RUNX1 and RUNX3 Genes Expression Level in Adult Acute Lymphoblastic Leukemia-A Case Control Study.

The genetic factors of adult acute lymphoblastic leukemia (ALL) development are only partially understood. The Runt-Related Transcription Factor (RUNX) gene family play a crucial role in hematological malignancies, serving both a tumor suppressor and promoter function. The aim of this study was the assessment of relative RUNX1 and RUNX3 genes expression level among adult ALL cases and a geographically and ethnically matched control group. The relative RUNX1 and RUNX3 genes expression level was assessed by qPCR. The investigated group comprised 60 adult patients newly diagnosed with ALL. The obtained results were compared with a group of 40 healthy individuals, as well as clinical and hematological parameters of patients, and submitted for statistical analysis. ALL patients tend to have significantly higher RUNX1 gene expression level compared with controls. This observation is also true for risk group stratification where high-risk (HR) patients presented higher levels of RUNX1. A higher RUNX1 transcript level correlates with greater leukocytosis while RUNX3 expression is reduced in Philadelphia chromosome bearers. The conducted study sustains the hypothesis that both a reduction and increase in the transcript level of RUNX family genes may be involved in leukemia pathogenesis, although their interaction is complex. In this context, overexpression of the RUNX1 gene in adult ALL cases in particular seems interesting. Obtained results should be interpreted with caution. Further analysis in this research field is needed.

Clonal evolution in CLL patients as detected by FISH versus chromosome banding analysis, and its clinical significance.

The acquisition of new aberrations during the course of chronic lymphocytic leukemia (CLL) named clonal evolution (CE) is usually detected by one of the two methods: chromosome banding analysis (CBA) and interphase fluorescence in situ hybridization (I-FISH). The purpose of this study was to compare the usefulness of FISH and CBA for detecting CE and to evaluate its influence on clinical outcome. FISH and CBA were performed at two time points: baseline and follow-up. Thirty-eight previously untreated patients with CLL were included in this study. CBA and I-FISH revealed CE in 15 (39.5%) and 10 (26.3%) patients, respectively. High-risk CE was detected in six cases by CBA and in five cases by I-FISH. In four cases with CE-dependent 17p abnormalities detected by CBA, metaphase FISH was needed for the confirmation of 17p13.1 deletion. Time from first-line to second-line treatment (TTST) and overall survival (OS) did not differ between patients with and without CE, irrespective of the CE-detecting method used. However, shorter OS (P = 0.043) and TTST (P = 0.006) were observed for the patients with potentially relevant CE (rCE) detected by CBA, in which acquired aberrations were present in at least 20% of undivided cells and/or changed baseline karyotype to abnormal or complex and were not resulting from 13q deletion. Our results suggest that some, but not all, CE-dependent aberrations detected by CBA influence clinical outcome. Moreover, I-FISH, which was aimed at detecting aberrations of prognostic significance, was found to be more precise than CBA in their detection, especially TP53 deletion.

Cytotoxic and apoptosis-inducing effects of bendamustine used alone and in combination with rituximab on chronic lymphocytic leukemia cells in vitro.

AIM: The aim of our study was to compare the cytotoxic effects of bendamustine (BENDA) and rituximab (RIT) used either alone or in combination and to evaluate the influence of the above mentioned drugs on apoptosis measured as changes in mitochondrial transmembrane potential (Δψm), expression of caspases and selected apoptosis-regulating proteins in freshly isolated peripheral blood mononuclear cells of chronic lymphocytic leukemia (CLL) patients. MATERIALS/METHODS: Cytotoxic effect of tested drugs, as well as induction of apoptosis, drop in Δψm and expression of selected proteins involved in regulation of apoptosis were assessed in 48 hour cultures containing autologous serum (AS) using flow cytometry. BENDA was used at the concentration of 40 µg/ml and RIT at the concentration of 10 µg/ml. Control cultures were incubated without drugs. RESULTS: BENDA used either alone or in combination with RIT strongly induced apoptosis as well as enhanced expression of selected apoptotic proteins, especially those involved in the intrinsic apoptotic pathway: P53, PUMA and BAX, which cause mitochondrial transmembrane potential changes leading to activation of caspase-9 and -3. CONCLUSIONS: Our results indicate that both BENDA and RIT participate in the induction of apoptosis of CLL lymphocytes in vitro in the presence of AS in the culture medium. The drug-induced apoptosis occurs mainly via intrinsic pathway and activation of P53 and PUMA proteins, however the extrinsic pathway is likely to be involved as well. We also found that the combination of these drugs induces the expression of P53, caspase-8 and -9 more potently than either of them used separately.

Type of serum influences the rituximab dependent cytotoxicity and apoptosis of chronic lymphocytic leukemia cells in vitro.

PURPOSE: The aim of the study was to compare the influence of types of serum on the in vitro viability and on either spontaneous or rituximab (RIT)-induced apoptosis of chronic lymphocytic leukemia (CLL) cells. METHODS: The influence of fetal calf serum (FCS), patients' autologous serum (AS) and human AB-serum (ABS), used alone and in combinations consisting of two of them (v/v-1:1), on RIT-dependent cytotoxicity, apoptosis, detection of active forms of caspases-3,-9,-8 and disruption of mitochondrial membrane potential (ΔΨm) were assessed by flow cytometry. RIT was used at the concentration of 10 µg/ml. The spontaneous apoptosis was assessed in culture without RIT. RESULTS: AS revealed the protective action on CLL cells, however this serum added in vitro to the culture either alone or in combination with FCS was the only one to allow RIT to exert its cytotoxic action against CLL cells. RIT-induced apoptosis involved changes in ΔΨm and activation of caspases-3,-8,-9 when AS+FCS was applicated. Drug induced apoptosis (DIA) was 6.02 and 0.34, when FCS+AS and FCS alone were used, respectively (p<0.01). The RIT-dependent cytotoxic effect decreased when FCS+AS or FCS+ABS were used, as compared to effect of AS used separately. The cytotoxic effect of RIT did not depend on drug concentration, but on the type of serum added to the culture. CONCLUSIONS: The strongest cytotoxic effect of RIT in the presence of AS suggests that this drug activity towards CLL cells is enhanced by known cytotoxic mechanisms, caspase-dependent apoptotic pathway and possible influence of other extracellular factors present in the patients' sera.

The influence of venetoclax, used alone or in combination with cladribine (2-CdA), on CLL cells apoptosis in vitro: Preliminary results.

BACKGROUND: Venetoclax (VEN), a highly selective BCL-2 inhibitor, is successfully used in the treatment of chronic lymphocytic leukemia (CLL). The purine analogue - cladribine (2-CdA) - is also administered to CLL patients, especially as a part of chemoimmunotherapy. OBJECTIVES: To compare the effects of the VEN+2-CdA regimen with that of the 2 drugs used alone on the apoptosis of CLL lymphocytes in vitro. MATERIAL AND METHODS: Mononuclear cells were collected from 103 previously untreated CLL patients. They were incubated with VEN (40 nM) or/and 2-CdA (16 µM) for 48 h. Cytotoxicity, overall apoptosis, mitochondrial transmembrane potential changes (ΔΨm), and expression of selected apoptosis-involved proteins were measured. RESULTS: The cytotoxicity, overall apoptosis, caspase-3 or caspase-9 expression, and ΔΨm were significantly higher after VEN+2-CdA addition compared to both drugs used alone, with a very strong synergistic effect observed. The percentage of BCL-2-positive cells decreased after VEN and VEN+2-CdA addition compared to controls. The TP53-expressing cells increased under the influence of all tested regimens. The VEN+2-CdA increased the expression of BIM, BAX and NOXA compared to either controls or VEN or 2-CdA alone. Similar increases in PUMA expression were observed after VEN, 2-CdA and VEN+2-CdA addition. The FAS-associated death-domain protein (FADD) expression was significantly higher after 2-CdA and 2-CdA+VEN addition as compared to control. CONCLUSIONS: Our results confirm the involvement of both VEN and 2-CdA in the intrinsic apoptotic pathway. They also demonstrate that these agents have a synergistic effect on CLL cells in vitro. Further studies are needed to assess the influence of VEN+2-CdA on the expression of apoptosis-involved genes.

The safety of available chemo-free treatments for mantle cell lymphoma.

INTRODUCTION: Conventional treatment for mantle cell lymphoma (MCL) patients includes regimens combining rituximab with other cytotoxic drugs, followed or not by consolidation with autologous stem cell transplantation and rituximab maintenance. However, older, unfit, and relapsed/refractory patients are often ineligible for intense treatment. Currently, available new targeted treatment options seem to offer hope in this group of patients. AREAS COVERED: This article reviews the safety profiles of new therapeutic chemotherapy-free options for MCL patients. Publications in English from 2010 through June 2020 were surveyed on the MEDLINE database for articles. Proceedings of the American Society of Hematology during the last 5 years were also included. EXPERT OPINION: MCL is a clinically heterogenous disease predominantly affecting elderly patients. Its variable clinical course requires personalization and individualization of treatment to achieve optimal survival and acceptable safety profiles, especially in poor prognosis patients. Results of clinical trials performed in the past decade indicated that novel drugs used as a single agent or as part of a conventional chemotherapeutic treatment offer promise in minimalizing the relapse rate for MCL and may allow more effective and safer treatment options by reducing the risk of adverse events, especially cytopenias and infections.

Venetoclax in the treatment of chronic lymphocytic leukemia.

INTRODUCTION: Venetoclax, an antagonist of BCL-2 protein plays an important role in the pathogenesis of chronic lymphocytic leukemia (CLL). It has been approved by the FDA for the treatment of relapsed/refractory CLL with del17p, and by the EMA for patients with del17p/TP53 mutation who have failed a BCR inhibitor, or in patients without those aberrations who have failed previous therapy, regardless of their genetic/molecular profile. Venetoclax in combination with rituximab has been also approved for the treatment of CLL after at least 1 prior therapy, regardless of del17p. Areas covered: This article reviews the chemical structure, mechanisms of action, pharmacokinetic, and the clinical applications of venetoclax in monotherapy and in combined treatment of CLL. Publications dated 2010 through March 2019 were obtained from the MEDLINE database. The proceedings of the American Society of Hematology held during the last five years were also included. Expert opinion: Venetoclax shows high efficacy, a favorable toxicity profile, and a high rate of minimal residual disease negativity, which is thought to have an impact on overall survival. It is efficient in patients with del17p/TP53 mutations, the incidence of which increases during clonal CLL evolution, and after the failure of BCR pathway inhibitors.

Bendamustine alone or with rituximab modifies expression of apoptosis-regulating genes and proteins of CLL cells, depending on IGVH mutational status.

We studied whether bendamustine (BENDA) alone or with rituximab (RIT) modifies in vitro expression of apoptosis-involved genes and proteins of chronic lymphocytic leukemia (CLL) cells depending on IGVH mutational status. Circulating lymphocytes from 34 untreated patients (18 IGVH-MUT and 16 IGVH-UNMUT) were incubated with above drugs to evaluate proteins expression. Microarray analysis of 93 genes was performed in 14 patients. BENDA and BENDA + RIT increased expression of BAX and BBC3 in IGVH-MUT and IGVH-UNMUT groups, and significant differences in expression of above genes after BENDA + RIT were observed between both groups. Additionally, BENDA + RIT decreased NFκB and BCL-2 genes in IGVH-UNMUT patients and increased expression of P53, BAX and PUMA proteins in IGVH-MUT and UNMUT subjects. However, no significant differences were found between these groups. In conclusion, BENDA + RIT modified gene expression profile in CLL cells and affected expression of some apoptosis-regulating proteins in vitro. Expression of BAX and BBC3 depends on action of drugs and IGVH mutational status.

The safety profile of monoclonal antibodies for chronic lymphocytic leukemia.

INTRODUCTION: Monoclonal antibodies (MoAbs), non-chemotherapeutic agents targeting the antigens present on chronic lymphocytic leukemia (CLL) lymphocytes, are being implemented increasingly more often as treatment options. Areas covered: This article reviews the similarities and differences in the structure, mechanism of action, efficacy and safety profile of commercially-available MoAbs and prevents new agents potentially useful for CLL treatment. Publications in English before June 2016 were surveyed on the MEDLINE database for articles. Proceedings of the American Society of Hematology held during the last five years were also included. Expert opinion: MoAbs, especially those targeting CD20, are highly effective biological options for first-line and salvage treatment of CLL, particularly in chemoimmunotherapy, and possibly also as maintenance therapy. Treatment with MoAbs is associated with reduced risk of such adverse events as cytopenias, infections and secondary neoplasias and is generally well tolerated. Depending on antibody type, the most common adverse events are usually transient and limited to grade 1 and 2 infusion-related reactions. In addition to commercially available MoAbs, several other antibodies exist which are targeted against different antigens studied in the clinical trials.

Ofatumumab for treating chronic lymphocytic leukemia: a safety profile.

INTRODUCTION: Ofatumumab, the first fully human IgG1κ, belongs to the second generation of the first class of anti-CD20 monoclonal antibodies. The drug used alone and in combination with drugs having different mechanisms of action has shown a favorable toxicity profile and significant benefit especially in relapsed/refractory chronic lymphocytic leukemia (CLL) patients in doses up to 2000 mg. AREAS COVERED: This article reviews pharmacokinetic, clinical application for CLL treatment, and safety profile of ofatumumab as well as differences and similarity between ofatumumab and rituximab. Publications in English from 2010 through October 2015 were surveyed on the MEDLINE database for articles. Proceedings of the American Society of Hematology held during the last 5 years were also included. EXPERT OPINION: Ofatumumab more effectively than rituximab enhanced complement-dependent cytotoxicity playing the crucial role for its therapeutic activity. The drug is highly effective in the first-line and salvage treatment of CLL, essentially as a part of immunochemotherapy, and probably also as maintenance therapy. Its safety profile is very advantageous, since adverse events are usually limited to grade 1 and 2 infusion-related reactions, which tend to decrease throughout the treatment. Its advantage over the other anti-CD20 monoclonal antibodies in the treatment of CLL remains to be determined in the direct head-to-head trials.

Pharmacodynamic considerations of small molecule targeted therapy for treating B-cell malignancies in the elderly.

INTRODUCTION: Small molecule inhibitors are currently in various stages of preclinical and clinical trials and are expected to revolutionize the treatment of many neoplastic diseases, including B-cell lymphoid malignancies. AREAS COVERED: This article reviews the chemical structure, mechanisms of action, pharmacodynamic and pharmacokinetic properties, as well as clinical applications of small molecules in the treatment of elderly patients with B-cell hematological malignancies. Bibliographic research covering mainly the period from 2010 until February 2015 was conducted on the MEDLINE database for articles in English. Proceedings of the American Society of Hematology, European Hematology Association and American Society of Clinical Oncology conferences held during the last 5 years were also included. EXPERT OPINION: In the last few years, several preclinical and clinical trials have evaluated many small weight organic molecules which downregulate B-cell receptor (BCR) signaling and act via inhibition of either BCR-associated kinases or cyclin-dependent kinases, or which are antagonists of members of the B-cell lymphoma 2 protein family. Pharmacokinetic profiles of these agents as well as dosage used and adverse events in patients with lymphoid malignancies have been established. Some of these inhibitors satisfy therapeutic modalities as suitable for the elderly patients, including those with chronic lymphocytic leukemia and non-Hodgkin's lymphoma.

New insights into biology, prognostic factors, and current therapeutic strategies in chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is characterized by the clonal proliferation and accumulation of mature B lymphocytes. CLL cells show an antiapoptotic profile, suggesting the important role of apoptosis inhibition in the disease development. However, there is some population of proliferating CLL cells, which may also play a role in progression of the disease. There are several newer, biological prognostic factors in CLL. Currently, cytogenetic abnormalities with different prognostic values seem to be the most biologically relevant. During the last decades, the treatment of CLL has been significantly changed. Different strategies such as monotherapy with chlorambucil and purine nucleoside analogues (PNA) used alone or in combination with cyclophosphamide have been introduced. Most recently, immunochemotherapy with anti-CD20 monoclonal antibody, rituximab, combined with fludarabine and cyclophosphamide, became a gold standard of first-line treatment in eligible CLL patients. Currently, new treatment strategies including new monoclonal antibodies, bendamustine, lenalidomide, or inhibitors of several cell signaling pathways are under clinical studies in resistant/relapsed CLL patients. Moreover, allogeneic stem cell transplantation has to be considered, especially in younger high risk patients, for example, those who are resistant to PNA or those with 17p deletion. In this paper, we present the most important recent advances in CLL biology and treatment.

Correlations between concentrations of interleukin (IL)-17A, IL-17B and IL-17F, and endothelial cells and proangiogenic cytokines in systemic lupus erythematosus patients.

Systemic lupus erythematosus (SLE) is an autoimmune disease of multifactorial pathoaetiology. Different organs and blood vessels may be affected by chronic inflammation. A direct cause of the disease has not yet been found, so research is being carried out to this effect. The role of the recently identified helper T lymphocyte CD4+, described as Th17, and its dependent cytokines have been of particular interest. The aim of the study was to evaluate IL-17A, IL-17B, IL-17F and IL-23 in 60 SLE patients and 26 age-matched, healthy volunteers and also to investigate the correlation between levels of the investigated cytokines and VEGF, PIGF, as well as number of endothelial cells. IL-17A, IL-17B, IL-17BR and IL-17F levels were found to be higher in SLE patients than in the control group. However, only IL-17F levels showed a statistically significant correlation with the number of endothelial cells (aCEC) and disease activity. Correlations between levels of IL-17F and VEGF and PIGF as well as VEGF and IL-17A and IL-23 were statistically significant. Increased levels of the selected cytokines from the IL-17 family in SLE patients suggest a role for them not only in the inflammatory process but also in angiogenesis. This also highlights the role of IL-17F in activating vascular endothelial cells and consequently blood vessel formation, and in the relationship between the inflammatory reaction and angiogenesis in the development of SLE.

Pharmacokinetic evaluation and therapeutic activity of bendamustine in B-cell lymphoid malignancies.

INTRODUCTION: Bendamustine , a cytotoxic agent comprising structural features of both an alkylating drug and a purine nucleoside analog, was approved by the US FDA for treatment of chronic lymphocytic leukemia (CLL) and indolent B-cell non-Hodgkin's lymphoma (NHL). Additionally, in Europe the drug has been also approved for treatment of multiple myeloma (MM) and in Asia, especially in Japan for refractory/relapsed NHL and mantle cell lymphoma. AREAS COVERED: The authors present the chemical structure, mechanism of action, pharmacokinetic properties and clinical application of bendamustine in hematological malignancies. Publications in English related to the above, up to June 2012, have been surveyed in the MEDLINE database. Conference proceedings reports from the last 5 years are also included. Additional relevant publications have been obtained by reviewing the references from the chosen articles. EXPERT OPINION: The availability of bendamustine provides an effective treatment option for patients with lymphoid malignancies. Several recent clinical trials have documented the activity of bendamustine in CLL, NHL and MM, both as a single agent and in combination with other cytotoxic drugs. However, doses, schedules and also the role of bendamustine in treatment of patients with hematological malignancies should be further investigated to establish its real place in the management of these diseases.

Apoptotic gene expression under influence of fludarabine and cladribine in chronic lymphocytic leukemia-microarray study.

BACKGROUND: A deep insight into gene expression profiling (GEP) is a key to understanding the background of disease. It can lead to identification of diagnostic and prognostic factors and then to a selection of the most appropriate therapy. The aim of this study was to evaluate differences in apoptotic gene expression in chronic lymphocytic leukemia (CLL) cells influenced by fludarabine (FA) or cladribine (2-CdA). METHODS: GEP was performed in cells obtained from 10 untreated CLL patients and cultured in vitro with FA or 2-CdA. Ninety-three selected apoptotic genes were analyzed using 384 TaqMan® Low Density Arrays in pooled RNA. RESULTS: Relevant results were found in a set of 27 genes, however, the most striking differences between FA and 2-CdA were observed in the following 5 genes: BAD, TNFRSF21, DAPK1, CARD 6 and CARD 9. CONCLUSION: We have found some differences in apoptotic gene expression between FAand 2-CdA. These findings give prominence to genes qualifying for further studies currently conducted in our Department.

Changes in the apoptotic gene expression profile in CLL patients treated with rituximab combined with cladribine and cyclophosphamide-preliminary results.

The study was aimed to investigate modifications of apoptotic gene expression profile by microarray technique in 10 patients with chronic lymphocytic leukemia by treatment with rituximab, cladribine and cyclophosphamide (RCC) according to IGHV mutational status. The TaqMan Low Density Array for 96 gene transcripts was used. Those modifications followed two distinctive patterns largely overlapping the IGHV mutational status. In the IGHV-mutated group, the expression of many proapoptotic genes increased after treatment as compared to initial value. Our results suggest that RCC drugs may act through influence on the expression of some apoptosis-involved genes dependently on the IGVH mutational status.

Chromosomal aberrations in chronic lymphocytic leukemia detected by conventional cytogenetics with DSP30 as a single agent: comparison with FISH.

The aim of our study was to estimate the usefulness for conventional cytogenetics (CC) of DSP30 as a single agent (CC-DSP30) for detecting the most important chromosomal aberrations revealed in CLL by FISH and to find other abnormalities possibly existing but undetected by FISH with standard probes. Using CC-DSP30, the metaphases suitable for analysis were obtained in 90% of patients. CC-DSP30 and FISH were similarly efficacious for detecting del(11)(q22) and trisomy 12, whereas FISH was more sensitive for del(13)(q14). Sole del(13)(q14) detected by FISH, in 50% of patients was associated with other aberrations revealed by CC-DSP30. Additionally, the most recurrent anomaly detected by CC-DSP30 were structural aberrations of chromosome 2.

The effects of intracoronary delivery of mononuclear bone marrow cells in patients with myocardial infarction: a two year follow-up results.

BACKGROUND: Transplantation of bone marrow stem cells (BMSC) is a new method of prevention of left ventricular (LV) remodelling in post-infarction patients. Studies published to date point to LV systolic and diastolic function improvement following this therapy however only a few studies assessed the long-term effects of BMSC. AIM: To assess the 2 year prognosis in patients with anterior myocardial infarction (MI) treated with BMSC transplantation in the acute phase. METHODS: The study group consisted of 60 patients with first anterior ST-segment elevation MI (STEMI), treated with primary percutaneous angioplasty, with baseline LV ejection fraction (LVEF) 〈 40%, who were randomly assigned to undergo BMSC transplantation on day 7 of the STEMI (40 patients, BMSC group) or to receive standard treatment (20 patients, control group). In all the patients echocardiography was performed at baseline and after 1, 3, 6, 12 and 24 months. The composite end-point (death, MI, admission for heart failure or repeat revascularisation) was assessed after 2 years of follow-up. RESULTS: Absolute increase of LVEF compared to baseline values was higher in the BMSC group than in the control group. The LVEF increase in BMSC group at 1 month was 7.1% (95% CI 3.1-11.1%), at 6 months - 9.3% (95% CI 5.3-13.3%), at 12 months - 11.0% (95% CI 6.2-13.3%) and at 24 months - 10% (95% CI 7.2-12.1%). In the control group, LVEF increase was 3.7% (95% CI 2.3-9.7%) at 1 month, 4.7% (95% CI 1.2-10.6%) at 6 months, 4.8% (95% CI 1.5-11.0%) at 12 months and 4.7% (95% CI 1.4-10.7%) at 24 months. The composite end-point occurred significantly more frequently in the control group (55%) than in the BMSC group (23%): OR 2.72; 95% CI 1.06-7.02, p = 0.015. CONCLUSIONS: Treatment with mononuclear bone marrow cells on day 7 of the first anterior MI in patients with significant baseline systolic dysfunction improves 2-year outcome.