Abstract Introduction Continuous treatment with a Bruton's tyrosine kinase inhibitor (BTKi) is a standard of care option for patients (pts) with previously untreated chronic lymphocytic leukemia (CLL). Ibrutinib (Ibr), the first-in-class BTKi, has demonstrated survival benefits comparable to an age-matched general population. In the RESONATE-2 trial with up to 10 years of follow-up, Ibr achieved sustained progression-free survival benefit. Furthermore, Ibr dose modifications improved or resolved adverse events (AEs) in ≥ 85% of pts in this study. In the real-world settings, Ibr dose modification approaches may enhance overall tolerability and decrease side effects without impacting efficacy. Previous research has shown that total BTK protein levels decrease during Ibr treatment, and a model-based approach demonstrated that reducing the dose of Ibr to 280 mg following an initial 28-day cycle at 420 mg can effectively sustain BTK inhibition (Ouerdani A et al, 2025). Supporting this, pilot clinical data with limited follow-up have shown that the biological activity of Ibr can be retained following a similar dose modification approach (Chen LS et al, 2018). Here we present the exploratory analyses of the pharmacokinetics and pharmacodynamics data from the randomized Ibr monotherapy cohorts of the prospective phase 2 trial TAILOR study in pts with previously untreated CLL. Methods In the phase 2 TAILOR (NCT05963074) study, pts with untreated CLL received either Ibr + venetoclax or Ibr monotherapy at physician's choice. Ibr monotherapy cohorts were randomized 1:1 to receive Ibr 420 mg once daily (QD) (Cohort 2a) or 1 cycle of Ibr 420 mg QD followed by Ibr 280 mg QD (Cohort 2b) until progression or intolerability. Blood samples from pts receiving Ibr monotherapy were collected at screening, Cycle 1 Day 14 (C1D14), and Cycle 4 Day 1 (C4D1) to assess predose BTK occupancy, and at C1D14/C4D1 to assess plasma Ibr concentration before and after dose administration (at predose, 2 hours [h], and 4h after dosing). Results were summarized using descriptive statistics; Ibr plasma concentrations were calculated as geometric means (gmeans) and BTK occupancy as median of data from the specified cohort/timepoint. Results Blood samples were evaluated from 21 pts: 10 from Cohort 2a and 11 from Cohort 2b. Pts had a median age of 76 years (range, 59-84) and 85.7% were aged ≥ 65 years. Of 21 pts, 7 had a TP53/del(17p) aberration and 14 of 18 assessed pts had unmutated IGHV. Data from additional patient samples will be included in the presentation. In Cohort 2a, predose gmean Ibr plasma concentration was similar at C4D1 compared with C1D14 (2.2 [standard deviation (SD): 1.8] and 2.1 [SD: 2.4] ng/mL) but was higher 2h post treatment at C4D1 compared with C1D14 (58.8 [SD: 42.2] and 32.1 [SD: 48.6] ng/mL), likely due to variability and possibly some slight accumulation of plasma Ibr. In Cohort 2b (proactively dose-reduced cohort), the gmean Ibr plasma concentration was lower at C4D1 compared with C1D14 at predose (1.7 [SD: 3.1] and 3.4 [SD: 14.1] ng/mL) and 2h post treatment (19.2 [SD: 46.2] and 34.0 [SD: 242.7] ng/mL). When comparing gmean Ibr plasma concentration at C4D1 between Cohorts 2a and 2b, the predose and 2h post treatment plasma concentrations were lower for Cohort 2b: 2.2 (SD: 1.8) versus 1.7 (SD: 3.1) ng/mL and 58.8 (SD: 42.2) versus 19.2 (SD: 46.2) ng/mL, respectively. Median predose BTK occupancy was similar between Cohorts 2a and 2b at C1D14 (99.4% [range, 98.7-99.9%] and 98.9% [range, 92.9-100.0%]), and C4D1 (99.5% [range, 98.3-100%] and 99.5% [range, 98.7-100.0%]). Median BTK occupancy for C1D14 and C4D1 combined was 99.4% for Cohort 2a and 99.2% for Cohort 2b. BTK occupancy was observed to be nearly complete for all Ibr plasma concentration levels measured at C1D14 and C4D1. Conclusions In this exploratory end point analysis of the prospective phase 2 TAILOR study, both study cohorts had > 99% median post-baseline BTK occupancy regardless of Ibr dosage group. These results indicate that high BTK occupancy levels can be maintained after proactively reducing Ibr dose to 280 mg QD after 1 cycle with 420 mg QD, with concomitant lower serum Ibr concentration in pts randomized to the proactively dose reduced Cohort 2b. Further analyses are needed to determine whether the observed BTK occupancy and lower Ibr serum concentrations are associated with similar efficacy and better safety outcomes.

Central nervous system (CNS) involvement in multiple myeloma (MM) is a rare but severe complication with a poor prognosis. The identification of malignant plasma cells in cerebrospinal fluid (CSF) is essential for early diagnosis and intervention. However, the sensitivity of traditional diagnostic methods like cytology is low, especially in samples with low-cell counts. This study aimed to develop a multidimensional radar dot-plot analysis using Kaluza software to enhance the sensitivity and specificity of flow cytometry for detecting abnormal plasma cells in CSF. One hundred and twenty-five CSF samples were sent for flow cytometric testing to investigate the central nervous system involvement of MM. Finally, 89 samples from 40 patients were included in our study. Multicolor flow cytometry was performed using an 8-color labeling method, and radar dot-plot analysis was developed using diagnostic bone marrow samples to distinguish normal plasma cells, abnormal plasma cells, and cellular debris. The sensitivity of the novel method was evaluated by diluting myeloma bone marrow cells in pooled CSF samples to simulate low cell counts. Of the 125 CSF specimens, 16 samples from 4 patients showed abnormal plasma cells using both conventional and multidimensional flow cytometry analysis. Discordant results were found in 32 samples (25%), where conventional analysis suggested the presence of abnormal cells, but these were ruled out by multidimensional analysis. Sensitivity testing showed that the multidimensional dot-plot method outperforms conventional two-dimensional dot-plot analysis, as the radar dot plot can be used to identify abnormal cells in samples diluted to 5 WBC/μL, where the cell count of abnormal plasma cells is < 1 cell/μL. Our results showed that the new radar dot-plot analysis can increase the sensitivity and specificity of flow cytometry in MM for the detection of CNS involvement, even in low-cell-count CSF samples, regardless of whether the sample was obtained in a tube containing special reagent or not (TransFix/EDTA CSF Sample Storage tubes). This approach improves diagnostic accuracy, reduces the number of false positive cases caused by antibodies adhering to cell debris, and provides a reliable tool for assessing neurological complications in MM. Further validation is needed in a larger number of cases and testing of the method on different antibody panels.

Background/Objectives: Despite therapeutic advances, managing relapsed/refractory multiple myeloma (RRMM) remains challenging. For patients with frailty, comorbidities, mobility limitations, or when treatment preference and drug accessibility are key considerations, the all-oral ixazomib-lenalidomide-dexamethasone (IRd) regimen offers a practical alternative. Methods: We performed a multicenter retrospective study of RRMM patients treated with IRd in Hungary between 1 January 2020 and 30 June 2025. Results: The median age at treatment initiation was 73.7 years. Treatment was initiated for clinical progression in 38.2%, biochemical progression in 53.3%, and for intolerance or toxicity of prior therapy in 8.6%. Median progression-free survival (PFS) was 18.7 months, and median overall survival (OS) was 34.7 months. Patients treated at biochemical progression had significantly longer PFS than those treated at clinical progression (24.3 vs. 15.6 months; p = 0.004), with additional benefit when IRd was initiated owing to intolerance or toxicity of previous therapy (p = 0.04). In the second-line setting, median PFS was 24.5 months, and median OS was not reached. Adverse events occurred in 68.3% of patients; dose reductions were required in 18.4%, and 21.6% discontinued treatment because of intolerance or toxicity. Most common toxicities were neutropenia (32.9%), thrombocytopenia (27.6%), diarrhoea (25%), peripheral neuropathy (25.3%), and infections (22.4%). Conclusions: IRd initiation at biochemical progression was associated with superior PFS compared with treatment at clinical progression. When compared with a recent Hungarian multicenter cohort treated with second-line daratumumab, lenalidomide, and dexamethasone, outcomes with IRd are not significantly inferior (36-month OS calculated from 2nd line treatment initiation: 65.5% for DRd vs. 60% in our cohort; p = 0.56). These real-world data support IRd as an effective, convenient, all-oral option for appropriately selected RRMM patients.

Background: Acquired factor FXIII (FXIII) deficiency can be immune- or non-immune mediated and may cause severe bleeding symptoms. The incidence of acquired FXIII deficiency and its etiology in patients with multiple myeloma (MM) are poorly understood.Objectives: To assess FXIII levels and the balance of fibrinolysis in newly diagnosed, untreated MM and monoclonal gammopathy of undetermined significance (MGUS) patients.Methods: FXIII activity, mixing studies, FXIII-A2B2 antigen, total FXIII-B antigen were measured in platelet-poor plasma from 17 untreated MM patients, 33 untreated MGUS patients, and 30 age and sex-matched healthy controls. Besides routine laboratory measurements, the balance of coagulation and fibrinolysis was evaluated using quantitative fibrin monomer (FM) test, thrombin-antithrombin assay, alpha 2-antiplasmin activity, plasmin-alpha 2antiplasmin (PAP) complex, D-dimer, plasmin generation assay, clot lysis assay, and ClotPro-TPA test.Results: FXIII-A2B2 levels were significantly lower in MM patients compared to controls [median (IQR):14.6 (11.2-19.4) vs. 21.8 (17.1-26.4) mg/L, p = 0.0015], whereas total FXIII-B did not differ between groups. Decrease in FXIII activity was parallel to the decrease in FXIII-A2B2. An immune-mediated inhibitory mechanism was ruled out. Free/total FXIII-B was significantly higher in MM patients compared to MGUS and healthy controls, suggesting an etiology of FXIII-A consumption. In MM and MGUS patients, FM, D-dimer, and PAP complex were significantly elevated compared to controls, indicating hypercoagulability and ongoing fibrinolysis.Conclusions: Low FXIII levels due to consumption were observed in MM patients at diagnosis. Hypercoagulability and ongoing fibrinolysis were detected in MM and MGUS, indicating that a disturbed hemostasis balance is already present in the latter benign condition.

Signaling pathways of Retinoblastoma (Rb) protein, Akt-kinase, and Erk-kinase (extracellular signal-regulated kinase) have an important role in the pathogenesis of acute myeloid leukemia. Constitutive activation of these proteins by phosphorylation contributes to cell survival by regulation of cell cycle, proliferation and proapoptotic signaling processes. According to previous data phosphorylated forms of these proteins represent a worse outcome for cancer patients. We investigated the presence of phosphorylated Rb (P-Rb), Akt (P-Akt) and Erk (P-Erk) proteins by Western blot technique using phospho-specific antibodies in bone marrow or peripheral blood samples of 69 AML patients, 36 patients with myelodysplastic syndrome (MDS) and 10 healthy volunteers. Expression level of PTEN (Phosphatase and tensin homolog) and PHLPP (PH domain and leucine-rich repeat Protein Phosphatase) phosphatases, the negative regulators of Akt kinase pathway were also examined. We tested the effect of these proteins on survival and on the correlation with known prognostic features in AML. We found 46.3% of AML patients had detectable P-Rb, 34.7% had P-Akt and 28.9% had P-Erk protein. 66.1% of patients expressing PTEN, 38.9% PHLPP, 37.2% both PTEN and PHLPP and 32.2% neither PTEN nor PHLPP phosphatases. Compared to nucleophosmin mutation (NPMc) negative samples P-Erk was significantly less in nucleophosmin mutated patients, P-Rb was significantly less in patients’ group with more than 30 G/L peripheral leukocyte count by diagnosis. PHLPP was significantly present in FAB type M5. The expression of P-Rb represented significant better overall survival (OS), while P-Akt represented significantly worse event-free survival (EFS) in unfavorable cytogenetics patients. The presence of both PHLPP and PTEN phosphatases contributes to better OS and EFS, although the differences were not statistically significant. We confirmed significant positive correlation between P-Akt and PHLPP. Assessing the phosphorylation of Rb, Akt and Erk may define a subgroup of AML patients who would benefit especially from new targeted treatment options complemented the standard chemotherapy, and it may contribute to monitoring remission, relapse or progression of AML.

TP53 aberrations predict chemoresistance and represent a contraindication for the use of standard chemoimmunotherapy in chronic lymphocytic leukaemia (CLL). Recent next-generation sequencing (NGS)-based studies have identified frequent low-burden TP53 mutations with variant allele frequencies below 10%, but the clinical impact of these low-burden TP53 mutations is still a matter of debate. In this study, we aimed to scrutinise the subclonal architecture and clinical impact of TP53 mutations using a sensitive, NGS-based mutation analysis in a 'real-world' cohort of 901 patients with CLL. In total, 225 TP53 mutations were identified in 17.5% (158/901) of the patients; 48% of these alterations represented high-burden mutations, while 52% were low-burden TP53 mutations. Low-burden mutations as sole alterations were identified in 39% (62/158) of all mutated cases with 82% (51/62) of these being represented by a single low-burden TP53 mutation. Patients harbouring low-burden TP53 mutations had significantly lower time to first treatment compared to patients with wild-type TP53. Our study has expanded the knowledge on the frequency, clonal architecture, and clinical impact of low-burden TP53 mutations. By demonstrating that patients with sole low-burden TP53 variants represent more than one-third of patients with TP53 mutations and have an increased risk for treatment initiation, our findings strengthen the need to redefine the threshold of TP53 variant reporting to below 10% in the routine diagnostic setting.

BevezetésA marginális zóna lymphomás betegekben a hepatitis C vírus pozitivitás viszonylag gyakori (39%). Krónikus hepatitis C vírus fertőzésben a non-Hodgkin lymphoma incidenciája 15%, ami magasabb, mint az átlagpopulációban (1,5%). Marginális zóna lymphomában az antivirális kezelés komplett remissziót eredményezhet.Betegek és módszerKét marginális zóna lymphomában szenvedő, hepatitis C vírussal fertőzött beteg esetét mutatjuk be.EredményekKét hepatitis C vírus fertőzött marginális zóna lymphomás betegünk antivirális kezelés hatására komplett remisszióba került, amely az egyik betegnél két éve tartósan fennáll. A másik betegnél az eredményes kezelést követően négy év után jelentkező inguinális lymphadenomegalia hátterében szekunder aktivált B-sejtes diffúz nagy B-sejtes lymphoma igazolódott, amely immunkemoterápia hatására komplett remisszióba került.MegbeszélésNon-Hodgkin lymphoma diagnózisakor minden esetben ajánlott a hepatitis C vírus szerológia elvégzése. A hepatitis C vírus pozitivitás mellett kialakult marginális zóna lymphoma kezelése immunkemoterápia nélkül, antivirális kezelés hatására is eredményes lehet.

BACKGROUND:Patients with multiple myeloma (MM) are at high risk of thrombosis especially when receiving immunomodulatory therapy. Thrombotic risk in patients with monoclonal gammopathy of undetermined significance (MGUS) may also be increased. Although activated protein C (APC) resistance has been linked to an increased risk of thrombosis in MM, little is known about how APC influences thrombotic risk in MGUS. We compared thrombin generation (TG) in MM and MGUS patients to that of healthy controls (HCs) and investigated the exogenous effect of APC on TG in these groups. METHODS:Hemostasis tests including factor VIII (FVIII) and von Willebrand factor (vWF) levels were measured in platelet-poor plasma in 14 untreated MM patients, 34 MGUS patients, and 30 age and sex-matched HCs. TG assay was performed with or without the addition of APC. RESULTS:Peak thrombin and velocity index were significantly higher in MM and MGUS patients compared to HCs, while MM patients also had elevated endogenous thrombin potential (ETP). In MGUS cases, ETP and peak thrombin values significantly correlated with FVIII and vWF levels. In the presence of APC, peak thrombin and ETP were reduced in MGUS and control plasmas whereas lagtime and time to peak were significantly prolonged. In contrast, adding APC to MM plasma had no effect on any TG parameters. CONCLUSIONS:Hypercoagulability was observed in MM and even in MGUS cases with very low monoclonal protein concentration. In MM patients, APC had no effect on TG, but it attenuated TG in MGUS patients.

Topic: 16. Myeloproliferative neoplasms - Clinical Background: Nearly 35% of MF pts have severe thrombocytopenia (TCP) with a shortened median overall survival of 7-15 months (mos) (Masarova 2018, 2020). Most pts present with high symptom burden due to dysregulated cytokines (Tefferi 2011), of which elevated IL-8 impairs megakaryocyte function (Emadi 2005). Treatment with JAK inhibitors (JAKi) can worsen TCP in MF pts, with no effective therapies to improve plt counts. TL-895 is a highly potent, selective, orally available, small molecule inhibitor of BTK and bone marrow tyrosine kinase X-linked (BMX) being studied in MF for its potential to (i) impair stromal adhesion, (ii) disrupt aberrant CD34+ cell trafficking (Nimmagadda 2019), (iii) reduce proinflammatory cytokine-mediated symptoms, and (iv) reverse dysfunctional megakaryopoiesis to improve plt counts. Aims: Safety, efficacy, and tolerability of TL-895 in MF pts with severe TCP. Methods: Cohort 3 of this open-label, global Phase (Ph) 2 study (NCT04640532) enrolled adult pts with JAKi-ineligible MF (plts <50 K/μL, ≥25 K/μL). Pts were randomized to TL-895 150 mg BID (Arm A) or 300 mg QD (Arm B). Eligible pts were symptomatic with intermediate/high-risk MF (DIPSS), ECOG £2 with splenomegaly. The primary objective was the recommended Ph 2 dose (RP2D). Key secondary objectives were Total Symptom Score improvement ³50% by MFSAF v4.0 at Week (Wk) 24 (TSS-50), spleen volume reduction ³35% at Wk 24 by central review (SVR-35) and safety. Plt response was assessed per modified IWG-MRT 2006 criteria (plt increase ≥50% from baseline and >50 K/μL for ≥8 wks independent of plt transfusion [Tefferi 2006]). Results: As of 27 Jan 2022, 11 pts were enrolled in Arm A and five pts in Arm B with median follow-up of 11.7 mos. Complete BTK occupancy (≥95%) was achieved in Arm A at trough (C1D8), but not Arm B which closed early. Arm A is described herein. In these 11 pts, baseline median plt count was 39 K/μL, median spleen volume was 1908 cm3, median TSS was 24.7, and 73% were previously treated with a JAKi (Table 1). Six pts (55%) remain on study and five discontinued due to Grade (Gr) 3 fatigue (n=1), progression (n=2) and investigator decision (n=2). Ten (91%) pts were alive at data cut (median 12.4 mos). At Wk 24, four pts (36%) achieved TSS-50 (Fig. 1), despite no pts achieving SVR-35 (median SVR -5.3%, range -18, 38)). Per modified IWG-MRT criteria, five pts (45%) achieved plt response (≥50% improvement for ≥8 weeks [Fig. 2]), two pts (18%) achieved ≥100% plt improvement with a third pending 8-week confirmation (27%). Median time to plt response was 2.9 mos; median duration was 6.7 mos (range 1.9, 13.1+). Median change in serum IL-8 levels from baseline to Wk 12 was -38% and was associated with plt and TSS response. Median change in circulating CD34+ cells from baseline to Wk 4 was +85%, demonstrating transient cell trafficking due to BTK inhibition. The most common treatment-emergent adverse events were anemia (55%), abdominal pain, nausea and TCP (27% each). Anemia and TCP were the most common Gr 3/4 AEs, regardless of causality, 46% and 27%, respectively (Table 1). Summary/Conclusion: In MF pts with severe TCP, TL-895 provided clinically meaningful improvements in TSS and plt counts that were associated with reductions in IL-8. This is the first clinical proof-of-concept for BTKi in the treatment of MF and supports further investigation in a recently commenced randomized, double blind, placebo-controlled Ph 2b study.Keywords: Thrombocytopenia, IL-8, Myelofibrosis, Bone Marrow Fibrosis

Patients with myelofibrosis (MF) who discontinue ruxolitinib due to progression/resistance have poor prognoses. JAK inhibitors control symptoms and reduce spleen volumes with limited impact on underlying disease pathophysiology. Murine double minute 2 (MDM2), a negative regulator of p53, is overexpressed in circulating malignant CD34+ MF cells. The oral MDM2 inhibitor navtemadlin (KRT-232) restores p53 activity to drive apoptosis of wild-type TP53 tumor cells by inducing expression of pro-apoptotic Bcl-2 family proteins. Navtemadlin demonstrated promising clinical and disease-modifying activity and acceptable safety in a phase II study in patients with relapsed/refractory MF. The randomized phase III BOREAS study compares the efficacy and safety of navtemadlin to best available therapy in patients with MF that is relapsed/refractory to JAK inhibitor treatment.

Invasive aspergillosis (IA) may occur as a serious complication of hematological malignancy. Delays in antifungal therapy can lead to an invasive disease resulting in high mortality. Currently, there are no well-established blood circulating microRNA biomarkers or laboratory tests which can be used to diagnose IA. Therefore, we aimed to define dysregulated miRNAs in hematology and oncology (HO) patients to identify biomarkers predisposing disease. We performed an in-depth analysis of high-throughput small transcriptome sequencing data obtained from the whole blood samples of our study cohort of 50 participants including 26 high-risk HO patients and 24 controls. By integrating in silico bioinformatic analyses of small noncoding RNA data, 57 miRNAs exhibiting significant expression differences (P < 0.05) were identified between IA-infected patients and non-IA HO patients. Among these, we found 36 differentially expressed miRNAs (DEMs) irrespective of HO malignancy. Of the top ranked DEMs, we found 14 significantly deregulated miRNAs, whose expression levels were successfully quantified by qRT-PCR. MiRNA target prediction revealed the involvement of IA related miRNAs in the biological pathways of tumorigenesis, the cell cycle, the immune response, cell differentiation and apoptosis.

Despite the introduction of novel agents, multiple myeloma remains incurable for most patients, necessitating further therapeutic options. Venetoclax, a selective BCL-2 inhibitor, had shown promising results in patients with translocation t(11;14), but questions remain open about its optimal use. We have contacted all Hungarian haematology centers for their experience treating t(11;14) myeloma patients with venetoclax. 58 patients were reported. 37 received venetoclax in the relapsed/refractory setting with few or no other therapeutic options available. 21 patients started venetoclax as salvage after failing to achieve satisfactory response to first line therapy. In the relapsed/refractory setting objective response rate (ORR) was 94%, median progression-free survival (PFS) 10.0 months and median overall survival (OS) 14.6 months. In reinduction patients, ORR was 100%, median PFS and OS were not reached. Importantly, we found no adverse effect of high risk features such as deletion 17p or renal failure, in fact renal failure ameliorated in 42% of the cases, including three patients who became dialysis independent. Our study also reports the highest number of plasma cell leukemia cases successfully treated with venetoclax published in literature, with refractory plasma cell leukemia patients achieving a median PFS of 10.0 and a median OS of 12.2 months.

Introduction Chronic lymphocytic leukemia (CLL) is a clinically heterogeneous disease with various genetic abnormalities that can affect the treatment and survival of patients. Recently, by the advent of next-generation sequencing (NGS) techniques recurrently mutated genes have been described in CLL. Driver gene mutations such as NOTCH1, SF3B1, TP53, and MYD88 have been considered to have prognostic impact in CLL. The aim of this study was to detect pathogenic variants of commonly mutated genes in CLL patients and to explore their associations with IGHV mutational status and cytogenetic abnormalities. Patients and Methods The samples were collected from bone marrow or peripheral blood of 124 CLL patients. Genomic DNA was isolated using QIAamp Blood Mini kit. NGS analysis was performed using Twist Custom Panel and bidirectional sequencing with a minimum coverage of 1000x on Illumina NextSeq according to the manufacturers’ instructions. Data were analyzed by NextGene software. The following 27 recurrently mutated CLL genes were studied: PIK3CD, PIK3CA, SF3B1, MYD88, FBXW7, BRAF, NOTCH1, PTEN, BIRC3, PTPN6, MAP2K1, TP53, STAT5B, STAT3, CD79B, TCF3, MAPK1, DDX3X,ATM, BCL2, BTK, KRAS, NRAS, CARD11, CXCR4, PLCG2, POT1. Variants below 5% variant allele frequency (VAF) were filtered out from the analysis. Variants were considered subclonal with 5-10% VAF and clonal when VAF was >10%. Classification of variants were performed according to the American College of Medical Genetics and Genomics (ACMG) criteria. Results Our NGS study revealed a total of 96 variants in 124 CLL patients. Overall, 63/124 patients (50.8%) harbored at least one mutation, the majority of them (60/96, 62.5%) was clonal. In 36 patients only one pathogenic variant was detected, while 88 patients carried two or more (2-4) genetic alterations. The most frequently mutated gene was the NOTCH1, followed by TP53, SF3B1, FBXW7, and BRIC3. Subclonal variants were also detected in 36/124 patients (29%), particularly in NOTCH1,FBXW7, BIRC3, and PIK3CA genes. TP53 and NOTCH1 variants occured mostly in unmutated IGHV CLL cases. Patients with mutated IGHV harbored mainly FBXW7 and BIRC3 mutationss. FBXW7 variants were detected in 9/125 (7%) patients, that is a higher frequency compared to other studies. Fluorescence in situ hybridization (FISH) was performed in 113 cases. The majority of these patients (84 cases, 74%) had one or more cytogenetic abnormality. In this cohort, 13q deletion was the most common aberration (65%, 55/84 patients), followed by 11q deletion (26%, 22/84 patients) and 17q deletion (14%, 12/84 patients). Deletion of 17p co-occured with TP53 single nucleotide variants in 10 of 12 patients. SF3B1, NOTCH1, BIRC3 and TP53 variants occured with 11q deletion in 10 cases. Trisomy 12 was detected with NOTCH1, BIRC3 and FBXW7 variants, and showed negative association with unfavorable SF3B1 mutations. Conclusions Our results confirmed the presence of high number of clonal and subclonal variants in the most frequently mutated genes (NOTCH1, TP53, SF3B1, FBXW7, BIRC3). TP53, NOTCH1 and SF3B1 mutations were enriched with unfavorable biologic factors, as they were found to be associated with unmutated IGHV and cytogenetic aberrations, such as del17p, del11q and trisomy 12. Cases with NOTCH1 variants harbored all types of the investigated cytogenetic alterations (del13q, del11q, de17p, trisomy 12) and were associated exclusively with unmutated IGHV. TP53 variants occured mostly in unmutated IGHV CLL cases together with del17p. Patients with SF3B1 variants showed del11q, del13q and mostly unmutated IGHV. FBXW7 variants enriched in both of unmutated and mutated IGHV cases and harbored del13q, de17p and trisomy 12. FBXW7 and NOTCH1 pathogenic variants have the same biological consequences in CLL, therefore presence of FBXW7 mutations may have clinical relevance regarding anti-CD20 therapy. Taken together, these NGS results complemented with the study of IGHV mutational status and cytogenetic data can contribute to better prognostic workup and management of our CLL patients.

SummaryThe Bruton's tyrosine kinase (BTK) inhibitor ibrutinib has revolutionised the therapeutic landscape of chronic lymphocytic leukaemia (CLL). Acquired mutations emerging at position C481 in the BTK tyrosine kinase domain are the predominant genetic alterations associated with secondary ibrutinib resistance. To assess the correlation between disease progression, and the emergence and temporal dynamics of the most common resistance mutation BTKC481S, sensitive (10−4) time‐resolved screening was performed in 83 relapsed/refractory CLL patients during single‐agent ibrutinib treatment. With a median follow‐up time of 40 months, BTKC481S was detected in 48·2% (40/83) of the patients, with 80·0% (32/40) of them showing disease progression during the examined period. In these 32 cases, representing 72·7% (32/44) of all patients experiencing relapse, emergence of the BTKC481S mutation preceded the symptoms of clinical relapse with a median of nine months. Subsequent Bcl‐2 inhibition therapy applied in 28/32 patients harbouring BTKC481S and progressing on ibrutinib conferred clinical and molecular remission across the patients. Our study demonstrates the clinical value of sensitive BTKC481S monitoring with the largest longitudinally analysed real‐world patient cohort reported to date and validates the feasibility of an early prediction of relapse in the majority of ibrutinib‐treated relapsed/refractory CLL patients experiencing disease progression.

Összefoglaló. Bevezetés: A krónikus lymphoid leukémia kezelésében jelentős előrelépést eredményezett az ibrutinibterápia bevezetése. A gyógyszer első vonalbeli és többed vonalbeli kezelése is magas remissziós arányt eredményezett, bár a terápia korai bevezetése és a kedvező genetikai eltérések esetén az eredmények jobbak. A progressziómentes túlélést befolyásoló egyéb tényezőkről azonban még kevés adat áll rendelkezésre. Célkitűzés: Krónikus lymphoid leukémiás betegek ibrutinibkezelése során a teljes hematológiai remisszió elérését és a progressziómentes túlélés időtartamát befolyásoló tényezők vizsgálata. Betegek és módszer: 47 krónikus lymphoid leukémiás beteg (életkor: 39–84 év, férfi 27, nő 20, követési idő 5–58 hónap, medián 15 hónap) klinikai és laboratóriumi adatainak retrospektív elemzése. Eredmények: A teljes hematológiai remisszió elérése független volt a betegek nemétől, életkorától, a betegség stádiumától, az immunglobulin gén nehézlánc-variábilis régió státuszától, a genetikai aberrációktól, az abszolút neutrophilszámtól, az abszolút monocytaszámtól és a vörösvértestnagyság-eloszlási görbe szélességétől. A progressziómentes túlélést a komplett remisszió elérése ( p = 0,00073) és a magasabb abszolút neutrophilszám (<4 G/l vs. ≥4 G/l, p = 0,022) befolyásolta szignifikánsan, a vörösvértestnagyság-eloszlási görbe szélességértékével való összefüggés pedig statisztikailag határértéken volt ( p = 0,065). A Cox-féle regressziós elemzésbe bevont változók közül csak a teljes hematológiai remisszió elérése mutatott szignifikáns hatást a progressziómentes túlélésre ( p = 0,0147). Következtetések: A teljes hematológiai remisszió elérése az egyéb vizsgált tényezőktől független, szignifikáns hatással bír a betegek progressziómentes túlélésére. Az abszolút neutrophilszám és a vörösvértestnagyság-eloszlási görbe szélessége szintén hasznos kiegészítő prognosztikus marker lehet. Az elemzett esetek száma még alacsony a komolyabb következtetések levonására, azonban így is elmondható, hogy az eredmények egy része már a szakirodalom korábbi eredményeit tükrözi. Summary. Introduction: The introduction of ibrutinib therapy has led to significant advances in the treatment of chronic lymphocytic leukemia. Both first-line and multiple-lines treatments of the drug resulted in high remission rates, although results were better with early initiation of therapy and favorable genetic abnormalities. However, little data are available on other factors influencing progression-free survival. Objective: To investigate factors influencing the achievement of complete hematological remission and progression-free survival with ibrutinib treatment in patients with chronic lymphocytic leukemia. Patients and metods: Retrospective analysis of clinical and laboratory data from 47 chronic lymphoid leukemia patients (age: 39–84 years, male 27, female 20, follow-up 5–58 months, median 15 months). Results: Achieving complete hematologic remission was independent of patient gender, age, disease stage, immunoglobulin heavy chain variable region status, genetic aberrations, absolute neutrophil count, absolute monocyte count, and red blood cell distribution width. Progression-free survival was significantly affected by complete remission ( p = 0.00073), and higher absolute neutrophil counts (<4 G/l vs. ≥ 4 G/l, p = 0.022), the red blood cell distribution width was statistically less significant ( p = 0.065). Of the variables included in the Cox regression analysis, only the achievement of complete hematologic remission had a significant effect on progression-free survival ( p = 0.0147). Conclusions: Achieving complete hematologic remission, independent of the other factors studied, has a significant effect on patients’ progression-free survival. Absolute neutrophil count and red blood cell distribution width can also be a useful additional prognostic marker. The number of analyzed cases is still low to draw more serious conclusions, but it can still be said that some of the results already reflect previous results in the literature.

Background Despite therapeutic advances, multiple myeloma remains incurable. Venetoclax, a selective bcl-2 inhibitor may be a step toward personalised therapy for t(11;14) patients, but questions remain about its optimal use. Methods We retrospectively evaluated hematologic response, survival and safety after venetoclax treatment in t(11;14) myeloma patients in Hungary. Results Overall, 49 patients from seven clinical centers were reported. We divided patients into two groups based on the clinical setting: 32 relapsed/refractory patients, who received venetoclax after multiple lines of therapy, often in an ultimate effort; and 17 frontline patients, who achieved unsatisfactory response to standard first line therapy and were treated with venetoclax as reinduction before intended ASCT. We observed remarkably good hematological response rates (ORR): 94% in the relapsed/refractory group and 100% in the frontline setting. This translated into a median PFS of 9.6 months and a median OS of 14.6 months in the relapsed group; median PFS and OS were not reached in the reinduction group. Known adverse prognostic factors, such as 17p deletion, kidney failure or 1q21 amplification did not convey significantly worse prognosis in our study. Almost one third of patients had impaired kidney function during venetoclax therapy, including three patients requiring dialysis. Clinically relevant improvement was observed in 42% of these patients; dialysis could be stopped in all three cases. Notably, our study also included six plasma cell leukemia patients, with a remarkable median PFS of 10 months and over one year median OS in relapsed disease. Vulnerable patients with PCL, renal failure or refractory disease reported more adverse events, requiring supportive measures or dose adjustment, but cessation of venetoclax therapy did not become necessary. Conclusion Venetoclax therapy is a promising option with few side effects and very good response rates for t(11;14) patients, both in the frontline and in the relapsed setting Despite therapeutic advances, multiple myeloma remains incurable. Venetoclax, a selective bcl-2 inhibitor may be a step toward personalised therapy for t(11;14) patients, but questions remain about its optimal use. We retrospectively evaluated hematologic response, survival and safety after venetoclax treatment in t(11;14) myeloma patients in Hungary. Overall, 49 patients from seven clinical centers were reported. We divided patients into two groups based on the clinical setting: 32 relapsed/refractory patients, who received venetoclax after multiple lines of therapy, often in an ultimate effort; and 17 frontline patients, who achieved unsatisfactory response to standard first line therapy and were treated with venetoclax as reinduction before intended ASCT. We observed remarkably good hematological response rates (ORR): 94% in the relapsed/refractory group and 100% in the frontline setting. This translated into a median PFS of 9.6 months and a median OS of 14.6 months in the relapsed group; median PFS and OS were not reached in the reinduction group. Known adverse prognostic factors, such as 17p deletion, kidney failure or 1q21 amplification did not convey significantly worse prognosis in our study. Almost one third of patients had impaired kidney function during venetoclax therapy, including three patients requiring dialysis. Clinically relevant improvement was observed in 42% of these patients; dialysis could be stopped in all three cases. Notably, our study also included six plasma cell leukemia patients, with a remarkable median PFS of 10 months and over one year median OS in relapsed disease. Vulnerable patients with PCL, renal failure or refractory disease reported more adverse events, requiring supportive measures or dose adjustment, but cessation of venetoclax therapy did not become necessary. Venetoclax therapy is a promising option with few side effects and very good response rates for t(11;14) patients, both in the frontline and in the relapsed setting

Fungal infections represent a worrisome complication in hematologic cancer patients and in the absence of disease specific symptoms, it is important to establish new biological indicators, which can be used during mould-active prophylaxis. Recently, miRNAs have appeared as candidate diagnostic and prognostic markers of several diseases. A pilot clinical study was performed to evaluate the diagnostic utility of 14 microRNAs which can be related to invasive fungal infections. Based on our data miR-142-3p, miR-142-5p, miR-26b-5p and miR-21-5p showed significant overexpression (p < 0.005) due to invasive aspergillosis in hemato-oncology patients with profound neutropenia. A tetramiR assay was designed to monitor peripheral blood specimens. Optimal cut-off was estimated by using the median value (fold change 1.1) of the log10 transformed gene expressions. The biomarker panel was evaluated on two independent sample cohorts implementing different antimicrobial prophylactic strategies. The receiver operating characteristic analysis with area under the curve proved to be 0.97. Three miRNAs (miR-142-5p, miR-142-3p, miR-16-5p) showed significant expression alterations in episodes with sepsis. In summary, the tetramiR assay proved to be a promising diagnostic adjunct with sufficient accuracy and sensitivity to trace invasive aspergillosis in hemato-oncology patients.

Background. Myeloma patients reaped immense benefit from the introduction of new classes of drugs over the last decades. This improvement, however, was much less marked in patients with translocation 11;14 [t(11;14)], a group in which immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs) - the two most important pillars of current myeloma care - are less effective. This subgroup of patients used to be known for their relatively slow pace of progression often experiencing long plateau phases following autologous stem cell transplantation (ASCT), whereas at the same time t(11;14) is also disproportionately prevalent in difficult to treat clinical entities such as plasma cell leukemia or AL amyloidosis. Venetoclax, a selective bcl-2 inhibitor first approved for CLL was investigated for the treatment of relapsed myeloma patients and although it failed to show benefit for myeloma patients as a whole, t(11;14) patients showed exceptional responses, thus paving the way towards the first genetically targeted treatment in myeloma. As a result, its off label use is on the rise, even though clinicians have to face unanswered questions regarding the right dosage and therapy length, as well as the potential for adverse events (AEs), especially infections. Real world data could help elucidate its optimal use, but is as of yet very limited. Aims and methods. We addressed all Hungarian centers treating myeloma to evaluate the efficacy and safety of venetoclax, collecting data about the treatment duration, AEs, dose modifications and treatment discontinuations, and analysed response rates as well as progression free survival (PFS). Results. 33 patients were reported from 7 Hungarian sites. After the initial analyses, we identified two distinct rationales for venetoclax treatment. 22 patients were relapsed and heavily pretreated with an average of 4.5 prior lines; here venetoclax was chosen as ultimum refugium. In this group, combination partners were bortezomib-dexamethasone (VelDex) in 14 patients, 5 had dexamethasone only, one VRd, one DRd and one Kd. Considering the highly pretreated nature of this group, the overall response rate was a remarkably high 95% with 40.9% partial, 31.8% very good partial, and 22.7% complete responses. Treatment mostly continued until progression. The median PFS and OS calculated from venetoclax initiation were 299 and 437 days. The most common AEs were cytopenias and infections reported in 8 and 6 patients with 1 fatal infection. In the second group, 11 patients received venetoclax after a suboptimal initial response (6 PR, 4 SD, 1 PD) to their first line IMiD+PI combination with the goal of further tumor elimination preceding ASCT. Remarkably, although the length of venetoclax treatment was short - median 2 cycles -, all 11 patients deepened their response to at least VGPR and 7 to CR. 9 patients had ASCT converting 2 further VGPRs into CR, so at the end of the planned protocol 10 of the 11 patients had CR. Venetoclax was combined with VelDex in 9 and VTD in 2 cases, the one year PFS and OS were 91 and 100%, with no venetoclax related AEs reported. An important aspect of our analysis was the question of venetoclax dosing, as the appropriate dose in this indication is not yet clear. Reflecting this uncertainty, as well as funding difficulties with this off-label drug, only one patient received 800 mg dose as seen in the Bellini trial; one received 600 mg daily, with all others taking 400 mg or less. To counteract this lower daily dose available, some centers employed a combination with clarithromycin, a strong CYP3A inhibitor known to increase venetoclax serum levels two- to threefold. Where available, serum venetoclax levels were monitored to ensure serum levels comparable to regular dosing. Another point to emphasize is that 5 patients in the relapsed, and another 2 in the frontline group had deletion 17p, usually resulting in refractoriness to standard treatments. Among these patients however, 5 reached VGPR, 1 PR, and only one progressed on venetoclax treatment. Some responses proved lasting especially in the frontline group. Conclusion. Our results highlight the importance of targeted treatments in multiple myeloma. We experienced lasting responses in quadruple-refractory patients. In the newly diagnosed group where the depth of pre-ASCT response has a big impact on PFS, venetoclax may have a role converting suboptimal responses into CRs by eliminating residual disease. Figure Disclosures Illés: Takeda, Seattle Genetics: Research Funding; Novartis, Janssen, Pfizer, Roche;: Other: Travel, Accommodations, Expenses; Celgene, Janssen, Novartis,Roche, Takeda: Consultancy; Janssen, Celgene, Takeda, Novartis Pharma SAS, Pfizer Pharmaceuticals Israel, Roche;: Consultancy, Honoraria. OffLabel Disclosure: venetoclax use in t(11;14) myeloma which is not yet licensed

Acute promyelocytic leukemia (APL) is generally characterized by t(15;17)(q24;q21). In some cases, the classic translocation cannot be identified by conventional methods, since the PML-RARA fusion protein results from complex, variant, or cryptic translocation. The diagnostic algorithm of APL starts with screening methods, such as flow cytometry (FC), followed by fluorescence in situ hybridization or polymerase chain reaction to confirm the diagnosis. Our aim was to develop a novel protocol for analyzing APL samples based on multidimensional dot-plots that can provide comprehensive information about several markers at the same time. The protocol included four optimized multidimensional dot-plots, which were tested by retrospective reanalysis of FC results in APL (n = 8) and non-APL (n = 12) acute myeloid leukemia (AML) cases. After predicting the potential position of hypergranular- and microgranular-type aberrant promyelocytes, the percentages of blast populations were examined within the gates in all AML cases. The percentage of blasts in each predefined gate was well above the cut-off value (95%) in APL cases in all tubes. In non-APL AML cases, the percentage of blasts in the same gates never reached the cut-off value in all investigated tubes, and even when it did in a single tube, the pattern was markedly different from that observed in APL cases. In conclusion, multidimensional dot-plots can be used for screening APL even in cryptic APL cases, although reproducibility across several laboratories would require standardization of antibodies and fluorochromes. This easy-to-use and quick method can support the diagnosis of APL and the prompt initiation of the appropriate treatment.