridge between Lys271 and Glu286 and the conserved Asp-Phe-Gly (DFG) motif is oriented Asp-In. In contrast, the structure of PP1 bound to the Src family kinase Hck is in the inactive conformation (6). Second, the strictly conserved Glu286 residue, not Lys271, hydrogen bonds to the hydroxyl group of AB129. Third, the conformation of the phosphate binding loop (P-loop) is extended. We provide kinetic and calorimetric data to support our rationale for the observed conformational changes.

Molecular Dynamics Simulations Show That Conformational Selection Governs the Binding Preferences of Imatinib for Several Tyrosine Kinases

Tyrosine kinases transmit cellular signals through a complex mechanism, involving their phosphorylation and switching between inactive and active conformations. The cancer drug imatinib binds tightly to several homologous kinases, including Abl, but weakly to others, including Src. Imatinib specifically targets the inactive, so-called "DFG-out" conformation of Abl, which differs from the preferred, "DFG-in" conformation of Src in the orientation of a conserved Asp-Phe-Gly (DFG) activation loop. However, recent x-ray structures showed that Src can also adopt the DFG-out conformation and uses it to bind imatinib. The Src/Abl-binding free energy difference can thus be decomposed into two contributions. Contribution i measures the different protein-imatinib interactions when either kinase is in its DFG-out conformation. Contribution ii depends on the ability of imatinib to select or induce this conformation, i.e. on the relative stabilities of the DFG-out and DFG-in conformations of each kinase. Neither contribution has been measured experimentally. We use molecular dynamics simulations to show that contribution i is very small, 0.2 +/- 0.6 kcal/mol; imatinib interactions are very similar in the two kinases, including long range electrostatic interactions with the imatinib positive charge. Contribution ii, deduced using the experimental binding free energy difference, is much larger, 4.4 +/- 0.9 kcal/mol. Thus, conformational selection, easy in Abl, difficult in Src, underpins imatinib specificity. Contribution ii has a simple interpretation; it closely approximates the stability difference between the DFG-out and DFG-in conformations of apo-Src. Additional calculations show that conformational selection also governs the relative binding of imatinib to the kinases c-Kit and Lck. These results should help clarify the current framework for engineering kinase signaling.

A conserved protonation-dependent switch controls drug binding in the Abl kinase

In many protein kinases, a characteristic conformational change (the "DFG flip") connects catalytically active and inactive conformations. Many kinase inhibitors--including the cancer drug imatinib--selectively target a specific DFG conformation, but the function and mechanism of the flip remain unclear. Using long molecular dynamics simulations of the Abl kinase, we visualized the DFG flip in atomic-level detail and formulated an energetic model predicting that protonation of the DFG aspartate controls the flip. Consistent with our model's predictions, we demonstrated experimentally that the kinetics of imatinib binding to Abl kinase have a pH dependence that disappears when the DFG aspartate is mutated. Our model suggests a possible explanation for the high degree of conservation of the DFG motif: that the flip, modulated by electrostatic changes inherent to the catalytic cycle, allows the kinase to access flexible conformations facilitating nucleotide binding and release.

Analysis of binding energy activity of imatinib and Abl tyrosine kinase domain based on simple consideration for conformational change: An explanation for variation in imatinib effect in mutated type

Imatinib is a clinically well-tolerated small molecule inhibitor that exerts selective, dual inhibition on the transforming growth factor beta and platelet-derived growth factor pathways. The recognition of an inactive conformation of Abl, in which a catalytically important Asp-Phe-Gly motif is flipped by approximately 180 degrees with respect to the active conformation, underlies the specificity of the cancer drug imatinib, which is used to treat chronic myelogenous leukemia. However, conformational analysis shows that the effect of the drug depends on the potential energy, which varies due to the alpha rotatable angles of the Abl tyrosine kinase domain. In this study, the author determines the change of binding energy between the Abl tyrosine kinase domain, due to the variation in rotatable angles, and bond lengthening. According to this study, the ratio between the required binding energy between the wild and mutated types is equal to 1: 1.16.

Turning promiscuous kinase inhibitors into safer drugs

Burgeoning interest in multi-target drugs to treat complex diseases and malignancies has motivated a reassessment of the therapeutic value of promiscuity. Although drug efficacy might not correlate with specificity, it would be risky to welcome promiscuous compounds without a rational strategy to control therapeutic impact. This situation might change as novel selectivity filters are incorporated into drug design. For example, cardiotoxic side effects of the cancer drug imatinib might be curbed by applying such premises. Here, we survey approaches to control the therapeutic impact of cross-reactive kinase inhibitors and advocate the application of a novel selectivity filter by illustrating its cleaning efficacy. Finally, we evaluate the possibility of turning multi-target kinase inhibitors into clinical opportunities through judicious chemical modifications.

C-Abl Tyrosine Kinase and Inhibition by the Cancer Drug Imatinib (Gleevec/STI-571)

The search for specific protein kinase inhibitors is an intense area of research because of the potential for drug development. The small-molecule inhibitor imatinib (Gleevec/STI-571) can specifically inactivate the tyrosine kinase c-Abl, whose normal mechanism of autoinhibition is disrupted in chronic myelogenous leukemia. Crystallographic analysis of c-Abl reveals that imatinib recognizes a distinct inactive conformation of the Abl kinase domain that relies on the mechanism of autoinhibition achieved in the context of a larger fragment of the protein. This mechanism is distinct from that seen in the related Src family kinases, where autoinhibition is achieved through the internal engagement of a C-terminal phosphotyrosine residue by the Src homology 2 domain (SH2) domain. Notably, this phosphotyrosine residue is lacking in c-Abl, where instead autoinhibition is mediated by an interaction between the kinase domain and the N-terminal myristoyl modification. Within the framework of these 2 distinct modes of autoinhibition, the SH3-SH2 unit is structurally conserved between Abl and Src, leading to large conformational differences in their kinase domains. These differences help explain the ability of imatinib to preferentially inhibit Abl over Src.

C-Src Binds to the Cancer Drug Imatinib with an Inactive Abl/c-Kit Conformation and a Distributed Thermodynamic Penalty

The cancer drug imatinib inhibits the tyrosine kinases c-Abl, c-Kit, and the PDGF receptor. Imatinib is less effective against c-Src, which is difficult to understand because residues interacting with imatinib in crystal structures of Abl and c-Kit are conserved in c-Src. The crystal structure of the c-Src kinase domain in complex with imatinib closely resembles that of Abl*imatinib and c-Kit*imatinib, and differs significantly from the inactive "Src/CDK" conformation of the Src family kinases. Attempts to increase the affinity of c-Src for imatinib by swapping residues with the corresponding residues in Abl have not been successful, suggesting that the thermodynamic penalty for adoption of the imatinib-binding conformation by c-Src is distributed over a broad region of the structure. Two mutations that are expected to destabilize the inactive Src/CDK conformation increase drug sensitivity 15-fold, suggesting that the free-energy balance between different inactive states is a key to imatinib binding.

The cancer drug imatinib can cause cardiotoxicity

The cancer drug imatinib can cause cardiotoxicity

In silico profiling of tyrosine kinases binding specificity and drug resistance using Monte Carlo simulations with the ensembles of protein kinase crystal structures

The molecular basis of the tyrosine kinases binding specificity and drug resistance against cancer drugs Imatinib and Dasatinib is elucidated using Monte Carlo simulations of the inhibitor-receptor binding with the ensembles of protein kinase crystal structures. In silico proteomics analysis unravels mechanisms by which structural plasticity of the tyrosine kinases is linked with the conformational preferences of Imatinib and Dasatinib in achieving effective drug binding with a distinct spectrum of the tyrosine kinome. The differences in the inhibitor sensitivities to the ABL kinase mutants are rationalized based on variations in the binding free energy profiles with the conformational states of the ABL kinase. While Imatinib binding is highly sensitive to the activation state of the enzyme, the computed binding profile of Dasatinib is remarkably tolerant to the conformational state of ABL. A comparative analysis of the inhibitor binding profiles with the clinically important ABL kinase mutants has revealed an excellent agreement with the biochemical and proteomics data. We have found that conformational adaptability of the kinase inhibitors to structurally different conformational states of the tyrosine kinases may have pharmacological relevance in acquiring a specific array of potent activities and regulating a scope of the inhibitor resistance mutations. This study outlines a useful approach for understanding and predicting the molecular basis of the inhibitor sensitivity against potential kinase targets and drug resistance.

A Src-Like Inactive Conformation in the Abl Tyrosine Kinase Domain

The improper activation of the Abl tyrosine kinase results in chronic myeloid leukemia (CML). The recognition of an inactive conformation of Abl, in which a catalytically important Asp-Phe-Gly (DFG) motif is flipped by approximately 180° with respect to the active conformation, underlies the specificity of the cancer drug imatinib, which is used to treat CML. The DFG motif is not flipped in crystal structures of inactive forms of the closely related Src kinases, and imatinib does not inhibit c-Src. We present a structure of the kinase domain of Abl, determined in complex with an ATP–peptide conjugate, in which the protein adopts an inactive conformation that resembles closely that of the Src kinases. An interesting aspect of the Src-like inactive structure, suggested by molecular dynamics simulations and additional crystal structures, is the presence of features that might facilitate the flip of the DFG motif by providing room for the phenylalanine to move and by coordinating the aspartate side chain as it leaves the active site. One class of mutations in BCR–Abl that confers resistance to imatinib appears more likely to destabilize the inactive Src-like conformation than the active or imatinib-bound conformations. Our results suggest that interconversion between distinctly different inactive conformations is a characteristic feature of the Abl kinase domain.