Raman spectroscopy can recognize the KMT2A rearrangement as a distinct subtype of leukemia.

Acute lymphoblastic leukemia (ALL) and acute myeloid leukemia (AML) are the two most common hematologic malignancies, challenging to treat and associated with high recurrence and mortality rates. This work aims to identify specific Raman biomarkers of ALL cells with the KMT2A gene rearrangement (KMT2A-r), representing a highly aggressive subtype of childhood leukemia with a poor prognosis. The proposed approach combines the sensitivity and specificity of Raman spectroscopy with machine learning and allows us to distinguish not only myelo- and lymphoblasts but also discriminate B-cell precursor (BCP) ALL with KMT2A-r from other blasts of BCP-ALL. We have found that KMT2A-r ALL cells fixed with 0.5% glutaraldehyde exhibit a unique spectroscopic profile that enables us to identify this subtype from other leukemias and normal cells. Therefore, a rapid and label-free method was developed to identify ALL blasts with KMT2A-r based on the ratio of the two Raman bands assigned to phenylalanine - 1040 and 1008 cm-1. This is the first time that a particular group of leukemic cells has been identified in a label-free way. The identified biomarker can be used as a screening method in diagnostic laboratories or non-reference medical centers.

Raman classification of selected subtypes of acute lymphoblastic leukemia (ALL).

B-cell precursor acute lymphoblastic leukemia (BCP-ALL) with chromosome translocations like KMT2A gene rearrangement (KMT2A-r) and BCR-ABL1 fusion gene have been recognized as crucial drivers in both BCP-ALL leukemogenesis and treatment management. Standard diagnostic protocols for proliferative diseases of the hematopoietic system, like KMT2A-r-ALL, are genetically based and strongly molecularly oriented. Therefore, an efficient diagnostic procedure requires not only experienced and multidisciplinary laboratory staff but also considerable instrumentation and material costs. In recent years, a Raman spectroscopy method has been increasingly used to detect subtle chemical changes in individual cells resulting from stress or disease. Therefore, the objective of this study was to identify Raman signatures for the molecular subtypes and to develop a classification method based on the unique spectroscopic profile of in vitro models that represent specific aberrations aimed at KMT2A-r (RS4;11, and SEM) and the BCR-ABL1 fusion gene (SUP-B15, BV-173, and SD-1). Data analysis was based on chemometric methods, i.e. principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and support vector machine (SVM). The PCA-based multivariate model was used for pattern recognition of each investigated group of cells while PLS-DA and SVM were used to build models for the discrimination of spectra from the studied BCP-ALL molecular subtypes. The results showed that the studied molecular subtypes of ALL have characteristic spectroscopic profiles reflecting their peculiar biochemical state. The content of lipids (1600 cm-1), nucleic acids (789 cm-1), and haemoproteins (754, 1130, and 1315 cm-1), which are crucial in cell metabolism, was indicated as the main source of differentiation between subtypes. Identification of spectroscopic markers of cells with BCR-ABL1 or KMT2A-r may be useful in pharmacological studies to monitor the effectiveness of chemotherapy and further to understand differences in molecular responses between leukemia primary cells and cell lines.

Reliable cell preparation protocol for Raman imaging to effectively differentiate normal leukocytes and leukemic blasts.

Leukemias are a remarkably diverse group of malignancies originating from abnormal progenitor cells in the bone marrow. Leukemia subtypes are classified according to the cell type that has undergone neoplastic transformation using demanding and time-consuming methods. Alternative is Raman imaging that can be used both for living and fixed cells. However, considering the diversity of leukemic cell types and normal leukocytes, and the availability of different sample preparation protocols, the main objective of this work was to verify them for leukemia and normal blood cell samples for Raman imaging. The effect of glutaraldehyde (GA) fixation in a concentration gradient (0.1 %, 0.5 %, and 2.5 % GA) on the molecular structure of T-cell acute lymphoblastic leukemia (T-ALL) and peripheral blood mononuclear cells (PBMCs) was verified. Changes in the secondary structure of proteins within cells were indicated as the main effect of fixation, as shown by an increase in band intensity at 1041 cm-1, characteristic for in-plane δ(CH) deformation in phenylalanine (Phe). Different sensitivity of mononuclear and leukemic cells to fixation was observed. While the 0.1 % concentration of GA was too low to preserve the cell structure for an extended period of time, a GA concentration of 0.5 % seemed optimal for both normal and malignant cells. Chemical changes in PBMCs samples stored for 11 days were also investigated, which manifested in numerous modifications in the secondary structure of proteins and the content of nucleic acids. The impact of cell preculturing for 72 h after unbanking was verified, and there was no significant effect on the molecular structure of cells fixed with 0.5 % GA. In summary, the developed protocol for the preparation of samples for Raman imaging allows for the effective differentiation of fixed normal leukocytes from malignant T lymphoblasts.

The kinetics of blast clearance are associated with copy number alterations in childhood B-cell acute lymphoblastic leukemia.

We analyzed the pattern of whole-genome copy number alterations (CNAs) and their association with the kinetics of blast clearance during the induction treatment among 195 pediatric patients with B-cell precursor acute lymphoblastic leukemia (BCP-ALL) who displayed intermediate or high levels of minimal residual disease (MRD). Using unsupervised hierarchical clustering of CNAs > 5 Mbp, we dissected three clusters of leukemic samples with distinct kinetics of blast clearance [A - early slow responders (n=105), B - patients with persistent leukemia (n=24), C - fast responders with the low but detectable disease at the end of induction (n=66)] that corresponded with the patients' clinical features, the microdeletion profile,the presence of gene fusions and patients survival. Low incidence of large CNAs and chromosomal numerical aberrations occurred in cluster A which included ALL samples showing recurrent microdeletions within the genes encoding transcription factors (i.e., IKZF1, PAX5, ETV6, and ERG), DNA repair genes (XRCC3 and TOX), or harboring chromothriptic pattern of CNAs. Low hyperdiploid karyotype with trisomy 8 or hypodiploidy was predominantly observed in cluster B. Whereas cluster C included almost exclusively high-hyperdiploid ALL samples with concomitant mutations in RAS pathway genes. The pattern of CNAs influences the kinetics of leukemic cell clearance and selected aberrations affecting DNA repair genes may contribute to BCP-ALL chemoresistance.

Raman-based spectrophenotyping of the most important cells of the immune system.

INTRODUCTION: Human peripheral blood mononuclear cells (PBMCs) are a heterogeneous population of cells that includes T and B lymphocytes. The total number of lymphocytes and their percentage in the blood can be a marker for the diagnosis of several human diseases. Currently, cytometric methods are widely used to distinguish subtypes of leukocytes and quantify their number. These techniques use cell immunophenotyping, which is limited by the number of fluorochrome-labeled antibodies that can be applied simultaneously. OBJECTIVE: B and T lymphocytes were isolated from peripheral blood obtained from healthy human donors. METHODS: The immunomagnetic negative selection was used for the enrichment of B and T cells fractions, and their purity was assessed by flow cytometry. Isolated cells were fixed with 0.5% glutaraldehyde and measured using confocal Raman imaging. K-means cluster analysis, principal component analysis and partial least squares discriminant methods were applied for the identification of spectroscopic markers to distinguish B and T cells. HPLC was the reference method for identifying carotene in T cells. RESULTS: Reliable discrimination between T and B lymphocytes based on their spectral profile has been demonstrated using label-free Raman imaging and chemometric analysis. The presence of carotene in T lymphocytes (in addition to the previously reported in plasma) was confirmed and for the first time unequivocally identified as ß-carotene. In addition, the molecular features of the lymphocytes nuclei were found to support the discriminant analysis. It has been shown that although the presence of carotenoids in T cells depends on individual donor variability, the reliable differentiation between lymphocytes is possible based on Raman spectra collected from individual cells. CONCLUSIONS: This proves the potential of Raman spectroscopy in clinical diagnostics to automatically differentiate between cells that are an important component of our immune system.

Towards Raman-Based Screening of Acute Lymphoblastic Leukemia-Type B (B-ALL) Subtypes.

Acute lymphoblastic leukemia (ALL) is the most common type of malignant neoplasms in the pediatric population. B-cell precursor ALLs (BCP-ALLs) are derived from the progenitors of B lymphocytes. Traditionally, risk factors stratifying therapy in ALL patients included age at diagnosis, initial leukocytosis, and the response to chemotherapy. Currently, treatment intensity is modified according to the presence of specific gene alterations in the leukemic genome. Raman imaging is a promising diagnostic tool, which enables the molecular characterization of cells and differentiation of subtypes of leukemia in clinical samples. This study aimed to characterize and distinguish cells isolated from the bone marrow of patients suffering from three subtypes of BCP-ALL, defined by gene rearrangements, i.e., BCR-ABL1 (Philadelphia-positive, t(9;22)), TEL-AML1 (t(12;21)) and TCF3-PBX1 (t(1;19)), using single-cell Raman imaging combined with multivariate statistical analysis. Spectra collected from clinical samples were compared with single-cell spectra of B-cells collected from healthy donors, constituting the control group. We demonstrated that Raman spectra of normal B cells strongly differ from spectra of their malignant counterparts, especially in the intensity of bands, which can be assigned to nucleic acids. We also showed that the identification of leukemia subtypes could be automated with the use of chemometric methods. Results prove the clinical suitability of Raman imaging for the identification of spectroscopic markers characterizing leukemia cells.

New Insights into Red Blood Cell Microcytosis upon mTOR Inhibitor Administration.

The aim of this study was to evaluate the effect of everolimus, a mammalian target of rapamycin (mTOR) inhibitor, on red blood cell parameters in the context of iron homeostasis in patients with tuberous sclerosis complex (TSC) and evaluate its effect on cell size in vitro. Everolimus has a significant impact on red blood cell parameters in patients with TSC. The most common alteration was microcytosis. The mean MCV value decreased by 9.2%, 12%, and 11.8% after 3, 6, and 12 months of everolimus treatment. The iron level declined during the first 3 months, and human soluble transferrin receptor concentration increased during 6 months of therapy. The size of K562 cells decreased when cultured in the presence of 5 µM everolimus by approximately 8%. The addition of hemin to the cell culture with 5 µM everolimus did not prevent any decrease in cell size. The stage of erythroid maturation did not affect the response to everolimus. Our results showed that the mTOR inhibitor everolimus caused red blood cell microcytosis in vivo and in vitro. This effect is not clearly related to a deficit of iron and erythroid maturation. This observation confirms that mTOR signaling plays a complex role in the control of cell size.

Gene expression of ASNS, LGMN and CTSB is elevated in a subgroup of childhood BCP-ALL with PAX5 deletion.

Resistance to L-asparaginase (L-asp) is a major contributor to poor treatment outcomes of several subtypes of childhood B cell precursor acute lymphoblastic leukemia (BCP-ALL). Asparagine synthetase (ASNS), legumain (LGMN) and cathepsin B (CTSB) serve a key role in L-asp resistance. The association between genetic subtypes of BCP-ALL and the expression of ASNS, LGMN and CTSB may elucidate the mechanisms of treatment failure. Bone marrow samples of 52 children newly diagnosed with BCP-ALL were screened for major genetic abnormalities and ASNS, LGMN and CTSB gene expression levels. The cohort was further divided into groups corresponding to the key genetic aberrations occurring in BCP-ALL: Breakpoint cluster region and Abelson murine leukemia viral oncogene homolog 1 fusion; hyperdiploidy, hypodiploidy, ETS variant 6 and runt-related transcription factor 1 fusion and other BCP-ALL with no primary genetic aberration identified. A subgroup analysis based on the differences in copy number variations demonstrated a significant increase of ASNS, LGMN and CTSB median expression in other BCP-ALL cases with paired box 5 (PAX5) deletion (P=0.0117; P=0.0036; P<0.0001, respectively) compared with those with wild-type PAX5. Patients with high ASNS expression exhibited longer relapse-free survival (RFS) compared with those with low ASNS levels (P=0.0315; HR, 0.19; 95% CI, 0.04-0.86); the 5-year RFS for patients in the high ASNS expression group was 90.15% (95% CI, 87.90-92.40%). Despite the impact on ASNS, LGMN and CTSB expression, PAX5 deletion did not influence RFS in the other BCP-ALL group (P=0.6839). Therefore, the results of the present study revealed high levels of ASNS, LGMN and CTSB expression in the other BCP-ALL group with concomitant PAX5 deletion and no subsequent deterioration in 5-year RFS. High ASNS expression level, as a single factor, was strongly associated with an improved outcome.

Air Contamination by Mercury, Emissions and Transformations-a Review.

The present and future air contamination by mercury is and will continue to be a serious risk for human health. This publication presents a review of the literature dealing with the issues related to air contamination by mercury and its transformations as well as its natural and anthropogenic emissions. The assessment of mercury emissions into the air poses serious methodological problems. It is particularly difficult to distinguish between natural and anthropogenic emissions and re-emissions from lands and oceans, including past emissions. At present, the largest emission sources include fuel combustion, mainly that of coal, and "artisanal and small-scale gold mining" (ASGM). The distinctly highest emissions can be found in South and South-East Asia, accounting for 45% of the global emissions. The emissions of natural origin and re-emissions are estimated at 45-66% of the global emissions, with the largest part of emissions originating in the oceans. Forecasts on the future emission levels are not unambiguous; however, most forecasts do not provide for reductions in emissions. Ninety-five percent of mercury occurring in the air is Hg0-GEM, and its residence time in the air is estimated at 6 to 18 months. The residence times of its HgII-GOM and that in Hgp-TPM are estimated at hours and days. The highest mercury concentrations in the air can be found in the areas of mercury mines and those of ASGM. Since 1980 when it reached its maximum, the global background mercury concentration in the air has remained at a relatively constant level.

Mercury in Marine and Oceanic Waters-a Review.

Mercury contamination in water has been an issue to the environment and human health. In this article, mercury in marine and oceanic waters has been reviewed. In the aquatic environment, mercury occurs in many forms, which depend on the oxidation-reduction conditions. These forms have been briefly described in this article. Mercury concentrations in marine waters in the different parts of the world have been presented. In the relevant literature, two models describing the fate and behavior of mercury in saltwater reservoirs have been presented, a conceptual model which treats all the oceans as one ocean and the "ocean margin" model, providing that the ocean margins manifested themselves as the convergence of continents and oceans, covering such geological features, such as estuaries, inland seas, and the continental shelf. These two conceptual models have been summarized in the text. The mercury content in benthic sediments usually reflects is level in the water reservoir, particularly in reservoirs situated in contaminated areas (mines, metallurgical plants, chemically protected crops). The concentrations of mercury and its compounds determined in the sediments in surface waters in the different parts of the world have been presented. Due to the fact that the pollution caused by mercury is a serious threat for the marine environment, the short paragraph about mercury bioaccumulation in aquatic organisms has been included. The cited data demonstrated a large scatter of mercury contents both between the fish species and the water areas. Mathematical models, valuable tools which provide information about the possible responses of ecosystems, developed to simulate mercury emissions, both at a small scale, for local water reservoirs, and at a global scale, as well as to model mercury bioaccumulation in the chain web of aquatic systems have been described.

Phenotypic diversity of patient-derived melanoma populations in stem cell medium.

Melanomas are highly heterogeneous tumors and there is no treatment effective at achieving long-term remission for metastatic melanoma patients. Thus, an appropriate model system for studying melanoma biology and response to drugs is necessary. It has been shown that composition of the medium is a critical factor in preserving the complexity of the tumor in in vitro settings, and melanospheres maintained in stem cell medium are a good model in this respect. In the present study, we observed that not all nodular melanoma patient-derived cell populations grown in stem cell medium were capable of forming melanospheres, and cell aggregates and anchorage-independent single-cell cultures emerged instead. Self-renewing capacity and unlimited growth potential indicated the presence of cells with stem-like properties in all patient-derived populations but immunophenotype and MITF expression exhibited variability. Enhanced MITF expression and activity was observed in melanospheres in comparison with cell aggregates and single-cell culture, and hypoxic-like conditions that increased the ability of single-cell population to form melanospheres enhanced MITF expression and cell pigmentation as well. Thus, MITF seems to be a critical transcription factor for formation of both patient-derived and hypoxia-induced melanospheres. After 2 years of continuous culturing, melanospheres progressively underwent transition into cell aggregates that was accompanied by changes in expression of several MITF-dependent genes associated with melanogenesis and survival and alterations in the composition of subpopulations but not in the frequency of ABCB5-positive cells. Several biological properties of parent tumor are well preserved in patient-derived melanospheres, but during prolonged culturing the heterogeneity is substantially lost when the melanospheres are substituted by cell aggregates. This should be considered when cell aggregates instead of melanospheres are used in the study of melanoma biology and cell response to drugs.

Sphere formation and self-renewal capacity of melanoma cells is affected by the microenvironment.

Melanomas contain subsets of cancer stem-like cells with tumor-initiating capacity. The frequency of these cells in the tumor is still a topic of debate. We investigated the phenotypic plasticity of cancer cells grown as melanospheres to elucidate the influence of the microenvironment on some features of melanoma stem-like cells. Cells from surgical specimens of nodular melanoma were grown as anchorage-independent melanospheres in a stem cell medium and as adherent monolayer cultures in the presence of serum. Proliferation and viability were measured by cell counting and an acid phosphatase assay; surface marker expression was evaluated by flow cytometry, and the clonogenic potential of single cells was assessed by growth in soft agar. Patient-derived melanoma cells could be maintained in cell culture for more than 16 months when grown as melanospheres. In the presence of serum, melanospheres completely changed their growth characteristics and formed adherent monolayers. The transition from melanospheres to monolayers was accompanied by an apparent loss of clonogenic potential, an increased proliferation rate, and altered expressions of cell surface markers ABCB5, CD133, and CD49f. These changes, however, were reversible. Compared with adherent monolayer cultures, melanospheres are enriched in cells with clonogenic potential, reflecting the self-renewing capacity of cancer stem-like cells. This clonogenic potential can be lost and regained depending on the growth conditions. Our results demonstrate how easily melanoma cells change their function upon exposure to external stimuli and suggest that the frequency of melanoma stem-like cells strongly depends on the microenvironment.

STI571 and morpholine derivative of doxorubicin collaborate in inhibition of K562 cell proliferation by inducing differentiation and mitochondrial pathway of apoptosis.

Differentiation therapy is considered as a supplementary approach to the currently applied treatments for leukemia. We have previously shown that a morpholine derivative of doxorubicin (DOXM) appeared to be a more efficient inducer of erythroid differentiation of K562 cells than the parent drug [Czyz, M., Szulawska, A., Bednarek, A.K., Duchler, M., 2005. Effects of anthracycline derivatives on human leukemia K562 cell growth and differentiation. Biochem. Pharmacol. 70, 1431-1442.; Szulawska, A., Arkusinska, J., Czyz, M., 2007. Accumulation of gamma-globin mRNA and induction of irreversible erythroid differentiation after treatment of CML cell line K562 with new doxorubicin derivatives. Biochem. Pharmacol. 73, 175-184.]. In the current study we used this compound in combination with STI571, a front-line drug in therapy of chronic myelogenous leukemia (CML), to evaluate possible benefits of the combined treatment on the cellular level. Using K562 cells, we analyzed the response of CML cells to low concentrations of DOXM when Bcr-Abl activity was reduced to various levels by its specific inhibitor, STI571. Differentiation was significantly enhanced with the combination of 150 nM STI571 and 100 nM DOXM as compared to the levels obtained with either drug alone. A higher concentration of STI571 was required to diminish Bcr-Abl activity to the level which was sufficient to stimulate apoptotic cell death pathway in K562. Apoptosis induced by 250 nM STI571 was markedly enhanced by DOXM in the combined treatment. Mitochondrial transmembrane potential dissipation and translocation of phosphatydylserine to the outer plasma membrane were increased by 50%. Our results clearly indicate that differentiation and apoptosis, both reducing cellular proliferation, could be substantially enhanced by the combined treatment. We provide experimental evidence implicating that the diversification of cellular effects obtained in the combined treatment employing non-toxic approaches to enhance efficacy of STI571 might be considered as an alternative therapeutic strategy against CML, especially for apoptosis-reluctant cells.

STI571/doxorubicin concentration-dependent switch for diverse caspase actions in CML cell line K562.

UNLABELLED: We examined the response of the apoptosis-reluctant CML cell line K562 to doxorubicin alone or in combination with the tyrosine kinase inhibitor STI571. We found that at clinically relevant concentrations, doxorubicin induced differentiation and senescence, but did not induce apoptosis. Doxorubicin induced G(2)/M arrest and mitochondrial transmembrane potential dissipation. Interestingly, drug-induced differentiation could be diminished by caspase inhibitors. STI571 caused a graded response characterized by differentiation at low concentrations and apoptosis at higher. STI571 was not observed to induce senescence. Combination of STI571 and caspase inhibitors protected cells from apoptosis but did not influence differentiation. The diverse mode of action of both drugs contributed to the response observed during combination treatment. An additive effect on proliferation was obtained. The mechanisms contributing to inhibition of cellular proliferation were complex and strongly dependent on the applied drug concentrations. Differentiation or apoptosis were enhanced by combined treatment only in narrow ranges of concentrations. CONCLUSION: DOX and STI571 along diverse mechanisms contributed to elevated levels of activated caspases which might be then responsible for a switch from differentiation to apoptosis.

Combined effects of doxorubicin and STI571 on growth, differentiation and apoptosis of CML cell line K562.

STI571 (imatinib mesylate; Gleevec) is an inhibitor that targets the tyrosine kinase activity of Bcr-Abl present in chronic myelogenous leukemia (CML) cells. Some preclinical studies have demonstrated that the combination of STI571 with chemotherapeutic drugs results in enhanced toxicity in Bcr-Abl-positive leukemias. We investigated the potential benefit of using STI571 to down-regulate Bcr-Abl activity for the enhancement of doxorubicin anti-proliferative action in K562 cell line derived from blast crisis of CML. At low concentrations of both drugs (40 nM doxorubicin combined with STI571 in the range of 100-150 nM), the antiproliferative effects were mainly due to cellular differentiation as assessed by benzidine staining for hemoglobin synthesis level and real-time PCR for gamma-globin expression. Higher concentrations of STI571 used in combinations with doxorubicin caused mainly apoptosis as shown by DNA degradation and nuclear fragmentation visualized by fluorescence microscopy after DAPI staining, changes in cell morphology observed after Giemza-May Grünwald staining and cellular membrane organization estimated by flow cytometry after Annexin V staining. As compared with either drug alone, cotreatment with STI571 and DOX induced stronger cellular responses. A low concentration of STI571 in combination with a low concentration of DOX might be tested as an alternative approach to increasing the efficacy of chemotherapy against CML.

[Influence of environmental tobacco smoking on different barium contents in pharyngeal tonsils]. / Wplyw biernego palenia na zrónicowanie wystepowania baru w migdalkach gardlowych dzieci.

The aim of this study was analysis of content of barium in pharyngeal tonsils from children living on Southern Poland in during exposure on ETS. Barium contents in pharyngeal tonsils were higher for no exposure ETS children (0.099 microg/g) in comparison to exposure by ETS (0.033 microg/g), p=0.0042. The interaction between Ba and Ca, Fe, K or higher arithmetic and geometric mean, statistical range of changes contributed the role ETS in relation to children.

[STI571: a summary of targeted therapy]. / STI571--terapia celowana.

STI571 (imatinib; Gleevec) was developed as the first molecularly targeted therapy. It was the result of an extensive search for molecules to block the aberrant activity of Abl kinase in the fusion protein Bcr-Abl. In addition, it can specifically inhibit the activity of c-Kit and PDGF receptors. This orally bioavailable drug has a low toxicity profile. It is approved to treat the patients with chronic myelogenous leukemia (CML) or gastrointestinal stromal tumor (GIST). It produces hematological, cytogenetic, and molecular remission with significant efficacy, particularly in patients with chronic-phase CML. However, there is well-documented proof of primary and secondary resistance to STI571 with progression of leukemia. More evidence indicates that this single drug may not be sufficient to completely eradicate BCR-ABL-positive stem cells. A variety of strategies has already been developed to improve the effectiveness of CML treatment, including targeting the expression or stability of the Bcr-Abl kinase itself, targeting signaling pathways activated by this kinase, as well as designing novel Abl inhibitors. In this review the molecular mechanisms of STI571 action, its effectivenes against CML, GIST, and melanoma, as well as new approaches to improve its efficacy, mainly by overcoming STI571 resistance, are discussed.

[Novel inhibitors of Bcr-Abl]. / Alternatywne dla STI571 inhibitory kinazy Bcr-Abl.

STI571 (imatinib; Gleevec) was developed to specifically target the tyrosine kinase activity of the Bcr-Abl protein in Philadelphia chromosome-positive chronic myeloid leukemia (CML). It also inhibits the activity of c-Kit and PDGFR. It is the first-line drug for newly diagnosed CML, with remarkable efficacy to patients in the chronic phase of this cancer. However, CML patients in the accelerated phase or blast crisis often relapse due to drug resistance. STI571 fails to eradicate leukemic stem cells, and BCR-ABL(+). cells remain detectable in the majority of patients. The necessity for alternative or additional treatment for STI571-resistant leukemia resulted in the development of a second generation of drugs for targeted therapies. In this review a literature overview of the alternative inhibitors which were designed to override STI571 resistance and decrease the aberrant kinase activity of Bcr-Abl protein with higher efficiency is presented.

[The changes of barium and strontium content in gallstones from smoking and non-smoking women]. / Zmiany zawartosci baru i strontu w zlogach kamieni zólciowych kobiet palacych i niepalacych tyton.

The problem of statistical characteristics of occurrence barium and strontium content in gallstones from smoking and non-smoking women living in southern Poland is presented in the work. The subjects of the research were gallstones, gained intraoperatively from 146 women (49 smoking, 97 non-smoking). The content of barium and strontium was determined using atomic emission spectroscopy with inductive coupled plasma (ICP-AES). The statistical characteristic of barium and strontium content in gallstones shows that smoking does not decide about content level of these elements in gallstones. Content of barium (2.32 microgBa/g) as well as strontium (3.23 microgSr/g) in gallstones from non smoking women are higher in comparison to content of these elements in gallstones from smoking women (1.91 microgBa/g and 2.76 microgSr/g).