Imatinib In Chronic Myeloid Leukemia Patients Research Articles

Outcomes of Dose Escalation of Imatinib in Chronic Myeloid Leukemia Patients: A Retrospective Analysis From an Indian University Teaching Hospital.

Chronic myeloid leukemia (CML) treatment in low- and middle-income countries faces significant financial and logistical constraints. In scenarios where second-line tyrosine kinase inhibitors (TKIs) are unavailable or unaffordable, dose escalation of imatinib provides an alternative. This study evaluates the efficacy, safety, and progression-free survival (PFS) outcomes of dose escalation of imatinib in CML patients who experienced suboptimal response or progression on standard doses. A retrospective analysis of 123 CML patients treated at an Indian university teaching hospital from 2013 to 2016 was conducted. Patients who showed progression on a 400 mg dose of imatinib were escalated to 600 mg, and further to 800 mg if required. Demographic data, progression, and toxicity were analyzed. Out of 123 patients, 78 (63.4%) showed a complete hematologic response after dose escalation. The median PFS was 48 months, with a three-year PFS rate of 67%. Notable toxicities included Grade 3/4 neutropenia in 15% and gastrointestinal disturbances in 12%. Comparatively, studies suggest that switching to a second-line TKI in similar settings results in a higher PFS; however, our findings underscore that dose escalation of imatinib remains a viable alternative when financial constraints limit access to second-line therapies. In resource-constrained settings, dose escalation of imatinib can be an effective strategy for managing CML patients who progress on standard doses.

Analysis of factors predicting the efficacy of Imatinib in patients with Chronic Myeloid Leukemia: A retrospective analysis

Objective: Imatinib is a commonly used first generation tyrosine kinase inhibitor for patients with chronic myeloid leukemia (CML). The efficacy has been reported as very high even in recent studies. Material and methods: A retrospective analysis was made of newly diagnosed CML patients treated with Imatinib as a first-line agent from January 2010 to January 2020. The patients were classified as those who obtained an adequate response and those for whom treatment was discontinued due to inadequate efficacy. The two groups were compared to analyze factors predicting the efficacy of the agent. Results: Evaluation was made of a total of 47 CML patients, comprising 20 females (42.6%) and 27 males (57.4%) with a median age of 55 years. Imatinib was discontinued in 19 patients because of inadequate response, and 28 patients were still continuing the treatment at the end of median 33.3 months follow-up duration. At the end of follow-up, there were 44 survivors (93.6%), and 3 non-survivors (6.4%). Median Bcr-Abl (IS, %) at the time of diagnosis in patients with response was higher than patients in discontinued group (67.6 [0.0-291.4] vs 41.9 [0.0-208.5], p=0.022). All other disease and demographic characteristics were similar in both groups (p>0.05). Conclusion: Almost 10 years of follow-up demonstrated that there is still an unmet need to determine factors predicting the response to Imatinib in CML patients. Larger population-based studies are required to specify patients with high risk at the time of diagnosis to monitor closely.

Concomitant Medications and Comorbidities Affect Early Response in CML Patients Treated with Imatinib: A Retrospective Analysis of 340 Patients from the Polish Adult Leukemia Group (PALG) Registry

Background : It has been consistently shown that early molecular response is associated with positive long-term outcome in terms of overall survival and progression-free survival in chronic myeloid leukemia (CML) patients treated with tyrosine kinase inhibitors (TKIs). Achieving a BCR-ABL(IS) <10% at three months has thereby become an important goal of treatment. According to current European LeukemiaNet recommendations, the optimal treatment response is defined as BCR-ABL(IS) ≤ 10% at three months and BCR-ABL(IS)<1% and/or complete cytogenetic response (CCyR) at six months. Factors influencing the early response are still largely unknown.Our previous study demonstrated that more than half of newly diagnosed CML patients in Poland had comorbid conditions and received concomitant medications. TKIs are metabolized by the CYP3A4 isoenzyme and have been reported to be substrates of ABCB1 (MDR1) and ABCG2 (BCRP) transporters. Importantly, numerous commonly used drugs are transported and metabolized through the same pathways as TKIs. Drug interactions should therefore be considered when administering inhibitors or inducers of CYP3A4, ABCB1 and ABCG2 in combination with TKIs. Furthermore, it has been shown that the presence of comorbidities and the necessity of additional therapies may significantly affect treatment compliance.Objectives: The primary objective of the study was to investigate whether the presence of comorbidities and additional therapies affected early response in CML patients treated with imatinib.Patients and methods : We retrospectively analyzed patients with newly diagnosed CML who were receiving imatinib as the initial therapy for CML at nine Polish hematological tertiary care centers. Inclusion criteria were as follows: 1) age ≥18 years and 2) diagnosis of chronic phase CML between January 1st 2005 and December 31st 2014. Data on presence of comorbid conditions, concomitant drugs, cytogenetic and molecular response were anonymously collected through electronic case reports forms. Patients in whom therapy was changed because of imatinib intolerance were excluded. Influence of potential prognostic factors on probability of response at three and six months was tested by univariate and multivariate logistic regression.Results : We enrolled 340 patients with CML in the chronic phase, with median age 53 years (range, 41-62) where 50 % were males. According to the Sokal score, 190 patients (55.9%) were classified as low risk, 107 (31.5%) as intermediate risk and 43 (12.5%) as high risk. Overall, 142 (41.8%) patients reported no comorbidities, 121 (35.6%) reported one or two comorbidities and 77 (22.7%) had more than two comorbid conditions. The most common comorbidities were: hypertension (30%), hypercholesterolemia (16%), ischemic heart disease (10%) and diabetes (9%). One hundred thirty (38.2%) patients received no additional therapies, 110 (32.4%) patients received one or two therapies and 100 patients (29.4%) more than two. The most frequently used drugs were: beta blockers (22% of patients), ACE inhibitors (19%) and statins (15%). At three months, a molecular analysis was done in 194 patients and 144 (74%) achieved BCR ABL(IS) of less than 10%. At six months, 217 patients underwent molecular analysis and 305 had cytogenetic assessment. One hundred thirty five patients (62%) achieved BCR-ABL(IS) < 1% and 198 (65%) achieved CCyR. Multivariate analysis revealed that at three months patients who had a low Sokal score, more than two comorbidities and received more than two concomitant therapies, significantly more often achieved BCR-ABL <1% (p=0.013, p=0.054 and p=0.005 respectively). At six months, patients who received more than two concomitant medications still tended to have a higher probability of optimal response: BCR-ABL(IS) <1% (p=0.062) and CCyR (p=0.035). Other potential prognostic factors analyzed including patients' age, sex and BMI had no impact on the probability of optimal response.Conclusions: We found that comorbidities and concomitant therapies did not adversely affect early response to imatinib in CML patients. In contrast, patients with more than two additional therapies and comorbidities seem to more often achieve treatment goals. The potential explanations for these findings are drug interactions that increase imatinib exposure and better adherence to therapy of patients accustomed to systematic use of medications. DisclosuresGora Tybor:BMS: Honoraria, Speakers Bureau; Incyte: Honoraria, Speakers Bureau; Pfizer: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau. Sacha:Incyte: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Pfizer: Honoraria, Speakers Bureau; BMS: Honoraria, Speakers Bureau. Lewandowski:BMS: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Incyte: Honoraria, Speakers Bureau; Pfizer: Honoraria, Speakers Bureau. Warzocha:BMS: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau. Jamroziak:Janssen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Celgene: Honoraria, Speakers Bureau; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Takeda: Honoraria, Membership on an entity's Board of Directors or advisory committees; Abbvie: Honoraria.

Influence of genetic polymorphisms in glutathione-S-transferases gene in response to imatinib among Brazilian patients with chronic myeloid leukemia

Polymorphism in metabolizing enzymes can influence drug response as well as the risk for adverse drug reactions. Nevertheless, there are still few studies analyzing the consequence of polymorphisms for the Glutathione-S-transferases (GST) gene to drug response in chronic myeloid leukemia (CML). This study reports, the influence of GSTP1*B and GSTT1/GSTM1null polymorphisms in response to imatinib in CML patients in a Brazilian population. One hundred thirty-nine CML patients from the Clinical Hospital of Goiânia, Goiás, Brazil, treated with imatinib were enrolled in this study. Genotyping of GSTT1 and GSTM1 genes deletions were performed by qPCR and of GSTP1 gene was performed by RFLP-PCR. The frequency of GSTP1*1B, GSTT1 and GSTM1null polymorphisms were determined for all patients. The influence of each patient's genotypes was analyzed with the patient's response to imatinib treatment. Brazilian CML patients revealed GSTT1 and GSTM1 genes deletions. GSTT1 deletion was found in 19.3% of patients and GSTM1 deletion in 48.7% of patients with CML. GSTT1/GSTM1 deletion was found in 11.7% in Brazilian CML patients. The "G allele" of GSTP1*B, is associated with later cytogenetic response in imatinib therapy. While, the gene presence combined with GG genotype (GSTM1 present/GSTPI-GG) conferred a tend to a later cytogenetic response to patients. GSTP1*B and GSTT1/GSTM1null polymorphisms influence treatment response in CML. Brazilian CML patients presenting GSTP1 AA/AG genotypes alone and in combination with GSTT1 null reach the cytogenetic response faster, while patients presenting GSTP1-GG and GSTMI positive genotypes may take longer to achieve cytogenetic response. As a result, it allows a better prognosis, with the use of an alternative therapy, other than reducing treatment cost.

Lack of Association between C1236T, G2677T/A and C3435T Variants of the ABCB1 Gene and Imatinib Response in Iranian Chronic Myeloid Leukemia Patients

Imatinib introduction caused to improve the clinical outcomes of chronic myeloid leukemia (CML) patients. Despite the significant effects of Imatinib, pharmacogenetic variables induced treatment resistant is also observed.. Imatinib is known as a P-glycoprotein (P-gp) efflux pump substrate encoded by the ABCB1 gene. The ABCB1 C1236T, G2677T/A and C3435T variants are possibly correlated with interindividual variation in pharmacokinetic response to Imatinib therapy. The present study aimed to examine the effect of ABCB1 gene variants on the therapeutic response of Imatinib in CML patients. Sixty-nine Iranian CML patients treated with Imatinib or Nilotinib were selected and divided into two groups of sensitive and resistant to Imatinib. C1236T and G2677T/A variants were genotyped by high resolution melting (HRM) analysis, and C3435T variant was genotyped using polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP). Then, the results were compared between the two groups of patients. Our results showed that there were no significant differences between C1236T, G2677T/A and C3435T variants of ABCB1 gene and clinical response to Imatinib in the Iranian CML patients. According to the results of this study, genotyping of ABCB1 C1236T, G2677T/A and C3435T variants couldn’t help to predict the outcomes of Imatinib treatment in CML patients. So, these variants are not useful to make decisions about treatment, but it is suggested to do further investigations.

Use of generic imatinib as first-line treatment in patients with chronic myeloid leukemia (CML): the GIMS (Glivec to Imatinib Switch) study.

BackgroundGeneric formulations of imatinib mesylate have been introduced in Western Europe since 2017 to treat patients with chronic myeloid leukemia (CML). However, results on the safety and efficacy of generic formulations are contrasting. The aim of this study was to investigate the safety and efficacy of generic imatinib in CML patients treated in 12 Italian institutes.MethodsThis is an observational, retro-prospective analysis of patients with CML for whom the treatment was switched from brand to generic imatinib. We analyzed and compared the variation in quantitative PCR values before and after the switch, and the proportion of patients who maintained molecular response after changing from brand to generic imatinib. Adverse events (AEs) were also evaluated.ResultsTwo hundred patients were enrolled. The median PCR value after the switch was reduced by 0.25 compared to the values before the switch. A significant difference was found between median PCR values before and after the switch in favor of generic imatinib (P= 0.003). Molecular responses remained stable in 69.0%, improved in 25.5%, and worsened in 5.5% of patients. AEs were similar in the pre- and post-switch periods; however, a significant difference was found in favor of generic imatinib for muscular cramps (P< 0.0001), periorbital edema (P=0.0028), edema of the limbs (P<0.0001), fatigue (P=0.0482), and diarrhea (P=0.0027).ConclusionOur data indicate that generic imatinib does not have deleterious effects on CML control and present an acceptable safety profile, similar or better than brand imatinib.

Tumor suppressor p53 gene codon 72 polymorphism and imatinib cytogenetic response in chronic myeloid leukemia

Objective To study p53 codon 72 polymorphism in relation to cytogenetic response to imatinib treatment in patients with chronic myeloid leukemia (CML). Background P53 polymorphism involves the substitution of an arginine for a proline at codon position 72. Many studies have investigated a genetic link between this variation and response to treatment in cancer. Patients and methods This study was conducted on 54 CML patients presented to the Clinical Oncology Department, Menoufia University during the period from June 2013 to April 2015. They were classified according to their cytogenetic response to imatinib therapy into 40 CML patients, cytogenetic responders to imatinib and 14 CML patients who are cytogenetic nonresponders to imatinib. Patients were genotyped for p53 codon 72 polymorphism using PCR. Follow up of the patients should be done after 3, 6, 9, 12, and 18 months after diagnosis, and was done by complete blood count, conventional cytogenetic, and fluorescence in-situ hybridization. Results Age, sex, hematologic, and cytogenetic response to imatinib in CML patients did not differ significantly among p53 codon 72 genotypes (arg/arg, arg/pro, and pro/pro) (P = 0.44, P = 0.45, and P = 0.11, respectively). P53 codon 72 polymorphism did not significantly alter the risk to imatinib cytogenetic unresponsiveness (P = 0.9221). Conclusion It could be concluded that p53 codon 72 polymorphism is not associated with imatinib unresponsiveness in CML.

The efficacy of generic imatinib in patients with chronic myeloid leukemia - a single centre experience

Background/Aim. The treatment of chronic myeloid leukemia (CML) has changed dramatically with the advent of targeted therapies. This study aimed to assess the efficacy of generic imatinib in CML patients treated in our center. Methods. The study was retrospective. It included 101 patients diagnosed with CML ? chronic phase (CP). The patients were divided into two groups. Group 1 included 55 patients initially treated with branded imatinib and then switched to generic imatinib. Group 2 consisted of 46 newly diagnosed patients who received only generic imatinib from the beginning of therapy. Results. The patients were treated with branded imatinib for the mean of 42 months (range 6?132 months) before switching to generic imatinib. Treatment with generic imatinib lasted for 25 months on average (range 3?66 months). A quarter of the patients from the group 1 lost their cytogenetic reponse after being switched to generic imatinib, but with-out signs of transformation to acute leukemia. The patients treated with branded imatinib had a significantly longer event-free survival (EFS) and failure-free survival (FFS) (log-rank p = 0.01 and p = 0.03, respectively). These results could have been influenced by frequent changes of the brand and dosage formulation of generic imatinib. Conclusions. Our study showed a significantly longer EFS and FFS in the patients who were initially treated with branded imatinib, compared to those treated with generic imatinib only. These results provide useful information, but have to be interpreted within the context of the crossover study.

Low ABCB1 and high OCT1 levels play a favorable role in the molecular response to imatinib in CML patients in the community clinical practice

Despite the favorable clinical evolution of patients with chronic myeloid leukemia (CML), resistance or intolerance to imatinib is present in approximately 35% of patients. Sokal score is a widely used risk factor, however efflux and influx transporters are provisional risk factors implicated in imatinib resistance. This study analyzed Sokal score, ABCB1, ABCG2 and OCT1 mRNA transporter expression levels as well as P-glycoprotein expression and efflux transporters activity to seek a possible correlation between these factors and the molecular response at 12 months from imatinib start as well as 8-year overall survival (OS). Low plus intermediate Sokal score correlated to optimal imatinib responses, as well as OS at 8-years, thus confirming the established role of Sokal score as a prognostic factor in CML patients. Low ABCB1 and high OCT1 mRNA levels were associated with an optimal molecular response, while the inverse levels were associated with non-responders (warning and failure) patients. Our results suggest that ABCB1 and OCT1 mRNA expressions may present biological relevance to identify responder and non-responder patients to imatinib treatment.

PBA2, a novel inhibitor of imatinib-resistant BCR-ABL T315I mutation in chronic myeloid leukemia

Chronic Myeloid Leukemia (CML) is largely caused by the Philadelphia (Ph) chromosome carrying the Break point Cluster Region-Abelson (BCR-ABL) oncogene. Imatinib is a BCR-ABL-targeted therapy and considered the standard of care in CML management. Resistance to imatinib therapy often develops because of mutations in the BCR-ABL kinase domain. In this study, we evaluated PBA2, a novel BCR-ABL inhibitor, for its anti-cancer activity against BCR-ABL expressing BaF3 cells. PBA2 shows potent activity against wild-type and T315I mutated BaF3 cells as compared with imatinib. PBA2 inhibited the phosphorylation of BCR-ABL and its downstream signaling in BaF3/WT and BaF3/T315I cells. PBA2 inhibited the mRNA expression of BCR-ABL in BaF3/WT and BaF3/T315I cells. Mechanistically, PBA2 increased the cell population in sub G1 phase of the cell cycle, induced apoptosis and elevated ROS production in both BaF3/WT and BaF3/T315I cells. Taken together, our results indicate that PBA2 exhibits anti-proliferative effects and inhibits the imatinib-resistant T315I BCR-ABL mutation. PBA2 may be a novel drug candidate for overcoming the resistance to imatinib in CML patients.

MATE1 regulates cellular uptake and sensitivity to imatinib in CML patients.

Although imatinib is highly effective in the treatment of chronic myeloid leukemia (CML), 25–30% patients do not respond or relapse after initial response. Imatinib uptake into targeted cells is crucial for its molecular response and clinical effectiveness. The organic cation transporter 1 (OCT1) has been proposed to be responsible for this process, but its relevance has been discussed controversially in recent times. Here we found that the multidrug and toxin extrusion protein 1 (MATE1) transports imatinib with a manifold higher affinity. MATE1 mainly mediates the cellular uptake of imatinib into targeted cells and thereby controls the intracellular effectiveness of imatinib. Importantly, MATE1 but not OCT1 expression is reduced in total bone marrow cells of imatinib-non-responding CML patients compared with imatinib-responding patients, indicating that MATE1 but not OCT1 determines the therapeutic success of imatinib. We thus propose that imatinib non-responders could be identified early before starting therapy by measuring MATE1 expression levels.

Coexistence of P190 and P210 BCR/ABL transcripts in chronic myeloid leukemia blast crisis resistant to imatinib.

IntroductionPhiladelphia chromosome (Ph) is a hallmark of chronic myeloid leukemia (CML), which exists in more than 90% CML and in 3% to 40% acute lymphoblastic leukemia (ALL).Case descriptionA 25-year-old man was diagnosed with CML in chronic phase. He first received treatment with hydroxyurea, achieving hematological remission and following imatinib mesylate for main treatment. A year later, he began to appear unexplained high fever with ineffective antibiotic treatment and bone morrow and blood tests indicated blast crisis. Both BCR/ABL 210 and BCR/ABL 190 fusion transcript were positive. Imatinib resistance was confirmed by a screening for ABL kinase domain E255K mutations, and dasatinib was administered. After two months, the patient went on to hematological remission.Discussion and evaluationDuring medical treatment for CML, we experienced a relatively rare case with co-expression of the p210 and p190 encoding BCR-ABL transcripts in blastic phase. Imatinib resistance was confirmed and remission wasn’t easily obtained, yet dasatinib was helpful. When resistance emerges, the treatment options include increasing the daily dose of imatinib, or combining imatinib with other agents. Of course, dasatinib, nilotinib and bone marrow transplantation are good choice as well.ConclusionsThe presence of p-190 transcript in CML may be related to progression of the disease. Thus monitoring the resistance of imatinib in CML patients, especially for advanced phase CML and BCR-ABL ALL, may be meaningful to guide clinical treatment and predict the prognosis.

ABCB1 polymorphisms predict imatinib response in chronic myeloid leukemia patients: a systematic review and meta-analysis.

Imatinib mesylate, a competitive tyrosine kinase inhibitor, is considered the first-line therapy drug for Ph+ chronic myeloid leukemia (CML). Three single-nucleotide polymorphisms (SNPs) in the ATP-binding cassette, subfamily B (MDR/TAP), member 1 gene (ABCB1/MDR1), c.1236C>T, c.2677G>T/A and c.3435C>T, have been shown to affect cellular transport/metabolism of imatinib. The associations between these SNPs and imatinib response in CML patients have been widely evaluated, but the results were inconsistent. To derive a conclusive assessment of the associations, we performed a meta-analysis by combining data from a total of 12 reports including 1826 patients. The results showed that the 2677G allele or 3435T allele predicted a worse response to imatinib in CML patients, whereas 1236CC genotype was associated with better response in CML patients from Asian region. In conclusion, this meta-analysis suggests that c.1236C>T, c.2677G>T/A and c.3435C>T can be served as predictive markers for the therapeutical use of imatinib in CML patients.

Functional polymorphism of CYP2B6 G15631T is associated with hematologic and cytogenetic response in chronic myeloid leukemia patients treated with imatinib

In the spite of the impressive results achieved with imatinib in chronic myeloid leukemia (CML) patients, differences in patient's response are observed, which may be explained by interindividual genetic variability. It is known that cytochrome P450 enzymes play a major role in the metabolism of imatinib. The present study aimed to understand the functional impact of CYP2B6 15631G>T polymorphism on the response of imatinib in CML patients and its relation to CML susceptibility. We have genotyped CYP2B6 G15631T in 48 CML patients and 64 controls by PCR-RFLP. CYP2B6 15631G>T was not found to be a risk factor for CML (OR 95 % CI, 1.12, 0.6-2, p > 0.05). Hematologic response loss was higher in patients with 15631GG/TT genotype when compared with 15631GT (36.8 vs. 13.8 %; X (2) = 3.542, p = 0.063). Complete cytogenetic response was higher in 15631GG/GT genotype groups when compared with 15631TT (X (2) = 3.298, p = 0.024). Primary cytogenetic resistance was higher in patients carrying 15631GG/TT genotype when compared with 15631GT carriers (52.6 vs. 17.2 %; X (2) = 6.692, p = 0.010). Furthermore, side effects were more common for patients carrying 15631GG genotypes when compared with GT/TT carriers (36 vs. 13.8 %; X (2) = 8.3, p = 0.004). In light of our results, identification of 15631G>T polymorphism in CML patients might be helpful to predict therapeutic response to imatinib.

Chronic myeloid leukemia and imatinib: Experience at the Lome Campus teaching hospital (Togo)

Chronic myeloid leukemia (CML) is a clonal malignant myeloproliferative disorder characterized by the expansion of hematopoietic cells carrying the Philadelphia chromosome (t 9.22). Our main objective was to assess the efficacy of imatinib in CML patients, measured by their survival. Over a six-year period (June 2003 through May 2009), 25 patients were seen regularly for CML at the Lomé Campus teaching hospital. Patients received imatinib after diagnosis and underwent regular laboratory monitoring (quantification of BCR-ABL ratio by RT-PCR). Patients' survival and treatment response were evaluated. Patients' mean age at diagnosis was 40 years (range: 9 to 72 years). Men predominated (17 compared with 7 women). Splenomegaly was found in 80% of cases. The mean leukocyte level was 188.71 g/L (24.4-350). Six patients (24%) had thrombocytosis with a mean platelet count of 491.15 g/L (108-2000). Six patients (24%) died after developing accelerated-phase CML or blast crisis. Estimated overall survival of patients at 6 years was 60%. Molecular biology monitoring detected a secondary G250E mutation with resistance to imatinib in one patient. Standard hematological side effects led to reduction in imatinib doses. The principal nonhematological side effects were weight gain and transient digestible disorders. At six years after diagnosis, imatinib was effective in treating patients with CML, even in sub-Saharan Africa. Mutation-induced resistance required regular molecular biological monitoring to determine the need to switch to later-generation tyrosine kinase inhibitors.

Abstract 814: P-glycoprotein transport of the active imatinib metabolite, CGP74588, in chronic myeloid leukemia cells

Abstract The tyrosine kinase inhibitor (TKI) imatinib is, despite the introduction of second generation TKI's, the standard first-line therapy in chronic myeloid leukemia (CML). Early response to therapy is correlated to long-term therapeutic effect in CML. To be able to switch to second generation TKI's in an early stage of CML, it would be advantageous to identify predictive markers for failure on imatinib. Imatinib is metabolized by hepatic CYP3A4 and CYP3A5, forming approximately 30 metabolites. The main metabolite, CGP74588, is pharmacologically active with similar potency to that of imatinib. CGP74588 is present in ∼20% of imatinib plasma concentrations but with a large inter-individual variation. We have previously reported that high CYP3A activity is associated with a better therapeutic outcome of imatinib, indicating a clinical significance of imatinib metabolites (Gréen et al. 2010). Furthermore, imatinib is a substrate for the efflux transporter P-glycoprotein which is the product of the ABCB1 gene. Several ABCB1 polymorphisms have been described and some have been shown to influence the therapeutic response to imatinib in CML patients. The aim of this study was to investigate the effects of ABCB1 over-expression on the in vitro resistance to imatinib and its CYP3A metabolite CGP74588. ABCB1 wild type human cDNA was infected to the CML cell line K562 using a retroviral system. Co-expression of ABCB1 and the reporter gene for enhanced yellow fluorescent protein (EYFP) was used for the assessment of ABCB1 expression by the analysis of EYFP in FACS. The influence of ABCB1 expression on the cytotoxic effects of imatinib and CGP74588 was assessed using MTT assays. FACS analysis of EYFP confirmed an over-expression of ABCB1 in infected K562 cells with a mean fluorescence intensity of 23.5 compared to 0.55 in parental K562 cells. ABCB1-expressing cells (K562/ABCB1) were slightly, but not significantly, more resistance to imatinib than K562 cells (IC50 K562 = 0.41µM, K562/ABCB1 = 0.50µM). However, ABCB1 expression induced a 12-fold increase in resistance when cells were treated with the imatinib metabolite CGP74588 (IC50 K562 = 0.72µM, K562/ABCB1 = 8.53µM, p = &amp;lt;0.000). Recent reports have shown that CGP74588 accumulates in cell lines with acquired multi-drug resistance and high P-gp expression. Our studies on cells with ABCB1 expression as a single resistance mechanism proves that P-gp is indeed confer resistance to CGP74588 in CML cells. Furthermore, CGP74588 cytotoxicity is affected by ABCB1 expression to a greater extent than imatinib itself, indicating that ABCB1 activity could be important for CGP74588 plasma and target cell concentrations in vivo and might contribute to the large intra-individual variation seen in patient plasma concentrations. Future studies will be needed to determine the clinical significance of P-gp activity and CGP74588 pharmacokinetics in relation to TKI therapy of CML. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 814. doi:1538-7445.AM2012-814

Monitoring Response to Imatinib by Fluorescence In Situ Hybridization (FISH) and Real-Time Quantitative Polymerase Chain Reaction (RQ-PCR) in Chronic Myeloid Leukemia (CML) Patients in Chronic Phase. Experience of Argentina and Uruguay.

Introduction: The evaluation of response to imatinib in CML patients (pts) as guide to clinical management is undergoing substantial changes of criteria. The degree of reduction of total leukemia cell mass by imatinib as measured by FISH and RQ-PCR studies, correlates with progression-free survival. Therapeutic decisions, as dose scalation, is mandatory in pts with rising level of bcr/abl transcripts.Purpose: To determine the potential of RQ-PCR to predict the duration of response in pts in complete cytogenetic remission (CCyR).Patients and Methods: A total of 160 CML pts in first chronic phase, in CCyR treated with imatinib 400mg daily were studied prospectively since November 2005 (147 pts from 30 hematology centers in Argentina and 13 pts from 2 centers in Uruguay). According to previous treatment they were divided in two groups: 83 pts (51%) received as first line treatment IFN/Cytarabine and 77 pts (49%) received imatinib as first line. According to Sokal Index, 72% were low risk, 19% intermediate risk and 9% high risk. At baseline all pts were first evaluated by FISH and RQ-PCR under EAC program protocol and recommendations of T.P. Hughes, et al (Blood 108:28–37, 2006). Only RQ-PCR is being performed at 6 and 18 months from the beginning of the study. The median duration since beginning with imatinib treatment was 28 months (range 6–58).Results: At the time of first RQ-PCR evaluation, 93% had undetectable bcr/abl fusion gene by FISH and 7% had low level FISH positivity ( 0,1 – 5%). The distribution of the type of molecular responses (MR) in the 160 pts was: Complete (CMR)≥4 log red, U/<0.01% bcr-abl/c-abl : 23%; Major (MaMR) ≥ 3log red, <0,1%bcr-abl/c-abl : 17%; Minor (MiMR) >2 log red, 1–0,1 % bcr-abl/c-abl : 34%; MiMR < 2log red, 10–1% bcr-abl/c-abl : 15%; Nule (NuMR) <1 log red, > 10%bcr-abl/c-abl : 11%. Pts with Fish (+) showed association with MiMR. Thirty eight percent of pts with a follow-up between 6–24 months since the beginning of imatinib treatment achieved MaMR and CMR, similar rate as the observed in the group with > 24 months on imatinib (42%). Up to July 2006, 53 pts underwent a 2nd RQ-PCR evaluation at 6 months: 15% of pts improved molecular response, 68% maintained it while 17% showed worse response. The 87.5% of pts with decreasing bcr-abl transcripts level at 2nd evaluation corresponded to imatinib treatment as first line. The 78% of pts with rising levels at 2nd evaluation showed MiMR at beginning of this study. Up to now, no pts showed hematological relapse.Conclusion: This is an on-going study in CML pts in CCyR, where 93% showed a negative FISH study, and 7% had low FISH positivity. By RQ-PCR, 40% of pts presented CMR or MaMR (≥3 log red). In 2nd RQ-PCR evaluation, 83% of pts improved or maintained the molecular response. Up to now all pts remain in hematological remission.

IFNα Therapy Up-Regulates Serum Soluble TRAIL Levels and Induces Marked TRAIL mRNA Expressions in Polymorphonuclear Neutrophils in Chronic Myeloid Leukemia Patients.

[Purpose] Tumor necrosis factor related apoptosis-inducing ligand (TRAIL) is an interferon α (IFN)-induced, apoptosis-inducing molecule. TRAIL is one of the promising reagents for therapeutic use in combination with imatinib in chronic myeloid leukemia (CML). A previous report showed that IFN-induced TRAIL is only weakly expressed on cell surface but is mainly secreted as a soluble form (sTRAIL) by polymorphonuclear neutrophils (PMNs) (and also by monocytes) in healthy individuals. Up to now, however, whether sTRAIL is actually released into serum in CML patients and whether its serum level is up-regulated during in vivo IFN therapy are unsolved issues. To assess the significance of sTRAIL, we examined serum sTRAIL levels by ELISA in total 40 (36 in CP, 4 in BP) CML patients either prior to or undergoing IFN therapy.[Results] Serum sTRAIL were detectable in untreated CML patients (1258±560 pg/ml, n=25), which levels were substantially comparable to those in healthy donors (1159±343, n=10). Serum sTRAIL levels significantly increased in patients during IFN therapy (1801±712, n=18) compared to those in untreated patients (P=0.009). This IFN-induced up-regulation of serum sTRAIL was further confirmed in all seven patients whom we could analyze both before and during the therapies (before 1228±396 vs. during 1956±914, P=0.037). As for initial sTRAIL levels, among the restricted patients group who later showed any cytogenetic response (n=15), there was a tendency that initially higher serum sTRAIL levels accompanied better cytogenetic responses (CCyR: 1873±719, PCyR: 1130±204, minor CyR: 830±267, P=0.03), indicating that the initial sTRAIL level may have some relation with the later response to IFN therapy. Next, to investigate the source of the sTRAIL secretion into serum, we assessed TRAIL mRNA expressions in PMNs in the peripheral blood by RQ-PCR. Again, TRAIL mRNA expressions in patients during IFN therapy (n=22) significantly increased compared to those in untreated patients (n=16) (TRAIL/GAPDH: before 1.4 vs. during 8.0, P=0.005). This IFN-induced up-regulation of TRAIL mRNA expression in PMNs was confirmed in all 16 patients whom we could analyze both before and during IFN therapies (before 3.7 vs. during 21.3, P=0.018). In vitro IFN stimulation of PMNs in five untreated patients showed marked induction (10 to 30 fold) of TRAIL mRNA at 12h, while induction of FAS mRNA stayed rather low (1 to 2 fold). Furthermore, IFN concentration-dependent accumulations of sTRAIL into the culture supernatants were observed for up to 48 h. Next, we assessed TRAIL mRNA expressions in purified CD34-positive cells in the bone marrow. TRAIL mRNA expressions were also detectable, and the mRNA levels significantly increased in patients during IFN therapy (n=37), compared to those in untreated patients (n=17) (P=0.02). This IFN-induced up-regulation of TRAIL mRNA was confirmed in nine patients whom we could analyze both before and during the therapies (P=0.01), indicating that even immature CML elements are responsive to IFN and may contribute to sTRAIL release in the microenvironment.[Conclusion] This is the first evidence showing that IFN, in vivo, provokes the release of sTRAIL when administered systematically in CML patients. The main source of the serum sTRAIL is probably PMNs in CML patients. We suspect that one of the mechanisms of anti-proliferative action of IFN on CML is sTRAIL secreted into serum. Our findings also give rationale for the use of IFN (or recombinant sTRAIL) in combination with imatinib in CML patients.

High Catalase Activity: Another Possible Pathway for Imatinib-Resistant CML Patients.

Imatinib (imatinib mesylate: Novartis Pharmaceuticals, Basel, Switzerland) is a well-known selective inhibitor of BCR-ABL tyrosine kinase activity, a hallmark enzyme in the pathogenesis of chronic myeloid leukemia (CML). Resistance to imatinib in CML patients has been associated with different and complex types of mechanisms ranging from nonspecific multidrug resistance protein, preventing the drug to reach its target, to BCR-ABL kinase domain point mutations, reducing the affinity of the enzyme to Imatinib. More recently, it was described that modulation of reduced glutathione (GSH) intracellular levels restores sensitivity to Imatinib in Imatinib-resistant cell line. This suggests that induction of hydrogen peroxide (H2O2), a well-known apoptotic reactive oxygen species (ROS), might be another Imatinib anti-leukemic mechanism action. Additionally, it is known that catalase activity is enhanced through phosphorilation by the c-abl tyrosine kinase (TK). In a preliminary work, our group found normal levels of GSH but high levels of antioxidant enzyme catalase in CML patients. The aim of this work is to evaluate the possible role of catalase as another pathway to Imatinib resistance. White blood cells from blood donors, CML patients and patients with reactional leukocytosis caused by infectious bacterian diseases were obtained using Ficoll-Hypaque. Catalase activity was measured by monitoring decomposition of 10mM H2O2 at 240nm according to the method described by Aebi. The viability of eritroleukemia cell lineage K562 previously treated or not with 100μM 3-aminotriazole (catalase inhibitor) was evaluated by MTT method with Imatinib 1, 5 and 25μM for 24 and 48 hours. Imatinib induced intracellular ROS generation was monitored by flow cytometry with 40μM 2′, 7′-dichlorofluorescein diacetate (DCFH-DA), a well-known ROS capturing reagent. Catalase activity was increased up to 30x in CML patients but not in reactional leukocytosis. When BCR-ABL positive cell line K562 was treated with Imatinib (50μM), catalase activity levels decreased in 38% compared with untreated cells. Also, we found that 3-aminotriazole can overcome the resistance to Imatinib in K562 cells resulting in 25% enhancement of dead cells. The protection of catalase against Imatinib-ROS generation was confirmed by DCFH-DA/FACs in cells incubated with PEG-catalase (monomethoxypolyethylene glycol covalent attached to catalase). Taken together, these results suggest that catalase might be another attractive target for overcoming resistance to Imatinib in Imatinib-resistant CML cells with high catalase activity.

A phase I study of BMS-354825 in patients with imatinib-resistant and intolerant chronic phase chronic myeloid leukemia (CML): Results from CA180002

6519 Background: Resistance to imatinib in CML patients is often associated with Bcr-Abl point mutations which interfere with imatinib binding. BMS-354825 is a novel, orally available, dual SRC/ABL kinase inhibitor with more than 300-fold greater potency than imatinib and has preclinical activity against 14 of 15 imatinib resistant Bcr-Abl mutants (Shah et al, Science, 305:399, 2004). Methods: CA180002 is a phase I, dose-escalation study of BMS-354825 in CML patients with hematologic resistance or intolerance to imatinib. Intrapatient dose escalation is permitted. Patient samples are analyzed for pharmacokinetics (PK), Bcr-Abl kinase domain mutations, and CRKL, HCK, and LYN phosphorylation. Results: 36 chronic phase CML patients have been treated (31 resistant, 5 intolerant) with total daily doses ranging from 15 - 180 mg (given as single or divided doses). Mutations associated with imatinib resistance were identified in 27 patients. Thirty-one patients (86%) have had complete hematologic response (CHR). Only 2 patients have had progression of their disease. Thirteen of 29 patients had cytogenetic improvement including 5 complete and 4 partial cytogenetic responses (i.e. major cytogenetic responses (MCyR) in 9 of 29 patients. MCyR rate = 31%). Hematologic and cytogenetic responses have occurred regardless of mutation status, or whether the mutation was in the P-loop, catalytic or activation region, with the exception of one patient who progressed with T315I. Responses are durable, and 33 of 36 patients remain on study for 1+ to 13+ months. BMS-354825 has been well tolerated. Three patients had grade 4 thrombocytopenia and 2 patients had grade 4 neutropenia, all of which were reversible and easily managed with dose modification. One patient developed a duodenal ulcer possibly related to BMS-354825. Mild QTc prolongation has been noted. Conclusions: BMS-345825 appears to be safe and effective therapy for patients with imatinib-resistant and imatinib-intolerant CML producing hematologic and cytogenetic responses. The phase I study continues to accrue patients to determine the maximally tolerated dose. No significant financial relationships to disclose.