Immuno-Stimulating Activity of 1,25-Dihydroxyvitamin D in Blood Cells from Five Healthy People and in Blasts from Five Patients with Leukemias and Pre-Leukemic States.

(1) Hematological malignancies are characterized by an immortalization, uncontrolled proliferation of blood cells and their differentiation block, followed by the loss of function. The primary goal in the treatment of leukemias is the elimination of rapidly proliferating leukemic cells (named blasts). However, chemotherapy, which removes proliferating blasts, also prevents the remaining immune cells from being activated. Acute leukemias affect elderly people, who are often not fit to survive aggressive chemotherapy. Therefore, there is a need of milder treatment, named differentiation therapy, which might simulate the immune system of the patient. 1,25-Dihydroxyvitamin D, or low-calcemic analogs of this compound, were proposed as supporting therapy in acute leukemias. (2) Bone marrow blasts from patients with hematological malignancies, and leukocytes from healthy volunteers were ex vivo exposed to 1,25-dihydroxyvitamin D, and then their genomes and transcriptomes were investigated. (3) Our analysis indicates that 1,25-dihydroxyvitamin D regulates in blood cells predominantly genes involved in immune response, such as CAMP (cathelicidin antimicrobial peptide), CP (ceruloplasmin), CXCL9 (C-X-C motif chemokine ligand 9), CD14 (CD14 molecule) or VMO1 (vitelline membrane outer layer 1 homolog). This concerns blood cells from healthy people, as well as blasts from patients with hematological malignancies. In addition, in one patient, 1,25-dihydroxyvitamin D significantly downregulated transcription of genes responsible for cell division and immortalization. (4) In conclusion, the data presented in this paper suggest that addition of 1,25-dihydroxyvitamin D to the currently available treatments would stimulate immune system, inhibit proliferation and reduce immortal potential of blasts.

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.

BTLA Expression in CLL: Epigenetic Regulation and Impact on CLL B Cell Proliferation and Ability to IL-4 Production.

In our previous study, while chronic lymphocytic leukemia (CLL) cases showed higher levels of B and T lymphocyte attenuator (BTLA) mRNA compared to controls, lower BTLA protein expression was observed in cases compared to controls. Hence we hypothesize that micro RNA (miR) 155-5p regulates BTLA expression in CLL. In line with earlier data, expression of BTLA mRNA and miR-155-5p is elevated in CLL (p = 0.034 and p = 0.0006, respectively) as well as in MEC-1 cell line (p = 0.009 and 0.016, respectively). Inhibition of miR-155-5p partially restored BTLA protein expression in CLL patients (p = 0.01) and in MEC-1 cell lines (p = 0.058). Additionally, we aimed to evaluate the significance of BTLA deficiency in CLL cells on proliferation and IL-4 production of B cells. We found that secretion of IL-4 is not dependent on BTLA expression, since fractions of BTLA positive and BTLA negative B cells expressing intracellular IL-4 were similar in CLL patients and controls. We demonstrated that in controls the fraction of proliferating cells is lower in BTLA positive than in BTLA negative B cells (p = 0.059), which was not observed in CLL. However, the frequency of BTLA positive Ki67+ B cells in CLL was higher compared to corresponding cells from controls (p = 0.055) while there were no differences between the examined groups regarding frequency of BTLA negative Ki67+ B cells. Our studies suggest that miR-155-5p is involved in BTLA deficiency, affecting proliferation of CLL B cells, which may be one of the mechanisms responsible for CLL pathogenesis.

NF-κB1 -94del/del ATTG polymorphic variant maintains CLL at an early, mildest stage.

BACKGROUND: NF-κB is an essential player in cancer biology, especially in tumor development, due to its constitutive activation, and because a four-base deletion (ATTG) in the NF-κB1 promoter region at site -94, alters mRNA stability and regulates translation efficiency. This polymorphism is a good candidate risk marker and modulator of clinical course in chronic lymphocytic leukemia (CLL). As the effect of this NF-κB1 gene polymorphism has not been studied in patients with CLL so far, the present study was undertaken to find out whether the NF-κB1 promoter -94 ins/del ATTG polymorphism might be an essential genetic risk factor and/or modulatory disease player associated with CLL. OBJECTIVES: The NF-κB1 -94 ins/del ATTG (rs28362491) polymorphism was investigated as a potential CLL susceptibility and progression factor, along with demonstration of potential modulation of the stage of clinical disease based on Rai classification. MATERIAL AND METHODS: The associations of NF-κB1 -94 ins/del ATTG polymorphism with CLL and its clinical manifestation in 282 Polish individuals, including 156 CLL patients, were analyzed using polymerase chain reaction (PCR) with primers including a labeled forward primer, followed by capillary electrophoresis. RESULTS: A higher occurrence of the del/del homozygosity was observed among patients when compared to controls, resulting in an increase in CLL risk of more than twofold in patients carrying this homozygous genotype (OR = 2.23, p = 0.02, 95% CI = 1.14-4.37). Moreover, the del/del-positive patients more frequently presented the less aggressive disease phenotype (Rai 0), suggesting a low probability of progression to more advanced disease. CONCLUSIONS: The NF-κB1 -94 del/del genetic variant, although associated with increased risk of CLL disease, may be associated with maintenance of disease severity in the early, mildest stage. The likelihood of disease progression may increase as the frequency of wild-type (insertion) alleles for this polymorphism increases.

Association of Common Variants of TNFSF13 and TNFRSF13B Genes with CLL Risk and Clinical Picture, as Well as Expression of Their Products-APRIL and TACI Molecules.

Interactions between APRIL (TNFSF13) and its receptor TACI (TNFRSF13B) are implicated in providing survival benefits for chronic lymphocytic leukaemia (CLL) cells. Here we explored the relationship between TNFSF13 and TNFRSF13B SNPs and expression of APRIL and TACI molecules and performed extended case-control study to evaluate earlier observations. Expression of APRIL and TACI was detected by FACS for 72 and 145 patients, respectively, and soluble APRIL was measured by ELISA in plasma of 122 patients. Genotypes were determined in 439 CLL patients and 477 control subjects with TaqMan Assays or restriction fragment length polymorphism (RFLP). The rs4968210GG genotype of TNFSF13 was associated with a lower percentage of CD19+APRIL+ cells in CLL patients when compared to (AA + GA) genotypes (p-value = 0.027). Homozygosity at rs11078355 TNFRSF13B was associated with higher CD19+ TACI+ cell percentage in CLL patients (p-value = 0.036). The analysis of extended groups of patients and healthy controls confirmed the association of TNFSF13 rs3803800AA genotype with a higher CLL risk (OR = 2.13; CI95% = 1.21; 3.75; p-value = 0.007), while the possession of TNFRSF13B rs4985726G allele (CG + GG) genotype was associated with lower risk of CLL (OR = 0.69; CI95% = 0.51; 0.95; p-value = 0.02). Genetic variants of TNFSF13 and TNFRSF13B may have an impact on APRIL and TACI expression and may be considered as possible CLL risk factors.

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.

Analysis of the rRNA methylation complex components in pediatric B-cell precursor acute lymphoblastic leukemia: A pilot study.

BACKGROUND: Dysregulation of ribosome biogenesis and alteration of ribosome composition, including alteration in ribosomal RNA (rRNA) 2'-O-ribose methylation, can play a role in malignant transformation and cancer progression. Several studies recently reported that components of the rRNA methylation complex are associated with leukemogenesis. However, no study ever investigated the alteration of ribosome biogenesis factors in the most common pediatric malignancy - B-cell precursor acute lymphoblastic leukemia (BCP-ALL). OBJECTIVES: The objective of this study was to examine the expression of factors building the rRNA methylation complex, either the protein components (1 methyl-transferase (FBL), NOP56, NOP58, NHP2L1) or some RNA components (box C/D snoRNAs: SNORD35B, SNORD65, SNORD46, SNORD50A, SNORD38B), as well as CMYC, and nucleolin (NCL) - a protein involved in rRNA synthesis. Clinical effects in children with BCP-ALL were also investigated. MATERIAL AND METHODS: The factors involved in ribosome biogenesis were studied in 28 children with BCP-ALL with the use of real-time polymerase chain reaction (RT-PCR) using the BioMark HD System (Fluidigm, San Francisco, USA) in cDNA prepared from the bone marrow samples collected at diagnosis. RESULTS: Strong correlations were observed between NOP56, NOP58 and NCL, and multiple weaker correlations were observed in the box C/D snoRNA category, and between box C/D snoRNA and transcripts coding proteins of the rRNA methylation complex. The expression of analyzed transcripts did not correlate with the initial white blood cells count (WBC) or with bone marrow blast percentage. Ribosome biogenesis upregulation with overexpression of FBL and NOP56, and CMYC was found in patients who subsequently relapsed and the upregulation signature was not associated with known risk predictors. CONCLUSIONS: This is the first report on the clinical aspect of ribosome biogenesis in pediatric BCP-ALL, and it shows that overexpression of CMYC and C/D box nucleoproteins FBL and NOP56 is an antecedent event in patients who subsequently relapse. The dysregulation pattern is different from the previous reports in acute myeloid leukemia (AML), suggesting that dysregulation of ribosome biogenesis is specific for BCP-ALL.

Safety and Efficacy of Bendamustine Monotherapy in the First-Line Treatment of Patients with Chronic Lymphocytic Leukemia: Polish Lymphoma Research Group Real-Life Analysis.

Bendamustine is a cytostatic drug with a unique structure, combining the features of purine nucleoside analogs and alkylating agents. In patients with chronic lymphocytic leukemia (CLL) it is commonly used in combination with rituximab (BR protocol) both in the first-line as well as subsequent lines of therapy, and in clinical trials it is often combined with new targeted therapies. Therefore, the data on its real-life safety and efficacy are of clinical significance. As the Polish Lymphoma Research Group (PLRG), we retrospectively analyzed the efficacy and tolerability of bendamustine monotherapy in 96 patients with CLL. The median number of bendamustine cycles was 5, and 44 patients did not complete the planned 6 cycles (46%). Among the adverse events associated with the earlier termination of bendamustine treatment, infections were the most common (20.5%), followed by neutropenia (15.9%) and thrombocytopenia (15.9%). Dose reductions and/or delays occurred in 31% of treatment cycles (132 of 425) with neutropenia (17.9%) as the most frequent cause. Efficacy analysis showed an overall response rate of 88.2% with complete remission and partial remission achieved in 43.8 and 41.7% of patients, respectively. At the 24th month of follow-up, progression-free survival was 52% and overall survival was 69.7%. Bendamustine in monotherapy was found to be safe and efficacious, at least in terms of early response. Special attention should be paid to infectious complications, and especially that immune disorders are characteristic in the clinical course of CLL. Our observations suggest efforts must be made to ensure the proper timing and proper dose in the administration of the drug, and to avoid the premature termination of the treatment.

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.

Hodgkin lymphoma transformation of chronic lymphocytic leukemia-A real life data from the Polish Lymphoma Research Group.

Richter transformation (RT) of chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) to Hodgkin lymphoma (HL) is a rare and unexpected event in the course of the disease and data on this phenomenon is still limited. To better understand the clinical and histological characteristics and the outcomes of HL variant of RT (HvRS) the Polish Lymphoma Research Group performed a nationwide survey which identified 22 patients with histologically proven HvRS diagnosed between 2002 and 2016. There were 16 (73%) males. The median age at CLL/SLL and HvRS diagnosis was 59 (39-77) and 64 (40-77) years, respectively. The median interval between CLL/SLL and HvRS diagnosis was 38 months (range: 0-187). All patients had an advanced stage HL, and majority, 17 (77%), presented with B symptoms. The predominant subtypes of HL were nodular sclerosis (12; 55%) and mixed cellularity (9; 41%). Eighteen patients received non-palliative treatment, including 13 who received driamycin, bleomycin, vinblastine, and dacarbazine (ABVD) regimen first line. Objective response was: 50%, with 33% complete remissions (61% and 46% for ABVD, respectively). Median overall survival reached 13.3 months (95% CI, 3.7-NA). The only adverse prognostic factor for survival was a higher number (≤1 versus ≥2) of prior lines of treatment given for CLL/SLL with HR 3.57 (95% CI, 1.16-10.92). We conclude, HvRS harbors a poor prognosis, especially in patients heavily pretreated for CLL/SLL. Response to standard first-line anti-HL chemotherapy is unsatisfactory, and new agents should be tested to improve the outcome.

Expression of the PIM2 gene is associated with more aggressive clinical course in patients with chronic lymphocytic leukemia.

BACKGROUND: The PIM2 gene belongs to the PIM family, which encodes serine/threonine kinases involved in cell survival and apoptosis. The relation between the expression of the PIM2 gene and the course of chronic lymphocytic leukemia (CLL) has not been fully determined. OBJECTIVES: The aim of the study was to evaluate the role of the PIM2 gene as a marker of CLL malignancy and its importance as a predictive and prognostic factor. MATERIAL AND METHODS: Sixty-seven patients, 35 females and 32 males, aged 49-90 years, with de novo CLL, and 14 healthy individuals were enrolled in the study. Expression of the PIM2 gene was analyzed using TaqMan RQ-PCR assay and western blot test. RESULTS: Median PIM2 gene expression in CLL patients was higher than in controls. Patients with high expression of the PIM2 gene had shorter progression-free survival and time to first treatment than patients with low PIM2 expression. It was found that patients with CR had lower expression of the PIM2 gene than patients without complete remission (CR). Notably, associations between high PIM2 expression and rapid lymphocyte doubling time, the percentage of malignant lymphocytes with ZAP70 expression and the Rai stage were revealed. CONCLUSIONS: We found that the PIM2 gene is associated with a more aggressive clinical course of CLL.

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.

Elevated serum concentrations of metalloproteinases (MMP-2, MMP-9) and their inhibitors (TIMP-1, TIMP-2) in patients with Graves' orbitopathy.

BACKGROUND: Graves' orbitopathy (GO), also known as thyroid-associated ophthalmopathy, is characterized by dramatic tissue reactivity. Both inflammation and tissue remodeling characterize the clinical course of GO. Some data has been found regarding the association of MMPs and TIMPs in GO. MATERIAL AND METHODS: Serum concentrations of MMP-9, MMP-2, TIMP-1, and TIMP-2 were determined by ELISA method. OBJECTIVES: Forty-eight patients (34 females, 14 males, with median age 51.5 years) with GD and hyperthyroidism were enrolled in the study. In 28 patients, active, moderate-to-severe grade orbitopathy was diagnosed. The aim of this study was to assess the serum concentrations of MMP-2, MMP-9, TIMP-1, and TIMP-2 in patients with Graves' disease (GD), with and without GO, and their relationship with disease severity, as well as to evaluate how these concentrations change after successful treatment. RESULTS: Median serum concentrations of MMP-2 and MMP-9 were significantly higher in all patients with GD as well as in the subgroup with GO than in the control group. Median serum concentrations of TIMP-1 and TIMP-2 were significantly higher in all patients with GD than in controls. The same significant differences were observed in the subgroups with and without GO in comparison with controls. The GO subgroup showed a significant positive correlation between the MMP-9 concentration and the serum level of TSHRAb antibodies, and a clinical activity score ≥4 according to EUGOGO. CONCLUSIONS: In our study we found that only MMP-9 differentiates the patients with and without GO, and may be used as a marker of the disease severity in patients with this manifestation of GD.

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.

Association of variants in BAFF (rs9514828 and rs1041569) and BAFF-R (rs61756766) genes with the risk of chronic lymphocytic leukemia.

The B-cell activator factor (BAFF)/BAFF receptor (BAFF-R) axis seems to play an important role in the development and progression of chronic lymphocytic leukemia (CLL). Here, we investigated the association of eight single nucleotide polymorphisms (SNPs) in the BAFF (TNFSF13B) and BAFF-R (TNFRSF13C) genes with risk of sporadic CLL in a group of 439 CLL patients and 477 controls. We also examined the correlation between selected SNPs and CLL clinical parameters as well as BAFF plasma levels and intracellular BAFF expression. Our results point to a possible association between the rs9514828 (CT vs. CC + TT; OR = 0.74; CI 95 % = 0.57; 0.97; p = 0.022) and rs1041569 (AT vs. AA + TT; OR = 0.72; CI 95 % = 0.54; 0.95; p = 0.021) of BAFF gene and rs61756766 (CC vs. CT; OR = 2.03; CI 95 % = 1.03; 3.99; p = 0.03) of BAFF-R gene and CLL risk. Additionally, we observed that homozygotes rs1041569 AA and TT had a slightly higher risk (HR = 1.12) for the need of treatment in comparison to AT heterozygotes. In conclusion, our results indicate that SNPs in BAFF and BAFF-R genes may be considered as potential CLL risk factors.

Increased expression of metalloproteinase-2 and -9 (MMP-2, MMP-9), tissue inhibitor of metalloproteinase-1 and -2 (TIMP-1, TIMP-2), and EMMPRIN (CD147) in multiple myeloma.

INTRODUCTION: Activity of metalloproteinases (MMP) is controlled both by specific tissue inhibitors (TIMP) and activators (extracellular matrix metalloproteinase inducer, EMMPRIN). There are few data available concerning concentration the bone marrow of MMP-2, MMP-9, TIMP-1, and TIMP-2, or EMMPRIM expression by bone marrow mesenchymal stromal cells (BMSCs) in patients with multiple myeloma (MM). PATIENTS AND METHODS: We studied 40 newly diagnosed, untreated patients: 18 males and 22 females with de novo MM and 11 healthy controls. Bone marrow was collected prior to therapy. BMSCs were derived by culturing bone marrow cells on MesenCult. Protein concentrations were determined in bone marrow plasma and culture supernatants by ELISA. EMMPRIN expression by BMSCs was assessed by flow cytometry. RESULTS: The median concentrations of MMP-9, TIMP-1, and TIMP-2 in both marrow plasma and culture supernatants were significantly higher in MM patients than controls. CONCLUSION: EMMPRIN expression and ratios MMP-9/TIMP-1 and MMP-2/TIMP-2 were higher in MM patients, our results demonstrate that in MM patients MMP-2 and MMP-9 are secreted in higher amounts and are not balanced by inhibitors.

Patients with chronic lymphocytic leukaemia (CLL) differ in the pattern of CTLA-4 expression on CLL cells: the possible implications for immunotherapy with CTLA-4 blocking antibody.

Recently, systemic administration of a human monoclonal antibody directed against cytotoxic T lymphocyte-associated antigen 4 (CTLA-4) expressed on circulating T cells in patients with chronic lymphocytic leukaemia (CLL) has been considered. Also, CLL cells have been shown to express CTLA-4, increased levels of which in the leukaemic compartment are a predictor of good clinical outcome. Since both CLL and Treg microenvironment cells can be targeted by the CTLA-4 blocking antibody in this immunotherapy approach, the investigation of the functional effect of CTLA-4 blockade on CLL cells might be of potential clinical relevance. The main aim of this study was to examine the effect of CTLA-4 blockade on proliferation activity and apoptosis of CLL cells in patients with low and high CTLA-4 expression. We found that in the high CTLA-4-expressing CLL group, CTLA-4 blockade on the CLL cell surface resulted in a significant increase in the median percentages of Ki67(+) cells and a tendency to decrease in the proportion of apoptotic cells. In contrast, in the low CTLA-4 expressors, CTLA-4 blockade did not affect the proliferation activity or the frequency of apoptosis. This study reports for the first time the different effect of CTLA-4 blockade on CLL cells in CLL patients depending on the levels of CTLA-4 expression. CTLA-4 blockade seems to induce pro-survival signals in leukaemic cells from CLL patients exhibiting high CTLA-4 expression, suggesting that an immunotherapy approach based on the systemic use of monoclonal anti-CTLA-4 antibodies could be an unfavourable strategy for some CLL patients.

CTLA-4 affects expression of key cell cycle regulators of G0/G1 phase in neoplastic lymphocytes from patients with chronic lymphocytic leukaemia.

Previously, we showed that cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) is overexpressed in chronic lymphocytic leukaemia (CLL) and its expression is correlated with the expression of the major regulators of G1 phase progression: cyclins D2 and D3, and cyclin-dependent kinase inhibitory protein 1 (p27 (KIP1) ). In the present study, we blocked CTLA-4 on the surface of both CLL cells and normal B lymphocytes to investigate the impact of CTLA-4 on the expression of the mentioned G1 phase regulators. We found that in CLL patients and in healthy individuals, the median proportions of cyclin D2-positive cells as well as cyclin D3(+) cells significantly decreased following CTLA-4 blockade. Moreover, CTLA-4 blockade led to an increase in the median frequencies of p27 (KIP1) -positive cells, although this increase was marked only in CLL patients. Our study showed that CTLA-4 affects the expression of the key regulators of G1 phase progression in CLL cells as well as in normal B lymphocytes and may contribute to a better understanding of the role of CTLA-4 in the regulation of G1 phase progression.

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.