Amyloid-ß Homeostasis Bridges Inflammation, Synaptic Plasticity Deficits and Cognitive Dysfunction in Multiple Sclerosis.

Cognitive deficits are frequently observed in multiple sclerosis (MS), mainly involving processing speed and episodic memory. Both demyelination and gray matter atrophy can contribute to cognitive deficits in MS. In recent years, neuroinflammation is emerging as a new factor influencing clinical course in MS. Inflammatory cytokines induce synaptic dysfunction in MS. Synaptic plasticity occurring within hippocampal structures is considered as one of the basic physiological mechanisms of learning and memory. In experimental models of MS, hippocampal plasticity is profoundly altered by proinflammatory cytokines. Although mechanisms of inflammation-induced hippocampal pathology in MS are not completely understood, alteration of Amyloid-ß (Aß) metabolism is emerging as a key factor linking together inflammation, synaptic plasticity and neurodegeneration in different neurological diseases. We explored the correlation between concentrations of Aß1-42 and the levels of some proinflammatory and anti-inflammatory cytokines (interleukin-1ß (IL-1ß), IL1-ra, IL-8, IL-10, IL-12, tumor necrosis factor α (TNFα), interferon γ (IFNγ)) in the cerebrospinal fluid (CSF) of 103 remitting MS patients. CSF levels of Aß1-42 were negatively correlated with the proinflammatory cytokine IL-8 and positively correlated with the anti-inflammatory molecules IL-10 and interleukin-1 receptor antagonist (IL-1ra). Other correlations, although noticeable, were either borderline or not significant. Our data show that an imbalance between proinflammatory and anti-inflammatory cytokines may lead to altered Aß homeostasis, representing a key factor linking together inflammation, synaptic plasticity and cognitive dysfunction in MS. This could be relevant to identify novel therapeutic approaches to hinder the progression of cognitive dysfunction in MS.

High platelet count at diagnosis is a protective factor for thrombosis in patients with essential thrombocythemia.

To assess the role of platelet (PLT) count for thrombotic complications in Essential Thrombocythemia (ET), 1201 patients followed in 11 Hematological centers in the Latium region were retrospectively evaluated. At multivariate analysis, the following factors at diagnosis were predictive for a worse Thrombosis-free Survival (TFS): the occurrence of previous thrombotic events (p=0.0004), age>60years (p=0.0044), spleen enlargement (p=0.042) and a lower PLT count (p=0.03). Receiver Operating Characteristic (ROC) analyses based on thrombotic events during follow-up identified a baseline platelet count of 944×109/l as the best predictive threshold: thrombotic events were 40/384 (10.4%) in patients with PLT count >944×109/l and 109/817 (13.3%) in patients with PLT count <944×109/l, respectively (p=0.04). Patients with PLT count <944×109/l were older (median age 60.4years. vs 57.1years., p=0.016), had a lower median WBC count (8.8×109/l vs 10.6×109/l, p<0.0001), a higher median Hb level (14.1g/dl vs 13.6g/dl, p<0.0001) and a higher rate of JAK-2-V617F positivity (67.2% vs 41.6%, p<0.0001); no difference was observed as to thrombotic events before diagnosis, spleen enlargement and concomitant Cardiovascular Risk Factors. In conclusion, our results confirm the protective role for thrombosis of an high PLT count at diagnosis. The older age and the higher rate of JAK-2 V617F positivity in the group of patients with a baseline lower PLT count could in part be responsible of this counterintuitive finding.

Thrombosis and survival in essential thrombocythemia: a regional study of 1,144 patients.

To identify prognostic factors affecting thrombosis-free survival (TFS) and overall survival (OS), we report the experience of a Regional cooperative group in a real-life cohort of 1,144 patients with essential thrombocythemia (ET) diagnosed from January 1979 to December 2010. There were 107 thrombotic events (9.4%) during follow-up [60 (5.3%) arterial and 47 (4.1%) venous thromboses]. At univariate analysis, risk factors for a shorter TFS were: age >60 years (P < 0.0054, 95% CI 1.18-2.6), previous thrombosis (P < 0.0001, 95% CI 1.58-4.52) and the presence of at least one cardiovascular risk factor (P = 0.036, 95% CI 1.15-3.13). Patients with a previous thrombosis occurred ≥24 months before ET diagnosis had a shorter TFS compared to patients with a previous thrombosis occurred <24 months (P = 0.0029, 95% CI 1.5-6.1); furthermore, patients with previous thrombosis occurred <24 months did not show a shorter TFS compared with patients without previous thrombosis (P = 0.303, 95% CI 0.64-3.21). At multivariate analysis for TFS, only the occurrence of a previous thrombosis maintained its prognostic impact (P = 0.0004, 95% CI 1.48-3.79, RR 2.36). The 10-year OS was 89.9% (95% CI 87.3-92.5): at multivariate analysis for OS, age >60 years (P < 0.0001), anemia (P < 0.0001), male gender (P = 0.0019), previous thromboses (P = 0.0344), and white blood cell >15 × 10(9) /l (P = 0.0370) were independent risk factors. Previous thrombotic events in ET patients are crucial for TFS but their importance seems related not to the occurrence per se but mainly to the interval between the event and the diagnosis.

Rapid detection of IDH2 (R140Q and R172K) mutations in acute myeloid leukemia.

NADP-dependent enzyme isocitrate dehydrogenase (IDH) mutations, IDH1 and IDH2, have been described in acute myeloid leukemia (AML) using next generation sequencing approaches. IDH2 mutations are heterozygous; they alter a single arginine residue at position 140 or 172 and have distinct prognostic significance. The current detection methods of IDH2 mutations are laborious and time consuming as they require DNA sequencing. Herein, we report a new allele-specific oligonucleotide-polymerase chain reaction (ASO-PCR) method to detect the IDH2 mutations. Analysis of leukemic DNA samples from 120 AML patients enabled to identify IDH2 mutations in 22 cases which were confirmed by direct DNA sequencing. Of these, 17 harbored IDH2 (R140Q) and 5 IDH2 (R172K) mutations. Serial dilution experiments showed that the assay enable to detect mutations in 10⁻³ dilutions. Our ASO-PCR method appears useful for routine diagnostic screening of these prognostically relevant alterations in AML and may be conveniently included in the diagnostic workup.

Identification of emerging FLT3 ITD-positive clones during clinical remission and kinetics of disease relapse in acute myeloid leukaemia with mutated nucleophosmin.

FLT3 internal tandem duplication (ITD) mutations are frequently detected at diagnosis in cytogenetically normal acute myeloid leukaemia (CN-AML) and predict unfavourable outcome. FLT3 ITD is an unstable aberration and may be lost or acquired at relapse. Recent whole genome sequencing studies have suggested that FLT3 ITD(+)ve AML relapse may evolve from small subclones undetectable at diagnosis by routine polymerase chain reaction (PCR). We developed a patient-specific real-time quantitative-PCR (RQ-PCR) to implement FLT3 ITD detection in six AML patients whose blasts carried wild-type FLT3 at diagnosis and who relapsed with FLT3 ITD by routine PCR. Patient-specific forward primers were designed after cloning and sequencing the FLT3 ITD in each case. The assay allowed retrospective detection of FLT3 ITD in diagnostic samples of 4/6 cases and to establish the kinetics of clonal evolution preceding relapse. After conventional chemotherapy, all patients had early relapse despite having been classified as NPM1(+)ve/FLT3 ITD(-)ve at presentation, with shorter remissions being observed in four patients re-classified as FLT3 ITD(+)ve by the new assay. Notably, FLT3 ITD clone became detectable by conventional PCR in three patients tested during remission after initial treatment. Our data underscore the need of identifying low FLT3 ITD levels, which are probably associated with relapse in otherwise good prognosis CN-AML.

Analysis of mutational status, SNP rs16754, and expression levels of Wilms tumor 1 (WT1) gene in acute promyelocytic leukemia.

Overexpression, polymorphisms, and mutations of the WT1 gene have been reported in several human tumors including acute myeloid leukemia (AML) and variably correlated with prognosis. Acute promyelocytic leukemia (APL) represents the AML subset disclosing higher WT1 expression levels; however, no WT1 studies specifically focused on APL have been conducted. We screened for the presence of mutations, SNP rs16754, and expression levels of WT1 gene in 103 adult patients with newly diagnosed APL. Fms-like tyrosine kinase (FLT3) mutations were analyzed as well. WT1 mutations were identified in four (4 %) patients. At least one copy of the minor SNP rs16754 allele (WT1(AG) or WT1(GG)) was detected in 30 (29 %) patients. Six patients (6 %) were homozygous for the minor allele (WT1(GG)) and this genotype was associated with higher WT1 mRNA copies (p = 0.018). FLT3 mutations were found in 37 % of patients and correlated with high WT1 mRNA expression (p = 0.004). Patients heterozygous or homozygous for the minor allele and patients homozygous for major (WT1(AA)) allele did not differ in terms of presenting features. In adult APL, WT1 gene mutational and polymorphic profile shows similarities with pediatric AML rather than with adult AML.

Symptomatic mucocutaneous toxicity of hydroxyurea in Philadelphia chromosome-negative myeloproliferative neoplasms: the Mister Hyde face of a safe drug.

BACKGROUND: The current study was conducted to evaluate severe mucocutaneous toxicity during treatment with hydroxyurea (HU) in a large cohort of patients with Philadelphia chromosome-negative myeloproliferative neoplasms (MPN). METHODS: Among 993 consecutive patients newly diagnosed with MPN at 4 centers in Rome between January 1980 and December 2009, 614 patients (277 men and 337 women with a median age of 64.4 years [interquartile range (IR), 54.4 years-72.7 years]) received HU. HU was administered as first-line treatment in 523 patients (85.2%) and as ≥ second-line treatment in 91 patients (14.8%). RESULTS: Mucocutaneous toxicity was reported in 51 patients (8.3%) after a median period from the initiation of HU treatment of 32.1 months (IR, 10.5 months-74.6 months) and a mean HU dose of 1085 mg (± 390 mg); 30 patients (58.8%) developed a painful ulcerative skin toxicity, mainly located in the perimalleolar area; 11 patients (21.6%) had oral aphthous ulcers; and 10 patients (19.6%) developed a nonulcerative skin toxicity with erythema and skin infiltration. After the mucocutaneous toxicity occurred, HU treatment was continued at the same dose in 5 patients (9.8%), reduced in 12 patients (23.5%), and temporarily discontinued in 7 patients (13.7%); the remaining 27 patients (52.9%) required a permanent drug discontinuation. After a median period of 4.3 months (IR, 2.4 months-9.0 months) from the onset of the skin toxicity, 39 patients (76.5%) had a complete resolution and 12 patients (23.5%) had improvement without complete resolution. CONCLUSIONS: Mucocutaneous toxicity during HU treatment is more common than expected and may present with different clinical features. Moreover, it often requires a permanent drug discontinuation and only a partial resolution is reported to occur in approximately 25% of patients.