be a result of the fact that the drugs are taken up in cells by different mechanisms: imatinib influx is dependent on oct1 but nilotinib is not. nilotinib is also not a substrate for the efflux transporter p-glycoprotein pump, unlike imatinib. although the two dimensional molecular structures of these two drugs might look similar, they are dissimilar in terms of spatial structure and molecular properties. crystal structure of abl kinase domain (blue) in complex with nilotinib (red) binding nilotinib binds to the inactive conformation of the abl kinase domain, largely through lipophilic interactions and thus blocks its catalytic activity. nilotinib binds to the kinase domain by making four hydrogen bond interactions involving the pyridyl-n and the backbone nh of met-318, the anilino-nh and the side chain oh of thr-315, the amido-nh and side chain carboxylate of glu-286 and the amido carbonyl with the backbone nh of the asp-381. the anilino segment of nilotinib has close binding interactions with met-318, phe-317 and thr-315 residues of a region within the atp binding site. the remaining half of the compound extends beyond the thr-315 gatekeeper residue to bind within an additional pocket. the 3-methylimidazole and trifluoro-methyl groups of nilotinib make important interactions with the abl kinase domain. these groups also make the shape of nilotinib very different from that of imatinib. nilotinib also binds to the kinase through a large number of weak van der waals interactions. resistance nilotinib has shown effect against most mutations (32/33) that are associated with imatinib resistance but the t315i mutant remains resistant to nilotinib. its ineffectiveness against the t315i mutant seems to be a consequence of the loss of an h-bond interaction between threonine-o and aniline-nh on nilotinib and a steric clash between the isoleucine-methyl group and 2-methylphenyl phenyl group of nilotinib. on the other hand, resistance to nilotinib is associated with a limited spectrum of bcr-abl kinase mutations that mostly affect the p-loop and t315i. however all mutations except t315i were effectively suppressed by increasing nilotinib concentration. although nilotinib is more potent than imatinib it is possible that its specific mode of binding to abl may make other sites vulnerable to new kinds of drug resistance. dasatinib (bms-354825) dasatinib in its binding site development dasatinib is a thiazolylaminopyrimidine developed as the hydrochloride salt. it was discovered with a program directed towards immunosuppressive drugs and is 325-fold more potent against cells expressing wild type bcr-abl than imatinib. dasatinib is a multi targeted inhibitor of bcr-abl and src family kinases. it also has inhibitory activity against additional downstream kinases. crystal structure (pdb 2gqg) of abl kinase domain (blue) in complex with dasatinib (red). binding dasatinib binds to abl with less stringent conformational requirements than imatinib so it exhibits increased potency but reduced selectivity compared to imatinib. dasatinib binds both the active and inactive conformation of abl kinase, contrary to the binding of most other tkis to the active form only. compounds that target the active conformation have been identified but the binding site in all the hundreds of human protein kinases is very similar. therefore, there is a considerably greater scope for dissimilarities between the inactive conformations so the efforts to discover highly selective kinase inhibitors are being directed towards molecules that bind to the inactive conformation. dasatinib has some structural elements in common with nilotinib, in particular the juxtaposition of the aminopyrimidine and the carboxamide groups. the aminothiazole segment of dasatinib makes a bi-dentate h-bonding interaction with the backbone co and nh of met-318 and the amide-nh makes an h-bond with the side chain oxygen of thr-315. resistance since dasatinib is an inhibitor of src family kinases, it can overcome resistance due to src family kinase activation. because it does not bind to bcr-abl with the same stringent conformational requirements as imatinib, it can inhibit all bcr-abl kinase domain mutants except for t315i. dasatinib is also not a substrate of multidrug p-glycoprotein efflux pumps like imatinib. because of this dasatinib may be active in some patients after failure with both imatinib and nilotinib. although dasatinib is much more potent than imatinib it is possible, like with nilotinib, that its specific mode of binding to abl may lead to new vulnerable sites that could confer new kinds of drug resistance. mutations have been found on phe317 so that is a potential vulnerable site for this drug. development bosutinib bosutinib's structure is based on a quinoline scaffold and is structurally related to the astrazeneca quinazoline template. src kinase dependent yeast screening led to characterization of a 4-anilino-3-quinolinecarbonitrile as an src inhibitor. combination of the features of this hit and a related compound, and attachment of solubilizing groups, led to the discovery of bosutinib. it was suggested to be an abl kinase inhibitor and when tested as such it turned out to be slightly more potent against abl than src (ic50 1,4 nm vs. 3,5 nm). bosutinib's activity was first described in 2001 and it was disclosed as an abl kinase inhibitor in 2003. at first it was believed that bosutinib was a selective src kinase inhibitor but now it is known that its kinase inhibition profile is far less restricted than originally thought. bosutinib inhibits src, abl and a wide range of both tyrosine and serine-threonine kinases. resistance bosutinib inhibited cells expressing a variety of mutations, some of which led to imatinib resistance, but the t315 mutation was completely resistant to bosutinib. in contrast to imatinib, nilotinib and dasatinib, bosutinib is not an efficient substrate for multidrug resistance (mdr) transporters that promotes efflux of foreign molecules from cells. bosutinib even inhibits these transporter proteins in higher concentrations. ponatinib (ap24534) ariad pharmaceuticals, inc. announced on september 10, 2010 that ponatinib, an orally active bcr-abl tki effective against the t315i mutation had been approved for a phase ii clinical trial. the road to discovery can be linked to ap23464, one of the first of ariad's atp competitive dual src/abl inhibitors. ap23464 was identified using structure base drug design and focused synthetic libraries of trisubstituted purine analogs. the substance potently inhibits, on nanomolar scale, src and bcr-abl kinases including many common imatinib resistant bcr-abl mutations. ap23464 does not inhibit the t315i mutation, however, whereas ap24534 (ponatinib) does. development ponatinib development history ariad used the highly potent drug lead, ap23464 to further investigate inhibitory possibilities of purine cored templates for dual src/abl inhibitors. first, searching for substances effective on the inactive conformation of abl, the side chain bound to the nitrogen on the purine core was replaced with a diarylamide structure, that was known to have a high affinity to the inactive conformation by forming crucial hydrogen bonds and filling hydrophobic pockets on the kinase. furthermore, it was determined that the cyclopentyl group on the purine core clashed with a glycine rich p-loop in that confirmation and was thus removed from the molecule. then with in-vitro testing on inhibitory activity and in-vivo oral absorption assays a more lipophilic, amide bound, cyclopropyl group on c6 on the purine core was found to display both satisfactory pharmacokinetics and efficacy. finally modifications on the diarylamide side chain by adding imidazole appendages were inspired by then newly released nilotinib structure. those modifications resulted in what was called ap24163. during this development cycle, ariad tested several substances against cells transfected with t315i mutated bcr-abl kinase and, surprisingly, found ap24163 demonstrated reasonable inhibitory action on top of potent inhibition of native bcr-abl. following up on that breakthrough ariad began further research to increase the efficacy of compound ap24163 against the t315i mutation. docking of the molecule into the atp binding site of t315i mutated bcr-abl kinase revealed that the expected steric clash with isoleucine was not present due to a lesser sterically demanding vinyl linkage between the purine core and the diarylamide side chain compared to other tkis. the first step was to try to find an even less sterically demanding structure. first an acetylene linkage was tested, that resulted in higher potency but unfavorable pharmacokinetics. later, a more stable 2-butyne linkage was selected. to achieve this linkage an imidazolpyridine core was used as a starting material for a sonogashira reaction; but the pharmacokinetics were still poor. when developing ap24163, adding a cyclopropane side chain on c8 in the purine core resulted in favorable pharmacokinetics. several different side chains were then tested, but the best results were obtained with no side chain at all, resulting a substance with satisfactory pharmacokinetics, but now with reduced potency against t315i also. the first step in increasing the potency again was to look at other tki's. imatinib has a terminal methyl piperazine group which has been shown to form a hydrogen bond with the carbonyl oxygen atom of residue ile-360 in the activation loop of the abl kinase. the piperazine ring is also a common solubilizing group that could further improve the pharmacokinetic properties of the molecule. those speculations were confirmed with a two-fold increase in inhibitory action against bcr-abl t315i mutated kinase and the silver lining was the plasma protein binding of the substance (named '19a') appeared to have decreased, allowing for smaller doses with the same potency. whilst '19a' exhibited good oral pharmacokinetics in both mice and rats, it also retained high partition coefficient at 6.69. so, in attempts to reduce the molecule's lipophilicity further, substitution of a single carbon atom on the imidazopyridine core was made; which resulted in what is now known as the compound ponatinib. ponatinib in its binding site binding x-ray crystallographic analysis of ponatinib and t315i bcr-abl mutated kinase display that the imidazopyridazine core rests in the adenine pocket of the enzyme. the methylphenyl group occupies a hydrophobic pocket behind i315, the ethynyl linkage forms favorable van der waals interactions with the amino acid and the trifluoromethyl group binds to a pocket induced by the inactive conformation kinase. also in the conformation of the kinase that ponatinib rests in, additional favorable van der waals interactions between the drug and tyr-253 and phe-382. five hydrogen bonds are generated, with the backbone of met-318 in the hinge region, with the backbone of asp-381, with the side chain of glu-286 and the protonated methylpiperazine with the backbone-carbonyl atoms of ile-360 and his-361. with this structure ponatinib has been shown to have a relatively broad kinase specificity profile which can probably be linked to the linearity of the linkage section of the molecule. with this linear structure the drug appears to avoid steric clashes with hydrophobic tk gatekeeper residues. despite, or even because of this, ponatinib is a potent drug and targets not just most of the known mutations on the bcr-abl tk but, most importantly of all, t315i. this mutation is emerging as a common pathway to failure of both first and second line treatments. unlike other t315i targeting inhibitors in development, ponatinib does not target aurora kinases, which clearly distinguishes it from them and emphasizes the significance of its discovery. bafetinib (inno-406) with the emerging resistance to imatinib treatment after its launch alternative treatment was highly sought after. bafetinib was the offspring of an attempt to create a more potent drug than imatinib, with efficacy against various point mutations in the bcr-abl kinase, with fewer adverse effects and with narrower kinase spectra, namely just lyn and bcr-abl. development in the search for a substance that fit the criteria mentioned, the crystal structure of imatinib bound to abl was examined. this revealed a hydrophobic pocket around the phenyl ring adjacent to the piperazinylmethyl group of imatinib. attempts to utilize this pocket to increase efficacy led to the addition of various hydrophobic groups including single fluoro, bromo and chloro substituents. finally a trifluoromethyl group at position 3 was found to give the best results, with approximately 36-fold improvement over imatinib. the addition of a hydrophobic group now needed to be countered to sustain the solubility of the substance. closer examination of the crystal structure of imatinib-kinase complex revealed tyr-236 was in close proximity to the pyridine ring of imatinib, suggesting there was little or no room for a larger group there. with that in mind a more hydrophilic pyrimidine ring was substituted for the pyridine, which was found to increase solubility while leaving efficacy the same or even slightly greater. finally to improve the hydrogen bonding of the piperazine ring of imatinib with ile-360 and his-361, pyrrolidine and azetidine derivatives were introduced. the most promising substance from these