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Membrane transporters are important determinants of drug bioavailability. Their expression and activity affect the intracellular drug concentration in leukemic cells impacting response to therapy. Pharmacogenomics represents genetic markers that reflect allele arrangement of genes encoding drug transporters associated with treatment response. In previous work, we identified SNP rs460089 located in the promotor of SLC22A4 gene encoding imatinib transporter OCTN1 as influential on response of patients with chronic myeloid leukemia treated with imatinib. Patients with rs460089-GC pharmacogenotype had significantly superior response to first-line imatinib treatment compared to patients with rs460089-GG. This study investigated whether pharmacogenotypes of rs460089 are associated with sustainability of treatment-free remission (TFR) in patients from the EUROpean Stop Kinase Inhibitor (EURO-SKI) trial. In the learning sample, 176 patients showed a significantly higher 6-month probability of molecular relapse free survival (MRFS) in patients with GC genotype (73%, 95% CI: 60–82%) compared to patients with GG (51%, 95% CI: 41–61%). Also over time, patients with GC genotype had significantly higher MRFS probabilities compared with patients with GG (HR: 0.474, 95% CI: 0.280–0.802, p = 0.0054). Both results were validated with data on 93 patients from the Polish STOP imatinib study. The SNP rs460089 was found as an independent predictor of TFR.

Lower gastrointestinal (GI) graft versus host disease (GVHD) represents a severe complication in allogeneic hematopoietic stem cell transplant (HSCT) recipients with high rates of transplant-related mortality. Deregulated innate immunity reactions are the features of its pathogenesis. Cellular senescence has been considered a program of the innate immunity. We focused on lower GI GVHD from the perspective of cellular senescence.We analyzed the impact of p16Fifty-two single-center patients who presented with symptoms of lower GI GVHD were analyzed in a retrospective manner. Two SNPs located in the NFKB1 gene regions (rs3774937 C/T and rs3774959 A/G) were genotyped from the peripheral blood samples collected before the start of the conditioning. All patients underwent proctosigmoidoscopy with biopsy of the mucosa. The expression of p16Fifty-two patients (50% male) received HSCT for hematological diseases (acute leukemias in 67%) and developed lower GI symptoms. Patients with p16Our results address possible new mechanisms that may lead to better understanding of HSCT-related immune complications. Cellular senescence may bring novel approaches in GVHD diagnostics and therapy.

The purpose of this study was to analyze the results of second autologous hematopoietic stem cell transplantation (ASCT2) for patients with relapsed/refractory Hodgkin lymphoma (HL) after a first transplantation (ASCT1). Outcomes for 56 patients receiving an ASCT2 registered in the EBMT database were analyzed. The 4-year cumulative incidences of non-relapse mortality and disease relapse/progression were 5% and 67%, respectively. The 4-year overall survival (OS) and progression-free survival (PFS) were 62% and 28%. In univariate analysis, relapse of HL within 12 months of ASCT1 was associated with a worse OS (35% versus 76%

Graft-versus-host disease (GVHD) represents a significant cause of mortality after allogeneic hematopoietic stem cell transplantation (HSCT). NF-kB system is a master regulator of innate immunity responses. It controls the expression of various cytokines and chemokines many of which are involved in GVHD pathogenesis. Chemo(radio) therapy administered during conditioning induces DNA damage and activates DNA damage response (DDR) signaling resulting in irreversible cell cycle arrest - cellular senescence which has been described to be associated with robust pro-inflammatory secretion mostly controlled by NF-kB. The NFKB1 gene encodes the DNA-binding subunit of the NF-kB complex. Using the candidate gene approach, we analyzed possible association of two single-nucleotide polymorphisms (SNPs) rs3774937 C/T and rs3774959 A/G of the NFKB1 gene with GVHD and transplant-related mortality (TRM) occurrence in 109 recipients allografted from HLA-identical donor. Both SNPs in recipients were found to be strongly associated with acute GVHD. Nevertheless, no significant association with chronic GVHD and TRM was found. Presented pilot results contribute to pre-clinical observations and suggest that NF-kB may be an important regulator of HSCT-related inflammatory reactions such as acute GVHD. Novel pathogenic mechanisms of GVHD may arise from perspectives of DDR and cellular senescence where NF-kB plays an essential role.

Infectious complications during induction chemotherapy of acute myeloid leukaemia are very common. Prophylactic use of antibiotics however is an ongoing challenge in this situation due to bacterial multi-drug resistance. The aim of this study was to provide a comprehensive overview of the incidence of infectious complications in patients with AML undergoing induction therapy using the "7+3" protocol without routine antibiotic prophylaxis at one clinical site providing specialised haematological care in the Czech Republic, over a period of 15 years. The study also evaluates the aetiological spectrum of causative agents and the development of antibiotic resistance in the context of the use of the various classes of antibiotics. The analysis includes evaluation of the importance of risk factors for infectious complications and their impact on treatment of the underlying disease. The data are compared with published figures for similar cohorts of patients.This study presents a retrospective analysis of infectious complications in 242 patients with acute myeloid leukaemia undergoing the first cycle of induction therapy without routine antibiotic prophylaxis in one clinical site in Czech Republic during years 2006-2020.A total of 363 febrile episodes (FE) were recorded. At least 1 FE during the induction was detected in 229 (94.6%) patients. Clinically defined infection was the cause in 96 (26.4%) FEs and blood stream infection in 69 (19.0%) FEs. Both complications occurred simultaneously in 29 (8.0%) FEs. 169 (46.6%) FEs were evaluated as fever of unknown origin (FUO). The achievement of complete remission had a significant effect on the duration of the FE (6 vs. 9 days, P=0.0005) and on the overall survival duration (79.3 vs. 6.5 months, P0.0001). Patients diagnosed with infection or FUO at diagnosis were significantly more likely to suffer from colonisation by multi-drug resistant bacterial strains at discharge (29.2% vs. 16.3%, P=0.022). This group of patients used antibiotic therapy for a significantly longer time (35 vs. 23 days, P0.0001). Infection was a contributing cause of death in 18 (7.4%) patients. Mortality was significantly related to the failure to achieve complete remission (P0.0001).Infectious mortality during induction treatment without routine antibiotic prophylaxis was comparable to the published cohorts with prophylaxis. Regular microbiology surveillance with adequate initial antibiotic treatment can compensate routine antibiotic prophylaxis with slower development of antibiotic resistance.

The treatment outcome in patients with chronic myeloid leukaemia (CML) in blast crisis (BC) is unsatisfactory despite the use of allogeneic stem cell transplantation (ASCT). Moreover, in some patients ASCT is contraindicated, with limited treatment options. We report the case series of two patients with lymphoid BC CML in whom ASCT was not approachable. The first patient developed BC two months after diagnosis in association with dic(7;9)(p11.2;p11.2) and T315I mutation. Blast crisis with central nervous system leukemic involvement and K611N mutation of the SETD2 gene developed abruptly in the second patient five years after ceasing treatment with nilotinib in major molecular response (MMR) at the patient’s request. Both underwent one course of chemotherapy in combination with rituximab and imatinib, followed by dasatinib and interferon α (INFα) treatment in the first and dasatinib alone in the second case. Deep molecular response (DMR; MR 4.0) was achieved within a short time in both cases. It is probable that DMR was caused by a specific immune response to CML cells, described in both agents. The challenging medical condition that prompted these case series, and the subsequent results, suggest a re-visit to the use of a combination of well-known drugs as an area for further investigation.

BACKGROUND There is no universally accepted opinion on the use of granulocyte transfusions collected using apheresis (GTA) in neutropenic patients and severe infection. PATIENTS AND METHODS The efficacy and safety of GTAs transfused at a single center over 10 years were analyzed retrospectively. GTAs were harvested from voluntary unrelated donors after priming with methylprednisolone using continuous apheresis and hydroxyethylstarch as sedimentation agent. RESULTS 41 patients with neutropenia and hematologic malignancy (15 females and 26 males aged 22-69 (median 45.5)) were given a median 3.5 GTAs per patient (range: 1-17) containing a median 1.39×1010 granulocyte/GTA (range: 0.65-2.81). The indications for GTA use were soft tissue inflammation, sepsis, and pneumonia in 30, 22, and 14 cases, respectively. After GTA complete (30 patients: 73.2%) or partial (6 patients: 14.6%) healing of the infection was achieved. The success rate was 91.7% in soft tissue infections, 66.7% in invasive fungal infections, and 68% in sepsis. Septic shock (documented in 12 cases) was associated with a poor response (P

Patients with internal tandem duplication in fms-related tyrosine kinase receptor gene 3 (FLT3-ITD)-mutated acute myeloid leukemia (AML) have a dismal prognosis and the only curative option seems to be allogeneic stem cell transplantation (alloSCT). However, its timing is still matter of debate.We retrospectively analyzed 73 consecutive AML patients with FLT3-ITD (median age 53, range 20-68 years) allografted with consistent policy to try to refer them all for upfront alloSCT in first complete remission (CR1).With a median follow-up of 44 (range, 5-135) months the 5-year overall survival (OS)/disease-free survival (DFS) probabilities were 49%/47%. The cumulative incidence of relapse and nonrelapse mortality (NRM) were 37% and 14%, respectively. The estimated 5-year OS for patients who received transplantation in CR1 was 62% versus 0% for patients who received transplantation beyond CR1. Multivariable analysis identified stem cell transplantation beyond CR1 as the key factor for poor OS (hazard ratio [HR], 5.41; P .0001), DFS (HR, 4.41; P = .0002), and high relapse incidence (HR, 8.08; P .0001). Acute graft versus host disease Grade ≥3 predicted higher NRM (HR, 3.80; P = .059) as well as inferior OS (HR, 2.04; P = .0079). No association of patient age, nucleophosmin status, donor type, conditioning, and other variables on the survival was detected.AlloSCT should be regarded with urgency as soon as CR1 is achieved in this subset of AML patients.

We retrospectively evaluated the role of age and dosage in 372 CML patients (170 women, 202 men) treated with first-line imatinib (IMA) from the records of the CAMELIA registry. The median follow-up of the patients was 82.3 (18.0-177.3) months. The treatment results of 80 elderly patients aged over 65 years at diagnosis were compared in analysis "A" with those of 292 younger patients and in analysis "B" with those of 90 patients younger than 40 and 202 patients aged 40-64. The elderly patients had statistically adverse values of the Sokal, ELTS, and ECOG scores and Charlson comorbidity index in both analyses (p from = 0.012 to ≤ 0.001). Despite a more frequent use of a daily dose lower than 400 mg - in 31 elderly patients (38.8%) than in 45 younger ones (15.4%) (p 0.001), there were no statistically significant differences in the achievement of optimal haematological, cytogenetic, and molecular responses according to the ELN criteria in both the analyses, A and B. The comparisons of overall survival with CML-related death (OS

Micro-Abstract Coronavirus disease 2019 (COVID-19) represents an important infectious complication in patients with hematologic diseases. Our study presents first single-center experience from the Czech Republic in 96 patients in whom COVID-19 was confirmed. The study results confirm the prognostic significance of age for achieving treatment response of hematologic disease as well as the severity and mortality of COVID-19 in hematology patients., Background Coronavirus disease 2019 (COVID-19) represents an important infectious complication associated with high mortality rates in patients with hematologic diseases. There have not been published any epidemiologic studies from Czech Republic so far. Patients and Methods This study is the first analysis of patients with hematologic malignancies and bone marrow failure syndromes treated at single hematology center in the Czech Republic between March 1 and December 31, 2020, in whom COVID-19 infection was confirmed. Results The sample comprised 96 patients aged 26 to 84 years (median, 66.0 years). At the time of their COVID-19 diagnosis, 75 patients (78.1%) were treated for hematologic diseases. Twenty-seven patients (28.1%) in the sample had complete remission (CR) of their hematologic disease. They were nonsignificantly more likely to have asymptomatic to moderate COVID-19 infection than those who failed to achieve CR (74.1% vs. 56.5%; P = .06). A more severe course of the infection was significantly correlated with older age (P = .047). Lung involvement was also statistically significantly associated with older age (P = .045). Over the study period, a total of 15 patients died. Age greater than 60 years was significantly associated with deaths from COVID-19 (P = .036), with failure to achieve CR having a statistically nonsignificant impact on mortality (P = .22). Conclusion These results confirm the prognostic significance of age for achieving treatment response of hematologic disease as well as the severity and mortality of COVID-19 in hematology patients.

T-cell prolymphocytic leukemia (T-PLL) is a poor prognostic disease with very limited options of efficient therapies. Most patients are refractory to chemotherapies and despite high response rates after alemtuzumab, virtually all patients relapse. Therefore, there is an unmet medical need for novel therapies in T-PLL. As the chemokine receptor CCR7 is a molecule expressed in a wide range of malignancies and relevant in many tumor processes, the present study addressed the biologic role of this receptor in T-PLL. Furthermore, we elucidated the mechanisms of action mediated by an anti-CCR7 monoclonal antibody (mAb) and evaluated whether its anti-tumor activity would warrant development towards clinical applications in T-PLL. Our results demonstrate that CCR7 is a prognostic biomarker for overall survival in T-PLL patients and a functional receptor involved in the migration, invasion, and survival of leukemic cells. Targeting CCR7 with a mAb inhibited ligand-mediated signaling pathways and induced tumor cell killing in primary samples. In addition, directing antibodies against CCR7 was highly effective in T-cell leukemia xenograft models. Together, these findings make CCR7 an attractive molecule for novel mAb-based therapeutic applications in T-PLL, a disease where recent drug screen efforts and studies addressing new compounds have focused on chemotherapy or small molecules., Funding for this work was by grant PI015/02085 and PI18/01163 from the Fondo de Investigaciones Sanitarias, Ministerio de Sanidad y Consumo, Spain to CMC who was also cofinanced by FEDER funds. CCM and FT were supported by RTC-2015-3318-1 (Ministerio de Economía y Competitividad, Spain). MH was funded by the German Research Foundation (DFG; HE3553/ 4–2) as part of the collaborative research group on mature T-cell lymphomas, “CONTROL-T” (FOR1961). The José Carreras Leukemia Foundation (03F/2016), the Köln Fortune Program, and the Fritz Thyssen foundation (10.15.2.034MN) supported MH and AS. The Köln Fortune Program also supported LW. SM was supported by the Finnish Cancer Organizations, the Finnish Cancer Institute, Academy of Finland and the Sigrid Juselius Foundation. SM and MH are supported by the ERAPerMed program ‘JAK- STAT-TARGET’. MLT was granted by SAF2016–75442-R (Agencia Estatal de Investigación/ European Regional Development Fund, European Union, Spain).

Prognostic scores support clinicians in selecting risk-adjusted treatments and in comparatively assessing different results. For patients with chronic-phase chronic myeloid leukemia (CML), four baseline prognostic scores are commonly used. Our aim was to compare the prognostic performance of the scores and to arrive at an evidence-based score recommendation. In 2949 patients not involved in any score development, higher hazard ratios and concordance indices in any comparison demonstrated the best discrimination of long-term survival with the ELTS score. In a second step, of 5154 patients analyzed to investigate risk group classification differences, 23% (n = 1197) were allocated to high-risk by the Sokal score. Of the 1197 Sokal high-risk patients, 56% were non-high-risk according to the ELTS score and had a significantly more favorable long-term survival prognosis than the 526 high-risk patients according to both scores. The Sokal score identified too many patients as high-risk and relatively few (40%) as low-risk (versus 60% with the ELTS score). Inappropriate risk classification jeopardizes optimal treatment selection. The ELTS score outperformed the Sokal score, the Euro, and the EUTOS score regarding risk group discrimination. The recent recommendation of the European LeukemiaNet for preferred use of the ELTS score was supported with significant statistical evidence.

We performed a prospective study to evaluate the types and characteristics of central nervous system (CNS) disorders in patients after hematopoietic stem cell transplantation. The study included 163 episodes of CNS disorders of which 58 (36%) were infections. Proven or probable infections were documented in 34 patients and included fungi (n = 10, 29%), viruses (n = 12, 35%), Toxoplasma spp. (n = 9, 27%) and bacteria (n = 3, 9%). Non-infectious neurological disorders (n = 105, 64%) frequently encompassed metabolic/drug-induced abnormalities (n = 28, 27%) or cerebral vascular events (n = 22, 21%). Median onset times were later for infectious (day + 101) vs non-infectious neurological disorders (day + 50, p = 0.009). An unremarkable cranial CT scan was found in 33% of infection episodes. Absence of cerebrospinal fluid pleocytosis despite a normal or increased peripheral blood white blood cell count occurred in 26% of infections. Day-30 mortality rates were significantly higher for fungal (87%) vs non-fungal infections (40%, p

Impaired glucose metabolism (IGM) with hyperglycemia represents one of the most frequently observed adverse events (AE) during nilotinib therapy of chronic myeloid leukemia (CML). The exact mechanism of IGM remains controversial. Although a case report has shown a decrease in insulin secretion1 , our previous pilot data suggested development of insulin resistance as a possible mechanism.2 In this prospective study we aimed to confirm results from our pilot study using a larger cohort of CML patients treated with nilotinib and to compare results with data obtained on control groups receiving imatinib and dasatinib.

Tyrosine kinase inhibitors (TKI) have dramatically improved the prognosis of CML patients with life expectancy close to individuals without CML [1], and therefore the longterm safety of TKIs has become especially important due to the potential for lifelong treatment. Nilotinib (NILO) tends to have a negative effect on lipid [2] and glucose [3] metabolism, while imatinib (IMA) seems to improve [4] these abnormalities. Although the data appear convincing, the real impact of these changes on cardiovascular (CV) risk and the development of CV complications during TKI therapy have not yet been confirmed.

Background. Treatment-free remission (TFR) has become a new treatment goal for chronic myeloid leukemia (CML) patients. However, usually abrupt tyrosine kinase inhibitors (TKIs) therapy discontinuation has been successful only in about half of eligible patients and it can cause burdening TKI withdrawal syndrome (TWS) in about 30% of them. Moreover, any robust clinical or biological factor predictive for successful TFR has not been identified yet. On top of that, sustainable deep molecular response (DMR) as the main prerequisite for TKI discontinuation attempt is achieved only in 20-40% of patients. The majority of CML patients, therefore, need to be treated with the effective and well-tolerated drug for a long time or even life-long. Study design and methods. With the recognition of all these aspects, we designed a nationwide prospective investigator-initiated phase II clinical trial HALF (ClinicalTrials.gov NCT04147533) in order to evaluate efficacy and safety of TKI discontinuation after previous two-step dose reduction in patients with CML in DMR (Fig. 1). Step-wise TKI dose reduction, i.e. half of the standard during the first 6 months after study entry, and the same dose given alternatively (every other day) during the next 6 months, was derived from pharmacokinetics and experimental data as well as from clinical trials' results. We assume that the step-wise and eventually meaningful TKI dose reduction enables a higher rate of patients achieving successful TFR with less pronounce TWS, or even would represent a more reasonable and safer alternative to the complete and sudden TKI interruption. This unique nationwide academic project has been facilitated by hematological patients care centralization in the Czech Republic. A primary study objective is to evaluate the proportion of patients in major molecular response (MMR) at 6 and 12 months and in TFR at 18, 24, and 36 months after the study enrollment, respectively, and molecular recurrence-free survival at all mentioned time points as well. Main secondary and exploratory objectives are: to evaluate the proportion of patients loosing MMR and in whom MMR and MR4.0 would be re-achieved after TKI re-introduction, time to MMR and MR4.0 re-achievement, FFS, PFS, OS, TWS, and QoL assessment, predictive factors for successful TFR identification, quantification of BCR-ABL1 using digital droplet PCR at both the DNA and mRNA levels, immunological profiling, BCR-ABL1 kinetics mathematical modeling, assessment of TKI pharmacokinetics, clonal hematopoiesis and pharmaco-economics. Results. The study was launched in December 2019; however, due to the COVID-19 outbreak, patients' recruitment started on June 16, 2020. Here, characteristics of the first 74 patients included in the study until April 2021 are presented. There were 37 males and 37 females, with median age at the time of diagnosis of 53 years (range, 23-74) and at the time of the study entry of 67 years (range, 35-86). A median time of CML disease, TKI treatment, and DMR duration before the study initiation was as follows: 9.9 years (range, 4.4-22.5), 9.8 years (range, 4.2-20.2), and 7.3 years (range, 3.2-18.3), respectively. The ELTS score was low, intermediate, high and unknown in 62.2%, 21.6%, 13.5%, and 2.7% of patients, respectively. At the time of study entry, 58 patients (79.5%) were treated with imatinib, 10 (13.7%) with nilotinib, and 5 (6.8%) with dasatinib, respectively, whereas in 63 patients (86.3%) it was in the first line of therapy. With almost half of patients (48.6%), the TKI dose was already reduced at the time of study entry. With 10 (13.5%) patients, interferon-α treatment preceded TKI administration. At the time of this abstract preparation, on July 26, 2021, altogether 102 patients (from planned 150) have been enrolled in the study; 48 of them (47.1%) have already moved to the second de-escalation phase and 9 (8.8%) patients to the TFR phase. There were 2 cases of confirmed MMR loss (both in month 8 after the study entry) and no patient experienced symptoms resembling TWS. Conclusions. Despite the COVID-19 pandemic, the HALF study was successfully launched and initiated in the majority of centers, with 102 already included patients and continuing intensive enrolment. Based on our very preliminary results, the step-wise dose reduction seems to be an effective and safe approach. More included patients, longer follow-up and further analyses are needed in order to reach all set up objectives. Figure 1 Figure 1. Disclosures Žácková: Angelini: Consultancy, Speakers Bureau; Novartis: Speakers Bureau. Faber: Angelini: Consultancy, Other: conference fees, Research Funding, Speakers Bureau; Bristol-Myers Squibb: Consultancy, Other: conference fees, Research Funding, Speakers Bureau; Novartis: Consultancy, Other: conference fees, Research Funding, Speakers Bureau; Pfizer: Other: conference fees; TERUMO: Other: conference fees. Bělohlávková: Novartis: Consultancy; BMS/Celgene: Consultancy. Horňák: Angelini: Honoraria. Svobodník: Roche: Speakers Bureau; Janssen-Cilag: Speakers Bureau. Machová Poláková: Incyte: Consultancy; Angelini: Consultancy; Novartis: Research Funding. Mayer: Principia: Research Funding.

Untargeted metabolite profiling using high-resolution mass spectrometry coupled with liquid chromatography (LC-HRMS), followed by data analysis with the Compound Discoverer 2.0™ software, was used to study the metabolism of imatinib in humans with chronic myeloid leukemia. Plasma samples from control (drug-free) and patient (treated with imatinib) groups were analyzed in full-scan mode and the unknown ions occurring only in the patient group were then, as potential imatinib metabolites, subjected to multi-stage fragmentation in order to elucidate their structure. The application of an untargeted approach, as described in this study, enabled the detection of 24 novel structurally unexpected metabolites. Several sulphur-containing compounds, probably originating after the reaction of reactive intermediates of imatinib with endogenous glutathione, were found and annotated as cysteine and cystine adducts. In the proposed mechanism, the cysteine adducts were formed after the rearrangement of piperazine moiety to imidazoline. On the contrary, in vivo S-N exchange occurred in the case of the cystine adducts. In addition, N-O exchange was observed in the collision cell in the course of the fragmentation of the cystine adducts. The presence of sulphur in the cysteine and cystine conjugates was proved by means of ultra-high resolution measurements using Orbitrap Elite. The detection of metabolites derived from glutathione might improve knowledge about the disposition of imatinib towards bioactivation and help to improve understanding of the mechanism of its hepatotoxicity or nephrotoxicity in humans.

Chronic myeloid leukemia (CML) is usually diagnosed in chronic phase, yet there is a small percentage of patients that is diagnosed in accelerated phase or blast crisis. Due to this rarity, little is known on the prognosis of these patients. Our aim was to identify prognostic factors for this cohort. We identified 283 patients in the EUTOS population-based and out-study registries that were diagnosed in advanced phase. Nearly all patients were treated with tyrosine kinase inhibitors. Median survival in this heterogeneous cohort was 8.2 years. When comparing patients with more than 30% blasts to those with 20-29% the hazard ratio (HR) was 1.32 (95%-confidence interval (CI): [0.7 - 2.6]). Patients with 20-29% had a significantly higher risk than patients with less than 20% blasts (HR: 2.24, 95%-CI: [1.2 - 4.0], p = 0.008). We found that the blast count was the most important prognostic factor; however, age, hemoglobin, basophils and other chromosomal aberrations should be considered as well. The ELTS score was able to define two groups (high risk vs. non-high risk) with an HR of 3.01 (95%-CI: [1.81 - 5.00], p

The discovery of tyrosine kinase inhibitors (TKIs) brought a major breakthrough in the treatment of patients with chronic myeloid leukemia (CML). Pathogenetic CML events are closely linked with the Bcr-Abl protein with tyrosine kinase activity. TKIs block the ATP-binding site; therefore, the signal pathways leading to malignant transformation are no longer active. However, there is limited information about the impact of TKI treatment on the metabolome of CML patients. Using liquid chromatography mass spectrometric metabolite profiling and multivariate statistical methods, we analyzed plasma and leukocyte samples of patients newly diagnosed with CML, patients treated with hydroxyurea and TKIs (imatinib, dasatinib, nilotinib), and healthy controls. The global metabolic profiles clearly distinguished the newly diagnosed CML patients and the patients treated with hydroxyurea from those treated with TKIs and the healthy controls. The major changes were found in glycolysis, the citric acid cycle, and amino acid metabolism. We observed differences in the levels of amino acids and acylcarnitines between those patients responding to imatinib treatment and those who were resistant to it. According to our findings, the metabolic profiling may be potentially used as an additional tool for the assessment of response/resistance to imatinib.

BACKGROUND With an increasing number of cancer patients receiving tyrosine kinase inhibitors (TKIs), therapeutic drug monitoring of these molecules is becoming more widespread today. It is mainly based on liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) methods with typical run times of several minutes. In an online solid phase extraction-MS/MS (SPE-MS/MS) system, the chromatography column is replaced with a reusable solid phase extraction (SPE) cartridge and the analysis time is shortened to less than half a minute. The aim of this study was to develop such a method and test the performance of this high-throughput system in the analysis of imatinib (IMA), nilotinib (NIL), and lapatinib (LAP) in human plasma. METHODS Samples were prepared by simple protein precipitation with methanol containing deuterated internal standards. After centrifugation, the supernatant was diluted 10 fold with a mixture of methanol and water (1:1). A C4 cartridge was used for SPE and the analytes were eluted by acetonitrile. All the analytes were measured within a wide calibration range (50-5000 ng/mL for nilotinib and imatinib, 100-10,000 ng/mL for lapatinib). The method was compared with the LC-MS/MS method by the analysis of 176 clinical samples. RESULTS Intraday and interday inaccuracies within 15% and a coefficient of variation less than 15% were achieved for all the TKIs that were measured. Even though the matrix effects were higher in comparison with LC-MS/MS methods, their effect on the performance of the method was eliminated by the usage of deuterated internal standards. The total run time of the new method was 29 seconds for one analysis and the results were fully comparable with LC-MS/MS. CONCLUSIONS Routine clinical practice requiring high-throughput methods for therapeutic drug monitoring of TKIs may benefit from the online SPE-MS/MS method that provides fast, low-cost analysis, and results that are comparable with conventional methods.

Background: Tyrosine kinase inhibitors (TKIs) have improved the survival of patients with chronic myeloid leukaemia. Many patients have deep molecular responses, a prerequisite for TKI therapy discontinuation. We aimed to define precise conditions for stopping treatment. Methods: In this prospective, non-randomised trial, we enrolled patients with chronic myeloid leukaemia at 61 European centres in 11 countries. Eligible patients had chronic-phase chronic myeloid leukaemia, had received any TKI for at least 3 years (without treatment failure according to European LeukemiaNet [ELN] recommendations), and had a confirmed deep molecular response for at least 1 year. The primary endpoint was molecular relapse-free survival, defined by loss of major molecular response (MMR; >0·1% BCR-ABL1 on the International Scale) and assessed in all patients with at least one molecular result. Secondary endpoints were a prognostic analysis of factors affecting maintenance of MMR at 6 months in learning and validation samples and the cost impact of stopping TKI therapy. We considered loss of haematological response, progress to accelerated-phase chronic myeloid leukaemia, or blast crisis as serious adverse events. This study presents the results of the prespecified interim analysis, which was done after the 6-month molecular relapse-free survival status was known for 200 patients. The study is ongoing and is registered with ClinicalTrials.gov, number NCT01596114. Findings: Between May 30, 2012, and Dec 3, 2014, we assessed 868 patients with chronic myeloid leukaemia for eligibility, of whom 758 were enrolled. Median follow-up of the 755 patients evaluable for molecular response was 27 months (IQR 21–34). Molecular relapse-free survival for these patients was 61% (95% CI 57–64) at 6 months and 50% (46–54) at 24 months. Of these 755 patients, 371 (49%) lost MMR after TKI discontinuation, four (1%) died while in MMR for reasons unrelated to chronic myeloid leukaemia (myocardial infarction, lung cancer, renal cancer, and heart failure), and 13 (2%) restarted TKI therapy while in MMR. A further six (1%) patients died in chronic-phase chronic myeloid leukaemia after loss of MMR and re-initiation of TKI therapy for reasons unrelated to chronic myeloid leukaemia, and two (

T-cell prolymphocytic leukemia (T-PLL) is a rare and aggressive neoplasm of mature T-cells with an urgent need for rationally designed therapies to address its notoriously chemo-refractory behavior. The median survival of T-PLL patients is

Presented are results of allogeneic hematopoietic stem cell transplantations (HSCTs) in 13 patients with high-risk acute lymphoblastic leukemia (ALL) in the first complete remission after a reduced intensity conditioning combining fludarabine (150 mg/m2) and melphalan (140 mg/m2) with thymoglobulin (4.5 mg/kg). The immunosuppressive effect of T-cell depletion reducing the risk of graft-versus-host disease (GVHD) and non-relapse mortality was compensated by early initiation of reduction and withdrawal of prophylactic immunosuppression aimed at maintaining effective immunological antileukemic control. The median post-transplant follow-up was 23 (range, 10-65) months. Stable engraftment of donor's hematopoiesis was achieved in all patients. Acute GVHD was observed in two cases (15.4%); the chronic form was not noted. Two patients (15.4%) relapsed with ALL at 3 and 16 months after transplantation. During the above post-transplant follow-up, all 13 recipients were alive, with a probability of 2-year disease-free survival of 76.9% (95% CI 51-100%). Although the results were obtained with a small pilot study group it may be assumed that, given the prognostic risk of most patients and the nearly 2-year median post-transplant follow-up, the approach may be considered as an alternative to HSCTs after traditional myeloablative or reduced conditioning regimens with standard GVHD prophylaxis.

UNLABELLED Acute promyelocytic leukemia is a unique entity among acute leukemias. Introduction of all-trans retinoic acid and, subsequently, arsenic trioxide in its treatment has markedly improved treatment outcomes for this once frequently fatal disease. Improved outcomes have also been observed in elderly patients, including those in whom standard intensive therapy is contraindicated because of comorbidities.In our center, a total of 60 APL patients were treated in 1993-2013, of whom 9 were aged 60 or more years. Although most of them had significant comorbidities at the time of diagnosis, eight achieved complete remission. At the time of the analysis, six patients were alive and in long-term remission; two patients died of causes other than APL. The median follow-up was 59 months.Included is case report of a patient with a high comorbidity score whose treatment was markedly reduced and individualized.Our experience shows that, in APL patients a curative approach is generally tolerated and should always be attempted regardless of age and comorbidities. KEYWORDS APL - elderly patients - comorbidity.

Presented are updated results of allogeneic hematopoietic stem cell transplantations (HSCTs) in 25 adult patients with acute lymphoblastic leukemia (ALL) in complete remission (CR) after a reduced intensity conditioning (RIC) combining fludarabine (150 mg/m2) and melphalan (140 mg/m2) with thymoglobulin (4.5 mg/kg or recently 4.0 mg/kg) followed by early initiation of reduction and withdrawal of prophylactic posttransplant immunosuppression. The median post-transplant follow-up was 32 (range, 4-87) months. Stable engraftment of donor's hematopoiesis was achieved in all patients. Acute graft versus host disease (GVHD) as well as the chronic one were equally observed in four cases (16%). Five patients (20%) relapsed with ALL in the median of 9 (range, 3-15) months after HSCT. During the above post-transplant follow-up, 4 recipients (16%) died. Disease progression and posttransplant complications were the cause of death in three (12%) and one (4%) of them, respectively. The probabilities of 2-year event-free (EFS) and overall survival (OS) were 70.3% (95% CI 51.9-88.7%) and 86.1% (95% CI 71.6-100%), respectively. Presented study confirmed our previously reported promising results and this approach may be considered as an alternative to traditional HSCTs performed in high-risk patients with ALL.

Aims. A retrospective analysis of patients with Hodgkin lymphoma (HL) was performed to assess their outcome regarding relative dose intensity (RDI) of chemotherapy administered in primary treatment. Methods. A total of 194 patients were divided into three groups with different RDI of primary chemotherapy (100%, 9099% and < 90%). Reduced RDI in two groups (90-99% and < 90%) was caused by the delay of the interval between the administration of some chemotherapeutic courses. The probability of complete remission (CR), disease relapse, eventfree survival (EFS) and overall survival (OS) as the basic parameters of patient outcome were statistically compared. Results. Multivariate analysis showed here were no significant differences in probability of CR (HR 0.9, 95% CI [0.751.08], P=0.5), risk of relapse (HR 1.34, 95% CI [0.92-1.94], P=0.11) or death (HR 1.52, 95% CI [0.94-2.5], P=0.13). There were also no significant differences in probability of EFS (mean 13 vs. 10 vs. 12 years, P=0.17; HR 1.54, 95% CI [0.91-2.6], P=0.22) or OS (mean 15 vs. 13 vs. 14 years, P=0.13; HR 1.52, 95% CI [0.93-2.5], P=0.13). Conclusion. We found no significant impact of primary chemotherapy delay resulting in reduced RDI on outcome in HL patients.

This study analyzed the prognostic significance of soluble interleukin-2 receptor α (sIL-2Rα) levels in 100 prospectively enrolled patients with previously untreated follicular lymphoma. It showed that sIL-2Rα level ≥ 115 pmol/L at the time of treatment initiation correlated with a high Follicular Lymphoma International Prognostic Index-2 (FLIPI-2), bulky disease, advanced clinical stage, number of involved lymph nodes, bone marrow involvement and elevated β2-microglobulin (B2M) level. When testing all patients, sIL-2Rα ≥ 115 pmol/L was associated with significantly shorter progression-free (PFS; p0.03, hazard ratio [HR] 2.04) but not overall (OS; p = 0.06, HR 2.36) survival rates. Subanalysis of patients receiving cyclophosphamide, doxorubicin, vincristine and prednisone (CHOP) ± rituximab showed higher predictive power for both PFS (HR 2.75, 95% confidence interval [CI] 1.24-6.11, p = 0.01) and OS (HR 3.33, 95% CI 1.15-9.63, p = 0.02). In the whole population (n = 100), only B2M proved a significant univariate predictor (p = 0.007, HR = 2.8) of PFS. When testing patients treated with CHOP ± rituximab, sIL-2Rα was found to be the best univariate predictor for PFS among all FLIPI-2 factors (HR = 2.68, p = 0.015). Serum IL-2Rα levels may help to refine risk assessment in the modern immunotherapy era complementary to FLIPI-2 factors.

Introduction: The single nucleotide polymorphism (SNP) rs460089 (G/C) located in the promotor of SLC22A4 (transporter hOCTN1) was identified as a prognostic factor for the outcome of chronic myeloid leukaemia patients treated with imatinib first line (Jaruskova et al. JECCR 2017). Patients with GC genotype had significantly higher probability of achievement of sustained major molecular response (MMR, BCR-ABL≤0.1% IS) as compared with patients with GG. We investigated differences in the outcome after imatinib cessation in EURO-SKI patients according to the genotypes of the SNP rs460089. Methods: DNA analysis was performed by TaqMan SNP genotyping assay using StepOnePlus RQ-PCR System (Thermofisher Scientific). In addition to the inclusion criteria defined for prognostic analysis in Saussele et al. (Lancet Oncology 2018), all patients with interferon pre-treatment were excluded. Data on sex, duration of IM treatment, of deep molecular response and age at time of imatinib discontinuation as well as molecular status at 6 months thereafter were available for 178 patients. Logistic regression was used to investigate factors affecting MMR maintenance at 6 months. Level of significance was 0.05. Results: Of 178 patients, 106 (60%) maintained MMR 6 months after imatinib stop. GC genotype was identified in 64 patients, GG in 96 and CC in 18. Most beneficial for MMR maintenance was genotype GC (72%, 95% confidence interval (CI): 60-82%), followed by CC (61%, CI: 38-80%) and GG (51%, CI: 41-61%). Overall, the SNP rs460089 was associated with MMR maintenance (p=0.0335) with a significantly higher odds ratio (OR) for maintenance for GC genotype vs. GG (2.451, CI: 1.247-4.819, p=0.0093) but not for CC vs. GG (1.507, CI: 0.539-4.216, p=0.4343). Only duration of TKI treatment was significant (OR: 1.157, CI: 1.014-1.319, p=0.0303) when added to genotypes in multiple regression. The OR of GC vs. GG was slightly modified to 2.311 (1.164-4.588, p=0.0166). Conclusions: Based on observed data we suppose that the GC genotype of the SNP rs460089 is associated with sufficient intracellular concentration of imatinib allowing more efficient targeting of CML cells during the treatment. This resulted in a higher proportion of patients who sustained MMR after imatinib stop as compared with patients with GG. Longer duration of imatinib treatment increased the probability of MMR maintenance after imatinib cessation also in patients with GG. The frequency of CC was low and outcome in between GC and GG. The SNP rs460089 may provide an independent prognostic factor for molecular response maintenance after imatinib cessation. Supported by MZCR 00023736. Disclosures Machova: Novartis: Consultancy; BMS: Consultancy, Research Funding; Incyte: Consultancy. Fabarius:Novartis: Research Funding. Brümmendorf:Pfizer: Consultancy, Research Funding; University Hospital of the RWTH Aachen: Employment; Merck: Consultancy; Ariad: Consultancy; Novartis: Consultancy, Research Funding; Janssen: Consultancy. Burchert:Novartis: Research Funding. Mustjoki:BMS: Honoraria, Research Funding; Novartis: Research Funding; Pfizer: Research Funding. Mayer:AOP Orphan Pharmaceuticals AG: Research Funding. Žáčková:Bristol Myers Squibb: Consultancy; Angelini: Consultancy; Incyte: Consultancy; Novartis: Consultancy. Panayiotidis:Bayer: Other: Support of clinical trial. Richter:Novartis: Consultancy; Pfizer: Consultancy, Research Funding. Hjorth-Hansen:BMS: Research Funding; Pfizer: Consultancy, Research Funding; Austrian Orphan Pharma: Consultancy, Research Funding. Saussele:Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Pfizer: Honoraria; Incyte: Honoraria, Research Funding.

Aims. We assessed the long-term outcome of consecutive patients in the chronic phase of chronic myeloid leukemia (CML) treated with interferon-alpha (INF-α) in Central and Northern Moravia between 1989 and 2006. Methods. A retrospective study focused on the response, prognostic factors and side-effects of INF-α. Results. 118 patients (67 males and 51 females, median age 50 years; range 18-71) were analyzed. The median followup was 82.6 months (12.4-212.6). Thirty-six patients (30.5%) achieved major cytogenetic response (CyR) in median of 18.3 months (3.7-47.3) and maintained it for a median of 64.0 months (7.0-176.0). Sixty-one patients treated with INF-α for more than 12 months had an overall survival (OS) of 137.0 months (95% CI 117.6-156.4). Eighteen (29.5%) achieved complete CyR (CCyR). 109 patients discontinued the treatment with INF-α because of hematologic or cytogenetic resistance in 53 (48.7%), progression of CML in 31 (28.4%) and intolerance to INF-α in 17 (15.6%) patients. The percentage of peripheral blasts, leukocyte count (>50x10 9 /L), splenomegaly, anemia (Hgb≤110 g/L) and Sokal score had statistical impact on the OS in univariate assessment but only the Sokal score remained significant in multivariate analysis. Additional cytogenetic abnormalities at diagnosis were associated with poor prognosis. Conclusions. In most patients, treatment with INF-α had to be stopped because of a failure to induce response, progression of CML or side-effects but nearly one third of patients treated at least for one year had a long-term benefit from INF-α. The best prognosis was associated with achievement of CCyR and negativity of BCR-ABL in nested RT-PCR.

Using the data of 723 chronic myeloid leukemia (CML) patients in the chronic phase, we analyzed the prognostic value of the Sokal, Euro, and EUTOS scores as well as the level of BCR-ABL1 and the achievement of complete cytogenetic response (CCgR) at 3 months of imatinib therapy in relation to the so-called current survival measures: the current cumulative incidence (CCI) reflecting the probability of being alive and in CCgR after starting imatinib therapy; the current leukemia-free survival (CLFS) reflecting the probability of being alive and in CCgR after achieving the first CCgR; and the overall survival. The greatest difference between the CCI curves at 5 years after initiating imatinib therapy was observed for the BCR-ABL1 transcripts at 3 months. The 5-year CCI was 94.3% in patients with BCR-ABL1 transcripts ≤ 10% and 57.1% in patients with BCR-ABL1 transcripts 10% (P = 0.005). Therefore, the examination of BCR-ABL1 transcripts at 3 months may help in early identification of patients who are likely to perform poorly with imatinib. On the other hand, CLFS was not significantly affected by the considered stratifications. In conclusion, our results indicate that once the CCgR is achieved, the prognosis is good irrespective of the starting prognostic risks.

Background. The Recommendations of the European Leukemia Net (ELN) have become an essential tool in the management and prognosis of patients with chronic myeloid leukemia (CML) treated with tyrosine kinase inhibitors (TKIs). However, the definition of suboptimal response remains under discussion. Methods. We used conventional cytogenetics for the detection of clonal changes in Ph-positive and negative clones. RT-PCR and sequencing were carried out on peripheral blood leukocytes to detect the type of BCR-ABL1 transcript. The BCR-ABL1 mutational status was assessed using sequencing of RT-PCR products. High performance capillary electrophoresis for determination of imatinib (IMA) plasma concentration was used. Results. A retrospective study of 110 patients diagnosed with chronic-phase (CP) CML treated with IMA or 2 nd generation TKIs in the years 2000-2009 focused on analysis of patients with suboptimal response according to ELN criteria. 40 patients were administered IMA as first-line therapy and 70 had been pretreated with interferon-alpha (IFN-α) with or without Ara-C and/or hydroxyurea (HU) for a median 12 months (range, 1-92 months). After adjusting for the ELN criteria, major molecular response (MMR) was achieved after median 34 and 39 months in 66.7% and 41.7% of patients after the first and second-line IMA therapy with suboptimal response defined as lack of achievement of MMR at the 18 th month of treatment, respectively. In comparison to patients with optimal response, patients with suboptimal response did not show significant differences in overall survival (OS) or progression-free survival (PFS). Cytogenetic assays demonstrated additional chromosome abnormalities (ACAs): chromosome 8 trisomy in a Ph-negative clone during the IMA treatment (in 1 case) and der(9q) in Ph-positive clone (in 2 cases); in patients receiving first-line IMA only chromosome 8 trisomy was observed which was associated with myelodysplastic syndrome – this was the only case where hematopoietic stem cell transplantation (HSCT) was performed. During the treatment with IMA in both subgroups no regulatory mutations in the ABL kinase domain were confirmed. Conclusion. We believe that the category of suboptimal response should be redefined or withdrawn from the ELN 2009 recommendations for management of CML patients treated with TKIs. Patients with suboptimal response who have no additional risks (additional cytogenetic abnormalities or BCR-ABL1 regulatory mutations) may remain on IMA treatment while patients with these risks should be switched to the 2 nd generation TKIs.

We evaluated responses to the treatment and long-term outcomes of chronic myeloid leukemia patients treated with imatinib as first-line treatment in routine clinical setting from two countries with centralized tyrosine kinase inhibitors (TKIs) treatment. We assessed prognostic significance of European LeukemiaNet (ELN) 2006- and 2009-defined responses and the prognostic value of molecular responses at defined time points on 5-year survivals. Among the cumulative rates of incidence of hematologic, cytogenetic, and molecular responses and all important survival parameters, we evaluated the prognostic significance of different BCR-ABL transcript-level ratios (≤1%; >1%-≤10%; >10%) at 3, 6, 12, and 18 months (n = 199). The ELN optimal response criteria and their predictive role were significantly beneficial for event-free survival at all given time points. We found significant improvement in survivals of patients with BCR-ABL lower than 10% in the 6th and 12th months. Significantly better outcome was found in patients who achieved major molecular response (MMR) in the 12th month. The cumulative incidences of complete cytogenetic response (CCyR) and MMR were significantly associated with the molecular response in the 3rd month. The ELN response criteria and their predictive role were helpful at given time points; however, the 2009 definition did not significantly alter the prognostic accuracy compared with that of the 2006 definition. The significant value was observed for cytogenetic responses at the 6th and 12th month. Moreover, progression-free and event-free survivals were improved with MMR at the 12th month.