The prognostic impact of non-driver gene mutations and variant allele frequency in primary myelofibrosis.

Prognostic impact of non-MPN driver gene mutations in primary myelofibrosis. MIPSS70: Mutation-Enhanced International Prognostic Score System.

New Mediators in the Crosstalk between Different Adipose Tissues.

Adipose tissue is a multifunctional organ that regulates many physiological processes such as energy homeostasis, nutrition, the regulation of insulin sensitivity, body temperature, and immune response. In this review, we highlight the relevance of the different mediators that control adipose tissue activity through a systematic review of the main players present in white and brown adipose tissues. Among them, inflammatory mediators secreted by the adipose tissue, such as classical adipokines and more recent ones, elements of the immune system infiltrated into the adipose tissue (certain cell types and interleukins), as well as the role of intestinal microbiota and derived metabolites, have been reviewed. Furthermore, anti-obesity mediators that promote the activation of beige adipose tissue, e.g., myokines, thyroid hormones, amino acids, and both long and micro RNAs, are exhaustively examined. Finally, we also analyze therapeutic strategies based on those mediators that have been described to date. In conclusion, novel regulators of obesity, such as microRNAs or microbiota, are being characterized and are promising tools to treat obesity in the future.

Gut Microbiota Dysbiosis in COVID-19: Modulation and Approaches for Prevention and Therapy.

Inflammation and oxidative stress are critical underlying mechanisms associated with COVID-19 that contribute to the complications and clinical deterioration of patients. Additionally, COVID-19 has the potential to alter the composition of patients' gut microbiota, characterized by a decreased abundance of bacteria with probiotic effects. Interestingly, certain strains of these bacteria produce metabolites that can target the S protein of other coronaviruses, thereby preventing their transmission and harmful effects. At the same time, the presence of gut dysbiosis can exacerbate inflammation and oxidative stress, creating a vicious cycle that perpetuates the disease. Furthermore, it is widely recognized that the gut microbiota can metabolize various foods and drugs, producing by-products that may have either beneficial or detrimental effects. In this regard, a decrease in short-chain fatty acid (SCFA), such as acetate, propionate, and butyrate, can influence the overall inflammatory and oxidative state, affecting the prevention, treatment, or worsening of COVID-19. This review aims to explore the current evidence regarding gut dysbiosis in patients with COVID-19, its association with inflammation and oxidative stress, the molecular mechanisms involved, and the potential of gut microbiota modulation in preventing and treating SARS-CoV-2 infection. Given that gut microbiota has demonstrated high adaptability, exploring ways and strategies to maintain good intestinal health, as well as an appropriate diversity and composition of the gut microbiome, becomes crucial in the battle against COVID-19.

Polyphenols and Triterpenes Combination in an In Vitro Model of Cardiac Damage: Protective Effects.

Olive products contain high levels of monounsaturated fatty acids as well as other minor components such as triterpenic alcohols and other pentacyclic triterpenes, which together form the main triterpenes of virgin olive oil. Olive fruits and leaves contain significant amounts of hydrophilic and lipophilic bioactives including flavones, phenolic acids and phenolic alcohols, amongst others. Several studies have shown the benefits of these substances on the cardiovascular system. Regardless, little is known about the specific combination of bioactive compounds in cardiovascular health. Thus, we aimed to test the combination of a triterpenes (TT70) and a polyphenols (HT60) olive oil bioactive extract in H9c2 cells under stress conditions: LPS and H2O2 stimulation. To evaluate the effectiveness of the combination, we measured cell viability, superoxide production and protein expression of caspase 3, eNOS, peNOS, TNF-α and Il-6. Overall, cells stimulated with LPS or H2O2 and co-incubated with the combination of triterpenes and polyphenols had increased cell survival, lower levels of superoxide anion, lower protein expression of eNOS and higher expression of peNOS, increased protein expression of SOD-1 and lower protein expression of TNF-α and Il-6. The specific combination of HT60+TT70 is of great interest for further study as a possible treatment for cardiovascular damage.

Relevance of mitochondrial dysfunction in heart disease associated with insulin resistance conditions.

Insulin resistance plays a key role in the development and progression of obesity, diabetes, and their complications. Moreover, insulin resistance is considered the principal link between metabolic diseases and cardiovascular diseases. Heart disease associated with insulin resistance is one of the most important consequences of both obesity and diabetes, and it is characterized by impaired cardiac energetics, diastolic dysfunction, and finally heart failure. Mitochondrion plays a key role in cell energy homeostasis and is the main source of reactive oxygen species. Obesity and diabetes are associated with alterations in mitochondrial function and dynamics. Mitochondrial dysfunction is characterized by changes in mitochondrial respiratory chain with reduced ATP production and elevated reactive oxygen species production. These mitochondrial alterations together with inflammation contribute to the development and progression of heart disease under insulin resistance conditions. Finally, numerous miRNAs participate in the regulation of energy substrate metabolism, reactive oxygen species production, and apoptotic pathways within the mitochondria. This notion supports the relevance of interactions between miRNAs and mitochondrial dysfunction in the pathophysiology of metabolic heart disease.

Autologous stem-cell transplantation as consolidation of first-line chemotherapy in patients with peripheral T-cell lymphoma: a multicenter GELTAMO/FIL study.

Peripheral T-cell lymphomas (PTCL) are a heterogeneous group of rare lymphoid malignancies that mostly have poor prognoses with currently available treatments. Upfront consolidation with autologous stem cell transplantation (ASCT) is frequently carried out, but its efficacy has never been investigated in randomized trials. We designed a multicenter, international, retrospective study with the main objective of comparing progression-free survival and overall survival of patients with PTCL who underwent ASCT in complete remission (CR) after first-line chemotherapy with a control group who did not undergo ASCT. From the initial population of 286 registered patients, 174 patients with PTCL other than anaplastic large cell lymphoma, ALK-positive, deemed fit for ASCT at the time of diagnosis, and who were in CR or uncertain CR after induction therapy (CR1) were included in our analysis. one hundred and three patients underwent ASCT, whereas 71 did not, in most cases (n=53) because the physician decided against it. With a median follow-up of 65.5 months, progression-free survival was significantly better in the transplanted patients than in the non-transplanted group: 63% versus 48% at 5 years (P=0.042). Overall survival was significantly longer for ASCT patients in the subgroup with advanced stage at diagnosis (5-year overall survival: 70% vs. 50%, P=0.028). In the multivariate analysis, first-line ASCT was associated with significantly prolonged progression-free survival (HR=0.57, 95% CI: 0.35-0.93) and overall survival (HR=0.57, 95% CI: 0.33-0.99). In conclusion, our study supports the use of ASCT as a consolidation strategy for patients with PTCL in CR1. These results should be confirmed in a prospective randomized study.

Proanthocyanidins Maintain Cardiac Ionic Homeostasis in Aldosterone-Induced Hypertension and Heart Failure.

Excess aldosterone promotes pathological remodeling of the heart and imbalance in cardiac ion homeostasis of sodium, potassium and calcium. Novel treatment with proanthocyanidins in aldosterone-treated rats has resulted in downregulation of cardiac SGK1, the main genomic aldosterone-induced intracellular mediator of ion handling. It therefore follows that proanthocyanidins could be modulating cardiac ion homeostasis in aldosterone-treated rats. Male Wistar rats received aldosterone (1 mg kg-1 day-1) +1% NaCl for three weeks. Half of the animals in each group were simultaneously treated with the proanthocyanidins-rich extract (80% w/w) (PRO80, 5 mg kg-1 day-1). PRO80 prevented cardiac hypertrophy and decreased calcium content. Expression of ion channels (ROMK, NHE1, NKA and NCX1) and calcium transient mediators (CAV1.2, pCaMKII and oxCaMKII) were reduced by PRO80 treatment in aldosterone-treated rats. To conclude, our data indicate that PRO80 may offer an alternative treatment to conventional MR-blockade in the prevention of aldosterone-induced cardiac pathology.

Supplementation with an insoluble fiber obtained from carob pod (Ceratonia siliqua L.) rich in polyphenols prevents dyslipidemia in rabbits through SIRT1/PGC-1α pathway.

PURPOSE: To investigate the mechanism implicated in the effect of an insoluble fiber (obtained from carob pod) rich in polyphenols (IFCP) in lipid metabolism in the liver. METHODS: Male New Zealand rabbits were fed with the following diets for 8 weeks: control diet (CT group), dyslipidemic diet supplemented with 0.5% cholesterol + 14% coconut oil (DL group) and dyslipidemic diet containing 0.5% cholesterol + 14% coconut oil plus 3% IFCP (DL + IFCP group). RESULTS: Dyslipidemic diet with IFCP was able to reduce development of mixed dyslipidemia, liver relative weight and collagen I protein expression compared to DL rabbits. Analyses of the main enzymes implicated in cholesterol and triglycerides metabolism revealed that IFCP increased hepatic concentration of 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase) and cytochrome P450, family 7, subfamily a, polypeptide 1C (CYP7A1) (82.34, 114.42%, respectively) as well as protein expression of LDL receptor (42.48%) in DL rabbits. Importantly, IFCP also increased hepatic lipase (HL) levels (91.43%) and decreased glycerol phosphate acyltransferase (GPAT) and sterol regulatory element-binding protein 1C (SREBP1c) liver expression levels (20.38 and 41.20%, respectively). Finally, sirtuin 1 (SIRT1) and peroxisome proliferator-activated receptor gamma coactivator-1alpha (PGC-1α) hepatic expression increased in DL + IFCP group compared with DL (159.81 and 48.00%, respectively). CONCLUSIONS: These findings show that IFCP is able to abrogate the deleterious effects of hepatic dyslipidemia by modulating SIRT1 and PGC-1α pathways.

In vivo bioavailability of polyphenols from grape by-product extracts, and effect on lipemia of normocholesterolemic Wistar rats.

BACKGROUND: The direct use of phenolic extracts from grape by-products can be useful when formulating functional food to improve consumer health. The use of phenolic extracts instead of pure polyphenols as an ingredient is relevant in this context. The present study investigated the bioavailability and absorption of polyphenols from grape by-product extracts and their health effect on cholesterolemia, by adding the extract (GE) to Wistar rats diet (50 g kg-1 ) in vivo. RESULTS: GE caused the appearance of (+)-catechin, myricetin and quercetic acid in plasma and liver. (+)-Catechin was the most abundant compound (6 µg mL-1 in plasma and 0.7 µg mg-1 protein in liver), whereas no phenolic compounds were detected in plasma or liver in the control group. Similarly, 3,4-hydroxyphenylacetic, a major product of polyphenol digestion, was detected in the plasma, liver and urine of the GE-group only. GE-group had significantly lower cholesterol level and lower total cholesterol/high-density lipoprotein ratio in plasma. Total bile acid content significantly increased in fecal matter after 24 h administration of the GE-enriched diet. CONCLUSION: Grape extract polyphenols are partially bioavailable and showed improvement in lipid metabolism. Thus, the results suggest that GE is promising as a functional ingredient in the prevention of hypercholesterolemia. © 2018 Society of Chemical Industry.

Mutations in TP53 and JAK2 are independent prognostic biomarkers in B-cell precursor acute lymphoblastic leukaemia.

BACKGROUND: In B-cell precursor acute lymphoblastic leukaemia (B-ALL), the identification of additional genetic alterations associated with poor prognosis is still of importance. We determined the frequency and prognostic impact of somatic mutations in children and adult cases with B-ALL treated with Spanish PETHEMA and SEHOP protocols. METHODS: Mutational status of hotspot regions of TP53, JAK2, PAX5, LEF1, CRLF2 and IL7R genes was determined by next-generation deep sequencing in 340 B-ALL patients (211 children and 129 adults). The associations between mutation status and clinicopathological features at the time of diagnosis, treatment outcome and survival were assessed. Univariate and multivariate survival analyses were performed to identify independent prognostic factors associated with overall survival (OS), event-free survival (EFS) and relapse rate (RR). RESULTS: A mutation rate of 12.4% was identified. The frequency of adult mutations was higher (20.2% vs 7.6%, P=0.001). TP53 was the most frequently mutated gene (4.1%), followed by JAK2 (3.8%), CRLF2 (2.9%), PAX5 (2.4%), LEF1 (0.6%) and IL7R (0.3%). All mutations were observed in B-ALL without ETV6-RUNX1 (P=0.047) or BCR-ABL1 fusions (P<0.0001). In children, TP53mut was associated with lower OS (5-year OS: 50% vs 86%, P=0.002) and EFS rates (5-year EFS: 50% vs 78.3%, P=0.009) and higher RR (5-year RR: 33.3% vs 18.6% P=0.037), and was independently associated with higher RR (hazard ratio (HR)=4.5; P=0.04). In adults, TP53mut was associated with a lower OS (5-year OS: 0% vs 43.3%, P=0.019) and a higher RR (5-year RR: 100% vs 61.4%, P=0.029), whereas JAK2mut was associated with a lower EFS (5-year EFS: 0% vs 30.6%, P=0.035) and a higher RR (5-year RR: 100% vs 60.4%, P=0.002). TP53mut was an independent risk factor for shorter OS (HR=2.3; P=0.035) and, together with JAK2mut, also were independent markers of poor prognosis for RR (TP53mut: HR=5.9; P=0.027 and JAK2mut: HR=5.6; P=0.036). CONCLUSIONS: TP53mut and JAK2mut are potential biomarkers associated with poor prognosis in B-ALL patients.

Hyperdiploidy as a rare event that accompanies poor prognosis markers in CLL.

The presence of chromosomal gains other than trisomy 12 in chronic lymphocytic leukaemia (CLL) is unusual. However, some patients may show gains on several chromosomes simultaneously suggesting a hyperdiploid karyotype. OBJECTIVE: The objective of this study was to analyse by FISH the frequency and prognostic impact of hyperdiploidy in CLL. METHOD: A review of 1359 consecutive cases diagnosed with CLL referred for FISH analysis to a unique institution was carried out. Hyperdiploidy was considered when a gain of at least three of the five FISH probes used was observed. RESULTS: Seven cases (0.51%) with hyperdiploidy were found, confirming that it is a rare event in this disease. Although most patients presented with early Binet stages at diagnosis, six of seven (86%) shortly progressed. The median of time to the first therapy (TTFT) and overall survival (OS) for the patients with hyperdiploidy were short (1.4 months and 20 months, respectively). Moreover, comparing them with a control group of patients (non-hyperdiploid) with completed follow-up data, TTFT and OS of the patients with hyperdiploidy were significantly shorter than the control group. CONCLUSION: The presence of hyperdiploidy is uncommon and probably associated with poor prognostic markers in CLL.

Role of Mitochondrial Dysfunction in Hypertension and Obesity.

Mitochondria are essential for the maintenance of normal physiological function of tissue cells. Mitochondria are subject to dynamic processes in order to establish a control system related to survival or cell death and adaptation to changes in the metabolic environment of cells. Mitochondrial dynamics includes fusion and fission processes, biogenesis, and mitophagy. Modifications of mitochondrial dynamics in organs involved in energy metabolism such as the pancreas, liver, skeletal muscle, and white adipose tissue could be of relevance for the development of insulin resistance, obesity, and type 2 diabetes. Mitochondrial dynamics and the factors involved in its regulation are also critical for neuronal development, survival, and function. Modifications in mitochondrial dynamics in either agouti-related peptide (AgRP) or pro-opiomelanocortin (POMC), circuits which regulates feeding behavior, are related to changes of food intake, energy balance, and obesity development. Activation of the sympathetic nervous system has been considered as a crucial point in the pathogenesis of hypertension among obese individuals and it also plays a key role in cardiac remodeling. Hypertension-related cardiac hypertrophy is associated with changes in metabolic substrate utilization, dysfunction of the electron transport chain, and ATP synthesis. Alterations in both mitochondrial dynamics and ROS production have been associated with endothelial dysfunction, development of hypertension, and cardiac hypertrophy. Finally, it might be postulated that alterations of mitochondrial dynamics in white adipose tissue could contribute to the development and maintenance of hypertension in obesity situations through leptin overproduction. Leptin, together with insulin, will induce activation of sympathetic nervous system with consequences at renal, vascular, and cardiac levels, driving to sodium retention, hypertension, and left ventricular hypertrophy. Moreover, both leptin and insulin will induce mitochondrial alterations into arcuate nucleus leading to signals driving to increased food intake and reduced energy expenditure. This, in turn would perpetuate white adipose tissue excess and its well-known metabolic and cardiovascular consequences.

Bosutinib shows low cross intolerance, in chronic myeloid leukemia patients treated in fourth line. Results of the Spanish compassionate use program.

The role of bosutinib as rescue treatment of Philadelphia chromosome-positive chronic myeloid leukemia (CML) patients after failing three previous tyrosine kinase inhibitors (TKIs) is currently unknown. We report here the largest series (to our knowledge) of patients treated with bosutinib in fourth-line, after retrospectively reviewing 30 patients in chronic phase, and pretreated with imatinib, nilotinib, and dasatinib. With a median follow up of 11.1 months, the probability to either maintain or improve their CCyR response was 56.6% (17/30) and 11 patients (36.7%) achieved or maintained their baseline MMR. In patients not having baseline CCyR, the probabilities of obtaining CCyR, MMR, and MR4.5 were 13, 11, and 14%, respectively. The probabilities of obtaining MMR and deep molecular response MR4.5 in patients with baseline CCyR were 40.0% (6/15) and 20.0% (3/15). At 20 months, progression-free survival was 73%. Grade 3-4 hematological toxicities were more frequent in resistant than intolerant patients (45.4 vs. 0.0%). Nonhematological toxicities were also more frequent in resistant patients, being diarrhea the most conspicuous one. Bosutinib seems to be an appropriate treatment option for patients resistant or intolerant to three prior TKI's.

Antagonistic effect of TNF-alpha and insulin on uncoupling protein 2 (UCP-2) expression and vascular damage.

BACKGROUND: It has been reported that increased expression of UCP-2 in the vasculature may prevent the development of atherosclerosis in patients with increased production of reactive oxygen species, as in the diabetes, obesity or hypertension. Thus, a greater understanding in the modulation of UCP-2 could improve the atherosclerotic process. However, the effect of TNF-α or insulin modulating UCP-2 in the vascular wall is completely unknown. In this context, we propose to study new molecular mechanisms that help to explain whether the moderate hyperinsulinemia or lowering TNF-α levels might have a protective role against vascular damage mediated by UCP-2 expression levels. METHODS: We analyzed the effect of insulin or oleic acid in presence or not of TNF-α on UCP-2 expression in murine endothelial and vascular smooth muscle cells. At this step, we wondered if some mechanisms studied in vitro could be of any relevance in vivo. We used the following experimental models: ApoE-/- mice under Western type diet for 2, 6, 12 or 18 weeks, BATIRKO mice under high-fat diet for 16 weeks and 52-week-old BATIRKO mice with o without anti-TNF-α antibody pre-treatment. RESULTS: Firstly, we found that TNF-α pre-treatment reduced UCP-2 expression induced by insulin in vascular cells. Secondly, we observed a progressive reduction of UCP-2 levels together with an increase of lipid depots and lesion area in aorta from ApoE-/- mice. In vivo, we also observed that moderate hyperinsulinemic obese BATIRKO mice have lower TNF-α and ROS levels and increased UCP-2 expression levels within the aorta, lower lipid accumulation, vascular dysfunction and macrovascular damage. We also observed that the anti-TNF-α antibody pre-treatment impaired the loss of UCP-2 expression within the aorta and relieved vascular damage observed in 52-week-old BATIRKO mice. Finally, we observed that the pretreatment with iNOS inhibitor prevented UCP-2 reduction induced by TNF-α in vascular cells. Moreover, iNOS levels are augmented in aorta from mice with lower UCP-2 levels and higher TNF-α levels. CONCLUSIONS: Our data suggest that moderate hyperinsulinemia in response to insulin resistance or lowering of TNF-α levels within the aorta attenuates vascular damage, this protective effect being mediated by UCP-2 expression levels through iNOS.

Carob pod insoluble fiber exerts anti-atherosclerotic effects in rabbits through sirtuin-1 and peroxisome proliferator-activated receptor-γ coactivator-1α.

The aim of this study was to evaluate the potential effects of an insoluble dietary fiber from carob pod (IFC) (1 g ⋅ kg(-1) ⋅ d(-1) in the diet) on alterations associated with atherosclerosis in rabbits with dyslipidemia. Male New Zealand rabbits (n = 30) were fed the following diets for 8 wk: 1) a control diet (SF412; Panlab) as a control group representing normal conditions; 2) a control supplemented with 0.5% cholesterol + 14% coconut oil (DL) (SF302; Panlab) for 8 wk as a dyslipidemic group; and 3) a control containing 0.5% cholesterol + 14% coconut oil plus IFC (1 g ⋅ kg(-1) ⋅ d(-1)) (DL+IFC) for 8 wk. IFC was administered in a pellet mixed with the DL diet. The DL-fed group developed mixed dyslipidemia and atherosclerotic lesions, which were associated with endothelial dysfunction, inflammation, and fibrosis. Furthermore, sirtuin-1 (SIRT1) and peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) protein expression in the aorta were reduced to 77% and 63% of the control group, respectively (P < 0.05), in these rabbits. Administration of IFC to DL-fed rabbits reduced the size of the aortic lesion significantly (DL, 15.2% and DL+IFC, 2.6%) and normalized acetylcholine-induced relaxation (maximal response: control, 89.3%; DL, 61.6%; DL+IFC, 87.1%; P < 0.05) and endothelial nitric oxide synthase expression (DL, 52% and DL+IFC, 104% of the control group). IFC administration to DL-fed rabbits also reduced cluster of differentiation 36 (DL, 148% and DL+IFC, 104% of the control group; P < 0.05), plasminogen activator inhibitor-1 (DL, 141% and DL+IFC, 107% of the control group), tumor necrosis factor-α (DL, 166% and DL+IFC, 120% of the control group), vascular cell adhesion molecule-1 (DL, 153% and DL+IFC, 110% of the control group), transforming growth factor-ß (DL, 173% and DL+IFC, 99% of the control group), and collagen I (DL, 157% and DL+IFC, 112% of the control group) in the aorta. These effects were accompanied by an enhancement of SIRT1 and PGC-1α (160% and 121% of the control group, respectively; P < 0.05) vascular expression. In summary, we demonstrated for the first time, to our knowledge, that administration of IFC reduces the development of atherosclerosis in rabbits. This effect seems to be related to an improvement in endothelial function and a reduction of inflammation and fibrosis, most probably as a consequence of the reduction of serum concentrations of cholesterol and triglycerides. Increased expression of aortic SIRT1 and PGC-1α could play an important role in the observed effects of IFC in rabbits with dyslipidemia.

Potential protective role of let-7d-5p in atherosclerosis progression reducing the inflammatory pathway regulated by NF-κB and vascular smooth muscle cells proliferation.

The prevalence of cardiovascular diseases (CVDs) is increasing in the last decades, even is the main cause of death in first world countries being atherosclerosis one of the principal triggers. Therefore, there is an urgent need to decipher the underlying mechanisms involved in atherosclerosis progression. In this respect, microRNAs dysregulation is frequently involved in the progression of multiple diseases including CVDs. Our aim was to demonstrate that let-7d-5p unbalance could contribute to the pathophysiology of atherosclerosis and serve as a potential diagnostic biomarker. We evaluated let-7d-5p levels in vascular biopsies and exosome-enriched extracellular vesicles (EVs) from patients with carotid atherosclerosis and healthy donors. Moreover, we overexpressed let-7d-5p in vitro in vascular smooth muscle cells (VSMCs) to decipher the targets and the underlying mechanisms regulated by let-7d-5p in atherosclerosis. Our results demonstrate that let-7d-5p was significantly upregulated in carotid plaques from overweight patients with carotid atherosclerosis. Moreover, in EVs isolated from plasma, we found that let-7d-5p levels were increased in carotid atherosclerosis patients compared to control subjects specially in overweight patients. Receiver Operating Characteristic (ROC) analyses confirmed its utility as a diagnostic biomarker for atherosclerosis. In VSMCs, we demonstrated that increased let-7d-5p levels impairs cell proliferation and could serve as a protective mechanism against inflammation by impairing NF-κB pathway without affecting insulin resistance. In summary, our results highlight the role of let-7d-5p as a potential therapeutic target for atherosclerosis since its overexpression induce a decrease in inflammation and VSMCs proliferation, and also, as a novel non-invasive diagnostic biomarker for atherosclerosis in overweight patients.

The clinical utility and prognostic value of multiparameter flow cytometry immunophenotyping in light-chain amyloidosis.

The clinical value of multiparameter flow cytometry (MFC) immunophenotyping in primary or light chain amyloidosis (AL) remains unknown. We studied 44 consecutive bone marrow samples from newly diagnosed patients with amyloidosis; 35 patients with AL and 9 with other forms of amyloidosis. Monoclonal plasma cells (PCs) were identifiable by MFC immunophenotyping in 34 of 35 (97%) patients with AL, whereas it was absent from all but 1 of the 9 (11%) patients with other forms of amyloidosis. Quantification of bone marrow plasma cells (BMPCs) by MFC immunophenotyping was a significant prognostic factor for overall survival (OS) (≤ 1% vs > 1% BMPC cutoff; 2-year OS rates of 90% vs 44%, P = .02). Moreover, detecting persistent normal PCs at diagnosis identifies a subgroup of patients with AL with prolonged OS (> 5% vs ≤ 5% normal PC within all BMPC cutoff, 2-year rates of 88% vs 37%, P = .01). MFC immunophenotyping could be clinically useful for the demonstration of PC clonality in AL and for the prognostication of patients with AL.

Analysis of the immune system of multiple myeloma patients achieving long-term disease control by multidimensional flow cytometry.

Multiple myeloma remains largely incurable. However, a few patients experience more than 10 years of relapse-free survival and can be considered as operationally cured. Interestingly, long-term disease control in multiple myeloma is not restricted to patients with a complete response, since some patients revert to having a profile of monoclonal gammopathy of undetermined significance. We compared the distribution of multiple compartments of lymphocytes and dendritic cells in the bone marrow and peripheral blood of multiple myeloma patients with long-term disease control (n=28), patients with newly diagnosed monoclonal gammopathy of undetermined significance (n=23), patients with symptomatic multiple myeloma (n=23), and age-matched healthy adults (n=10). Similarly to the patients with monoclonal gammopathy of undetermined significance and symptomatic multiple myeloma, patients with long-term disease control showed an expansion of cytotoxic CD8(+) T cells and natural killer cells. However, the numbers of bone marrow T-regulatory cells were lower in patients with long-term disease control than in those with symptomatic multiple myeloma. It is noteworthy that B cells were depleted in patients with monoclonal gammopathy of undetermined significance and in those with symptomatic multiple myeloma, but recovered in both the bone marrow and peripheral blood of patients with long-term disease control, due to an increase in normal bone marrow B-cell precursors and plasma cells, as well as pre-germinal center peripheral blood B cells. The number of bone marrow dendritic cells and tissue macrophages differed significantly between patients with long-term disease control and those with symptomatic multiple myeloma, with a trend to cell count recovering in the former group of patients towards levels similar to those found in healthy adults. In summary, our results indicate that multiple myeloma patients with long-term disease control have a constellation of unique immune changes favoring both immune cytotoxicity and recovery of B-cell production and homing, suggesting improved immune surveillance.

Prognostic impact of the number of methylated genes in myelodysplastic syndromes and acute myeloid leukemias treated with azacytidine.

The prognostic impact of the aberrant hypermethylation in response to azacytidine (AZA) remains to be determined. Therefore, we have analyzed the influence of the methylation status prior to AZA treatment on the overall survival and clinical response of myeloid malignancies. DNA methylation status of 24 tumor suppressor genes was analyzed by methylation-specific multiplex ligation-dependent probe amplification in 63 patients with myelodysplastic syndromes and acute myeloid leukemia treated with azacytidine. Most patients (73 %) showed methylation of at least one gene, but only 12 % of patients displayed ≥3 methylated genes. The multivariate analysis demonstrated that the presence of a high number (≥2) of methylated genes (P = 0.022), a high WBC count (P = 0.033), or anemia (P = 0.029) were independent prognostic factors associated with shorter overall survival. The aberrant methylation status did not correlate with the response to AZA, although four of the five patients with ≥3 methylated genes did not respond. By contrast, favorable cytogenetics independently influenced the clinical response to AZA as 64.7 % of patients with good-risk cytogenetic abnormalities responded (P = 0.03). Aberrant methylation status influences the survival of patients treated with AZA, being shorter in those patients with a high number of methylated genes.