Factors influencing overall survival and GvHD development after allogeneic hematopoietic stem cell transplantation - single centre experience.

BACKGROUNDS: Allogeneic hematopoietic stem cell transplantation (alloHSCT) is a substantial therapeutic procedure for the treatment of a wide spectrum of severe diseases. Despite advancements in treatment and supportive care, alloHSCT still carries a considerable mortality risk, primarily caused by graft-versus-host disease (GvHD). Our retrospective analysis aimed to identify the factors influencing overall survival and GvHD development in HLA-identical sibling alloHSCT. We have analyzed patients' and donors' age, AB0 compatibility, recipient-donor gender match, stem cell source, time from the diagnosis to alloHSCT, conditioning regimen type, GvHD prophylaxis, and relapse. PATIENTS AND METHODS: Our study included 96 patients (54 male, 42 female) who underwent HLA-identical sibling alloHSCT. The median follow-up was 64.5 months (range 1-218 months), and the median age of both recipients and donors was 34 years. Malignant hematological diseases were the most common indications for alloHSCT. RESULTS: GvHD and its complications accounted for the highest number of deaths (N = 24; 46.2%), followed by relapse (N = 18; 34.6%). Acute GvHD developed in 30 patients (31.3%), while chronic GvHD occurred in 25 patients (26.0%), resulting in a total of 45 patients (46.9%) experiencing GvHD. Male recipients with female donors had significantly worse overall survival compared to other patients (P = 0.01; HR = 2.33). Overall survival was better in patients transplanted within 1 year from the diagnosis compared to those transplanted after 1 year (P = 0.03; HR = 1.93). No factor reached statistical significance regarding the impact on acute GvHD, chronic GvHD, or overall GvHD. CONCLUSION: We confirmed that sex mismatch, specifically in the case of a female donor and a male recipient, significantly negatively affects overall survival after alloHSCT. Additionally, overall survival is significantly shorter when the interval between the diagnosis and alloHSCT exceeds one year.

Influence of different spectra of NOTCH3 variants on the clinical phenotype of CADASIL - experience from Slovakia.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common hereditary vascular disorder causing ischaemic attacks and strokes in middle-aged adults. Though the clinical spectrum includes some typical symptoms, recognition of the disease, especially at an earlier stage, is very difficult because of the highly variable manifestation and incomplete clinical picture. Characteristic brain MRI findings and the presence of pathogenic variants in the NOTCH3 gene are fundamental for CADASIL diagnosis. In this paper, we provide the first comprehensive report on CADASIL patients from Slovakia. Altogether, we identified 23 different pathogenic variants in 35 unrelated families. In our cohort of patients with clinical suspicion of CADASIL, we found a causal genetic defect and confirmed the diagnosis in 10.2% of cases. We present the case reports with up-to-date unpublished NOTCH3 variants and describe their phenotype-genotype correlation: p.(Cys65Phe), p.(Pro86Leu/Ser502Phe), p.(Arg156*), p.(Cys408Arg), p.(Tyr423Cys), p.(Asp1720His), and p.(Asp1893Thrfs*13). The most frequently described location for pathogenic variants was in exon 4, whereas the most common single variant was p.Arg1076Cys in exon 20. Based on the results of our study, we propose a re-evaluation of the criteria for the selection of patients suitable for NOTCH3 gene analysis. We hereby state that the currently used protocol of a high score requirement is not ideal for assessing molecular analysis, and it will be desirable to be less strict in criteria for genetic testing.

Cardiovascular events and atherogenic lipid profile in chronic myeloid leukemia patients treated with nilotinib versus imatinib.

OBJECTIVES: The aim of this study was to assess cardiotoxicity and potential adverse effects related to lipid metabolism during treatment with tyrosine kinase inhibitors (TKIs) imatinib and nilotinib in patients with chronic myeloid leukemia (CML). PATIENTS AND METHODS: Eighty-two consecutive patients with CML, who received nilotinib and/or imatinib in a single haemato-oncological Slovak center between years 2002-2018 were evaluated in a retrospective study. The mean age was 55.8 years (range 22-77 years). Median of follow-up was 61.3 months. RESULTS: A significantly higher incidence of dyslipidemia, significantly higher levels of potential risk markers of cardiovascular disease small dense LDL cholesterol (sdLDL-CH) and a significant increase in total cholesterol were found in the patients during treatment with nilotinib in comparison to imatinib. Dyslipidemia led to drug therapy in 22 % of the patients in the nilotinib group. Fourteen percent of the patients in the nilotinib group had one or more cardiovascular events, including peripheral artery disease (10 %), myocardial infarction (4 %) and stroke (4 %). CONCLUSION: A higher risk of cardiovascular events and atherogenic dyslipidemia were associated with nilotinib therapy. Patients treated with TKI, especially nilotinib, require an early modification of cardiovascular risk factors and a careful cardiologic surveillance so that antileukemic therapy with this highly effective agent could continue (Tab. 4, Fig. 3, Ref. 32). Text in PDF www.elis.sk Keywords: tyrosine kinase inhibitors, cardiovascular events, dyslipidemia, small dense LDL-cholesterol, nilotinib, imatinib.

The use of granulocyte colony stimulating factor after autologous hematopoietic stem cell transplantation.

Restrospective study to evaluate the efficacy of early vs. delayed initiation of G-CSF after autologous hematopoietic stem cell transplantation (AHSCT) in patients with lymphoid malignancies. BACKGROUND: Granulocyte colony stimulating factor (G-CSF) is commonly used after AHSCT to accelerate stem cell engraftment to minimize the morbidity and mortality associated with prolonged neutropenia. However, there is no consensus on the optimal timing of G-CSF after HSCT. METHODS: A total of 117 patients with lymphoid malignancies who underwent AHSCT were included. All patients received G-CSF (filgrastim 5 µg/kg s.c.) daily after AHSCT (43 patients on day 6-8 and 74 patients on day 3 or 4). All patients received standard conditioning regimen for the underlying disease, and standard supportive treatment, including treatment of febrile neutropenia. RESULTS: The incidence of severe neutropenia was 81 % vs 17 %, and very severe neutropenia 61 % vs 4 % in the delayed and early G-CSF groups, respectively (p < 0.0001). The rate of fungal infection was higher in the group of patients who received delayed G-CSF (p < 0.005). CONCLUSION: An early administration of G-CSF after AHSCT (on day 3 or 4) accelerates neutophil engraftment; decreases the incidence of severe neutropenia and the risk of infectious complications (especially fungal infections) (Tab. 1, Fig. 3, Ref. 22).

Diagnostic Tools of Waldenströms Macroglobulinemia - Best Possibilities for Non-invasive and Long-term Disease Monitoring.

Waldenströms macroglobulinemia (WM) is a B-cell malignancy characterized by high level of monoclonal immunoglobulin M (IgM) paraprotein in blood serum and associated with the bone marrow infiltration by malignant cells with lymphoplasmacytic differentiation. WM remains incurable advances in therapy. Most of WM cases are associated with a somatic point mutation L265P in MYD88. Significantly higher risk of progression from the IgM monoclonal gammopathy of undetermined significance (IgM MGUS) to WM for patients with mutated MYD88 gene suggests that this mutation is an early oncogenic event and plays a central role in development of malignant clones. The second, most prevalent mutation in WM is found in the CXCR4 gene and is often associated with drug resistance and aggressive disease presentation. Therefore, detection of these mutations (MYD88L265P and CXCR4S338X) could be useful diagnostic and prognostic tool for the patients with WM. While detection of these mutations in bone marrow sample is common, the aim of our study was to compare sensitivity of detection of mutation from different cell fraction from peripheral blood and bone marrow. The results show possibility to describe MYD88 and CXCR4 mutation status even from peripheral blood sample (sensitivity for MYD88L265P was 100%, for CXCR4S338X 91%), which significantly facilitate material collection. Moreover, comparable detection sensitivity of these mutations in bone marrow and peripheral blood samples examined before and during the therapy offers a promising tool for more routine diagnostic and monitoring of disease progression.Key words: Waldenström macroglobulinemia - hematology - neoplasms - lymphoma - mutation - MYD88 - CXCR4.

Autologous hematopoietic stem cell transplantation for acute myeloid leukemia - single center experience.

We aimed to determine the effect of autologous hematopoietic stem cell transplantation (auto-HSCT) on acute myeloid leukemia (AML) patients as a valid alternative therapeutic option for patients without HLA-compatible donor. This retrospective single center study included 79 patients with AML older than 18 years. In this report, we describe the patient characteristics, engraftment, toxicity of treatment, complications, overall survival, and relapse incidence of 79 patients treated chemotherapy and followed by auto-HSCT. The descriptive statistics was used, and the method of Kaplan and Meier was applied to calculate the actuarial rate of overall survival. The patients achieved an absolute neutrophile count (ANC) of ≥ 0.5 x109/l in between 10 to 40 days; median was 14 days after auto-HSCT. The patients achieved platelet count ≥ 20 x109/l in between 10 to 209 days; median was 19 days after auto-HSCT. Hundred-day mortality after autologous transplant was 6.57% (5/76). The relapse rate was 39.5% (32 patients) and 7 patients (8.6%) were lost from follow-up. On the date of evaluation (April 30, 2016), 48 patients (60.8%) were alive, including 7 (8.6%) patients who are lost from follow-up (not responding to check-up request). The 5-year overall survival (OS) was 60.8%; median overall survival was not reached. The present clinical study has demonstrated safety and efficacy of myeloablative chemotherapy followed by auto-HSCT in the treatment of AML in first remission.

Tumors overexpressing RNF168 show altered DNA repair and responses to genotoxic treatments, genomic instability and resistance to proteotoxic stress.

Chromatin DNA damage response (DDR) is orchestrated by the E3 ubiquitin ligase ring finger protein 168 (RNF168), resulting in ubiquitin-dependent recruitment of DDR factors and tumor suppressors breast cancer 1 (BRCA1) and p53 binding protein 1 (53BP1). This ubiquitin signaling regulates pathway choice for repair of DNA double-strand breaks (DSB), toxic lesions whose frequency increases during tumorigenesis. Recruitment of 53BP1 curbs DNA end resection, thereby limiting homologous recombination (HR) and directing DSB repair toward error-prone non-homologous end joining (NHEJ). Under cancer-associated ubiquitin starvation conditions reflecting endogenous or treatment-evoked proteotoxic stress, the ubiquitin-dependent accrual of 53BP1 and BRCA1 at the DNA damage sites is attenuated or lost. Challenging this current paradigm, here we identified diverse human cancer cell lines that display 53BP1 recruitment to DSB sites even under proteasome inhibitor-induced proteotoxic stress, that is, under substantial depletion of free ubiquitin. We show that central to this unexpected phenotype is overabundance of RNF168 that enables more efficient exploitation of the residual-free ubiquitin. Cells with elevated RNF168 are more resistant to combined treatment by ionizing radiation and proteasome inhibition, suggesting that such aberrant RNF168-mediated signaling might reflect adaptation to chronic proteotoxic and genotoxic stresses experienced by tumor cells. Moreover, the overabundant RNF168 and the ensuing unorthodox recruitment patterns of 53BP1, RIF1 and REV7 (monitored on laser micro-irradiation-induced DNA damage) shift the DSB repair balance from HR toward NHEJ, a scenario accompanied by enhanced chromosomal instability/micronuclei formation and sensitivity under replication stress-inducing treatments with camptothecin or poly(ADP-ribose) polymerase (PARP) inhibitor. Overall, our data suggest that the deregulated RNF168/53BP1 pathway could promote tumorigenesis by selecting for a more robust, better stress-adapted cancer cell phenotype, through altered DNA repair, fueling genomic instability and tumor heterogeneity. Apart from providing insights into cancer (patho)biology, the elevated RNF168, documented here also by immunohistochemistry on human clinical tumor specimens, may impact responses to standard-of-care and some emerging targeted cancer therapies.

Graft-versus-host disease management.

Graft-versus-host disease (GVHD) remains a major problem of allogeneic hematopoietic-stem cell transplantation (HSCT) and an obstacle for successful outcome. Clinically significant acute GVHD (grade II or higher) developed in 20 to 65 percent of the patients. Death due to this complication accounts for approximately 50 percent of the deaths that are not due to a relapse of the neoplasm. Up to 70 % of patients who survive beyond day 100 develop chronic GVHD and it is the leading cause of nonrelapse mortality more than 2 years after allogeneic HSCT. In addition, chronic GVHD is associated with decreased quality of life, impaired functional status, and ongoing need for immunosuppressive medications. The incidence of chronic GVHD is increasing because of expansion of the donor population beyond HLA-identical siblings, older recipient age, use of peripheral blood cells as the graft source, and infusion of donor lymphocytes for treatment of recurrent malignancy after HSCT. With the current rush in new findings related to GVHD, we see a significant advancement in its management. Given these various new options and challenges, it is important to identify the minimal requirements for diagnosis and treatment of GVHD, as access to the most sophisticated advances may vary depending on local circumstances (Tab. 4, Fig. 1, Ref. 51).

Inflammatory environment created by fibroblast aggregates induces growth arrest and phenotypic shift of human myeloma cells.

Multiple myeloma (MM) is characterized by accumulation of clonal plasma cells (PCs) predominantly in the bone marrow but tumor cells appear in the circulation in significant numbers as the disease progress. The occurrence of circulating multiple myeloma cells raises question concerning interactions between these cells and stroma of peripheral organs specifically under certain pathophysiological conditions, e.g., inflammation. Therefore, in the present study we exposed three human multiple myeloma cell lines to sterile inflammation produced in a culture dish by clusters of cell-cell contact-activated dermal fibroblasts. We now observed that myeloma cells responded differently to this particular type of stromal cell activation, nemosis. Two cell lines U-266 and LP-1 were minimally affected by the proinflammatory signalling, while the third cell line RPMI 8226 responded with growth arrest and altered expression of three phenotypic markers CD38, CD45, and CD138, indicating dedifferentiation shift of these cells to less mature PC-like phenotype. In a preliminary study we identified a subclone of cells having similar phenotype in 14 out of 23 analysed specimens of MM patients. This set of data indicates that the observed phenomenon might be clinically relevant. Our results emphasize the potential role of activated stromal fibroblasts and subsequent inflammation in altering phenotype of PCs and directing myeloma progression towards dormancy. Given the significant implication of dormant myeloma cells that might serve as a major cellular basis for the relapse, understanding their unique biology and precise elucidation of the underlying molecular mechanisms for the maintenance of quiescence is important. Therefore, we consider this study as a particular contribution to development of experimental model for in vitro studies of cancer dormancy.

SETD2 loss-of-function promotes renal cancer branched evolution through replication stress and impaired DNA repair.

Defining mechanisms that generate intratumour heterogeneity and branched evolution may inspire novel therapeutic approaches to limit tumour diversity and adaptation. SETD2 (Su(var), Enhancer of zeste, Trithorax-domain containing 2) trimethylates histone-3 lysine-36 (H3K36me3) at sites of active transcription and is mutated in diverse tumour types, including clear cell renal carcinomas (ccRCCs). Distinct SETD2 mutations have been identified in spatially separated regions in ccRCC, indicative of intratumour heterogeneity. In this study, we have addressed the consequences of SETD2 loss-of-function through an integrated bioinformatics and functional genomics approach. We find that bi-allelic SETD2 aberrations are not associated with microsatellite instability in ccRCC. SETD2 depletion in ccRCC cells revealed aberrant and reduced nucleosome compaction and chromatin association of the key replication proteins minichromosome maintenance complex component (MCM7) and DNA polymerase δ hindering replication fork progression, and failure to load lens epithelium-derived growth factor and the Rad51 homologous recombination repair factor at DNA breaks. Consistent with these data, we observe chromosomal breakpoint locations are biased away from H3K36me3 sites in SETD2 wild-type ccRCCs relative to tumours with bi-allelic SETD2 aberrations and that H3K36me3-negative ccRCCs display elevated DNA damage in vivo. These data suggest a role for SETD2 in maintaining genome integrity through nucleosome stabilization, suppression of replication stress and the coordination of DNA repair.

Collection of peripheral hematopoietic stem/progenitor cells.

OBJECTIVES: Several variables possibly affecting collection of peripheral hematopoietic stem/progenitor cells (PBSC) were evaluated: type of apheresis machine (Amicus version 2.5, Baxter vs Cobe Spectra version 7.0, Terumo BCT), venous access (peripheral vein vs central venous catheter, i.g. CVC), and apheresis regimen (standard vs large volume leukapheresis, i.g. SVL vs LVL) with the objective to increase collection efficacy at the site. BACKGROUND: Peripheral blood represents the currently preferred source of hematopoietic stem/progenitor cells (HSCs) for transplantation. METHODS: Data regarding 169 collection procedures performed in healthy donors and patients between January 2008 and December 2011 at the Clinics of Haematology and Transfusiology in St Cyril and Method Hospital in Bratislava (Slovakia) were analysed. RESULTS: With Cobe Spectra apheresis machine it was possible to process larger blood volumes per procedure with higher CD34+ cell collection efficiency (p = 0.0229) and lower RBC contamination of the harvest than with Amicus (p = 0.0116). On the other hand, Amicus helped to limit PLT contamination of the harvest (p < 0.0001), thus minimizing post-procedural decrease in patient´s PLT count. The highest detected advantage of CVC usage was higher flow rate of procedure, thus processing larger blood volumes per unit of time. Interesting finding was the tendency to lower harvest PLT contamination (p = 0.054). When LVL was performed, significantly higher HSCs yields were collected, even in "poor mobilizers" when the pre-run parameters were low. CONCLUSION: Management of PBSC collection requires a particular approach in each subject. Institutionally and individually optimized collection may help to improve the transplantation outcome and decrease the financial costs (Tab. 8, Ref. 15).

[Pomalidomide in the treatment of relapsed and refractory multiple myeloma]. / Pomalidomid v liecbe relabujúceho a refraktérneho mnohopocetného myelómu.

Despite improvements in multiple myeloma therapy, the vast majority of patients continue to suffer relapses. Unfortunately, many patients event. develop disease that is refractory to lenalidomide and bortezomib and have few treatment options. Pomalidomide is a potent second-generation immunomodulatory agent with direct antiproliferative, pro-apoptotic, and antiangiogenic effects, as well as modulatory effects on bone resorption and on the immune system. Pomalidomide exhibited more potent anti-myeloma activity compared with thalidomide and lenalidomide. The optimal starting dose of pomalidomide is 4 mg given orally on days 1-21 of each 28-day cycle and combination with dexamethasone produces synergistic effects. In clinical trials, pomalidomide plus low-dose dexamethasone has shown better responses, progression-free and overall survival than high-dose dexamethasone or pomalidomide alone. Pomalidomide has limited cross-resistance with lenalidomide, and the overall response rates of pomalidomide in lenalidomide/bortezomib dual-refractory patients ranged from 26 to 31%. The most common grade 3 or 4 adverse events are hematologic, consisting of neutropenia, thrombocytopenia and anemia. Pomalidomide was approved by the FDA and the EMA in patients with multiple myeloma who have received at least two prior therapies including lenalidomide and bortezomib and have demonstrated disease progression on their last therapy. We review pomalidomide mechanism of action, clinical trials in relapsed and refractory patients, and novel pomalidomide-based combinations.

Treatment difficulty with acute GVHD - frequent cause of mortality after allogeneic hematopoietic stem cell transplantation.

OBJECTIVE: Acute graft-versus-host disease (aGvHD) remains a significant cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT). METHODS: In this study, we have retrospectively evaluated the major risk factors for the development of aGvHD in 100 patients who underwent allogeneic transplantation at the University Hospital in Bratislava between January 2007 and December 2011. RESULTS: 29 patients acquired acute GvHD (Grade I - 12 patients, G II - 5 , G III - 3, G IV - 9). We proved a higher incidence of developing aGvHD in patients with unrelated donor type, TBI conditioning and cyclosporine (CsA) replacement with mycophenolate mofetil due to CsA nephrotoxicity, while other risk factors such as older patient age, the use of peripheral blood progenitor cells and donor/recipient sex mismatch were without statistical significance. The average time of onset of aGvHD has been 57 days (range 13-260) after HSCT. Corticosteroids were used as standard initial therapy with 52 % complete response (CR) rate, although the likelihood of response rapidly decreased with increasing severity of disease (G IV - 100 % refracterness). The response to primary therapy also correlated with overall survival. Patients with steroid-refractory aGvHD received a different second-line therapies (antithymocyte globulin, anti-TNFα antibody, anti CD52 antibody) with response rate 45 % (CR - 18 %, PR - 27 %). CONCLUSION: Outcome for the patients with steroid-refractory aGvHD was poor, disease very often returned or progressed with one year mortality rate 81 % , that represents an important therapeutic problem (Tab. 2, Ref. 10).

[Lenalidomide maintenance therapy in patients with multiple myeloma]. / Udrziavacia liecba lenalidomidom u pacientov s mnohopocetným myelómom.

Maintenance therapy was recently defined as any treatment administered after the completion of induction therapy in patients whose disease is either responsive or nonprogressive at the time, with the goal of prolonging survival. Several studies have evaluated the use of novel agents as part of posttransplantation maintenance to improve progression free survival and overall survival. As shown by recent clinical trials, thalidomide and bortezomib are more indicated as consolidation agents, increasing the response rates. In contrast, lenalidomide showed a low toxicity profile and a benefit from prolonged treatment, making the drug one of the best choices for maintenance treatment. The role of lenalidomide in the maintenance therapy was evaluated in three randomized clinical trials. Lenalidomide was associated with a significantly increased progression free survival and in one study with a significant survival benefit. An unexpected finding from these trials was a modest increase in the incidence of secondary cancers. Whether lenalidomide maintenance should be routinely offered to patients is controversial among experts. For now, lenalidomide maintenance should be considered standard for patients not achieving a complete remission after stem cell transplantation, and for highrisk patients. For patients with low risk disease, the risk/ benefits of lenalidomide maintenance versus watchful waiting should be discussed until we have more data regarding survival benefits. Further analysis of lenalidomide maintenance therapy to determine the longterm risk of secondary malignancies and longer followup to assess the impact on overall survival is required.

Is there still a role for autologous stem cell transplantation in acute myeloid leukemia?

BACKGROUND: the role of autologous stem cell transplantation (ASCT) in treatment of acute myeloid leukemia (AML) remains unsettled. AIMS: retrospective analysis to evaluate the role of ASCT in patients with AML without HLA-matched donor. METHODS: between December 19, 1994 and August 1, 2012, a total of 63 patients with AML without HLA-matched donor in the department of Hematology and Transfusion Medicine, University Hospital, Bratislava, received an ASCT. Median age was 41 years (20-61 years). There were 35 (56%) males and 28 (44%) females. At the time of ASCT, 50 (79%) patients were in first complete remission (CR), 11 (18%) patients were in second CR and 2 (3%) patients were in relapse. RESULTS: with a median follow-up of 115 months (34-214 months), the 10 year overall survival (OS) and disease free survival (DFS) of all patients was 55% and 51%, respectively. Transplant-related mortality was 6%. The relapse rate was 38% and 9 years probability of relapse was 44%. CONCLUSION: ASCT is still an effective post-remission treatment in AML patients without HLA-matched donor; with the possibility of long-term survival or even cure in remarkable proportion of patients with AML, particularly in patients with favorable and intermediate cytogenetic risk. .

The feasibility of an early hospital discharge following chemotherapy for the acute myeloid leukemia.

OBJECTIVE: The hospitalization of the patient during the critical myelosuppressive period after chemotherapy is often complicated by infections caused by nosocomial pathogens, what is associated with a high antibiotics consumption and with prolongation of the period of hospitalization. These findings have led many centres to change their policy from "in-hospital" to "out-hospital care". In this retrospective study we tried, on the basis of our experiences, to identify the feasibility and safety of this approach. PATIENTS AND METHODS: We studied 56 patients with the acute myeloid leukemia treated in our clinic with the consolidation chemotherapy. We compared two groups of patients. In the first group, the patients were discharged upon completion of chemotherapy, consequently followed up as outpatients. Patients in the second group were observed in hospital during the entire nadir. Following 41 courses, patients were discharged and instructed to return immediately if fever or any other change of their clinical status occurred. RESULTS: In 24 cases after chemotherapy, the patients returned to the hospital after a discharge (in 23 cases because of fever), in 17 cases of nadir periods the hospitalisation was not necessary at all. Seven patients were readmitted in septic shock, but rapidly recovered. Two other patients died, one due to an irreversible shock within 12 hours of readmission and one due to bacterial meningitis within 48 hours after readmission. In 10 cases of rehospitalization, patients responded to the first line of antibiotics. In the second group of the patients, only 2 courses of consolidation from a total of 15 were not complicated. In contrast to the first group, we detected only poor effectiveness of broad-spectrum antibiotics in the group of inpatients. CONCLUSIONS: For AML patients in a good clinical status without any complicating medical conditions, the early discharge is feasible, safe and cost saving option (Tab. 2, Fig. 2, Ref. 7).

Abnormal cardiomarkers in leukemia patients treated with allogeneic hematopoietic stem cell transplantation.

OBJECTIVE: Clinical cardiac complications in oncologic patients may develop from subclinical myocardial damage. Biomarkers N-terminal pro brain natriuretic peptide (NT-proBNP) and troponin T (cTnT) have been hypothesized to reflect preclinical cardiotoxicity earlier than echocardiography. The aim of this study was to assess prospectively the serial values of these cardiomarkers in leukemia patients treated with allogeneic hematopoietic stem cell transplantation (HSCT). PATIENTS: Twenty-one patients who were treated with allogeneic HSCT for acute leukemia at mean age of 32.8 years (range: 19-58) were studied. The conditioning regimen included high-dose cyclophosphamide in combination with total body irradiation (TBI) or busulphan. All patients were treated with anthracyclines earlier (median cumulative dose 250 mg/m, range: 150-580). METHODS: Cardiomarkers were measured before the preparative regimen (PR) and on days 1, 14 and 30 after HSCT. Their cardiac systolic function was assessed before PR, and 1-2 months after HSCT by echocardiography. RESULTS AND CONCLUSION: The differences in NT-proBNP before PR and after HSCT were statistically significant (p<0.001). The values of cTnT before and after HSCT were also significantly different (p=0.005). Persistent abnormalities (30 days after HSCT) of NT-proBNP levels were found in 19/21 patients (90.5 %) and of cTnT levels in 10/21 patients (47.6 %). The median cTnT concentrations were higher in patients treated with TBI than in patients without TBI (p=0.013). The median NT-proBNP values were higher in patients pretreated with higher cumulative doses of anthracyclines (>250 mg/m vs ≤250 mg/m) Cardiac symptoms developed in 3/21 (14.3 %) patients (Tab. 1, Fig. 3, Ref. 36).

Granulocyte transfusions.

Neutrophils play an essential role in the defense of the body against bacterial and fungal infections. Disorders of their number or function significantly increase the risk of life-threatening infection. In spite of the development of growth factors, new broad spectrum antibiotics and antifungal drugs against nearly all known pathogens, severe neutropenia associated with bacterial or invasive fungal infections remains a major cause of morbidity and mortality in patients undergoing aggressive cancer chemotherapy or hematopoietic stem cell transplantation. Lately, an interest about granulocyte transfusions was renewed, what is a logical approach in the management of patients with prolonged 'reversible' severe neutropenia and severe infection, which is not controlled with appropriate antimicrobial and supportive treatment, including recombinant hematopoietic growth factors. It was a consequence of advances in the field of apheresis science, use of sedimenting agents and especially advances in mobilization of granulocytes to the peripheral blood. It became now possible to collect large numbers of neutrophils. Therefore, the clinical use of granulocyte transfusions, as a potential life saving treatment option in patients with severe neutropenia and uncontrolled infection in spite of appropriate antimicrobial therapy should be considered, with regard to possible benefits and risks (Ref. 74).

Use of direct sequencing for detection of mutations in the BCR-ABL kinase domain in Slovak patients with chronic myeloid leukemia.

The presence of BCR-ABL oncogene mutations in patients with chronic myeloid leukemia (CML) may be responsible for the failure of tyrosine kinase inhibitor (TKI) treatment. The aim of the study was to evaluate the frequency of BCR-ABL gene mutations in patients with CML treated with tyrosine kinase inhibitors. Our lab received 64 samples (34 women, 30 men) from patients with CML who failed or had suboptimal response to TKI treatment. The mutation analysis was performed in 61 patients with CML, 3 patients could not be tested because of inadequate RNA quality. An 866 base pair fragment containing the ABL kinase domain was amplified in a seminested RT (reverse transcriptase)-PCR and then sequenced using Applied Biosystems BigDye Terminator chemistry with two pairs of primers. We analyzed 61 patients with CML, 11 mutations were detected in 13 (21%) patients and SNP (single nucleotide polymorphism) in 6 patients (10%). In addition to 9 point mutations (G250E / F317L, F359V, L387M, Y253H, M388L, M244V, T315I, D276G), 35 bp insertion between exons 8 and 9 and deletion exon 7 were detected. Our results demonstrate that direct sequencing is suitable for routine clinical monitoring patients with CML and may be useful for optimizing therapy.