FLT3 Length Mutations as Marker for Follow-Up Studies in Acute Myeloid Leukaemia

Length mutations within the FLT3 gene (FLT3-LM) can be found in 23% of acute myeloid leukaemia (AML) and thus are the most frequent mutations in AML. FLT3-LM are highly correlated with AML with normal karyotype and other cytogenetic aberrations of the prognostically intermediate group. This group is supposed to be a mixed group of AML with differences in the underlying pathogenesis. For more individualized treatment options it would be helpful to better characterize this large AML group not only by molecular mutations but also use these markers for the definition of minimal residual disease (MRD). However, so far the cytogenetically intermediate AML has been lacking suitable markers for PCR-based MRD detection like the fusion genes in the prognostically favourable subgroups. The suitability of the FLT3-LM as a target for PCR-based MRD was discussed controversially as it seemed to be a rather unstable marker. Thus, we aimed at the evaluation of FLT3-LM as a marker for residual disease in a large cohort of AML. Paired samples of 97 patients with AML at diagnosis and at relapse were analyzed. It could be shown that in only four cases a loss of the length mutation was detected. This is in the range of other well-characterized AML relapsing with a different geno- and/or phenotype. In contrast, a change in the ratio of the mutated allele in comparison to the wild-type allele was frequently observed. In detail, the FLT3-LM showed a tendency to accumulate during disease progression and was found more frequently at relapse than at diagnosis. In addition, 45 patients were analyzed at different time points during and after therapy. Using conventional PCR it clearly could be shown that for most of the patients positive at presentation FLT3-LM is a reliable PCR marker for monitoring treatment response. Even an early detection of relapse was possible in some cases. Copyright (C) 2004 S. Karger AG, Basel

Molecular Diagnostics, Targeted Therapy, and the Indication for Allogeneic Stem Cell Transplantation in Acute Lymphoblastic Leukemia

In recent years, the panel of known molecular mutations in acute lymphoblastic leukemia (ALL) has been continuously increased. In Philadelphia-positive ALL, deletions of the IKZF1 gene were identified as prognostically adverse factors. These improved insights in the molecular background and the clinical heterogeneity of distinct cytogenetic subgroups may allow most differentiated therapeutic decisions, for example, with respect to the indication to allogeneic HSCT within genetically defined ALL subtypes. Quantitative real-time PCR allows highly sensitive monitoring of the minimal residual disease (MRD) load, either based on reciprocal gene fusions or immune gene rearrangements. Molecular diagnostics provided the basis for targeted therapy concepts, for example, combining the tyrosine kinase inhibitor imatinib with chemotherapy in patients with Philadelphia-positive ALL. Screening for BCR-ABL1 mutations in Philadelphia-positive ALL allows to identify patients who may benefit from second-generation tyrosine kinase inhibitors or from novel compounds targeting the T315I mutation. Considering the central role of the molecular techniques for the management of patients with ALL, efforts should be made to facilitate and harmonize immunophenotyping, cytogenetics, and molecular mutation screening. Furthermore, the potential of high-throughput sequencing should be evaluated for diagnosis and follow-up of patients with B-lineage ALL

Investigation of the kinetic regularities of the reaction of dry reforming of methane using nickel-containing catalysts based on cerium-zirconium oxides

In this study, we performed an investigation of the kinetics of the reaction of dry reforming of methane (DRM). For this work we used nickel-containing cerium-zirconium oxides. The catalysts were prepared in supercritical iso-propanol and characterized by means of XRD, TEM, XPS. It was found that the rate of DRM reaction becomes first order in methane and zero order in carbon dioxide. The dependences of the conversions of the reactants and product yields were obtained as a function of the contact time and the concentration of the initial mixture. In addition, the effective activation energy was calculated. It was shown that the most active and stable catalyst is 5wt% Ni/Ce0.5Zr0.5O[2]

A phase I study of a dual PI3-kinase/mTOR inhibitor BEZ235 in adult patients with relapsed or refractory acute leukemia

Background Combined inhibition of phosphatidylinositol 3-kinase (PI3K) and the mammalian target of rapamycin (mTOR) complexes may be an efficient treatment for acute leukemia. The primary objective of this phase I single center open label study was to determine the maximum tolerated dose (MTD) and recommended phase II dose (RP2D) of the dual pan-class I PI3K and mTOR inhibitor BEZ235 in patients with advanced leukemia. Methods Herein patients > 18 years of age who had relapsed or showed refractory leukemia were treated with BEZ235 (orally at 300–400 mg BID (cohort − 1/1)) to assess safety, tolerability, preliminary efficacy and pharmacokinetic (PK). Adverse events data and serious adverse events were analyzed and haematological and clinical biochemistry toxicities were assessed from laboratory test parameters. Response was assessed for the first time at the end of cycle 1 (day 29) and after every subsequent cycle. Pharmacokinetic and pharmacodynamic analyses of BEZ235 were also included (BEZ235 plasma levels, phosphorylation of AKT, S6 and 4EBP1). On statistics this trial is a multiple ascending dose study in which a following variant of the 3 + 3 rule (“Rolling Six”), a minimum of 6 and a maximum of 12 patients was recruited for the dose escalation and another 5 were planned for the expansion phase. Results Twenty-four patients with ALL (n = 11) or AML (n = 12) or CML-BP (n = 1) were enrolled. All patients had failed one (n = 5) or more lines of therapy (n = 5) and 14 patients were in refractory / refractory relapse. No formal MTD was defined, stomatitis and gastrointestinal toxicity at 400 mg BID dose was considered incompatible with prolonged treatment. The RP2D of BEZ235 was defined as 300 mg BID. Four of 24 patients showed clinical benefit. Twenty-two of 24 patients discontinued because of progression, (median time to progression 27 days (4d-112d). There was no association between PK parameters and efficacy or tolerability. Conclusions Combined inhibition of PI3K and mTOR inhibits a clinically meaningful driver pathway in a small subset of patients with ALL, with no benefit in patients with AML

CDKN1B, encoding the cyclin-dependent kinase inhibitor 1B (p27), is located in the minimally deleted region of 12p abnormalities in myeloid malignancies and its low expression is a favorable prognostic marker in acute myeloid leukemia

The online version of this article has a Supplementary Appendix. Background Alterations of the short arm of chromosome 12 (12p) occur in various hematologic malignancies and ETV6 and CDKN1B, which are located on 12p, have been implicated as leukemogenic genes of interest. Design and Methods We selected seven patients with myeloid malignancies and small 12p deletions detected by fluorescence in situ hybridization encompassing only the region centromeric of ETV6 and further evaluated them by single nucleotide polymorphism microarrays. Results The minimally deleted region contained only nine genes. These genes were subsequently analyzed by microarray expression profiling in an independent cohort of 781 patients, most, but not all, of whom had different hematologic malignancies CREBL2, MANSC1, and CDKN1B were expressed in more than 25% of cases, while the other six genes were expressed in only a minority of cases. As CDKN1B is a cell cycle regulator and functions as a tumor suppressor gene, this gene was selected for further expression studies in 286 patients with acute myeloid leukemia. When comparing patients with low CDKN1B expression (expression level <1,160; 1 st quartile) with those with intermediate or high expression (2 nd -4 th quartiles), certain mutations were observed more frequently in the former: RUNX1-RUNX1T1 (11/83, 13.3% versus 5/203; 2.5%; P=0.001), PML-RARA rearrangements (11/83, 13.3% versus 4/203, 2.0%; P<0.001), 11q23/MLL rearrangements (6/83, 7.2% versus 4/203, 2.0%; P=0.038), and FLT3-TKD mutations (7/63, 11.1% versus 6/167, 3.6%; P=0.047). The median overall survival of patients with low CDKN1B expression was longer than that of patients with intermediate/high expression (not reached versus 14.9 months; P=0.005). Likewise, patients with low CDKN1B expression had a longer event-free survival than those with intermediate/high expression (31.0 versus 9.7 months; P=0.013). Conclusions CDKN1B is an interesting candidate gene as a potential biomarker for prognostication in acute myeloid leukemia. Key words: 12p deletion, ETV6, CDKN1B, acute myeloid leukemia (AML), prognosis. leukemia. Haematologica 2011;96(6):829-836. doi:10.3324/haematol.2010 CDKN1B, encoding the cyclin-dependent kinase inhibitor 1B (p27), is located in the minimally deleted region of 12p abnormalities in myeloid malignancies and its low expression is a favorable prognostic marker in acute myeloid leukemia Citation: Haferlach C, Bacher U, Kohlmann A, Schindela S, Alpermann T, Kern W, Schnittger S, and Haferlach T. CDKN1B, encoding the cyclin-dependent kinase inhibitor 1B (p27), is located in the minimally deleted region of 12p abnormalities in myeloid malignancies and its low expression is a favorable prognostic marker in acute myeloi

Block of C/EBPα function by phosphorylation in acute myeloid leukemia with FLT3 activating mutations

Mutations constitutively activating FLT3 kinase are detected in ∼30% of acute myelogenous leukemia (AML) patients and affect downstream pathways such as extracellular signal–regulated kinase (ERK)1/2. We found that activation of FLT3 in human AML inhibits CCAAT/enhancer binding protein α (C/EBPα) function by ERK1/2-mediated phosphorylation, which may explain the differentiation block of leukemic blasts. In MV4;11 cells, pharmacological inhibition of either FLT3 or MEK1 leads to granulocytic differentiation. Differentiation of MV4;11 cells was also observed when C/EBPα mutated at serine 21 to alanine (S21A) was stably expressed. In contrast, there was no effect when serine 21 was mutated to aspartate (S21D), which mimics phosphorylation of C/EBPα. Thus, our results suggest that therapies targeting the MEK/ERK cascade or development of protein therapies based on transduction of constitutively active C/EBPα may prove effective in treatment of FLT3 mutant leukemias resistant to the FLT3 inhibitor therapies

The diagnosis of BCR/ABL-negative chronic myeloproliferative diseases (CMPD): a comprehensive approach based on morphology, cytogenetics, and molecular markers

Recent years showed significant progress in the molecular characterization of the chronic myeloproliferative disorders (CMPD) which are classified according to the WHO classification of 2001 as polycythemia vera (PV), chronic idiopathic myelofibrosis (CIMF), essential thrombocythemia (ET), CMPD/unclassifiable (CMPD-U), chronic neutrophilic leukemia, and chronic eosinophilic leukemia (CEL)/hypereosinophilic syndrome, all to be delineated from BCR/ABL-positive chronic myeloid leukemia (CML). After 2001, the detection of the high frequency of the JAK2V617F mutation in PV, CIMF, and ET, and of the FIP1L1–PDGFRA fusion gene in CEL further added important information in the diagnosis of CMPD. These findings also enhanced the importance of tyrosine kinase mutations in CMPD and paved the way to a more detailed classification and to an improved definition of prognosis using also novel minimal residual disease (MRD) markers. Simultaneously, the broadening of therapeutic strategies in the CMPD, e.g., due to reduced intensity conditioning in allogeneic hematopoietic stem cell transplantation and the introduction of tyrosine kinase inhibitors in CML, in CEL, and in other ABL and PDGRFB rearrangements, increased the demands to diagnostics. Therefore, today, a multimodal diagnostic approach combining cytomorphology, cytogenetics, and individual molecular methods is needed in BCR/ABL-negative CMPD. A stringent diagnostic algorithm for characterization, choice of treatment, and monitoring of MRD will be proposed in this review

Protein kinase c-β-dependent activation of NF-κB in stromal cells is indispensable for the survival of chronic lymphocytic leukemia B cells in vivo

Tumor cell survival critically depends on heterotypic communication with benign cells in the microenvironrnent. Here, we describe a survival signaling pathway activated in stromal cells by contact to B cells from patients with chronic lymphocytic leukemia (CLL). The expression of protein kinase C (PKC)-beta II and the subsequent activation of NF-kappa B in bone marrow stromal cells are prerequisites to support the survival of malignant B cells. PKC-beta knockout mice are insusceptible to CLL transplantations, underscoring the in vivo significance of the PKC-beta II-NF-kappa B signaling pathway in the tumor microenvironment. Upregulated stromal PKG-beta II in biopsies from patients with CLL, acute lymphoblastic leukemia, and mantle cell lymphoma suggests that this pathway may commonly be activated in a variety of hematological malignancies