Stroke and aspirin non-responder patients: relation with hypertension and platelet response to adenosine diphosphate.

Despite its widespread use, there are many concerns about the efficacy of aspirin in the secondary prevention of cardiovascular events after stroke, leading to the concept of aspirin non-response (ANR). Although the mechanisms of ANR remain uncertain, it is expected to be due to a combination of clinical, biological and genetic characteristics affecting platelet function. In this study, we investigated whether clinical and/or biological factors such as hypertension and platelet response to ADP could contribute to the ANR. As a secondary objective, we determine whether ANR and collagen/ADP closure time (CADP-CT) could be related to platelet glycoprotein single nucleotide polymorphisms (SNPs). One hundred patients on aspirin (160 mg/day) were enrolled. ANR was measured with a platelet function analyzer (PFA-100); genotyping of four SNPs (GP IIIa, GP Ia, P2Y12 and GP VI) was performed using a tetra-primer amplification refractory mutation system. Using a collagen/epinephrine-coated cartridge on the PFA-100, the prevalence of ANR was 15% (n = 15). In the ANR group, (i) CADP-CT was significantly shorter and (ii) hypertension was an independent clinical predictive factor of ANR (OR = 4.25; 95%CI: 1.06-17.11). No clear relation was found between CADT-CT and platelet gene polymorphism as well as ANR status and SNPs. In conclusion our study confirms the independent relationship between hypertension, platelet hypersensitivity to ADP and aspirin (160 mg/day) non-response. The differential sensitivity to aspirin may have potential clinical implications, where adaptation of antiplatelet therapy is necessary according to a patient's clinical and genetic characteristics.

Mutation status and clinical outcome of 89 imatinib mesylate-resistant chronic myelogenous leukemia patients: a retrospective analysis from the French intergroup of CML (Fi(phi)-LMC GROUP).

The emergence of ABL point mutations is the most frequent cause for imatinib resistance in chronic myelogenous leukemia (CML) patients and can occur during any phase of the disease; however, their clinical impact remains controversial. In this study, we retrospectively analyzed the predictive impact of 94 BCR-ABL kinase domain mutations (18 T315I, 26 P-loop, 50 in other sites) found in 89 imatinib-resistant CML patients. At imatinib onset, 64% of patients (57/89) were in chronic phase (CP), 24% (21/89) in accelerated phase (AP) and 12% (11/89) in blastic phase (BP). T315I and P-loop mutations were preferentially discovered in accelerated phase of BP CML, and other types of mutations in CP (P=0.003). With a median follow-up of 39.2 months (6.3-67.2), since imatinib initiation, overall survival (OS) was significantly worse for P-loop (28.3 months) and for T315I (12.6 months), and not reached for other mutations (P=0.0004). For CP only, multivariate analysis demonstrated a worse OS for P-loop mutations (P=0.014), and a worse progression-free survival (PFS) for T315I mutations (P=0.014). Therefore, P-loop and T315I mutations selectively impair the outcome of imatinib-resistant CML patients, in contrast to other mutations, which may benefit from dose escalation of imatinib, able to improve or stabilize disease response.

Aspirin resistance: definitions, mechanisms, prevalence, and clinical significance.

Aspirin is the most commonly used therapeutic agent in prevention of vascular ischemic events. Aspirin exerts its antithrombotic effect primarily by interfering with the biosynthesis of thromboxane A2 (TXA2) and inhibition of TXA2 -dependent platelet aggregation. A meta-analysis of secondary prevention trials indicated that aspirin reduced major cardiovascular or cerebral events by 25%. This led to the widespread use of aspirin for prevention of cardiovascular events. However, it appears that aspirin antiplatelet effect is not uniform in all patients and previous studies estimated that 8-45% of the population were aspirin resistant. Furthermore, (i) the optimal dosage of aspirin for complete inhibition of platelet aggregation by physiological agonists (i.e arachidonic acid) is subject to great interindividual variability, (ii) the tests to detect aspirin resistance in vitro are subject to debate and (iii) the mechanisms by which some patients are resistant to aspirin in vitro remain to be determined. Despite these unresolved questions, recent clinical studies provide the reliable evidence that aspirin resistance correlates with confirmed clinical unresponsiveness, highlighting the clinical interest of determining the aspirin inhibitory effects on patients' platelets. In conclusion, discovery of aspirin resistance in individuals might be important in order to devise better anti-platelet strategies and improve our ability to prevent acute thrombotic complication.

Resistance to aspirin in vitro is associated with increased platelet sensitivity to adenosine diphosphate.

BACKGROUND: Platelet activation plays an important role in arterial thrombosis and the widespread use of aspirin has reduced major events by 25% in the secondary prevention of cardiovascular diseases. However, it appears that aspirin antiplatelet effect is not uniform and 8-45% of the population are, in vitro, aspirin resistant, and it is well recognized that platelets can be activated by pathways that are not blocked by aspirin, such as adenosine diphosphate (ADP). OBJECTIVES: To investigate whether aspirin-resistant patients have a modified sensitivity to ADP-induced platelet activation MATERIALS AND METHODS: Seventy-two patients were enrolled. Platelet function was measured by the PFA-100(R) analyser; platelet GP IIb-IIIa activation by ADP 10 micro M was assessed by flow cytometry using PAC-1 MoAb. RESULTS: Using a collagen/epinephrine coated cartridge on the PFA-100(R), the prevalence of aspirin resistance was 29.2% (n=21). For aspirin-resistant patients, the collagen/ADP coated cartridge showed a closure time significantly shorter (p=0.004) compared to the sensitive and control groups. Platelets from aspirin-resistant patients bound PAC-1 significantly more (p=0.03) than the aspirin-sensitive patients and controls when activated with 10 micro M ADP. CONCLUSIONS: Platelets from aspirin-resistant patients appear to be more sensitive and activable by ADP. This hypersensitivity could provide a possible explanation for the so-called aspirin resistance, and this could justify therapeutic improvement with alternative antiplatelet agents.

A mesenchymal glioma stem cell profile is related to clinical outcome.

Recent studies have demonstrated a relationship between the expression of stem cell-associated genes and relapses in glioblastoma (GBM), suggesting a key role for tumor stem cells in this process. Although there is increasing interest in this field, glioma stem cells (GSCs) are still poorly characterized, their 'stemness' state and factors maintaining these properties remain largely unknown. We performed an expression profiling analysis of pluripotency in gliomaspheres derived from 11 patients. Comparative analysis between GSCs and H1 and H9 human embryonic stem cells as well as H9-derived neural stem cells indicates major variations in gene expression of pluripotency factors Nanog and OCT4, but a stable pattern for SOX2 suggesting its important function in maintaining pluripotency in GSCs. Our results also showed that all GSC lines have the capacity to commit to neural differentiation and express mesenchymal or endothelial differentiation markers. In addition, hierarchical clustering analysis revealed two groups of GSCs reflecting their heterogeneity and identified COL1A1 and IFITM1 as the most discriminating genes. Similar patterns have been observed in tumors from which gliomaspheres have been established. To determine whether this heterogeneity could be clinically relevant, the expression of both genes was further analyzed in an independent cohort of 30 patients with GBM and revealed strong correlation with overall survival. In vitro silencing of COL1A1 and IFTM1 confirmed the effect of these mesenchymal-associated genes on cell invasion and gliomasphere initiation. Our results indicate that COL1A1 and IFITM1 genes could be considered for use in stratifying patients with GBM into subgroups for risk of recurrence at diagnosis, as well as for prognostic and therapeutic evolution.

Evidence of ABL-kinase domain mutations in highly purified primitive stem cell populations of patients with chronic myelogenous leukemia.

To study the hierarchical levels of stem cell targets for ABL-kinase domain mutations in CML, highly purified CD34+CD38- and CD34+CD38+ cell populations and their LTC-IC-derived progeny were analyzed in four patients at diagnosis (n=1) or in advanced phases (n=3) of their disease. In the single patient with early phase CML who later developed an Imatinib Mesylate-resistance and a Y253H mutation, no mutation was detectable in purified cell fractions analyzed at diagnosis nor in their LTC-IC-derived progeny. In contrast, in three patients in advanced phase CML, ABL-kinase mutations demonstrated in peripheral blood cells by sequencing (Q252E and M351T) were detectable in the FACS-sorted cells and became amplified in the LTC-IC-derived progeny of the primitive cells. These findings demonstrate that in late CP or advanced CML, ABL-kinase mutations occur as an intraclonal event in the primitive Ph1+ stem cell compartments with progression of this clone towards IM-resistant blast phase.

Development of an immunomagnetic separation-polymerase chain reaction (IMS-PCR) assay specific for Enterocytozoon bieneusi in water samples.

AIMS: Microsporidia have become widely recognized as important human pathogens. Among Microsporidia, Enterocytozoon bieneusi is responsible for severe gastrointestinal disease. To date, no current therapy has been proven effective. Their mode of transmission and environmental occurrence are poorly documented because of the lack of detection methods that are both species-specific and sensitive. In this study, we developed a sensitive and specific molecular method to detect E. bieneusi spores in water samples. METHODS AND RESULTS: The molecular assay combined immunomagnetic separation (IMS) and polymerase chain reaction (PCR) amplification to detect E. bieneusi spores. A comparison was made of IMS magnetic beads coated with two different monoclonal antibodies, one specific for the Encephalitozoon genus that cross-reacts with E. bieneusi and the other specific only for the E. bieneusi species itself. CONCLUSIONS: Immunotech beads coated with the antibody specific for E. bieneusi were found to be the most effective combination. SIGNIFICANCE AND IMPACT OF THE STUDY: The highly specific IMS-PCR assay developed in this study provides a rapid and sensitive means of screening water samples for the presence of E. bieneusi spores.