Search

Real-time quantitative PCR (qPCR) is routinely used to detect minimal residual disease in chronic myeloid leukemia patients. The absolute quantification with droplet digital PCR (ddPCR) could reduce the inherent variability of qPCR. We established a duplex ddPCR assay using the Europe against Cancer (EAC) primer/probe system for breakpoint cluster region protein-tyrosine-protein kinase ABL1 (BCR-ABL1) and ABL1 and compared the results with qPCR. cDNA samples (n = 230) from patients with chronic myeloid leukemia were analyzed using both procedures. A second, commercially developed ddPCR assay for BCR-ABL1 was also evaluated. ABL1 and BCR-ABL1 transcript levels were similar with all assays, but the proportion of deep molecular responses was lower with ddPCR than with qPCR. The EAC ddPCR assay had a false-positive rate of 4% using a cutoff of three BCR-ABL1 copies per duplicate, compared with 2% without cutoff for the commercial ddPCR. The detection rate for molecular response 4.5 was 100, and a shift toward more minimal residual disease was seen in patient samples. In conclusion, using the EAC protocol for BCR-ABL1 quantification with ddPCR is feasible and shows low intra-assay and interassay variation but requires a cutoff that reduces sensitivity. The commercial ddPCR assay is highly sensitive and specific for BCR-ABL1. The use of either ddPCR assay resulted in a shift to lower molecular response classes compared with qPCR aligned to international scale., (Copyright © 2020 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

Ponatinib has potent activity against native and mutant BCR-ABL1, including BCR-ABL1 T315I The pivotal phase 2 Ponatinib Ph + ALL and CML Evaluation (PACE) trial evaluated efficacy and safety of ponatinib at a starting dose of 45 mg once daily in 449 patients with chronic myeloid leukemia (CML) or Philadelphia chromosome-positive acute lymphoblastic leukemia (ALL) resistant/intolerant to dasatinib or nilotinib, or with BCR-ABL1 T315I This analysis focuses on chronic-phase CML (CP-CML) patients (n = 270) with 56.8-month median follow-up. Among 267 evaluable patients, 60%, 40%, and 24% achieved major cytogenetic response (MCyR), major molecular response (MMR), and 4.5-log molecular response, respectively. The probability of maintaining MCyR for 5 years was 82% among responders. Dose reductions were implemented in October 2013 to decrease the risk of arterial occlusive events (AOEs); ≥90% of CP-CML patients who had achieved MCyR or MMR maintained response 40 months after elective dose reductions. Estimated 5-year overall survival was 73%. In CP-CML patients, the most common treatment-emergent adverse events were rash (47%), abdominal pain (46%), thrombocytopenia (46%), headache (43%), dry skin (42%), and constipation (41%). The cumulative incidence of AOEs in CP-CML patients increased over time to 31%, while the exposure-adjusted incidence of new AOEs (15.8 and 4.9 per 100 patient-years in years 1 and 5, respectively) did not increase over time. These final PACE results demonstrate ponatinib provides durable and clinically meaningful responses, irrespective of dose reductions, in this population of heavily pretreated CP-CML patients. This trial was registered at www.clinicaltrials.gov as #NCT01207440., (© 2018 by The American Society of Hematology.)

BCR-ABL1 kinase domain mutations can confer resistance to first- and second-generation tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia (CML). In preclinical studies, clinically achievable concentrations of the third-generation BCR-ABL1 TKI ponatinib inhibit T315I and all other single BCR-ABL1 mutants except T315M, which generates a single amino acid exchange, but requires 2 sequential nucleotide exchanges. In addition, certain compound mutants (containing ≥2 mutations in cis) confer resistance. Initial analyses based largely on conventional Sanger sequencing (SS) have suggested that the preclinical relationship between BCR-ABL1 mutation status and ponatinib efficacy is generally recapitulated in patients receiving therapy. Thus far, however, such analyses have been limited by the inability of SS to definitively identify compound mutations or mutations representing less than ~20% of total alleles (referred to as "low-level mutations"), as well as limited patient follow-up. Here we used next-generation sequencing (NGS) to define the baseline BCR-ABL1 mutation status of 267 heavily pretreated chronic phase (CP)-CML patients from the PACE trial, and used SS to identify clonally dominant mutants that may have developed on ponatinib therapy (30.1 months median follow-up). Durable cytogenetic and molecular responses were observed irrespective of baseline mutation status and included patients with compound mutations. No single or compound mutation was identified that consistently conferred primary and/or secondary resistance to ponatinib in CP-CML patients. Ponatinib is effective in CP-CML irrespective of baseline mutation status., (© 2016 by The American Society of Hematology.)

We studied the influence of comorbidities on remission rate and overall survival (OS) in patients with chronic myeloid leukemia (CML). Participants of the CML Study IV, a randomized 5-arm trial designed to optimize imatinib therapy, were analyzed for comorbidities at diagnosis using the Charlson Comorbidity Index (CCI); 511 indexed comorbidities were reported in 1519 CML patients. Age was an additional risk factor in 863 patients. Resulting CCI scores were as follows: CCI 2, n = 589; CCI 3 or 4, n = 599; CCI 5 or 6, n = 229; and CCI ≥ 7, n = 102. No differences in cumulative incidences of accelerated phase, blast crisis, or remission rates were observed between patients in the different CCI groups. Higher CCI was significantly associated with lower OS probabilities. The 8-year OS probabilities were 93.6%, 89.4%, 77.6%, and 46.4% for patients with CCI 2, 3 to 4, 5 to 6, and ≥7, respectively. In multivariate analysis, CCI was the most powerful predictor of OS, which was still valid after removal of its age-related components. Comorbidities have no impact on treatment success but do have a negative effect on OS, indicating that survival of patients with CML is determined more by comorbidities than by CML itself. OS may therefore be inappropriate as an outcome measure for specific CML treatments. The trial was registered at www.clinicaltrials.gov as #NCT00055874., (© 2015 by The American Society of Hematology.)

Advances in chronic myeloid leukemia treatment, particularly regarding tyrosine kinase inhibitors, mandate regular updating of concepts and management. A European LeukemiaNet expert panel reviewed prior and new studies to update recommendations made in 2009. We recommend as initial treatment imatinib, nilotinib, or dasatinib. Response is assessed with standardized real quantitative polymerase chain reaction and/or cytogenetics at 3, 6, and 12 months. BCR-ABL1 transcript levels ≤10% at 3 months, <1% at 6 months, and ≤0.1% from 12 months onward define optimal response, whereas >10% at 6 months and >1% from 12 months onward define failure, mandating a change in treatment. Similarly, partial cytogenetic response (PCyR) at 3 months and complete cytogenetic response (CCyR) from 6 months onward define optimal response, whereas no CyR (Philadelphia chromosome-positive [Ph+] >95%) at 3 months, less than PCyR at 6 months, and less than CCyR from 12 months onward define failure. Between optimal and failure, there is an intermediate warning zone requiring more frequent monitoring. Similar definitions are provided for response to second-line therapy. Specific recommendations are made for patients in the accelerated and blastic phases, and for allogeneic stem cell transplantation. Optimal responders should continue therapy indefinitely, with careful surveillance, or they can be enrolled in controlled studies of treatment discontinuation once a deeper molecular response is achieved.

Genomic instability is a hallmark of chronic myeloid leukemia in chronic phase (CML-CP) resulting in BCR-ABL1 mutations encoding resistance to tyrosine kinase inhibitors (TKIs) and/or additional chromosomal aberrations leading to disease relapse and/or malignant progression. TKI-naive and TKI-treated leukemia stem cells (LSCs) and leukemia progenitor cells (LPCs) accumulate high levels of reactive oxygen species (ROS) and oxidative DNA damage. To determine the role of TKI-refractory LSCs in genomic instability, we used a murine model of CML-CP where ROS-induced oxidative DNA damage was elevated in LSCs, including quiescent LSCs, but not in LPCs. ROS-induced oxidative DNA damage in LSCs caused clinically relevant genomic instability in CML-CP-like mice, such as TKI-resistant BCR-ABL1 mutations (E255K, T315I, H396P), deletions in Ikzf1 and Trp53, and additions in Zfp423 and Idh1. Despite inhibition of BCR-ABL1 kinase, imatinib did not downregulate ROS and oxidative DNA damage in TKI-refractory LSCs to the levels detected in normal cells, and CML-CP-like mice treated with imatinib continued to accumulate clinically relevant genetic aberrations. Inhibition of class I p21-activated protein kinases by IPA3 downregulated ROS in TKI-naive and TKI-treated LSCs. Altogether, we postulate that genomic instability may originate in the most primitive TKI-refractory LSCs in TKI-naive and TKI-treated patients.

Although the prognosis of chronic myeloid leukemia (CML) patients treated with imatinib is good, many fail to develop an optimal response or lose one. This heterogeneity could be attributed to the presence of human organic cation transporter-1 (hOCT1) single nucleotide polymorphisms (SNPs). In the present study, we analyzed the effect of 23 hOCT1 SNPs on imatinib treatment outcome in newly diagnosed CML patients using MassARRAY sequencing and pyrosequencing. The only SNP associated with outcome was M420del (rs35191146), with patients with the M420del demonstrating an increased probability of imatinib treatment failure. In CML cell lines transfected with M420del and/or M408V, M420del significantly decreased imatinib uptake, but this effect was countered if the M408V (rs628031) SNP was also present. A similar effect was seen for the uptake of the hOCT1 substrates TEA(+) and ASP(+). Finally, apparent hOCT1 mRNA levels were studied using both our earlier primers covering the M420del and another set that did not. Different mRNA expression was observed, explaining the disparity in published data on the prognostic importance of hOCT1 mRNA and highlighting the importance of avoiding common SNP sites in primer design. These data demonstrate that the common M420del SNP can modulate the outcome of imatinib treatment.

Molecular response to imatinib (IM) in chronic myeloid leukemia (CML) is associated with a biphasic but heterogeneous decline of BCR-ABL transcript levels. We analyzed this interindividual heterogeneity and provide a predictive mathematical model to prognosticate the long-term response and the individual risk of molecular relapse on treatment cessation. The parameters of the model were determined using 7-year follow-up data from a randomized clinical trial and validated by an independent dataset. Our model predicts that a subset of patients (14%) achieve complete leukemia eradication within less than 15 years and could therefore benefit from discontinuation of treatment. Furthermore, the model prognosticates that 31% of the patients will remain in deep molecular remission (MR(5.0)) after treatment cessation after a fixed period of 2 years in MR(5.0), whereas 69% are expected to relapse. As a major result, we propose a predictor that allows to assess the patient-specific risk of molecular relapse on treatment discontinuation and to identify patients for whom cessation of therapy would be an appropriate option. Application of the suggested rule for deciding about the time point of treatment cessation is predicted to result in a significant reduction in rate of molecular relapse.

The prognostic relevance of additional cytogenetic findings at diagnosis of chronic myeloid leukemia (CML) is unclear. The impact of additional cytogenetic findings at diagnosis on time to complete cytogenetic (CCR) and major molecular remission (MMR) and progression-free (PFS) and overall survival (OS) was analyzed using data from 1151 Philadelphia chromosome-positive (Ph(+)) CML patients randomized to the German CML Study IV. At diagnosis, 1003 of 1151 patients (87%) had standard t(9;22)(q34;q11) only, 69 patients (6.0%) had variant t(v;22), and 79 (6.9%) additional cytogenetic aberrations (ACAs). Of these, 38 patients (3.3%) lacked the Y chromosome (-Y) and 41 patients (3.6%) had ACAs except -Y; 16 of these (1.4%) were major route (second Philadelphia [Ph] chromosome, trisomy 8, isochromosome 17q, or trisomy 19) and 25 minor route (all other) ACAs. After a median observation time of 5.3 years for patients with t(9;22), t(v;22), -Y, minor- and major-route ACAs, the 5-year PFS was 90%, 81%, 88%, 96%, and 50%, and the 5-year OS was 92%, 87%, 91%, 96%, and 53%, respectively. In patients with major-route ACAs, the times to CCR and MMR were longer and PFS and OS were shorter (P < .001) than in patients with standard t(9;22). We conclude that major-route ACAs at diagnosis are associated with a negative impact on survival and signify progression to the accelerated phase and blast crisis.

T315I(+) Philadelphia chromosome-positive leukemias are inherently resistant to all licensed tyrosine kinase inhibitors, and therapeutic options remain limited. We report the outcome of allogeneic stem cell transplantation in 64 patients with documented BCR-ABL(T315I) mutations. Median follow-up was 52 months from mutation detection and 26 months from transplantation. At transplantation, 51.5% of patients with chronic myeloid leukemia were in the chronic phase and 4.5% were in advanced phases. Median overall survival after transplantation was 10.3 months (range 5.7 months to not reached [ie, still alive]) for those with chronic myeloid leukemia in the blast phase and 7.4 months (range 1.4 months to not reached [ie, still alive]) for those with Philadelphia chromosome-positive acute lymphoblastic leukemia but has not yet been reached for those in the chronic and accelerated phases of chronic myeloid leukemia. The occurrence of chronic GVHD had a positive impact on overall survival (P = .047). Transplant-related mortality rates were low. Multivariate analysis identified only blast phase at transplantation (hazard ratio 3.68, P = .0011) and unrelated stem cell donor (hazard ratio 2.98, P = .011) as unfavorable factors. We conclude that allogeneic stem cell transplantation represents a valuable therapeutic tool for eligible patients with BCR-ABL(T315I) mutation, a tool that may or may not be replaced by third-generation tyrosine kinase inhibitors.

Mutations in the Bcr-Abl kinase domain may cause, or contribute to, resistance to tyrosine kinase inhibitors (TKIs) in chronic myeloid leukemia patients. Recommendations aimed to rationalize the use of BCR-ABL mutation testing in chronic myeloid leukemia have been compiled by a panel of experts appointed by the European LeukemiaNet (ELN) and European Treatment and Outcome Study and are here reported. Based on a critical review of the literature and, whenever necessary, on panelists' experience, key issues were identified and discussed concerning: (1) when to perform mutation analysis, (2) how to perform it, and (3) how to translate results into clinical practice. In chronic phase patients receiving imatinib first-line, mutation analysis is recommended only in case of failure or suboptimal response according to the ELN criteria. In imatinib-resistant patients receiving an alternative TKI, mutation analysis is recommended in case of hematologic or cytogenetic failure as provisionally defined by the ELN. The recommended methodology is direct sequencing, although it may be preceded by screening with other techniques, such as denaturing-high performance liquid chromatography. In all the cases outlined within this abstract, a positive result is an indication for therapeutic change. Some specific mutations weigh on TKI selection.

Serial quantitation of BCR-ABL mRNA levels is an important indicator of therapeutic response for patients with chronic myelogenous leukemia and Philadelphia chromosome-positive acute lymphoblastic leukemia, but there is substantial variation in the real-time quantitative polymerase chain reaction methodologies used by different testing laboratories. To help improve the comparability of results between centers we sought to develop accredited reference reagents that are directly linked to the BCR-ABL international scale. After assessment of candidate cell lines, a reference material panel comprising 4 different dilution levels of freeze-dried preparations of K562 cells diluted in HL60 cells was prepared. After performance evaluation, the materials were assigned fixed percent BCR-ABL/control gene values according to the International Scale. A recommendation that the 4 materials be established as the first World Health Organization International Genetic Reference Panel for quantitation of BCR-ABL translocation by real-time quantitative polymerase chain reaction was approved by the Expert Committee on Biological Standardization of the World Health Organization in November 2009. We consider that the development of these reagents is a significant milestone in the standardization of this clinically important test, but because they are a limited resource we suggest that their availability is restricted to manufacturers of secondary reference materials.

This study examines the prognostic significance of early molecular response using an expanded dataset in chronic myeloid leukemia patients enrolled in the International Randomized Study of Interferon and STI571 (IRIS). Serial molecular studies demonstrate decreases in BCR-ABL transcripts over time. Analyses of event-free survival (EFS) and time to progression to accelerated phase/blast crisis (AP/BC) at 7 years were based on molecular responses using the international scale (IS) at 6-, 12-, and 18-month landmarks. Patients with BCR-ABL transcripts > 10% at 6 months and > 1% at 12 months had inferior EFS and higher rate of progression to AP/BC compared with all other molecular response groups. Conversely, patients who achieved major molecular response [MMR: BCR-ABL (IS) ≤ 0.1%] by 18 months enjoyed remarkably durable responses, with no progression to AP/BC and 95% EFS at 7 years. The probability of loss of complete cytogenetic response by 7 years was only 3% for patients in MMR at 18 months versus 26% for patients with complete cytogenetic response but not MMR (P < .001). This study shows a strong association between the degree to which BCR-ABL transcript numbers are reduced by therapy and long-term clinical outcome, supporting the use of time-dependent molecular measures to determine optimal response to therapy. This study is registered at www.clinicaltrials.gov as NCT00006343.

The role of allogeneic stem cell transplantation in chronic myeloid leukemia is being reevaluated. Whereas drug treatment has been shown to be superior in first-line treatment, data on allogeneic hematopoietic stem cell transplantation (allo SCT) as second-line therapy after imatinib failure are scarce. Using an interim safety analysis of the randomized German CML Study IV designed to optimize imatinib therapy by combination, dose escalation, and transplantation, we here report on 84 patients who underwent consecutive transplantation according to predefined criteria (low European Group for Blood and Marrow Transplantation [EBMT] score, imatinib failure, and advanced disease). Three-year survival after transplantation of 56 patients in chronic phase was 91% (median follow-up: 30 months). Transplantation-related mortality was 8%. In a matched pair comparison of patients who received a transplant and those who did not, survival was not different. Three-year survival after transplantation of 28 patients in advanced phase was 59%. Eighty-eight percent of patients who received a transplant achieved complete molecular remissions. We conclude that allo SCT could become the preferred second-line option after imatinib failure for suitable patients with a donor. The study is registered at the National Institutes of Health, http://clinicaltrials.gov: NCT00055874.

The BCR-ABL T315I mutation represents a major mechanism of resistance to tyrosine kinase inhibitors (TKIs). The objectives of this retrospective observational study were to estimate overall and progression-free survival for chronic myeloid leukemia in chronic-phase (CP), accelerated-phase (AP), or blastic-phase (BP) and Philadelphia chromosome-positive (Ph)(+) acute lymphoblastic leukemia (ALL) patients with T315I mutation. Medical records of 222 patients from 9 countries were reviewed; data were analyzed using log-rank tests and Cox proportional hazard models. Median age at T315I mutation detection was 54 years; 57% cases were men. Median time between TKI treatment initiation and T315I mutation detection was 29.2, 15.4, 5.8, and 9.1 months, respectively, for CP, AP, BP, and Ph(+) ALL patients. After T315I mutation detection, second-generation TKIs were used in 56% of cases, hydroxyurea in 39%, imatinib in 35%, cytarabine in 26%, MK-0457 in 11%, stem cell transplantation in 17%, and interferon-alpha in 6% of cases. Median overall survival from T315I mutation detection was 22.4, 28.4, 4.0, and 4.9 months, and median progression-free survival was 11.5, 22.2, 1.8, and 2.5 months, respectively, for CP, AP, BP, and Ph(+) ALL patients. These results confirm that survival of patients harboring a T315I mutation is dependent on disease phase at the time of mutation detection.

Dasatinib is a BCR-ABL inhibitor with 325-fold higher potency than imatinib against unmutated BCR-ABL in vitro. Imatinib failure is commonly caused by BCR-ABL mutations. Here, dasatinib efficacy was analyzed in patients recruited to phase 2/3 trials with chronic-phase chronic myeloid leukemia with or without BCR-ABL mutations after prior imatinib. Among 1043 patients, 39% had a preexisting BCR-ABL mutation, including 48% of 805 patients with imatinib resistance or suboptimal response. Sixty-threedifferent BCR-ABL mutations affecting 49 amino acids were detected at baseline, with G250, M351, M244, and F359 most frequently affected. After 2 years of follow-up, dasatinib treatment of imatinib-resistant patients with or without a mutation resulted in notable response rates (complete cytogenetic response: 43% vs 47%) and durable progression-free survival (70% vs 80%). High response rates were achieved with different mutations except T315I, including highly imatinib-resistant mutations in the P-loop region. Impaired responses were observed with some mutations with a dasatinib median inhibitory concentration (IC(50)) greater than 3nM; among patients with mutations with lower or unknown IC(50), efficacy was comparable with those with no mutation. Overall, dasatinib has durable efficacy in patients with or without BCR-ABL mutations. All trials were registered at http://www.clinicaltrials.gov as NCT00123474, NCT00101660, and NCT00103844.

A minority of patients treated with imatinib are either refractory to imatinib or eventually relapse. Relapse frequently depends on re-emergence of BCR-ABL kinase activity but may also indicate BCR-ABL-independent disease progression. Over 90 point mutations coding for single amino acid substitutions in the BCR-ABL kinase domain have been isolated from CML patients resistant to imatinib treatment. These mutations affect amino acids involved in imatinib binding or in regulatory regions of the BCR-ABL kinase domain, resulting in decreased sensitivity to imatinib while retaining aberrant kinase activity. The early detection of BCR-ABL mutants during therapy may aid in risk stratification as well as molecular-based treatment decisions. Therapeutic strategies of imatinib resistant disease include novel tyrosine kinase inhibitors with activity against imatinib-resistant mutations and/or with inhibition of alternative pathways, dose escalation to optimise imatinib levels, treatment interruption to stop selection of resistant cells and allogeneic stem cell transplantation in eligible patients.

Dasatinib is the most potent BCR-ABL inhibitor, with 325-fold higher potency than imatinib against unmutated BCR-ABL in vitro. Studies have demonstrated the benefits of dasatinib 70 mg twice daily in patients with accelerated-phase chronic myeloid leukemia intolerant or resistant to imatinib. A phase 3 study compared the efficacy and safety of dasatinib 140 mg once daily with the current twice-daily regimen. Here, results from the subgroup with accelerated-phase chronic myeloid leukemia (n = 317) with a median follow-up of 15 months (treatment duration, 0.03-31.15 months) are reported. Among patients randomized to once-daily (n = 158) or twice-daily (n = 159) treatment, rates of major hematologic and cytogenetic responses were comparable (major hematologic response, 66% vs 68%; major cytogenetic response, 39% vs 43%, respectively). Estimated progression-free survival rates at 24 months were 51% and 55%, whereas overall survival rates were 63% versus 72%. Once-daily treatment was associated with an improved safety profile. In particular, significantly fewer patients in the once-daily group experienced a pleural effusion (all grades, 20% vs 39% P < .001). These results demonstrate that dasatinib 140 mg once daily has similar efficacy to dasatinib 70 mg twice daily but with an improved safety profile. This trial is registered at www.clinicaltrials.gov as #CA180-035.

An international basis for comparison of BCR-ABL mRNA levels is required for the common interpretation of data derived from individual laboratories. This will aid clinical decisions for individual patients with chronic myeloid leukemia (CML) and assist interpretation of results from clinical studies. We aligned BCR-ABL values generated by 38 laboratories to an international scale (IS) where a major molecular response (MMR) is 0.1% or less. Alignment was achieved by application of laboratory-specific conversion factors calculated by comparisons performed with patient samples against a reference method. A validation procedure was completed for 19 methods. We determined performance characteristics (bias and precision) for consistent interpretation of MMR after IS conversion. When methods achieved an average BCR-ABL difference of plus or minus 1.2-fold from the reference method and 95% limits of agreement within plus or minus 5-fold, the MMR concordance was 91%. These criteria were met by 58% of methods. When not met, the MMR concordance was 74% or less. However, irrespective of precision, when the bias was plus or minus 1.2-fold as achieved by 89% of methods, there was good agreement between the overall MMR rates. This indicates that the IS can deliver accurate comparison of molecular response rates between clinical trials when measured by different laboratories.

Chronic myeloid leukemia (CML) is a clonal disease of hematopoietic stem cells caused by a reciprocal translocation of the long arms of chromosomes 9 and 22. In human leukocyte antigen A*0201(+) (HLA-A*0201(+)) individuals, response after interferon-alpha (IFN-alpha) was shown to be associated with the emergence of CML-specific cytotoxic T cells that recognize PR-1, a myeloblastin (MBN)-derived nonapeptide. In contrast, imatinib potently induces remissions from CML by specific inhibition of the ABL tyrosine kinase. Here, we explored molecular regulations associated with CML responses under different treatment forms using cDNA-array. Expression of MBN was found to be down-regulated in remission under imatinib therapy (0 of 7 MBN(+) patients). In contrast, MBN transcription was readily detectable in the peripheral blood in 8 of 8 tested IFN-alpha patients in complete remission (P =.0002). IFN-alpha-dependent MBN transcription was confirmed in vitro by stimulation of peripheral blood mononuclear cells (PBMCs) with IFN-alpha and by IFN-alpha-mediated activation of the MBN promoter in reporter gene assays. Finally, with the use of HLA-A*0201-restricted, MBN-specific tetrameric complexes, it was demonstrated that all of 4 IFN-alpha-treated patients (100%), but only 2 of 11 imatinib patients (19%), in complete hematological or cytogenetic remission developed MBN-specific cytotoxic T cells (P =.011). Together, the induction of MBN expression by IFN-alpha, but not imatinib, may contribute to the specific ability of IFN-alpha to induce an MBN-specific T-cell response in CML patients. This also implies that the character of remissions achieved with either drug may not be equivalent and therefore a therapy modality combining IFN-alpha and imatinib may be most effective.