Mass Spectrometry-Based Assessment of M-protein in Peripheral Blood During Maintenance Therapy in Multiple Myeloma.

Mass spectrometry (MS) can detect multiple myeloma-derived monoclonal proteins in peripheral blood (PB) with high sensitivity, potentially serving as a PB assay for measurable residual disease (MRD). This study evaluated the significance of PB MS MRD negativity during post-transplant therapy in patients with newly diagnosed multiple myeloma. Serum samples from 138 patients treated in the phase 3 ATLAS trial of post-transplant maintenance with either carfilzomib, lenalidomide, dexamethasone or lenalidomide alone were analyzed using EXENT MS methodology. We established feasibility of measuring MRD by MS in PB in the post-transplant setting, despite unavailability of pre-treatment calibration samples. There was high agreement between MRD by MS in PB and paired BM MRD results at the 10-5 threshold, assessed by either next generation sequencing (NGS) or multiparameter flow cytometry (MFC) (70% and 67%, respectively). Agreement between PB MS and both BM MRD methods was lowest early after transplant and increased with time. MS negativity was associated with improved progression-free survival (PFS), which in landmark analysis reached statistical significance after 18 cycles post-transplant. Combined PB/BM MRD negativity by MFC or NGS was associated with superior PFS compared to MRD negativity by only one modality. Sustained MS negativity carried similar prognostic performance to sustained BM MRD negativity at the 10-5 threshold. Overall, post-transplant MS assessment was feasible and provided additional prognostic information to BM MRD negativity. Further studies are needed to confirm the role and optimal timing of MS in disease evaluation algorithms. The ATLAS trial is registered at www.clinicaltrials.gov as #NCT02659293.

Acute Myeloid Leukemia Post Cytotoxic Therapy in Breast Cancer Survivors-Over 23 Years of Single Center Analysis.

Background: Acute myeloid leukemia post cytotoxic therapy (AML-pCT) among breast cancer (BC) survivors represents a life-threatening complication. This study aims to assess the clinical outcomes of AML-pCT post BC. Methods: An analysis of all AML patients treated at a single hematology center (2000-2023) was performed to select patients with AML-pCT post BC. We applied the 2022 ELN criteria to define the genetic risk. Results: Among 847 AML patients, 28 were diagnosed with AML-pCT following BC. Complex karyotype (CK) occurred in 23.8% of patients. The median overall survival (OS) was 40 months. The survival outcomes were better after allogenic hematopoietic stem cell transplantation (alloHCT) treatment compared to chemotherapy alone (median OS: 47 versus 7 months, p = 0.008). Patients demonstrating CK showed lower survival compared to those without CK (2-year OS: 25.0% versus 66.2%, p = 0.0048). The multivariable Cox proportional hazards regression model indicated that treatment with alloHCT emerged as a significant factor associated with improved OS. The treatment was associated with superior OS (HR = 0.07, 95% CI = 0.01-0.86, p = 0.04). Conclusions: Patients with AML-pCT following BC were characterized with the highest frequency of adverse genetic risk profiles and demonstrated worse survival rates. AlloHCT should be performed as early as possible in such patients. The growing need for studies on inherited cancer susceptibility underscores the importance of close AML-pCT development monitoring in BC survivors.

BCR::ABL1-like acute lymphoblastic leukaemia: a single institution experience on identification of potentially therapeutic targetable cases.

BACKGROUND: BCR::ABL1-like acute lymphoblastic leukaemia (BCR::ABL1-like ALL) is characterized by inferior outcomes. Current efforts concentrate on the identification of molecular targets to improve the therapy results. The accessibility to next generation sequencing, a recommended diagnostic method, is limited. We present our experience in the BCR::ABL1-like ALL diagnostics, using a simplified algorithm. RESULTS: Out of 102 B-ALL adult patients admitted to our Department in the years 2008-2022, 71 patients with available genetic material were included. The diagnostic algorithm comprised flow cytometry, fluorescent in-situ hybridization, karyotype analysis and molecular testing with high resolution melt analysis and Sanger Sequencing. We recognized recurring cytogenetic abnormalities in 32 patients. The remaining 39 patients were screened for BCR::ABL1-like features. Among them, we identified 6 patients with BCR::ABL1-like features (15.4%). Notably, we documented CRLF2-rearranged (CRLF2-r) BCR::ABL1-like ALL occurrence in a patient with long-term remission of previously CRLF2-r negative ALL. CONCLUSIONS: An algorithm implementing widely available techniques enables the identification of BCR::ABL1-like ALL cases in settings with limited resources.

The presence of additional cytogenetic aberrations in chronic myeloid leukemia cells at the time of diagnosis or their appearance on tyrosine kinase inhibitor therapy predicts the imatinib treatment failure.

Currently used treatment of CML dramatically improved the prognosis of disease. However, additional chromosome aberrations (ACA/Ph+) are still one of the adverse prognostic factors. OBJECTIVES: evaluation of the impact of ACA/Ph+ appearance during disease outcome on the response to treatment. THE STUDY GROUP: consisted of 203 patients. The median time of follow-up was 72 months. ACA/Ph+ was found in 53 patients. RESULTS: patients were divided into four groups: standard risk, intermediate, high and very high risk. When ACA/Ph+ presence was documented at diagnosis time the optimal response was observed in 41.2%, 25%, and 0% of pts with intermediate, high and very high risk, respectively. If ACA/Ph+ were detected during imatinib treatment the optimal response was in 4.8% of patients. The risk of blastic transformation for patients with standard risk, intermediate, high and very high risk was 2.7%, 18.4%, 20% and 50%, respectively. CONCLUSIONS: the presence of ACA/Ph+ at diagnosis time or their appearance on therapy seems to be clinically relevant not only in terms of the risk of blastic transformation but also in terms of the treatment failure. Gathering patients with various karyotypes and their responses to treatment would allow to set better guidelines and predictions.

Clinical outcomes of therapy-related acute myeloid leukemia: an over 20-year single-center retrospective analysis.

INTRODUCTION: Therapy­related acute myeloid leukemia (t­AML), a life­threatening complication of cytotoxic therapy, represents an emerging challenge of modern oncology. OBJECTIVES: We aimed to evaluate clinical outcomes of patients with t­AML, taking into consideration genetic changes and treatment intensity. PATIENTS AND METHODS: We conducted a retrospective analysis of all consecutive AML patients from a single hematology center (hospitalized between 2000 and 2021). The diagnosis of t­AML was established according to the 2016 World Health Organization criteria. Overall survival (OS) and progression­free survival (PFS) were used to evaluate treatment outcomes. Retrospective identification of 17p13 deletion and TP53 mutation was conducted. RESULTS: Among 743 patients with AML, 60 (8.1%) were diagnosed with t­AML (63.4% had previous solid tumors). A complex karyotype (CK) and 17p13 deletion were detected in 26.8% and 26.7% of the t­AML cases, respectively, while FLT3­ITD and TP53 mutations occurred in 15.4% and 12.5% of the patients with t­AML, respectively. Median OS and PFS were 13 and 8 months, respectively. The survival outcomes were superior in the patients who underwent an allogenic hematopoietic cell transplantation (alloHCT) than in those treated with intensive chemotherapy alone (median OS, 47 vs 7 months, respectively; P = 0.01). Patients with therapy­related acute promyelocytic leukemia did not reach the median OS, and worse survival was noted in CK than non­CK t­AML (median OS, 6 vs 24 months; P = 0.02). In intensively treated t­AML, the survival was better for the patients younger than 64 years (P = 0.03). In the multivariable Cox proportional hazards regression model, alloHCT was associated with longer OS (hazard ratio, 0.19; 95% CI, 0.04-0.91; P = 0.04). Moreover, we noted a high frequency of treatment­related complications of t­AML. CONCLUSIONS: Our study revealed that prognosis of t­AML varies. Hence, the treatment strategy should include performing alloHCT as soon as possible in the patients with an adverse genetic profile.

Comparative proteomic profiling of sera from patients with refractory multiple myeloma reveals potential biomarkers predicting response to bortezomib-based therapy.

INTRODUCTION: In the era of implementing novel agents in multiple myeloma (MM) regimens, drug resistance has become a key factor undermining the results of treatment. Identifying biomarkers allows the prediction of therapy outcomes with specific agents and may lead to the avoidance of resistance. OBJECTIVES: This study aimed to identify biomarkers in the pretreatment sera of patients with refractory/ relapsed MM that differ from those in the sera of patients who achieved a better depth of response with bortezomib-containing therapy. PATIENTS AND METHODS: Pretreatment serum samples were obtained from 61 proteasome inhibitor-naive, transplant-eligible patients who were eligible for salvage PAD (bortezomib, doxorubicin, and dexamethasone) or VTD (bortezomib, thalidomide, and dexamethasone) chemotherapy. Based on their response to therapy, patients were classified into 3 groups: complete or very good partial response, partial response, and progressive or stable disease. A comparative proteomic analysis of the groups was performed. RESULTS: The analyzed groups significantly differed in terms of both overall survival and progression­free survival. In total, 632 proteins were identified. The proteomic signature revealed 54 proteins that differentiated each analyzed experimental group. Functional analysis revealed that the main identified pathways (17 proteins) involved the regulation of hydrolase activity and cellular response to stimuli. The identified proteins included apolipoprotein C1, complement components, and sulfhydryl oxidase 1. CONCLUSIONS: Our results demonstrated that the label-free proteomic analysis is a useful method for describing proteins differentially expressed in the sera of patients with MM. Further studies are needed to analyze the use of identified proteins as biomarkers.

Glutaminase inhibitor CB-839 synergizes with carfilzomib in resistant multiple myeloma cells.

Curative responses in the treatment of multiple myeloma (MM) are limited by the emergence of therapeutic resistance. To address this problem, we set out to identify druggable mechanisms that convey resistance to proteasome inhibitors (PIs; e.g., bortezomib), which are cornerstone agents in the treatment of MM. In isogenic pairs of PI sensitive and resistant cells, we observed stark differences in cellular bioenergetics between the divergent phenotypes. PI resistant cells exhibited increased mitochondrial respiration driven by glutamine as the principle fuel source. To target glutamine-induced respiration in PI resistant cells, we utilized the glutaminase-1 inhibitor, CB-839. CB-839 inhibited mitochondrial respiration and was more cytotoxic in PI resistant cells as a single agent. Furthermore, we found that CB-839 synergistically enhanced the activity of multiple PIs with the most dramatic synergy being observed with carfilzomib (Crflz), which was confirmed in a panel of genetically diverse PI sensitive and resistant MM cells. Mechanistically, CB-839 enhanced Crflz-induced ER stress and apoptosis, characterized by a robust induction of ATF4 and CHOP and the activation of caspases. Our findings suggest that the acquisition of PI resistance involves adaptations in cellular bioenergetics, supporting the combination of CB-839 with Crflz for the treatment of refractory MM.

Concomitance of monosomal karyotype with at least 5 chromosomal abnormalities is associated with dismal treatment outcome of AML patients with complex karyotype - retrospective analysis of Polish Adult Leukemia Group (PALG).

Monosomal karyotype (MK) and complex karyotype (CK) are poor prognostic factors in acute myeloid leukemia (AML). A comprehensive analysis of cytogenetic and clinical factors influencing an outcome of AML-CK+ was performed. The impact of cladribine containing induction on treatment results was also evaluated. We analyzed 125 patients with AML-CK+ treated within PALG protocols. MK was found in 75 (60%) individuals. The overall complete remission (CR) rate of 66 intensively treated patients was 62% vs. 28% in CK+ MK- and CK+ MK+ group (p = .01). No difference in CR rate was observed between DA and DAC arms. The overall survival (OS) in intensively treated patients was negatively influenced by MK, karyotype complexity (≥5 abnormalities), and WBC >20 G/L in multivariate analysis. The addition of cladribine to DA regimen improved OS only in MK- but not in MK+ group. In conclusion, concomitance of MK with ≥5 chromosomal abnormalities is associated with dismal treatment outcome in AMK-CK+.

Comparative proteomic profiling of refractory/relapsed multiple myeloma reveals biomarkers involved in resistance to bortezomib-based therapy.

Identifying biomarkers of the resistance in multiple myeloma (MM) is a key research challenge. We aimed to identify proteins that differentiate plasma cells in patients with refractory/relapsed MM (RRMM) who achieved at least very good partial response (VGPR) and in those with reduced response to PAD chemotherapy (bortezomib, doxorubicin and dexamethasone). Comparative proteomic analysis was conducted on pretreatment plasma cells from 77 proteasome inhibitor naïve patients treated subsequently with PAD due to RRMM. To increase data confidence we used two independent proteomic platforms: isobaric Tags for Relative and Absolute Quantitation (iTRAQ) and label free (LF). Proteins were considered as differentially expressed when their accumulation between groups differed by at least 50% in iTRAQ and LF. The proteomic signature revealed 118 proteins (35 up-regulated and 83 down-regulated in ≥ VGPR group). Proteins were classified into four classes: (1) involved in proteasome function; (2) involved in the response to oxidative stress; (3) related to defense response; and (4) regulating the apoptotic process. We confirmed the differential expression of proteasome activator complex subunit 1 (PSME1) by enzyme-linked immunosorbent assay. Increased expression of proteasomes and proteins involved in protection from oxidative stress (eg., TXN, TXNDC5) plays a major role in bortezomib resistance.

The prognostic significance of surface dipeptidylpeptidase IV (CD26) expression in B-cell chronic lymphocytic leukemia.

A number of factors related to B-cell chronic lymphocytic leukemia (B-CLL) patients' prognosis have been identified. However, still some factors better reflecting disease activity in individual cases are explored. The study aimed to evaluate prognostic significance of dipeptidylpeptidase IV/CD26 expression on B-CLL cells and its relationship with other well established prognostic factors. The study included 94 patients with newly diagnosed B-CLL and involved analysis of clinical, laboratory, flow-cytometry and cytogenetic data. Detailed analysis showed that CD26 expression on B-CLL cells correlates with Rai's clinical stage of the disease at diagnosis (p=0.034), ß2-microglobulin concentration (p=0.012), lactic acid dehydrogenase activity (p=0.045) and absolute lymphocytes' count (p=0.027) in the blood. The multivariate analysis revealed that time to treatment (TTT) was significantly influenced by Rai clinical stage, LDH activity in blood and CD26 expression on B-CLL cell's. Moreover, in the multivariate analysis restricted to the group of patients with documented cytogenetic risk (n=36) CD26 expression, Rai clinical stage and cytogenetic profile remained their independent impact on TTT. The results of our study indicate that the CD26 expression should be incorporated in B-CLL patients risk assessment along with well known prognostic factors, since it seems to have a relationship with the tumor mass and influences TTT.

Megakaryocytic blast crisis in a chronic myeloid leukemia patient with a rare variant of Philadelphia rearrangement t(9;22;22) and a constitutional translocation t(3;7).

Megakaryocytic blast crisis occurs extremely rarely, accounting for <3% of cases of chronic myelogenous leukemia in blastic transformation. In chronic myeloid leukemia, a variant Philadelphia translocation is reported in 2-10% of cases. We report an unusual case of megakaryocytic blast crisis with the Philadelphia variant rearrangement t(9;22;22) and a constitutional translocation t(3;7). The breakpoint in the 22q13 region was involved in this translocation. The chromosome region 22q13 harbors MKL1 gene, which is engaged in a specific translocation associated with acute megakaryoblastic leukemia. Study of deregulation of these four genes could contribute to better understanding of the effects of the t(9;22;22) rearrangement in a megakaryocytic blast crisis.