In a group of newly diagnosed acute lymphocytic leukemia (ALL) children we evaluated a number of hemostatic and inflammatory markers at diagnosis and at different time points during chemotherapy for the remission induction to identify alterations in the plasma levels of prothrombotic markers before and during the course of chemotherapy. The following plasma markers were evaluated: thrombin-antithrombin complex (TAT), D-Dimer, plasminogen activator inhibitor 1 (PAI-1), antithrombin, fibrinogen, von Willebrand factor (VWF) antigen and high molecular weight VWF (HMW-VWF) multimers, P-selectin, tumor necrosis factor alpha (TNF-a), and interleukin 6 (IL-6). Plasma samples were collected at the following time points: at T0 (baseline) and T1 (124 days of therapy), T2 (136 days therapy), and T3 (164 days therapy). The results show that, at diagnosis, ALL children presented with laboratory signs of increased thrombin generation and fibrin formation (i.e. high TAT and D-dimer levels), fibrinolysis inhibition (i.e. high PAI-1 level), endothelial activation (i.e., high HMW-VWF and soluble P-selectin levels) and inflammation (i.e. high TNF-alpha and IL-6 levels). After starting induction therapy, the thrombin generation markers and inflammatory cytokines significantly decreased. To the opposite, PAI-1 and P-selectin significantly increased, suggesting an insult by chemotherapy on the vascular endothelium. These effects were more evident during steroid administration. Symptomatic venous thromboembolism (VTE) episodes developed in two cases during induction therapy, which did not allow the evaluation of the predictive value for VTE of laboratory markers. Am. J. Hematol. 85:325-330, 2010. V

Neuroblastoma (NB) and Ewing's sarcoma (ES) represent the most common extracranial solid tumors of childhood. Heat shock proteins (HSP) are elevated in cancer cells and their over-expression was correlated to drug-resistance. In this work we identified the HSP by a sensitive proteomic analysis of NB and ES cell lines, then, we studied the HSP response to doxorubicin. Some identified HSP were constitutively more expressed in NB than in ES cells. Doxorubicinstimulated HSP response only in NB cells. Quercetin was found to inhibit HSP expression depleting heat shock factor 1 (HSF1) cellular stores. Quercetin caused a higher antiproliferative effect in NB (IC 50 : 6.9 ± 5.8 lmol/L) than in ES cells (IC 50 : 85.5 ± 53.1 lmol/L). Moreover, quercetin caused a very pronounced doxorubicin sensitizing effect in NB cells (241 fold IC 50 decrease) and a moderate effect in ES cells. HSP involvement in NB cells sensitization was confirmed by the silencing of HSF1. Quercetin treatment and HSF1 silencing increased the pro-apoptotic effect of doxorubicin. In conclusion, the higher HSP levels, observed in NB cells, did not confer increased resistance to doxorubicin; on the contrary, HSP inhibition by quercetin or gene silencing caused higher sensitization to doxorubicin. These results may have a potential application in the treatment of NB. Keywords: doxorubicin, Ewing's sarcoma, gene silencing, heat shock proteins, neuroblastoma, quercetin. Neuroblastoma (NB) and Ewing's sarcoma (ES) are tumors of neuroectodermal origin. They are the prototypes of 'small, round blue-cell tumors', a broad category of pediatric tumors, and are the most common extra cranial solid tumors of childhood (Brodeur and Maris 2002;Ginsberg et al. 2002). Both arise from the neural crest though their cytogenetic and biological features differ (Nesbit et al. 1990;Ladenstein et al. 1994). Advanced NB and ES are still frequently fatal despite aggressive management (Blanc et al. 2003). NB is chemosensitive, but usually recurs and resists further treatment. p-Glycoprotein (pgp) and neurotrophic tyrosine kinase receptor type 2 expression appears to play a role in NB drug resistance (Ho et al. 2002). Heat shock proteins (HSP) role in neuroprotection following thermal/oxidative stress and ischemia/reperfusion injury has been described (Goldbaum and Richter-Landsberg 2001;Yenari 2002;Bando et al. 2003) but little is known about HSP functions and distribution in neuroectodermal tumors. A single report indicated chemotherapic sensitizing effect of quercetin in an ES cell line; the effect was correlated with HSP27 and HSP70 inhibition, (Debes et al. 2003), no data are still available about quercetin sensitizing effect in NB cells. HSP were often implicated in anti-cancer drug response (Volm et al. 1995), even though cancer cells presenting high levels of HSP were not demonstrated to be more resistant to anti-cancer-drugs than cancer cells with low HSP levels (Hettinga et al. 1996;Bedin et al. 2004). Few HSP inhibitors are known; between them, quercetin, a flavon...

Background: Autoimmune lymphoproliferative syndrome (ALPS) is a rare inherited disorder characterized by defective function of Fas, autoimmune manifestations that predominantly involve blood cells, polyclonal accumulation of lymphocytes in the spleen and lymph nodes with lymphoadenomegaly and/or splenomegaly, and expansion of TCRαβ+ CD4/CD8 double-negative (DN) T cells in the peripheral blood. Most frequently, it is due to Fas gene mutations, causing ALPS type Ia (ALPS-Ia). However, other mutations, namely of the FasL gene (ALPS-Ib) and the caspase-10 gene (ALPS-II) are occasionally detected, whereas some patients do not present any known mutations (ALPS-III). Recently, mutations of the NRAS gene have been suggested to cause ALPS-IV.

A multicentre study evaluating the presence of glycosil phosphatidyl-inositol (GPI)-negative populations was performed in 85 children with acquired aplastic anemia (AA). A GPI-negative population was observed in 41% of patients at diagnosis, 48% during immune-suppressive therapy (IST), and 45% in patients off-therapy. No association was found between the presence of a GPI-negative population at diagnosis and the response to IST. In addition, the response rate to IST did not differ between the patients who were GPI-positive at diagnosis and later developed GPI-negative populations and the 11 patients who remained GPI-positive. Two patients with a GPI-negative population >10%, and laboratory signs of hemolysis without hemoglobinuria were considered affected by paroxysmal nocturnal hemoglobinuria (PNH) secondary to AA; no thrombotic event was reported. Excluding the 2 patients with a GPI-negative population greater than 10%, we did not observe a significant correlation between LDH levels and GPI-negative population size. In this study monitoring for laboratory signs of hemolysis was sufficient to diagnose PNH in AA patients. The presence of minor GPI-negative populations at diagnosis in our series did not influence the therapeutic response. As occasionally the appearance of a GPI-negative population was observed at cyclosporine (CSA) tapering or AA relapse, a possible role of GPI-negative population monitoring during IST modulation may need further investigation.

Mycophenolate mofetil (MMF) has been shown to be effective in children with immune thrombocytopenia (ITP) and Evans syndrome (ES), but data from larger series and details on the timing of the response are lacking. We evaluated 56 children treated with MMF for ITP (n = 40) or ES (n = 16), which was primary or secondary to autoimmune lymphoproliferative syndrome -related syndrome (ARS). Thirty-five of the 54 evaluable patients (65%) achieved a partial (18%) or complete (46%) response after a median (range) of 20 (7-137) and 37 (7-192) d, respectively. ITP and ES patients responded in 58% and 81% of cases (P = not significant, ns), with complete response in 32% and 81% (P = 0·01), respectively. 60% and 73% of children with primary disease and ARS responded (P = ns) with complete response in 34% and 68% of cases (P = 0·01), respectively. Six of 35 (17%) children relapsed after a median of 283 d (range 189-1036). Limited toxicity was observed in four patients. The median durations of treatment and follow-up were seven and 12·7 months, respectively. This is the largest reported cohort of patients treated with MMF for ITP/ES. The results show that MMF is effective and safe and provides a relatively quick response, suggesting that it has a potential role as an alternative to more aggressive and expensive second/further-line treatments.