Expression and putative biological roles of lipoxygenases and leukotriene receptors in leukemia and lymphoma.

This mini-review addresses lipoxygenases and receptors for leukotrienes in hematological malignancies. Potential novel biomarkers and drug targets in leukemia and B-cell lymphoma are discussed.

EMA Review of Panobinostat (Farydak) for the Treatment of Adult Patients with Relapsed and/or Refractory Multiple Myeloma.

On August 28, 2015, a marketing authorization valid through the European Union was issued for panobinostat, in combination with bortezomib and dexamethasone, for the treatment of adult patients with relapsed and/or refractory multiple myeloma who have received at least two prior regimens including bortezomib and an immunomodulatory agent (IMiD).Panobinostat is an orally available histone deacetylase (HDAC) inhibitor that inhibits the enzymatic activity of HDAC proteins at nanomolar concentrations. HDAC proteins catalyze the removal of acetyl groups from the lysine residues of histones and some nonhistone proteins. Inhibition of HDAC activity results in increased acetylation of histone proteins, an epigenetic alteration that results in a relaxing of chromatin, leading to transcriptional activation. The recommended starting dose of panobinostat is 20 mg, taken orally in a cyclical manner for up to 48 weeks.The use of panobinostat in combination with bortezomib and dexamethasone was studied in a randomized, double-blind, placebo-controlled, multicenter phase III study (PANORAMA I) in 768 patients with relapsed or relapsed and refractory multiple myeloma who had received one to three prior lines of therapies. In the subgroup of patients who have received at least two prior regimens including bortezomib and an IMiD, there was a difference of 7.8 months in the progression-free survival in favor of the experimental arm (12.5 months for panobinostat + bortezomib + dexamethasone vs. 4.7 months for placebo + bortezomib + dexamethasone; hazard ratio = 0.47, 95% confidence interal 0.31-0.72; log-rank p value = .0003). The incidence of grade 3-4 adverse events suspected to be related to study drug was 76.9% vs. 51.2%, for the panobinostat and the placebo group, respectively. The most common side effects (grade 3-4) associated with panobinostat included diarrhea (18.9%), fatigue (14.7%), nausea (4.5%), vomiting (5.5%), thrombocytopenia (43.6%), anemia (7.9%), neutropenia (16.5%) and lymphopenia (8.1%).This article summarizes the scientific review of the application leading to regulatory approval in the European Union. The full scientific assessment report and product information, including the Summary of Product Characteristics, are available on the European Medicines Agency website (http://www.ema.europa.eu/ema/index.jsp?curl=pages/includes/medicines/medicines_landing_page.jsp&mid=). IMPLICATIONS FOR PRACTICE: Farydak was approved in the European Union in combination with bortezomib and dexamethasone, for the treatment of adult patients with relapsed and/or refractory multiple myeloma who have received at least two prior regimens including bortezomib and an immunomodulatory agent (IMiD). The addition of panobinostat to bortezomib and dexamethasone resulted in a clinically meaningful and statistically significant improvement of progression-free survival compared with bortezomib and dexamethasone, and an additional therapeutic option with a new mechanism of action was considered valuable. Although the toxicity associated with panobinostat combination was significant, at the time of the marketing authorization of panobinostat, it was considered that it was acceptable and that it should be left to the clinician and the patient to decide whether the panobinostat combination is the preferred treatment option or not.

Human 15-lipoxygenase-1 is a regulator of dendritic-cell spreading and podosome formation.

Dendritic cells (DCs) involved in proinflammatory immune responses derive mainly from peripheral monocytes, and the cells subsequently mature and migrate into the inflammatory micromilieu. Here we report that suppressing of 15-lipoxygenase-1 led to a substantial reduction in DC spreading and podosome formation in vitro. The surface expression of CD83 was significantly lower in both sh-15-lipoxygenase-1 (15-LOX-1)-transduced cells and DCs cultivated in the presence of a novel specific 15-LOX-1 inhibitor. The T-cell response against tetanus-pulsed DCs was only affected to a minor extent on inhibition of 15-LOX-1. In contrast, endocytosis and migration ability of DCs were significantly suppressed on 15-LOX-1 inhibition. The expression of 15-LOX-1 in DCs was also demonstrated in affected human skin in atopic and contact dermatitis, showing that the enzyme is indeed expressed in inflammatory diseases in vivo. This study demonstrated that inhibiting 15-LOX-1 led to an impaired podosome formation in DCs, and consequently suppressed antigen uptake and migration capacity. These results indicated that 15-LOX-1 is a potential target for inhibiting the trafficking of DCs to lymphoid organs and inflamed tissues and decreasing the inflammatory response attenuating symptoms of certain immunologic and inflammatory disorders such as dermatitis.-Han, H., Liang, X., Ekberg, M., Kritikou, J. S., Brunnström, Å., Pelcman, B., Matl, M., Miao, X., Andersson, M., Yuan, X., Schain, F., Parvin, S., Melin, E., Sjöberg, J., Xu, D., Westerberg, L. S., Björkholm, M., Claesson, H.-E. Human 15-lipoxygenase-1 is a regulator of dendritic-cell spreading and podosome formation.

Early growth response gene (EGR)-1 regulates leukotriene D4-induced cytokine transcription in Hodgkin lymphoma cells.

Classical Hodgkin lymphoma (cHL) has a unique pathological feature characterized by a minority of malignant Hodgkin Reed-Sternberg (H-RS) cells surrounded by numerous inflammatory cells. Cysteinyl-leukotrienes (CysLTs) are produced by eosinophils, macrophages and mast cells in the HL tumor microenvironment. In the present study we have explored the signal transduction pathways leading to leukotriene (LT) D4 induced expression of cytokines in the Hodgkin lymphoma cell line L1236 and KM-H2. Stimulation of L1236 and KM-H2 cells with LTD4 led to a concentration- and time-dependent increase at the transcriptional level of tumor necrosis factor-alpha (TNF-α), interleukin (IL)-6, IL-8, chemokine (C-C motif) ligand 3 (CCL3) and CCL4. The expression of several transcription factors was induced upon stimulation of Hodgkin cell lines with LTD4. Among these, EGR-1 was required for cytokine production. Inhibition of EGR-1 expression using shEGR-1 transduced by lentivirus led to suppression of the expression of TNF-α and IL-6. The effect of LTD4 on the expression of transcription factors and cytokines were also blocked by the specific CysLT1 receptor antagonist zafirlukast. These results demonstrate that EGR-1 plays a critical role in LTD4-induced cytokine transcription in Hodgkin cell lines.

Progress in Hodgkin lymphoma: a population-based study on patients diagnosed in Sweden from 1973-2009.

In recent decades, attention has focused on reducing long-term, treatment-related morbidity and mortality in Hodgkin lymphoma (HL). In the present study, we looked for trends in relative survival for all patients diagnosed with HL in Sweden from 1973-2009 (N = 6949; 3985 men and 2964 women; median age, 45 years) and followed up for death until the end of 2010. Patients were categorized into 6 age groups and 5 calendar periods (1973-1979, 1980-1986, 1987-1994, 1994-2000, and 2001-2009). Relative survival improved in all age groups, with the greatest improvement in patients 51-65 years of age (P < .0005). A plateau in relative survival was observed in patients below 65 years of age during the last calendar period, suggesting a reduced long-term, treatment-related mortality. The 10-year relative survival for patients diagnosed in 2000-2009 was 0.95, 0.96, 0.93, 0.80, and 0.44 for the age groups 0-18, 19-35, 36-50, 51-65, and 66-80, respectively. Therefore, despite progress, age at diagnosis remains an important prognostic factor (P < .0005). Advances in therapy for patients with limited and advanced-stage HL have contributed to an increasing cure rate. In addition, our findings support that long-term mortality of HL therapy has decreased. Elderly HL patients still do poorly, and targeted treatment options associated with fewer side effects will advance the clinical HL field.

Multidrug resistance in relapsed acute myeloid leukemia: evidence of biological heterogeneity.

BACKGROUND: Studies of mechanisms mediating resistance to chemotherapy led to the discovery of the multidrug transporter ABCB1 (ATP-binding cassette, subfamily B, member 1), often expressed in leukemic cells of patients with acute myeloid leukemia (AML). Most clinical trials evaluating the strategy of inhibiting efflux-mediated chemotherapeutic resistance have been unsuccessful, clearly indicating the need for a better approach. METHODS: This study investigated the clinical relevance of 380 genes whose expression has been shown to affect the response to chemotherapy, mostly through in vitro studies, in 11 paired samples obtained at AML diagnosis and at relapse. The expression profiling of these 380 genes was performed using TaqMan-based quantitative reverse-transcription polymerase chain reaction. Patients had a median age of 58 years at diagnosis, a median duration of complete remission of 284.5 days, and a median overall survival of 563 days. Cytogenetic abnormalities were detected at diagnosis in 4 patients, whereas 5 displayed a normal karyotype and 2 were not investigated. RESULTS: Hierarchical clustering shows that samples taken at diagnosis and relapse clustered in pairs for 6 patients of the 11 studied, suggesting recurrence of the same leukemic blast, whereas for the other 5 patients, the data indicate their relapse blasts arose from different origins. A patient-by-patient analysis of the paired samples led to the striking observation that each had a unique gene signature representing different mechanisms of resistance. CONCLUSIONS: The data underline the need for personalized molecular analysis to tailor treatment for patients with AML.

The European Medicines Agency review of pixantrone for the treatment of adult patients with multiply relapsed or refractory aggressive non-Hodgkin's B-cell lymphomas: summary of the scientific assessment of the committee for medicinal products for human use.

On May 10, 2012, the European Commission issued a conditional marketing authorization valid throughout the European Union for pixantrone for the treatment of adult patients with multiply relapsed or refractory aggressive non-Hodgkin's B-cell lymphoma (NHL). Pixantrone is a cytotoxic aza-anthracenedione that directly alkylates DNA-forming stable DNA adducts and cross-strand breaks. The recommended dose of pixantrone is 50 mg/m(2) administered on days 1, 8, and 15 of each 28-day cycle for up to 6 cycles. In the main study submitted for this application, a significant difference in response rate (proportion of complete responses and unconfirmed complete responses) was observed in favor of pixantrone (20.0% vs. 5.7% for pixantrone and physician's best choice, respectively), supported by the results of secondary endpoints of median progression-free and overall survival times (increase of 2.7 and 2.6 months, respectively). The most common side effects with pixantrone were bone marrow suppression (particularly of the neutrophil lineage) nausea, vomiting, and asthenia. This article summarizes the scientific review of the application leading to approval in the European Union. The detailed scientific assessment report and product information, including the summary of product characteristics, are available on the European Medicines Agency website (http://www.ema.europa.eu).

Epigenetic and transcriptional control of the 15-lipoxygenase-1 gene in a Hodgkin lymphoma cell line.

Lipoxygenases oxidatively metabolize polyunsaturated fatty acids to a rich spectrum of biologically active metabolites. The present study aimed at delineating the transcriptional and epigenetic mechanisms leading to 15-lipoxygenase-1 (15-LOX-1) expression in the Hodgkin lymphoma (HL) cell line L1236. Examination of the 15-LOX-1 5' promoter region demonstrated three putative binding sites for signal transducer and activator of transcription (STAT6) within the proximal 1200 base pairs relative to the start codon. Analysis by serial promoter deletions and STAT6 binding site mutations indicated that all three STAT6 binding sites are required for full activation of the 15-LOX-1 promoter. Chromatin immunoprecipitation assay demonstrated that these regions were occupied by STAT6 in L1236 (15-LOX-1 positive) but not in L428 (15-LOX-1 negative) cultured HL cells. Furthermore, DNA hypomethylation and histone hyperacetylation were detectable within the core promoter region of 15-LOX-1 only in L1236 cells but not L428 cells. Taken together, our data indicate that STAT6 activation and chromatin remodeling by DNA demethylation and histone acetylation are crucial for transcriptional activation of 15-LOX-1 in cultured HL cells. These prerequisites are fulfilled in the L1236 cell line, but not in the L428 cell line.

How we treat elderly patients with Hodgkin lymphoma.

PURPOSE OF REVIEW: To review Hodgkin lymphoma in older adults with regard to epidemiology, disease characteristics, prognosis, treatment, and future developments. RECENT FINDINGS: Older Hodgkin lymphoma patients defined by chronological age represent a heterogeneous population in terms of life expectancy, morbidities, and functional status. Twenty-one percent of Swedish Hodgkin lymphoma patients are greater than 65 years (5%, >81 years) at diagnosis. In general, less than 10% of patients included in broad clinical trials are greater than 60 years. The proportion of mixed cellularity histopathology and EBV-genome-positive tumors is higher in older adults. Five-year relative survival of Swedish patients 66-80 and greater than 81 years is 0.58 and 0.26, respectively. Older patients have lower remission rates, but relapse-free survival is less impaired. No standard treatment recommendations exist. In older fit patients less than 65-70 years--go for 'young' treatment. Estimate thoroughly the individual patient's frailness/comorbidities in order to properly adjust treatment, thus saving patients from over/undertreatment. The use of early PET should be optimized in clinical practice. Representativeness of large clinical trials including evaluation of functional status and comorbidity remains crucial. SUMMARY: Elderly Hodgkin lymphoma patients still do poorly and improved prognostics, personalized and targeted treatment options associated with fewer side-effects will hopefully advance the clinical Hodgkin lymphoma field.

Combinations in the first-line treatment of patients with advanced/metastatic renal cell cancer: regulatory aspects.

The therapeutic landscape in the treatment of advanced/metastatic renal cell cancer has evolved over the last 2 years with the advent of immune checkpoint inhibitors. In 2018 and 2019, marketing authorisations valid throughout the European Union were issued for nivolumab and ipilimumab dual checkpoint inhibition and pembrolizumab or avelumab in combination with the tyrosine kinase inhibitor axitinib. These applications presented numerous regulatory challenges.In this paper, we summarise the main regulatory considerations, originating from the assessment of the dossiers submitted from the applicants for the three combinations. The regulatory issues are grouped in four sections: clinical pharmacology, efficacy, biomarkers and safety. In each section, we describe the issues raised during the regulatory evaluation performed by the Committee for Medicinal Products for Human Use (CHMP) assessors. The CHMP assessments determine whether the medicines concerned meet the necessary quality, safety and efficacy requirements, and whether the benefit-risk balance is positive.In summary, although the overall benefit-risk was considered positive for the three combinations, the immaturity of the outcome data and the absence of long-term safety data remain issues to be addressed. Postauthorisation efficacy studies have been required to confirm the effects of the new combinations.

Hodgkin Lymphoma Monozygotic Triplets Reveal Divergences in DNA Methylation Signatures.

We studied DNA methylation profiles in four different cell populations from a unique constellation of monozygotic triplets in whom two had developed Hodgkin Lymphoma (HL). We detected shared differences in DNA methylation signatures when comparing the two HL-affected triplets with the non-affected triplet. The differences were observed in naïve B-cells and marginal zone-like B-cells. DNA methylation differences were also detected when comparing each of the HL-affected triplets against each other. Even though we cannot determine whether treatment and/or disease triggered the observed differences, we believe our data are important on behalf of forthcoming studies, and that it might provide important clues for a better understanding of HL pathogenesis.

15-Lipoxygenase-1 induces expression and release of chemokines in cultured human lung epithelial cells.

15-Lipoxygenase-1 (15-LOX-1) has been proposed to be involved in various physiological and pathophysiological activities such as inflammation, atherosclerosis, cell maturation, and tumorigenesis. Asthma and chronic obstructive pulmonary disease are associated with increased expression of 15-LOX-1 in bronchial epithelial cells, but the potential functions of 15-LOX-1 in airway epithelial cells have not been well clarified. To study the function of 15-LOX-1 in bronchial epithelial cells, we ectopically expressed 15-LOX-1 in the human lung epithelial cell line A549. We found that overexpression of 15-LOX-1 in A549 cells leads to increased release of the chemokines MIP-1alpha, RANTES, and IP-10, and thereby to increased recruitment of immature dendritic cells, mast cells, and activated T cells. These results suggest that an increased expression and activity of 15-LOX-1 in lung epithelial cells is a proinflammatory event in the pathogenesis of asthma and other inflammatory lung disorders.

Expression of the Epstein-Barr virus-encoded Epstein-Barr virus nuclear antigen 1 in Hodgkin's lymphoma cells mediates Up-regulation of CCL20 and the migration of regulatory T cells.

In approximately 50% of patients with Hodgkin's lymphoma (HL), the Epstein-Barr virus (EBV), an oncogenic herpesvirus, is present in tumor cells. After microarray profiling of both HL tumors and cell lines, we found that EBV infection increased the expression of the chemokine CCL20 in both primary Hodgkin and Reed-Sternberg cells and Hodgkin and Reed-Sternberg cell-derived cell lines. Additionally, this up-regulation could be mediated by the EBV nuclear antigen 1 protein. The higher levels of CCL20 in the supernatants of EBV-infected HL cell lines increased the migration of CD4(+) lymphocytes that expressed FOXP3, a marker of regulatory T cells (Tregs), which are specialized CD4(+) T cells that inhibit effector CD4(+) and CD8(+) T cells. In HL, an increased number of Tregs is associated with the loss of EBV-specific immunity. Our results identify a mechanism by which EBV can recruit Tregs to the microenvironment of HL by inducing the expression of CCL20 and, by doing so, prevent immune responses against the virus-infected tumor population. Further investigation of how EBV recruits and modifies Tregs will contribute not only to our understanding of the pathogenesis of virus-associated tumors but also to the development of therapeutic strategies designed to manipulate Treg activity.

The telomerase reverse transcriptase (hTERT) gene is a direct target of the histone methyltransferase SMYD3.

Recent evidence has accumulated that the dynamic histone methylation mediated by histone methyltransferases and demethylases plays key roles in regulation of chromatin structure and transcription. In the present study, we show that SET and MYND domain-containing protein 3 (SMYD3), a histone methyltransferase implicated in oncogenesis, directly trans-activates the telomerase reverse transcriptase (hTERT) gene that is essential for cellular immortalization and transformation. SMYD3 occupies its binding motifs on the hTERT promoter and is required for maintenance of histone H3-K4 trimethylation, thereby contributing to inducible and constitutive hTERT expression in normal and malignant human cells. Knocking down SMYD3 in tumor cells abolished trimethylation of H3-K4, attenuated the occupancy by the trans-activators c-MYC and Sp1, and led to diminished histone H3 acetylation in the hTERT promoter region, which was coupled with down-regulation of hTERT mRNA and telomerase activity. These results suggest that SMYD3-mediated trimethylation of H3-K4 functions as a licensing element for subsequent transcription factor binding to the hTERT promoter. The present findings provide significant insights into regulatory mechanisms of hTERT/telomerase expression; moreover, identification of the hTERT gene as a direct target of SMYD3 contributes to a better understanding of SMYD3-mediated cellular transformation.

Evidence for a pathophysiological role of cysteinyl leukotrienes in classical Hodgkin lymphoma.

Classical Hodgkin lymphoma (cHL) is characterized histologically by a minority of malignant Hodgkin Reed-Sternberg cells surrounded by abundant inflammatory cells, generally believed to be of major importance in the pathophysiology of the disease. Here, we present data that link inflammatory cell-derived arachidonic acid metabolites, the cysteinyl leukotrienes (CysLT), to the pathogenesis of cHL. Two HL cell lines, L1236 and KMH2, were shown to express functional CysLT(1) receptors, responding with a robust calcium signal upon leukotriene (LT) D(4) challenge. LTD(4) stimulated protein release of tumor necrosis factor-alpha, interleukin-6 and -8 by L1236 cells and interleukin-8 by KMH2 cells. Importantly, all these LTD(4)-induced effects were blocked by the CysLT(1) receptor-specific antagonist zafirlukast. Immunohistochemical studies of cHL biopsies and microarray analysis of microdissected cells revealed that the CysLT(1) receptor is expressed also by primary Hodgkin Reed-Sternberg cells. As these cells are surrounded by CysLT-producing eosinophils, macrophages and mast cells, our results suggest the CysLTs as mediators in the pathogenesis of cHL, contributing to the aberrant cytokine network of this lymphoma.

Hodgkin Reed-Sternberg cells express 15-lipoxygenase-1 and are putative producers of eoxins in vivo: novel insight into the inflammatory features of classical Hodgkin lymphoma.

Classical Hodgkin lymphoma has unique clinical and pathological features and tumour tissue is characterized by a minority of malignant Hodgkin Reed-Sternberg cells surrounded by inflammatory cells. In the present study, we report that the Hodgkin lymphoma-derived cell line L1236 has high expression of 15-lipoxygenase-1 and that these cells readily convert arachidonic acid to eoxin C(4), eoxin D(4) and eoxin E(4). These mediators were only recently discovered in human eosinophils and mast cells and found to be potent proinflammatory mediators. Western blot and immunocytochemistry analyses of L1236 cells demonstrated that 15-lipoxygenase-1 was present mainly in the cytosol and that the enzyme translocated to the membrane upon calcium challenge. By immunohistochemistry of Hodgkin lymphoma tumour tissue, 15-lipoxygenase-1 was found to be expressed in primary Hodgkin Reed-Sternberg cells in 17 of 20 (85%) investigated biopsies. The enzyme 15-lipoxygenase-1, however, was not expressed in any of 10 biopsies representing nine different subtypes of non-Hodgkin lymphoma. In essence, the expression of 15-lipoxygenase-1 and the putative formation of eoxins by Hodgkin Reed-Sternberg cells in vivo are likely to contribute to the inflammatory features of Hodgkin lymphoma. These findings may have important diagnostic and therapeutic implications in Hodgkin lymphoma. Furthermore, the discovery of the high 15-lipoxygenase-1 activity in L1236 cells demonstrates that this cell line comprises a useful model system to study the chemical and biological roles of 15-lipoxygenase-1.

Differential expression of cysteinyl leukotriene receptor 1 and 15-lipoxygenase-1 in non-Hodgkin lymphomas.

BACKGROUND: Arachidonic acid metabolites have been suggested to play an important role in carcinogenesis. We have recently reported that the cysteinyl leukotriene receptor 1 (CysLT1) and 15-lipoxygenase-1 (15-LO-1) are expressed by the malignant Hodgkin Reed-Sternberg cells of Hodgkin lymphoma and certain Hodgkin lymphoma cell lines, and that these cells convert arachidonic acid to the novel proinflammatory eoxins. MATERIALS AND METHODS: The expression of the CysLT1 receptor and 15-LO-1 was investigated in a broad range of non-Hodgkin lymphomas (NHLs) by immunohistochemistry. The functionality of the CysLT1 receptor in primary mediastinal B-cell lymphoma (PMBCL) cell lines was studied by calcium mobilization assays. RESULTS: Primary mediastinal B-cell lymphoma was the only NHL entity showing tumor cells positive for the CysLT1 receptor (9 of 10 tumors), and the PMBCL cell line Med-B1 expressed functional CysLT1 receptors, responding with a robust calcium signal upon cysteinyl leukotriene challenge. Furthermore, the tumor cells in 1 of 4 T-cell-derived anaplastic large-cell lymphomas, in contrast to all other studied NHLs, strongly expressed 15-LO-1. CONCLUSION: Among the NHL entities included in this study, the CysLT1 receptor was exclusively expressed by the tumor cells of PMBCL. Thus, this further corroborates the pathologic overlap between PMBCL and classical Hodgkin lymphoma.