Survival Of Chronic Myeloid Leukemia Patients Research Articles

PBA2, a novel inhibitor of the β-catenin/CBP pathway, eradicates chronic myeloid leukemia including BCR-ABL T315I mutation

BackgroundBCR-ABL is a constitutively active tyrosine kinase that stimulates multiple downstream signaling pathways to promote the survival and proliferation of chronic myeloid leukemia (CML) cells. The clinical application of specific BCR-ABL tyrosine kinase inhibitors (TKIs) has led to significantly improved prognosis and overall survival in CML patients compared to previous treatment regimens. However, direct targeting of BCR-ABL does not eradicate CML cells expressing T315I-mutated BCR-ABL. Our previous study revealed that inhibiting CREB binding protein (CBP) is efficacious in activating β-catenin/p300 signaling, promoting cell differentiation and inducing p53/p21-dependent senescence regardless of BCR-ABL mutation status. We hypothesize that the specific inhibition of CBP may represent a novel strategy to promote β-catenin/p300-mediated differentiation and suppress cancer cell proliferation for treating CML patients.MethodsThe anticancer efficacy of PBA2, a novel CBP inhibitor, in CML cells expressing wild-type or T315I-mutated BCR-ABL was investigated in vitro and in vivo. Cell differentiation was determined by the nitroblue tetrazolium (NBT) reduction assay. The extent of cellular senescence was assessed by senescence-associated β-galactosidase (SA-β-Gal) activity. Cytotoxicity was measured by MTS assay. RNA interference was performed to evaluate the cell proliferation effects of CBP knockdown. The interaction of β-catenin and CBP/p300 was examined by co-immunoprecipitation assay.ResultsPBA2 exhibited significantly higher anticancer effects than imatinib in CML cells harboring either wild-type or T315I-mutated BCR-ABL both in vitro and in vivo. Mechanistically, PBA2 reduced CBP expression and promoted β-catenin-p300 interaction to induce cell differentiation and senescence.ConclusionOur data supported the rational treatment of CML by inhibiting the β-catenin/CBP pathway regardless of BCR-ABL mutation status.

Single Cell Analysis of Genes Involved in Asymmetric Hematopoietic Stem Cell (HSC) Division in Chronic Myeloid Leukemia (CML) : Evidence of the Regulation of Tetraspanin CD63 during Leukemia Progression

Tyrosine kinase inhibitor (TKI) therapies have profoundly changed the natural history and survival of CML patients. However, the majority of the patients remain on long-lasting therapies and switch one TKI to another, either because of intolerance or resistance. All TKI cessation trials showed approximately 50% of leukemia recurrence rates even after several years of TKI therapies, suggesting the persistence of residual leukemic stem cells in patients in deep molecular response (DMR). The resistance and persistence of CML HSC are related to intrinsic (BCR-ABL independent survival) or extrinsic (leukemic niche ) mechanisms some of which have been extensively studied. However, there has been no data with regard to the role of asymmetric cell division pathways in the persistence and progression of CML stem cell clone. Indeed, During asymmetric division process of hematopoietic stem cells, some proteins are differently segregated between daughter HSCs such as endosome associated tetraspanin proteins (CD53, CD63) and mitochondria-associated fission regulator Dynamin related protein-1(DRP1). Similarly, it has been shown that asymmetric organelle inheritance can influence HSC decisions towards differentiation versus self-renewal. Interestingly, the tetraspanin CD63 is known to be involved in the quiescence of human and murine HSC via activation of TGF-beta pathway, CD63 hi cells exhibiting increased self-renewal potential after the asymmetric division with reduced differentiation potential. Materials and Methods We have analyzed the transcriptome performed in a cohort of CML patients in chronic phase (CP) and blast phase (BP) (GSE4170) as well as in a validation cohort of RNA-sequencing (GSE100026). Similarly, single cell transcriptome analysis of Lin- CD34+ CD38- CML progenitors (GSE76312) were included in the study evaluating the expression of genes involved in asymmetric cell division. We have then generated an asymmetric hematopoietic stem cell risk score using a panel of genes involved in the asymmetric division of HSCs and correlated these results with the transcriptome of the CML cohorts above. A pseudotime analysis was performed to evaluate single cell trajectories of the genes identified. Results To determine the potential role of asymmetric cell division markers in the progression of CML, we have analyzed the expression of several of these genes during CML progression in two independent transcriptome cohorts. The regulation concordance between the two distinct cohorts was principally observed on asymmetric molecules which were found to be down-regulated during blast crisis as compared to chronic phase. The regulation concordance between the two distinct cohorts was principally observed on asymmetric molecules down regulated during blast crisis as compared to chronic phase suggesting less of asymmetric properties during advanced phase of the disease. The down regulation in patients in blast crisis was confirmed at single cell level for the expression of SELL, CD63, NUMB, HK2 and LAMP2. CD63 expression has also been found to be of interest in CML chronic phase. Indeed, single cell neural network and deep learning in CP samples showed that the CD63 expression was the best asymmetric gene marker predicting a poor prognosis and this appeared to be induced by opposite regulation of HK2 and NUMB. In patients in CP-CML, the single cell trajectory reconstitution revealed the major regulatory role of CD63 highlighting a trajectory cluster implicating HSPB1, PIM2, ANXA5, LAMTOR1, CFL1, CD52, RAD52, MEIS1, PDIA3, all genes known to be involved also in hematopoietic malignancies such as myelodysplasia, AML and ALL. Conclusion We show here for the first time the identification of the expression of genes involved in the asymmetric division of CML HSC. Among these genes we identified the tetraspanin CD63 as being down-regulated during CML progression. The expression of the genes involved in the asymmetric HSC division should require further studies in CML, either to predict persistence of CML or progressive disease. Figure 1 : Single cell transcriptome of Lin- CD34+ CD38- CML progenitors in CP links CD63 expression to progressive disease. A/ Piechart of counts for chronic phase cells analyzed according to prognosis; B/ pseudotime transformation of single cell transcriptome according patient prognosis; C/ Pseudotime expression heatmap; D/ Pseudotime expression of CD63 in CP.

Microrna-142 Deficiency Promotes Chronic Myeloid Leukemia (CML) Transformation from Chronic Phase (CP) to Blast Crisis (BC)

Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm resulting from the BCR-ABL1 fusion gene that encodes a constitutively activated tyrosine kinase (TK). Although TK inhibitors (TKIs) induce disease remission and prolonged survival in CML patients, a subset are resistant and progress from chronic phase (CP) to blast crisis (BC) with poor prognosis. Understanding the molecular mechanisms of transformation from CP to BC is necessary in the development of effective treatments. Here, we used the inducible SCLtTA/BCR-ABL transgenic CP CML model to study the molecular mechanism of disease evolution. Upon tetracycline withdrawal to induce BCR-ABL expression, both the SCLtTA/BCR-ABL homozygous (homo, i.e., SCLtTA+/+BCR-ABL+/+, hereafter called BCR-ABL) and heterozygous (het, i.e., SCLtTA+/-BCR-ABL+/-) mice developed and died of CP CML without developing BC CML, implying that BCR-ABL dosage is insufficient to induce transformation. MicroRNA (miR)-142 is highly expressed in hematopoietic cells with a critical role in normal hematopoiesis. In miR-142 knockout (KO)(miR-142−/−) mice, hematopoietic stem and progenitor cells expanded with a decrease of hematopoietic output. Loss of miR-142 function has been reported in lymphoma, acute lymphocytic leukemia and acute myeloid leukemia. Of note, we also observed lower levels of miR-142 in CD34+CD38- cells from patients with BC CML versus (vs) patients with CP CML. Thus, we hypothesized that miR-142 insufficiency may promote CML transformation from CP to BC. To test our hypothesis, we generated miR-142 KO BCR-ABL (i.e., miR-142−/−BCR-ABL) mice and observed increasing leukemic blasts over time after BCR-ABL induction in the blood and bone marrow (BM), but not in miR-142 wt (miR-142+/+)BCR-ABL controls even when the latter became moribund. MiR-142−/−BCR-ABL mice had larger spleens and significantly shorter survival [median: 26 vs 54 days (d); p<0.0001] than miR-142+/+BCR-ABL controls. Of note, while both homo (miR-142−/−) and het (miR-142+/−) miR-142 KO BCR-ABL mice eventually developed BC CML, the former had a significantly faster progression to BC and shorter survival (median: 26 vs 45 d; p=0.003) than the latter, suggesting miR-142 deficiency alone is sufficient to initiate BC transformation in the CP CML model in a dose-dependent manner. Importantly, all these features were recapitulated in congenic recipient mice transplanted with BM Lin-Sca-1+c-Kit+ cells (LSKs, 2000/mouse) from diseased miR-142−/−BCR-ABL mice, suggesting LSKs were enriched in leukemic stem cells and able to reproduce BC. Of note, in an RNA-seq analysis comparing LSKs from diseased miR-142−/−BCR-ABL (BC) and miR-142+/+ BCR-ABL(CP) mice, 504 genes were found differentially expressed. Gene set enrichment analysis (GSEA) showed only four pathways differentially expressed (upregulated); three [i.e., oxidative phosphorylation, glycolysis and adipogenesis] regulating cell metabolism and the fourth regulating protein secretion. Next, we developed a novel CpG-miR-142 mimic oligonucleotide, hereafter called CpG-M-miR-142, to restore miR-142 levels. Treatment with CpG-M-miR-142 (20mg/kg/day, iv, 4 weeks) on day 2 after BCR-ABL induction significantly prolonged survival of miR-142−/−BCR-ABL mice compared with CpG-scramble (SCR) (75% vs 33% survival rate at day 40 after BCR-ABL induction; median survival: not reached vs 25 d; p=0.03). Since we observed lower miR-142 levels in TKI-resistant vs TKI-sensitive CML patients (p=0.02), we selected LSKs from diseased miR-142−/−BCR-ABL and miR-142+/+BCR-ABL mice and exposed them to TKI nilotinib (NIL; 2µM) or vehicle for 72 hours to evaluate if downregulation of miR-142 was associated with TKI resistance. We observed lower apoptosis and higher cell growth in NIL-treated miR-142−/−BCR-ABL LSKs vs NIL-treated miR-142+/+BCR-ABL LSKs. The decreased sensitivity of miR-142−/−BCR-ABL LSKs to TKI was rescued by treatment with CpG-M-miR-142. CpG-M-miR-142 (2µM) plus NIL significantly increased apoptosis and reduced cell growth in miR-142−/−BCR-ABL LSKs compared with SCR+ NIL. We showed a key role of miR-142 deficiency in the transformation of CP CML to BC CML associated with deregulation of metabolic pathways. Restoring miR-142 expression in vivo with CpG-M-miR-142 significantly decreased the BC transformation rate, prolonged survival of miR-142−/−BCR-ABL mice and may increase sensitivity to TKIs. Disclosures Marcucci: Iaso Bio: Membership on an entity's Board of Directors or advisory committees; Abbvie: Speakers Bureau; Novartis: Speakers Bureau; Pfizer: Other: Research Support (Investigation Initiated Clinical Trial); Takeda: Other: Research Support (Investigation Initiated Clinical Trial); Merck: Other: Research Support (Investigation Initiated Clinical Trial).

Distribution of BCR-ABL1 Transcript Variants in Nigerians with Chronic Myeloid Leukemia.

The distribution of BCR-ABL1 transcript variants e13a2 ("b2a2") and e14a2 ("b3a2") in Nigerians with chronic myeloid leukemia (CML) had not been previously studied. In addition, there is paucity of data on the impact of BCR-ABL1 transcript variants on clinical presentation and survival in CML patients in Nigeria. The BCR-ABL1 transcript variants were analyzed in 230 Imatinib-treated CML patients at diagnosis. Patients with incomplete data (n = 28), e19a2 (n = 3) and e1a2 (n = 1) were excluded from analysis of transcript variant on disease presentation and survival leaving only 198. The frequencies of BCR-ABL1 transcript variants were 30 (13.0%), 114 (49.6%), 82 (35.7%), three (1.3%) and one (0.4%) for e13a2, e14a2, co-expression of e13a2/e14a2, e19a2 and e1a2, respectively. A significantly higher platelet count was found in patients with e13a2 variant (531.1 ± 563.4 × 109/L) than in those expressing e14a2 (488.2 ± 560.3 × 109/L) or e13a2/e14a2 (320.7 ± 215.8 × 109/L); p = 0.03. No significant differences were found between the variants with regards to gender, age, phase of disease at diagnosis, total white blood cell count, neutrophil percentage, hematocrit, splenomegaly or hepatomegaly. Overall survival was higher but not statistically significant (p = 0.4) in patients with e14a2 variant (134months) than in e13a2 (119months) and co-expression of e13a2/e14a2 (115months). Nigerian CML patients have the highest incidence of co-expression of e13a2 and e14a2. Distinct disease characteristics which contrast with findings from the Western countries were also identified in Nigerians which may be due to genetic factors.

Managing chronic myeloid leukemia for treatment-free remission: a proposal from the GIMEMA CML WP

AbstractSeveral papers authored by international experts have proposed recommendations on the management of BCR-ABL1+ chronic myeloid leukemia (CML). Following these recommendations, survival of CML patients has become very close to normal. The next, ambitious, step is to bring as many patients as possible into a condition of treatment-free remission (TFR). The Gruppo Italiano Malattie EMatologiche dell'Adulto (GIMEMA; Italian Group for Hematologic Diseases of the Adult) CML Working Party (WP) has developed a project aimed at selecting the treatment policies that may increase the probability of TFR, taking into account 4 variables: the need for TFR, the tyrosine kinase inhibitors (TKIs), the characteristics of leukemia, and the patient. A Delphi-like method was used to reach a consensus among the representatives of 50 centers of the CML WP. A consensus was reached on the assessment of disease risk (EUTOS Long Term Survival [ELTS] score), on the definition of the most appropriate age boundaries for the choice of first-line treatment, on the choice of the TKI for first-line treatment, and on the definition of the responses that do not require a change of the TKI (BCR-ABL1 ≤10% at 3 months, ≤1% at 6 months, ≤0.1% at 12 months, ≤0.01% at 24 months), and of the responses that require a change of the TKI, when the goal is TFR (BCR-ABL1 >10% at 3 and 6 months, >1% at 12 months, and >0.1% at 24 months). These suggestions may help optimize the treatment strategy for TFR.

TREATMENT OUTCOMES IN PATIENTS WITH CHRONIC MYELOID LEUKEMIA ACCORDING TO THE RUSSIAN PART OF THE EUTOS POPULATION-BASED STUDY

Background.The European Population Register EUTOS for CML includes data on adult patients (n = 2904) diagnosed with Ph-positive (Ph+) and/or BCR-ABL1-positive (BCR-ABL1+) chronic myeloid leukemia (CML) in 20 European countries during the period from 2008 to December 2012. Russia took part in this study, having contributed 6.8 % of CML patients to the total number of patients in the Register.Aim.To estimate long-term treatment outcomes in patients with newly diagnosed CML in the Russian Federation in comparison with the data obtained for a pan-European population cohort of patients.Patients and methods.The cohort under study consisted of 197 patients from 6 Russia regions, all of whom were diagnosed with a Ph+ / BCR-ABL1 + CML during the period from October, 1, 2009 to December, 31, 2012. The patients’ median age was 50 (18–82) years, with men and women being represented in approximately equal proportions.Results.In the first line, 97 % and 3 % of the patients received Imatinib and 2nd generation tyrosine kinase (TKI) inhibitors, respectively. The response dynamics was as follows: 12 months after the treatment, a complete cytogenetic response and a major molecular response were achieved in 40 % and 20 % of the patients. The overall survival (OS) and progression-free survival rates in patients in Russia following 12, 24 and 30 months were 93 %, 87 % and 84 %, and 92 %, 87 % and 87 %, respectively. In Russia, the study was prolonged. By 80 months of observation, the OS of patients in the chronic CML phase with a low and high risk of the disease progression had been 88 % and 56 %, respectively. In the acceleration phase, the 5-year overall survival rate was 39 %.Conclusion.An analysis of treatment outcomes in CML patients in a population-based non-selected sample indicates an increase in the survival of CML patients. However, problematic aspects of the therapy have been identified, along with a need for intensification of the treatment in patients with an unfavourable CML prognosis.

Research progress in treatment of chronic myeloid leukemia patients with imatinib resistance

Chronic myeloid leukemia (CML) is a malignant myeloproliferative neoplasm that occurs in pluripotent hematopoietic stem cells. Pathogenesis of CML is slow, and leukocytosis and splenomegaly are main clinical features of patients with CML. The introduction of tyrosine kinase inhibitor (TKI) imatinib mesylate (IM) has changed the natural progression of CML and significantly improved survival of CML patients. However, with the prolonged application of IM, IM resistance has become a major challenge in the treatment of CML. Many studies are currently working to elucidate the mechanisms of CML resistance and to research and develop new TKI. There is a urgent need to replace new drug targets and new therapeutic strategies for BCR/ABL1 fusion gene in response to IM-resistant CML patients. This article focuses on the development of CML-related TKI and non-TKI latest drugs for IM-resistant CML patients, and describes immunological approaches activating specific T cell responses against CML cells. Key words: Leukemia, myelogenous, chronic, BCR-ABL positive; Receptor protein-tyrosine kinases; Drug resistance, neoplasm; Immunotherapy; Therapeutic uses

Aberrant DNA methylation at HOXA4 and HOXA5 genes are associated with resistance to imatinib mesylate among chronic myeloid leukemia patients.

Imatinib mesylate is a molecularly targeted tyrosine kinase inhibitor drug. It is effectively used in the treatment of chronic myeloid leukemia (CML) patients. However, development of resistance to imatinib mesylate as a result of BCR-ABL dependent and BCR-ABL independent mechanisms has emerged as a daunting problem in the management of CML patients. Between these mechanisms, BCR-ABL independent mechanisms are still not robustly understood. To investigate the correlation of HOXA4 and HOXA5 promoter DNA hypermethylation with imatinib resistance among CML patients. Samples from 175 Philadelphia positive CML patients (83 good response and 92 BCR-ABL non-mutated imatinib resistant patients) were subjected to Methylation Specific High Resolution Melt Analysis for methylation levels quantification of the HOXA4 and HOXA5 promoter regions. Receiver operating characteristic curve analysis was done to elucidate the optimal methylation cut-off point followed by multiple logistic regression analysis. Log-Rank analysis was done to measure the overall survival difference between CML groups. The optimal methylation cut-off point was found to be at 62.5% for both HOXA4 and HOXA5. Chronic myeloid leukemia patients with ≥63% HOXA4 and HOXA5 methylation level were shown to have 3.78 and 3.95 times the odds, respectively, to acquire resistance to imatinib. However, overall survival of CML patients that have ≤62% and≥63% methylation levels of HOXA4 and HOXA5 genes were found to be not significant (P-value=0.126 for HOXA4; P-value=0.217 for HOXA5). Hypermethylation of the HOXA4 and HOXA5 promoter is correlated with imatinib resistance and with further investigation, it could be a potential epigenetic biomarker in supplement to the BCR-ABL gene mutation in predicting imatinib treatment response among CML patients but could not be considered as a prognostic marker.

Impact of polymorphisms of TP53 Arg72Pro, excision repair cross-complementing Protein 1 Asn118Asn (C118T) and deletion in Intron 2 of BIM in chronic myeloid leukemia patients on imatinib treatment

Background: Imatinib (IM) results in durable responses in a large number of patients with chronic myeloid leukemia (CML). However, a substantial number develops drug resistance. There could be several reasons for the heterogeneity of response. To survive genotoxic damage induced by IM, BCR-ABL1 can modulate DNA repair mechanisms, cell-cycle checkpoints, and Bcl-2 family members. Hence, we aimed to find out the impact of single-nucleotide polymorphisms mainly TP53Arg72Pro, excision repair cross-complementing protein 1 (ERCC1) Asn118Asn (C118T) and deletion in intron 2 of BIM genes in CML patients and their association with disease progression and treatment outcome. Materials and Methods: In the present study, 174 CML and 174 control samples were analyzed for polymorphisms of TP53Arg72Pro, ERCC1 C118T, and intron 2 deletion in BIM genes using polymerase chain reaction-restriction fragment length polymorphism method. Results: Genotyping of TP53Arg72Pro polymorphism showed a trend toward proline genotype/allele association with younger age (P = 0.031), advanced phase, high BCR-ABL1 expression levels and presence of tyrosine kinase domain (TKD) mutations compared to respective groups. Genotyping of ERCC1 C118T polymorphism showed a significant association of CT genotype and T allele with high BCR-ABL1 levels BIM deletion polymorphism was not observed in the present study (P = 0.030) and presence of TKD mutations (P = 0.013). The 72 proline allele and heterozygous 118CT genotype frequencies were significantly elevated in deceased patients compared to those on IM and other tyrosine kinase inhibitors treatment. Conclusion: our data suggested that proline genotype/allele of TP53Arg72Pro polymorphism possibly leading to inefficient apoptosis and 118CT genotype in ERCC1 gene with possibly less DNA repair efficiency might be responsible for the suboptimal responses to IM and poor survival in CML patients. Deletion in intron 2 of BIM gene was not observed in our study.

Imatinib reduces proliferation of leukemic cells in vitro

Introduction: Philadelphia chromosome is a cytogenetic marker for chronic myeloid leukemia (CML). The main aims of this study were to assess the positive responses, side-effects and survival of CML patients treated with imatinib mesylate.Methods: All recently diagnosed CML patients who were treated with imatinib were recruited to this study. We investigated hematological and cytogenetic parameters by CBC, FISH and RT-PCR individually.Results: Of the 10 cases, 7 (70%) were males and 3 (30%) were female. Four (40%) of the cases were analyzed retrospectively and 8 cases (80%) exhibited general exhaustion (75%), fever (80%), and splenomegaly (80%). Indications of bleeding and rashes were rarely seen at presentation. The majority of the patients had a generally low risk profile (70%), 30% had intermediate risk; with no subjects exhibiting high risk CML, 9 subjects (90%) were in remission. One patient (10%) had been in remission for 3 years, 4 (40%) had been in remission for 6 years, one was in remission after 7 years and 5 (50%) were in remission after 10 years. Most of the patients (90%) exhibited a deficient major molecular reaction, after 6 years of treatment and 42% of them had a deficient major molecular reaction after 10 years of treatment. No significant side effects associated with Imatinib treatment were reported by the patients. Imatinib treatment resulted in diminished expansion in CML CFU-GM cells.Conclusion: Imatinib mesylate is indicated for the treatment of Philadelphia chromosome-positive CP-CML with no significant adverse outcomes.Bangladesh Journal of Medical Science Vol.16(2) 2017 p.320-324

Presence of novel compound BCR-ABL mutations in late chronic and advanced phase imatinib sensitive CML patients indicates their possible role in CML progression

ABSTRACTBCR-ABL kinase domain (KD) mutations are well known for causing resistance against tyrosine kinase inhibitors (TKIs) and disease progression in chronic myeloid leukemia (CML). In recent years, compound BCR-ABL mutations have emerged as a new threat to CML patients by causing higher degrees of resistance involving multiple TKIs, including ponatinib. However, there are limited reports about association of compound BCR-ABL mutations with disease progression in imatinib (IM) sensitive CML patients. Therefore, we investigated presence of ABL-KD mutations in chronic phase (n = 41), late chronic phase (n = 33) and accelerated phase (n = 16) imatinib responders. Direct sequencing analysis was used for this purpose. Eleven patients (12.22%) in late-CP CML were detected having total 24 types of point mutations, out of which 8 (72.72%) harbored compound mutated sites. SH2 contact site mutations were dominant in our study cohort, with E355G (3.33%) being the most prevalent. Five patients (45%) all having compound mutated sites, progressed to advanced phases of disease during follow up studies. Two novel silent mutations G208G and E292E/E were detected in combination with other mutants, indicating limited tolerance for BCR-ABL1 kinase domain for missense mutations. However, no patient in early CP of disease manifested mutated ABL-KD. Occurrence of mutations was found associated with elevated platelet count (p = 0.037) and patients of male sex (p = 0.049). The median overall survival and event free survival of CML patients (n = 90) was 6.98 and 5.8 y respectively. The compound missense mutations in BCR-ABL kinase domain responsible to elicit disease progression, drug resistance or disease relapse in CML, can be present in yet Imatinib sensitive patients. Disease progression observed here, emphasizes the need of ABL-KD mutation screening in late chronic phase CML patients for improved clinical management of disease.

Abstract C16: Disparities in survival outcomes based on insurance status in CML: A national cancer database analysis

Abstract Purpose: The treatment of chronic myeloid leukemia (CML) has made major progress in the last decade. The survival of CML patients has improved with advent of tyrosine kinase inhibitors. The cost of medication, compliance of patients and access to care are important issues that can also affect outcome. The purpose of the present study is to investigate effects of insurance status on outcomes of CML by analyzing data from National cancer database. Methods: Data was analyzed from 18, 415 men and women (≥ 18 years of age) registered in the NCDB who were diagnosed with CML between 2004 and 2012 and had follow-ups to end of 2013. The primary predictor variable was insurance status and the outcome variable was overall survival. Additional variables addressed and adjusted for included sex, age, race, Charleston Co morbidity index, level of education, income, and distance traveled, facility type, diagnosing/treating facility and treatment delay. Results: The mean age of patients was 59 years with standard deviation of 18. In multivariate analysis, after adjusting for secondary predictor variables, insurance status was a statistically significant predictor of overall survival from CML. Relative to privately insured patients, patients with Medicaid had a 62.7% increased risk, Medicare had a 44.3% and no insurance had a 34.7% increased risk of mortality from CML. Among the factors analyzed, age, gender, race, and comorbidity index, were all found to be significant predictors of survival in CML patients, respectively. Conclusion: We observed that insurance status has a statistically significant relationship with overall survival from CML. This remained true after adjusting for other predictive factors. Further research is necessary how the disparities associated with different types of insurance result in inferior treatment outcomes and how they can be addressed. Citation Format: Samip R. Master, Nebu Koshy, Glenn Mills, Runhua R. Shi. Disparities in survival outcomes based on insurance status in CML: A national cancer database analysis. [abstract]. In: Proceedings of the Ninth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2016 Sep 25-28; Fort Lauderdale, FL. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2017;26(2 Suppl):Abstract nr C16.

Functional studies of miR-130a on the inhibitory pathways of apoptosis in patients with chronic myeloid leukemia.

The p53 mutation in chronic myeloid leukemia (CML) led to decreased overall survival and therapy resistance which was also closely correlated with the downstream proto-oncogenes BCL-2, TCL-1 and MCL-1. We in this study aimed to investigate the function of miR130a in p53 tumor suppressor signaling pathway. We performed microRNA (miRNA) expression profile analysis in CML cancer stem cells of 38 cases and extracted total RNA from peripheral blood of 143 cases. Standard curves of U6 and miRNA were made from 10-fold serial dilutions of the cDNA, which were quantified using real-time quantitative PCR with SYBR Green by ABI 7300. The p53 mutations and BCR/ABL mutation status analysis in CML patients were detected by PCR and direct sequencing. Candidate targets of miR130a of putative relevance in CML pathogenesis were analyzed by bioinformatics approach. We then used dual-luciferase activity assay to verify the target genes of miR130a and used western blot analysis to elucidate the mechanism of miR130a on modulating drug resistance. The levels of miR-130a expression in CML were significantly lower in poor prognostic subgroups, defined by prognostic factors including mutated BCR/ABL status, p53 and ATM deletions and p53 mutations. Furthermore, underexpression of miR-130a was significantly associated with shorter overall survival and treatment-free survival in CML patients. We demonstrated that miR130a function as tumor suppressors by inhibiting multiple anti-apoptosis proteins, including BCL-2, MCL-1 and XIAP. This was a direct effect because miR130a negatively regulated expression of a BCL-2/MCL-1/XIAP 3'untranslated region-based reporter construct. Transfection of miR130a mimics into CML cells from 30 patients without p53 aberrations led to significant increases in apoptosis compared with transfection with the miRNA control. Besides, enforced expression of miR130a had no significant drug-sensitization effect in CML cells from p53-attenuated patients. MiR-130a may have an important role in the pathogenesis of CML and may be useful for assessing prognosis in patients with CML. Moreover, miR130a may provide a possible therapeutic avenue and a sensitive indicator of the activity of the p53 in CML.

Discontinuation of tyrosine kinase therapy in CML.

Over the past decade, a broad array of drugs designed to selectively inhibit protein tyrosine kinases (tyrosine kinase inhibitors or TKIs) have emerged as novel therapies for cancer patients. Chronic myeloid leukemia (CML) is one of the best examples of successful targeted therapy with a TKI. The overall survival of CML patients who respond to treatment is close to that of the healthy population. The response in many patients is so profound that it is possible to consider stopping their treatment and with time, the number of patients in this group has increased to the point where the issue of treatment cessation has become of utmost importance. This has led to the development of a new concept in the evaluation of CML entitled treatment-free remission. It will be the criterion to evaluate the success of future clinical trials, especially if we want to improve the management of the disease to the point where we can claim to have cured CML.

Isolation, Characterization and Application of BCR-ABL-ve Mesenchymal Stem Cells Derived from Peripheral Blood of Chronic Myeloid Leukemia (CML) in Enhancing Potentiality of Bone Marrow Transplantation in CML Patients

Chronic Myeloid leukemia (CML) is a disorder causing uncontrolled growth of myeloproliferative blast cells. Tyrosine kinase inhibitors are not really useful in advanced stages and have their own side effects, reactions and resistance as well. Till date, CML can be fully cured only by bone marrow transplantation (BMT) which has its own limitations and thus need further studies to increase better survival of CML patients by this procedure. Our lab at Jaslok Hospital has been working on studying and characterizing mesenchymal stem cells in various types of hematological malignancies. We came up with a concept of isolating and characterizing mesenchymal stem cells (MSCs) from peripheral blood of a CML patient which can be used in combination with Bone Marrow Transplantation in the same CML patient. Our study has shown that the stem cells derived from peripheral blood of CML patient were found to be BCR/ABL -ve . We designated this cell line as a “BCR/ABL -ve ” cell line. Molecular characterization of these cells further confirmed their Mesenchymal phenotypes with distinct expression of CD105, CD13 and CD73 genes. Interestingly these cells also expressed pluripotency genes such as OCT4 and NANOG and cytokines i.e. IL6 and TNFα. We further confirmed the normal phenotype of BCR/ABL -ve MSCs by localizing expression of H-Ras, Rb, P53, P16, P21, and EGFR and Ki67 cancer related proteins in these cells by immunofluorescence Microscopy and by in vitro transformation assay. Thus, we suggest that these normal BCR/ABL -ve MSCs derived from CML patient’s peripheral blood can be used in addition to BMT procedure for a better recovery of CML disease in near future.

Therapy of Chronic Myeloid Leukemia: Twilight of the Imatinib Era?

Chronic myeloid leukemia (CML) results from the clonal expansion of pluripotent hematopoietic stem cells containing the active BCR/ABL fusion gene produced by a reciprocal translocation of the ABL1 gene to the BCR gene. The BCR/ABL protein displays a constitutive tyrosine kinase activity and confers on leukemic cells growth and proliferation advantage and resistance to apoptosis. Introduction of imatinib (IM) and other tyrosine kinase inhibitors (TKIs) has radically improved the outcome of patients with CML and some other diseases with BCR/ABL expression. However, a fraction of CML patients presents with resistance to this drug. Regardless of clinical profits of IM, there are several drawbacks associated with its use, including lack of eradication of the malignant clone and increasing relapse rate resulting from long-term therapy, resistance, and intolerance. Second and third generations of TKIs have been developed to break IM resistance. Clinical studies revealed that the introduction of second-generation TKIs has improved the overall survival of CML patients; however, some with specific mutations such as T315I remain resistant. Second-generation TKIs may completely replace imatinib in perspective CML therapy, and addition of third-generation inhibitors may overcome resistance induced by every form of point mutations.

BCR-ABL kinase domain mutations, including 2 novel mutations in imatinib resistant Malaysian chronic myeloid leukemia patients—Frequency and clinical outcome

Discovery of imatinib mesylate (IM) as the targeted BCR-ABL protein tyrosine kinase inhibitor (TKI) has resulted in its use as the frontline therapy for chronic myeloid leukemia (CML) across the world. Although high response rates are observed in CML patients who receive IM treatment, a significant number of patients develop resistance to IM. Resistance to IM in patients has been associated with a heterogeneous array of mechanisms of which point mutations within the ABL tyrosine kinase domain (TKD) are the frequently documented. The types and frequencies of mutations reported in different population studies have shown wide variability. We screened 125 Malaysian CML patients on IM therapy who showed either TKI refractory or resistance to IM to investigate the frequency and pattern of BCR-ABL kinase domain mutations among Malaysian CML patients undergoing IM therapy and to determine the clinical significance. Mutational screening using denaturing high performance liquid chromatography (dHPLC) followed by DNA sequencing was performed on 125 IM resistant Malaysian CML patients. Mutations were detected in 28 patients (22.4%). Fifteen different types of mutations (T315I, E255K, G250E, M351T, F359C, G251E, Y253H, V289F, E355G, N368S, L387M, H369R, A397P, E355A, D276G), including 2 novel mutations were identified, with T315I as the predominant type of mutation. The data generated from clinical and molecular parameters studied were correlated with the survival of CML patients. Patients with Y253H, M351T and E355G TKD mutations showed poorer prognosis compared to those without mutation. Interestingly, when the prognostic impact of the observed mutations was compared inter-individually, E355G and Y253H mutations were associated with more adverse prognosis and shorter survival (P=0.025 and 0.005 respectively) than T315I mutation. Results suggest that apart from those mutations occurring in the three crucial regions (catalytic domain, P-loop and activation-loop), other rare mutations also may have high impact in the development of resistance and adverse prognosis. Presence of mutations in different regions of BCR-ABL TKD leads to different levels of resistance and early detection of emerging mutant clones may help in decision making for alternative treatment. Serial monitoring of BCR-ABL1 transcripts in CML patients allows appropriate selection of CML patients for BCR-ABL1 KD mutation analysis associated with acquired TKI resistance. Identification of these KD mutations is essential in order to direct alternative treatments in such CML patients.

Importance of early and deeper responses to long-term survival in CML patients: Implications of BCR-ABL testing in management of CML in Indian setting.

The prognosis of patients with chronic myeloid leukemia (CML) has changed radically since the advent of imatinib mesylate, a selective inhibitor of BCR-ABL tyrosine kinase. Shortly thereafter, more potent BCR-ABL inhibitors (dasatinib and nilotinib) were introduced for use in patients resistant to or intolerant of imatinib. All three drugs are now approved for initial therapy for chronic phase CML. Response to tyrosine kinase inhibitor (TKI) treatment is assessed with standardized quantitative reverse transcriptase polymerase chain reaction (Q-RTPCR) and/or cytogenetics at 3, 6 and 12 months. Clinical trials have clearly demonstrated that early and deeper cytogenetic and molecular response to TKI therapy is associated with lower rate of disease progression and improved long-term outcomes. In recent times, molecular response as determined by BCR-ABL transcript levels at defined time points is rapidly gaining popularity as a predictive marker for subsequent outcomes in CML. Optimal response is defined as BCR-ABL transcript levels of ≤10% at 3 months, <1% at 6 months, and ≤0.1% from 12 months onward while >10% at 6 months and >1% from 12 months onward define failure. Patients who do not achieve molecular milestones at 3 or 6 months with 3 months being highly predictive are less likely to achieve cytogenetic responses eventually; early identification of such patients who have a low probability of achieving an adequate response are thus candidates for alternative treatment. Review of literature by electronic search of MEDline, Google Scholar was done using keywords and data was identified and systematically evaluated.

Leucémie myéloïde chronique : « archétype » de l’impact des traitements ciblés

Chronic myeloid leukemia (CML) is a chronic blood disorder characterized by a reciprocal translocation between chromosomes 9 and 22, leading to the creation of a chimeric gene encoding the BCR-ABL fusion protein with a constitutive tyrosine kinase activity. Although long known as a disease with an inexorable progression to acute leukemia, CML history has been significantly improved by the use of imatinib, a tyrosine kinase inhibitor. Imatinib has revolutionized the treatment of CML by transforming it from an invariably fatal disease to a chronic but manageable condition. In fact, the discovery of this class of targeted therapy had an impact not only on the survival of CML patients but also on other scientific and medical fields. This review illustrates the impact of imatinib, the first example of tyrosine kinase inhibitors on the treatment of CML, on the treatment of other cancers, the impact on health systems and on the scientific research in general.

Emerging Therapies in Chronic Myeloid Leukemia

Chronic myeloid leukemia (CML) therapy has dramatically changed in the last decade due to the introduction of tyrosine kinase inhibitors (TKIs) - imatinib, nilotinib and dasatinib. Despite the significant prolongation of overall survival of CML patients there is still room for improvement. Approximately 20-25% of patients initially treated with imatinib will need alternative therapy, due to drug resistance which is often caused by the appearance of clones expressing mutant forms of BCR-ABL. Second generation TKIs dasatinib and nilotinib have shown promising results in imatinibresistant or intolerant CML patients, but are not active against CML clones with highly resistant T315I mutation. In recent years special attention is placed on small pool of leukemic stem cells which may contribute to the persistence of the leukemia. This article provides a review of preclinical and clinical data concerning the most promising new directions in CML treatment, with special emphasis on new drugs active in T315I mutation and compounds affecting leukemic stem cells.