imatinib is the frontline therapy for chronic - phase chronic myeloid leukemia ( cml ) because it demonstrates high complete cytogenetic response ( ccyr ) and major molecular response ( mmr ) rates . however , a further 20%-25% of those who do achieve complete hematologic response ( chr ) or ccyr , or both , may eventually stop responding and acquire resistance during exposure to imatinib . in advanced - phase cml ( i.e. , accelerated phase and blast crisis ) , 38% of the patients treated with imatinib achieved major cytogenetic response ( mcr ) . dasatinib and nilotinib are potent tyrosine kinase inhibitors ( tkis ) with activity against many imatinib - resistant cml clones . for patients with cml who become or are inherently resistant to imatinib therapy , dose escalation and the second - generation tkis need to be considered based on the bcr - abl mutation profile and the patient 's disease history , with the notable exception of the t315i mutation . with the increasing use of tkis , it has been suggested that the spectrum of kinase domain mutations may change and possible selection of new resistant clones may occur . more recently , experimental and computational works suggest that any of the pair - wise cross - resistance of imatinib , dasatinib , and nilotinib could overcome and prevent resistance . in this case study , we report two patients showing resistance to imatinib and intolerance to recommended - dose dasatinib , who were treated on combined standard - dose imatinib and reduced - dose dasatinib with no significant extramedullary toxicity . our observations confirm a very recent report showing that combination of tkis may provide an additive / synergistic antileukemic effect . he commenced imatinib at an initial dose of 400 mg / day while the disease was progressing , with a platelet count of 100010/l , in april 2004 . after 18 months of treatment with imatinib , he achieved a partial cytogenetic response ( 30%ph+ metaphase ) and commenced imatinib at 600 mg / day . the dynamics of bcr - abl fusion clones by fluorescence in situ hybridization ( fish ) on a peripheral blood specimen using dual probes for the bcr and abl genes showed 0.6% in june 2006 and 36% in october 2006 . he then commenced dasatinib at 70 mg b.i.d . for 10 days , but experienced dyspnea and a feeling of impending death . although he had an hla - matched sibling , he declined allogeneic stem cell transplantation . at that point , direct sequencing of pcr - rflp - amplified bcr - abl1 products did not detect any mutation . five months later , the patient lost hematologic response , showing persistent high platelet values ( > 90010/l ) , administration of dasatinib of 70 mg q.o.d and imatinib of 600 mg / day was initiated in september 2007 . this resulted in a chr and ccyr after 1 month and 8 months , respectively , and the regimen was very well tolerated with no side effects . the bcr - abl1/abl1 ratio in peripheral blood decreased to less than 0.01% by rt - pcr ( four log sensitivity ) in july 2008 ( 10 months on combination imatinib with dasatinib ) . he remained in persistent mmr at the 40 month regular follow - up . in january 2011 , the bcr - abl1/abl1 ratio in peripheral blood dramatically increased to 31.1% when the bcr - abl t315i mutation proved positive by dna sequencing . the patient then received 2 cycles of treatment consisting of low - dose aclarubicin , cytarabine and g - csf , but had a poor outcome . the second patient was a 71-year - old man diagnosed with cml in september 2002 . following 3 years of hydroxyurea and alpha interferon therapy , he commenced imatinib at 400 mg / day and achieved a partial cytogenetic response ( 20%ph+ metaphases ) after 10 months . the disease progressed to the accelerated phase , which was detected by observing bone marrow morphology and cytogenetic analysis of 39% ph+ metaphases in october 2006 . after 3 , 6 , and 9 months of imatinib therapy , bcr - abl fusion clones by fish were 43% , 43.3% , and 4% , respectively . with no mutations detected , he commenced imatinib of 800 mg / day until september 2007 , at which time he had failed to achieve chr . white blood cell ( wbc ) count was 15.210/l with 89% myeloid blasts in the differential . meanwhile , bone marrow morphology and immunophenotyping showed malignant blasts positive for cd13 , cd117 , cd34 , and cd33 , without expressing other lymphoid markers . cytogenetic analysis showed 47 , xy , + 8 , t(9;22)(q34;q11 ) , i(17q ) in 20 metaphases . he commenced dasatinib at 70 mg b.i.d . , but 3 days later exhibited dyspnea and cardiopalmus . further examination by a computed tomography scan showed a great quantity of pleural effusion with an albumin level of 39.4 however , after 4 days of dasatinib therapy dyspnea recurred . at this point , myeloblasts decreased to 1% in the peripheral blood smear . subsequently , the patient was started with a combination of imatinib of 600 mg / day and dasatinib of 70 mg q.o.d . a ccyr and mmr were achieved at 3 and 12 months after combined therapy , respectively . on his last assessment in august 2013 ( 66 months on combination of imatinib with dasatinib ) , bcr - abl transcripts were undetectable in every 3 month exam . resistance to imatinib is likely to be a multifactorial process . the predominant cause of acquired resistance to imatinib is reactivation of bcr - abl kinase activity via kinase domain mutations , amplification of the bcr - abl genomic locus , and is independent of bcr - abl . secondary clonal abnormalities have been observed to develop in approximately 5% of patients achieving ccyr . it has been suggested that alternative genetic aberrations synergize with bcr - abl in the evolution of imatinib resistance . although the mechanism is not clear , the disease progression may be associated with clonal evolution , which was also verified in our second patient . the optimal treatment for patients failing imatinib treatment is imatinib dose escalation , a second - generation tki , allogeneic stem cell transplantation , or other antileukemia agents , such as homoharringtonine . a sequential use of imatinib , followed by either dasatinib or nilotinib on an empirical basis has provided therapeutic options for patients who presented with imatinib resistance . dasatinib is a synthetic small - molecule inhibitor of src - family kinases , which inhibits the binding of abl kinase to bcr - abl , irrespective of abl conformation . in vitro , dasatinib presents 325-fold greater activity against native bcr - abl , when compared with imatinib , and has shown efficacy against all imatinib - resistant bcr - abl mutations with the exception of t315i . it is also more potent than imatinib in inhibiting nonmutated bcr - abl kinase activity . different kinase domain mutation profiles have been recovered with imatinib , dasatinib , and nilotinib in mutagenesis assays . described a case of persisting primary resistance to three tkis used in sequence , and dynamically developed sequential mutant clones with disappearance of those previously detected . the patient developed m244v mutation after imatinib therapy , but acquisition of a new m351 t mutation and disappearance of m244v mutation after nilotinib therapy and acquisition of a new f317l mutation and disappearance of m351 t after dasatinib therapy . recently , cortes et al . reported that 29 of 112 patients ( 26% ) who received either dasatinib or nilotinib after imatinib failure developed new kinase domain mutation . mutations that confer resistance to dasatinib , but not to imatinib , have been identified in vitro . for example , v299l mutation , also involving a contact point , was reported in patients who failed dasatinib , but rarely in patients with imatinib therapy . therefore , the rational approach for the use of several tkis is to tailor these agents to tackle specific bcr - abl1 mutant clones according to the individual mutations detected at each time - point during the course of therapy . one attractive strategy for minimizing the onset of acquired drug resistance is to use a combined abl inhibitor approach . computational work has suggested that a combination of different drugs against cml could similarly overcome the resistance problem , even during advanced stages . further studies using computational method have shown that combining two of imatinib , dasatinib , and nilotinib can provide an advantage over using one drug alone . however , combining three tki drugs may not lead to any further advantage when compared with the combination of two drugs . recently , hiwase et al . found that the combination of low - dose dasatinib and nilotinib induced significantly higher cell death than that induced by each individual drug alone in vitro . thus , the most potential benefit of the new therapeutical approach that is it may target a wider range of resistant clones than a single agent , and thereby prohibit or delay the onset of acquired drug resistance . interestingly and surprisingly , imatinib works at all advanced stages of the disease because blast crisis is usually attributed to additional genetic changes conferring bcr - abl - independent pathways of proliferation and maturation arrest . for a combined abl inhibitor approach to work , it is crucial that imatinib does not interfere with the ability of the src / abl inhibitor to access its binding site within the bcr - abl kinase domain , even when coadministered with imatinib at concentrations above typical clinical levels . thus , owing to the enhanced inhibitory activity , combined abl inhibitor therapy could lead to eradication of a higher proportion of residual leukemic cells and , potentially , decreasing bcr - abl - dependent molecular disease persistence in cml ; this improves the molecular response rates in newly diagnosed patients and those with imatinib - resistant cml . thus , if acceptable , frontline concomitant combinations of different abl inhibitors in tkis nve patients are warranted , taking into consideration that the use of tkis combinations will lead to increased toxicity in normal tissues in the clinical setting . in view of efficacy and safety , the administration of standard dose of imatinib at 600 mg once daily combined with reduced dose of dasatinib at 70 mg q.o.d . to our two patients resulted in the rapid achievement of ccyr , which is still ongoing and accompanied by mmr . to the best of our knowledge , this is the first report on the use of imatinib - dasatinib combinations for patients with imatinib - resistant cml in advanced phase . the recommended daily dose of dasatinib for patients with chronic - phase cml is from 70 mg b.i.d . to 100 mg once daily . dasatinib of 100 mg once daily retains the efficacy of 70 mg twice daily with less toxicity ; however , survival was significantly better with dasatinib of 100 mg daily . a recent in vitro study by hiwase et al . showed that the combination of low - dose dasatinib and nilotinib may provide an additive / synergistic antileukemic effect , which may be of particular relevance to tki therapy and expressing abcb1 . based on our in vivo and other experimental findings , we speculate that a combination of imatinib and dasatinib may be synergistic , and imatinib - mediated inhibition of abcb1 may enhance the intracellular uptake and retention ( iur ) of dasatinib . this would be analogous to the described synergy between nilotinib and imatinib or dasatinib and nilotinib , which we speculated to be owing to an imatinib - mediated increase in nilotinib iur , a nilotinib - mediated increase in dasatinib iur . both our patients were rescued from switching to standard - dose imatinib and reduced - dose dasatinib combination therapy without new combination - related side effect and obtained longer - term benefit . as it has been recently pointed out in an excellent review , dasatinib was efficacious and the results did not differ significantly across dasatinib doses . intermittent target inhibition with tkis may preserve the efficacy and reduce adverse events . administering imatinib and dasatinib simultaneously produced a marked reduction in cells expressing bcr - abl or some mutant except normal cell and primary cml cells expressing mutant t315i . moreover , there does not appear to be cross - intolerance between dasatinib and imatinib . consistent with the study of o'hare et al , our results showed that the combined abl inhibitors do not exhibit increased toxicity when compared with single agents . based on our data , it can be postulated that imatinib affects malignant cells , while dasatinib has a critical role in inhibiting or eradicating the several subclones ( mutant abcb1-overexpressing cells , and/or resistant - imatinib malignant clone ) . in addition , malignant clones associated with resistance to dasatinib may be vulnerable to imatinib , thus decreasing the overall chances of resistance . additive / synergistic antiproliferative effects from imatinib - dasatinib combination could lead to eradication of a higher proportion of residual leukemic cells . thus , the data presented here suggest that a combined imatinib - dasatinib approach can be employed for further clinical trials in the subset of patients with tki resistance and stem cell refractoriness , in addition to opening a field for research on the use of any of the pairwise cross - resistant combinations of these powerful agents .
abstractfor patients with chronic myeloid leukemia ( cml ) failing imatinib therapy , second - generation tyrosine kinase inhibitors ( tkis ) are recommended . here , we describe two patients with advanced cml who failed imatinib therapy and did not tolerate the recommended dose of dasatinib , but then achieved a major molecular response with the combination of imatinib and dasatinib with no significant extramedullary toxicity . 
 our observations suggest that combination of tkis may provide an additive / synergistic antileukemic effect .