Cytogenetic response to imatinib in chronic myeloid leukaemia patients: concentration dependency and associated influences under field-conditions

Introduction: Imatinib, a first-line drug for chronic myeloid leukaemia (CML), has been increasingly proposed for therapeutic drug monitoring (TDM), as trough concentrations >=1000 ng/ml (Cmin) have been associated with improved molecular and complete cytogenetic response (CCyR). The pharmacological monitoring project of EUTOS (European Treatment and Outcome Study) was launched to validate retrospectively the correlation between Cmin and response in a large population of patients followed by central TDM in Bordeaux.¦Methods: 1898 CML patients with first TDM 0-9 years after imatinib initiation, providing cytogenetic data along with demographic and comedication (37%) information, were included. Individual Cmin, estimated by non-linear regression (NONMEM), was adjusted to initial standard dose (400 mg/day) and stratified at 1000 ng/ml. Kaplan-Meier estimates of overall cumulative CCyR rates (stratified by sex, age, comedication and Cmin) were compared using asymptotic logrank k-sample test for interval-censored data. Differences in Cmin were assessed by Wilcoxon test.¦Results: There were no significant differences in overall cumulative CCyR rates between Cmin strata, sex and comedication with P-glycoprotein inhibitors/inducers or CYP3A4 inhibitors (p >0.05). Lower rates were observed in 113 young patients <30 years (p = 0.037; 1-year rates: 43% vs 60% in older patients), as well as in 29 patients with CYP3A4 inducers (p = 0.001, 1-year rates: 40% vs 66% without). Higher rates were observed in 108 patients on organic-cation-transporter-1 (hOCT-1) inhibitors (p = 0.034, 1-year rates: 83% vs 56% without). Considering 1-year CCyR rates, a trend towards better response for Cmin above 1000 ng/ml was observed: 64% (95%CI: 60-69%) vs 59% (95%CI: 56-61%). Median Cmin (400 mg/day) was significantly reduced in male patients (732 vs 899ng/ml, p <0.001), young patients <30 years (734 vs 802 ng/ml, p = 0.037) and under CYP3A4 inducers (758 vs 859 ng/ml, p = 0.022). Under hOCT-1 inhibitors, Cmin was increased (939 vs 827 ng/ml, p = 0.038).¦Conclusion: Based on observational TDM data, the impact of imatinib Cmin >1000 ng/ml on CCyR was not salient. Young CML patients (<30 years) and patients taking CYP3A4 inducers probably need close monitoring and possibly higher imatinib doses, due to lower Cmin along with lower CCyR rates. Patients taking hOCT-1 inhibitors seem in contrast to have improved CCyR response rates. The precise role for imatinib TDM remains to be established prospectively.

Role of myosin V in actin cable organization in fission yeast

Functional specialization is tightly linked to the ability of eukaryotic cells to acquire a particular shape. Cell morphogenesis, in turn, relies on the capacity to establish and maintain cell "polarity", which is achieved by orienting the trafficking of signaling molecules and organelles towards specific cellular locations and/or membrane domains. The "oriented" transport is based upon cytoskeletal polymers, microtubules and actin filaments, which serve as tracks for molecular motors. These latter generate motion that is translated either into pulling forces or directed transport. Fission yeast, a rod-like unicellular eukaryote, shapes itself by restricting growth at cell tips through the concerted activity of microtubules and actin cables. Microtubules, which assemble into 2-6 bundles and run parallel to the long axis of the cell, serve to orient growth to the tips. Growth is supported by the actin cytoskeleton, which provides tracks, the cables, for motor-based transport of secretory vesicles. The molecular motors, which bind cargos and deliver them to the tips along cables, are also known as type V myosins (hereafter indicated as myosin V). How the bundles of parallel actin filaments, i.e. the cables, extend from the tips through the cell and whether they serve any other purpose, besides providing tracks, is poorly understood. It is also unclear how the crosstalk between the two cytoskeletal systems is achieved. These are the basic questions I addressed during my PhD. The first part of the thesis work (Chapter two) suggests that the sole function of actin cables in polarized growth is to serve as tracks for motors. The data indicate that cells may have evolved two cytoskeletal systems to provide robustness to the polarization process but in principle a unique cytoskeleton might have been able to direct and support polarized growth. How actin cables are organized within the cell to optimize cargo transport is addressed later on (Chapter three). The major finding, based on the actin cable defect of cells lacking myosin Vs, is that actin filaments self-organize through the activity of the transport motors. In fact, by delivering cargos to cell tips and exerting physical pulling forces on actin filaments, Myosin Vs contribute not only to polarize cargo transport but also actin tracks. Among the cargos transported by Myosin V, which may be relevant to its function in organizing cables, there is likely the endoplasmic reticulum (ER). Actin cables, which run parallel to cortical ER, may serve as tracks for Myosin V. Myosin V-driven displacement, in turn, may account for the dynamic expansion and organization of ER during polarized growth as suggested in Chapter four. The last part of the work (Chapter five) highlights the existence of a crosstalk between actin and microtubules. In absence of myosin V, indeed, microtubules contribute to actin cable organization, likely playing a scaffolding/tethering function. Whether or not the kinesin 1, Klp3, plays any role in such process has to be demonstrated. In conclusion the work proposes a novel role for myosin Vs in actin organization, besides its transport function, and provides molecular tools to further dissect the role of this type of myosin in fission yeast. - La spécialisation fonctionnelle est étroitement connectée à la capacité des cellules eucaryotes d'acquérir une forme particulière. La morphogenèse cellulaire à son tour, est basée sur la capacité d'établir et de maintenir la polarité cellulaire, polarité réalisée en orientant le trafic des molécules signales et des organelles vers des zones cellulaires spécifiques. Ce transport directionnel dépend des polymères du cytosquelette, microtubules et microfilaments, qui servent comme des voies pour les moteurs moléculaires. Ces derniers engendrent du mouvement, traduit soit en force de traction soit en transport directionnel. La levure fissipare, un eucaryote unicellulaire en forme de bâtonnet, acquière sa forme en limitant sa croissance aux extrémités par l'action concertée des microtubules et de l'actine. Les microtubules, qui s'assemblent de façon antiparallèle et parcourent la cellule parallèlement à l'axe longitudinal, servent à orienter la croissance aux extrémités. Cette croissance est permise par le cytosquelette d'actine, fournissant des voies, les câbles, pour le transport actif des vésicules de sécrétion. Les moteurs moléculaires, responsables de ce transport actif sont aussi appelés myosines de type V (par la suite appelés myosines V). La manière dont ces câbles s'étendent depuis l'extrémité jusqu'à l'intérieur de la cellule est peu connue. De plus, on ignore également si ces câbles présentent une fonction autre que le transport. L'interaction entre les deux cytosquelettes est également obscure. Ce sont ces questions de base auxquelles j'ai tenté de répondre lors de ma thèse. La première partie de cette thèse (chapitre II) suggère que les câbles d'actine, pendant la croissance polarisée, fonctionnent uniquement comme des voies pour les moteurs moléculaires. Les données indiqueraient que les cellules ont fait évoluer deux systèmes de cytosquelette pour assurer plus de robustesse au processus de polarisation, bien que, comme nous le verrons, un système unique est suffisant. Au chapitre III, nous verrons comment les câbles d'actine sont organisés à l'intérieur de la cellule afin d'optimiser le transport des cargo. La découverte majeure, réalisée en observant des cellules dont la myosine V fait défaut, est que ces filaments d'actine s'auto organisent grâce au passage des moteurs moléculaires le long de ces voies. En réalité, en délivrant les cargos aux extrémités de la cellule et en exerçant des forces de traction sur les câbles, les myosines V contribuent non seulement à polariser le transport mais également à polariser les voies elles mêmes. Nous verrons également au chapitre IV, que parmi les cargos importants pour l'organisation des câbles, il y aurait le réticulum endoplasmique (RE). En effet, les câbles d'actine, qui s'étalent parallèlement au RE cortical, pourraient servir comme voie pour la myosine V. Cette dernière en retour pourrait être responsable de l'expansion dynamique et de l'organisation du RE pendant la croissance polarisée.

Aging effects on selective attention-related electroencephalographic patterns during face encoding.

Previous electrophysiological studies revealed that human faces elicit an early visual event-related potential (ERP) within the occipito-temporal cortex, the N170 component. Although face perception has been proposed to rely on automatic processing, the impact of selective attention on N170 remains controversial both in young and elderly individuals. Using early visual ERP and alpha power analysis, we assessed the influence of aging on selective attention to faces during delayed-recognition tasks for face and letter stimuli, examining 36 elderly and 20 young adults with preserved cognition. Face recognition performance worsened with age. Aging induced a latency delay of the N1 component for faces and letters, as well as of the face N170 component. Contrasting with letters, ignored faces elicited larger N1 and N170 components than attended faces in both age groups. This counterintuitive attention effect on face processing persisted when scenes replaced letters. In contrast with young, elderly subjects failed to suppress irrelevant letters when attending faces. Whereas attended stimuli induced a parietal alpha band desynchronization within 300-1000 ms post-stimulus with bilateral-to-right distribution for faces and left lateralization for letters, ignored and passively viewed stimuli elicited a central alpha synchronization larger on the right hemisphere. Aging delayed the latency of this alpha synchronization for both face and letter stimuli, and reduced its amplitude for ignored letters. These results suggest that due to their social relevance, human faces may cause paradoxical attention effects on early visual ERP components, but they still undergo classical top-down control as a function of endogenous selective attention. Aging does not affect the face bottom-up alerting mechanism but reduces the top-down suppression of distracting letters, possibly impinging upon face recognition, and more generally delays the top-down suppression of task-irrelevant information.

Need of reduced and harmonized bureaucracy in multi-centre clinical research - a case-report from a Swiss trial

Introduction: We launched an investigator-initiated study (ISRCTN31181395) to evaluate the potential benefit of pharmacokinetic-guided dosage individualization of imatinib for leukaemiapatients followed in public and private sectors. Following approval by the research ethics committee (REC) of the coordinating centre, recruitment throughout Switzerland necessitatedto submit the protocol to 11 cantonal RECs.Materials and Methods: We analysed requirements and evaluation procedures of the 12 RECs with associated costs.Results: 1-18 copies of the dossier, in total 4300 printed pages, were required (printing/posting costs: ~300 CHF) to meet initial requirements. Meeting frequencies of RECs ranged between 2 weeks and 2 months, time from submission to fi rst feedback took 2-75 days. Study approval was obtained from a chairman, a subor the full committee, the evaluation work being invoiced by0-1000 CHF (median: 750 CHF, total: 9200 CHF). While 5 RECs gave immediate approval, the other 6 rose in total 38 queries before study release, mainly related to wording in the patient information, leading to 7 different fi nal versions approved. Submission tasks employed an investigator half-time over about 6 months.Conclusion: While the necessity of clinical research evaluation by independent RECs is undisputed, there is a need of further harmonization and cooperation in evaluation procedures. Current administrative burden is indeed complex, time-consuming and costly. A harmonized electronic application form, preferably compatible with other regulatory bodies and European countries, could increase transparency, improve communication, and encourage academic multi-centre clinical research in Switzerland.

Once-daily dolutegravir is superior to once-daily darunavir/ritonavir in treatment-naïve HIV-1-positive individuals: 96 week results from FLAMINGO.

INTRODUCTION: Dolutegravir (DTG) 50 mg once daily was superior to darunavir/ritonavir (DRV/r) 800 mg/100 mg once daily through Week 48, with 90% vs. 83% of participants achieving HIV RNA 50 c/mL (p=0.025) [1]. We present data through Week 96. MATERIAL AND METHODS: FLAMINGO is a multicentre, randomized, open-label, Phase IIIb non-inferiority study, in which HIV-1-positive ART-naïve adults with HIV-1 RNA≥1000 c/mL and no evidence of viral resistance were randomized 1:1 to receive DTG or DRV/r, with investigator-selected backbone NRTIs (TDF/FTC or ABC/3TC). Participants were stratified by screening HIV-1 RNA (≤100K c/mL) and NRTI backbone. RESULTS: A total of 484 adults were randomized and treated; 25% had baseline HIV RNA 100K c/mL. At Week 96, the proportion of participants with HIV RNA 50 c/mL was 80% in the DTG arm vs. 68% in the DRV/r arm (adjusted difference 12.4%; 95% CI 4.7, 20.2%; p=0.002). Secondary analyses supported primary results: per-protocol [(DTG 83% vs. DRV/r 70%), 95% CI 12.9 (5.3, 20.6)] and treatment-related discontinuation = failure [(98% vs. 95%), 95% CI 3.2 (-0.3, 6.7)]. Overall virologic non-response (DTG 8%; DRV/r 12%) and non-response due to other reasons (DTG 12%; DRV/r 21%) occurred less frequently on DTG. As at Week 48, the difference between arms was most pronounced in participants with high baseline viral load (82% vs. 52% response through Week 96) and in the TDF/FTC stratum (79% vs. 64%); consistent responses were seen in the ABC/3TC stratum (82% vs. 75%). Six participants (DTG 2, none post-Week 48; DRV/r 4, two post-Week 48) experienced protocol-defined virologic failure (PDVF; confirmed viral load 200 c/mL on or after Week 24); none had treatment-emergent resistance to study drugs. Most frequent drug-related adverse events (AEs) were diarrhoea, nausea and headache, with diarrhoea significantly more common on DRV/r (24%) than DTG (10%). Significantly more participants had Grade 2 fasting LDL toxicities on DRV/r (22%) vs. DTG (7%), p<0.001; mean changes in creatinine for DTG (~0.18 mg/dL) observed at Week 2 were stable through Week 96. CONCLUSIONS: Once-daily DTG was superior to once-daily DRV/r in treatment-naïve HIV-1-positive individuals, with no evidence of emergent resistance to DTG in virologic failure and relatively similar safety profiles for DTG and DRV/r through 96 Weeks.

Effect of an I.V. bolus of phenylephrine or ephedrine on skin blood flow during spinal anesthesia

Effet d'un bolus intraveineux de phénylephrine ou d'éphedríne sur le flux sanguin cutané lors d'une anesthésie rachidienne Introduction : La phénylephrine et l'éphedrine sont des substances vaso-actives utilisées de routine pour corriger des épisodes d'hypotension artérielle induits par l'anesthésie intrarachidienne. L'influence de ces deux vasopresseurs sur le flux sanguin cutané (FSC) dans ce contexte n'a jusqu'à maintenant pas été décrite. Cette étude évalue l'effet d'une injection intraveineuse de 75 µg de phénylephrine ou de 7.5 mg d'éphedrine sur le FSC mesuré par Laser Doppler, dans les zones concernées parle bloc sympathiqué induit par l'anesthésie intrarachidienne (membres inférieurs) et dans les zones non concernées (membres supérieurs). Méthode :Après acceptation par le Comité d'Éthique, et obtention de leur accord écrit, 20 patients devant subir une intervention chirurgicale élective en décubitus dorsal sous anesthésie. intrarachidienne ont été inclus dans cette étude randomisée en double insu. Le FSC a été mesuré en continu par deux sondes fixées l'une à la cuisse (zone avec bloc sympathique) et l'autre sur l'avantbras (zone sans bloc sympathique). Les valeurs de FSC ont été enregistrées après l'anesthésie rachidienne (valeur contrôle), puis après l'injection i.v. dè phénylephrine (10 patients) ou d'éphedrine (10 patients) pour corriger une hypotension définie comme une chute de 20 mmHg de la pression artérielle systolique. Les variations de FSC exprimées en pourcentage de la valeur contrôle moyenne (+/- écart type) ont été analysées par le test t de Student. Résultats :Les données démographiques des patients et le niveau sensitif induit par l'anesthésie rachidienne sont similaires dans les deux groupes. Aux doses utilisées, seule l'éphedrine restaure la pression artérielle aux valeurs précédant l'anesthésie rachidienne. La phénylephrine augmente le FSC de l'avant-bras de 44% (+/- 79%) et de la cuisse de 34% (+/-24%), alors que l'éphedrine diminue le débit sanguin cutané de l'avant-bras de 16% (+/- 15%) et de la cuisse de 22% (+/-11%). Conclusion : L'injection intraveineuse de phénylephrine et d'éphedrine ont des effets opposés sur le flux sanguin cutané, et cette réponse n'est pas modifiée par le bloc sympathique.. Cette différence peut s'expliquer par la distribution des sous-types de récepteurs adrénergiques alpha et leur prédominance relative dans les veines et les artères de différents diamètres perfusant le tissu sous-cutané et la peau. L'éphedrine, èn raison de sa meilleure efficacité pour traiter les épisodes d'hypotension artérielle après anesthésie intrarachidienne devrait être préféré à la phénylephrine, leurs effets opposés sur le flux sanguin cutané n'étant pas pertinents en pratique clinique. SUMMARY Background: Phenylephrine or ephedrine is routinely used to correct hypotensive episodes fallowing spinal anaesthesia (SA). The influence of these two vasopressors on skin blood flow (SBF) has not yet been described. We have therefore evaluated the effects of an i.v. bolus of 75 µg phenylephrine or 7.5 mg of ephedrine on SBF measured by laser Doppler flowmetry during sympathetic blockade induced by SA. Methods: With Ethical Committee approval and written consent, 20 patients scheduled for elective procedures in supine position under SA were enrolled in this double-blind randomized study. SBF was measured continuously by two probes fixed at the thigh (area with sympathic blockade) and forearm level (area without sympathic blockade) respectively. SBF values were recorded after SA (control values) and then after a bolus administration of phenylephriné (n=10) or ephedrine (n=10) when systolic blood pressure decreased by 20 mmHg. Changes were expressed as percentage of control SBF values and analysed by Student's paired t-test. Results: Patient characteristics and dermatomal sensory levels were similar in both groups. Phenylephrine increases mean SBF at the forearm level by 44% (79%) [mean (SD)j and at the thigh by 34% (24%). Ephedrine decreases SBF at the forearm level by 16% (15%) and at the thigh by 22% (il%). Ephedrine bolus restores arterial blood pressure to pre-anaesthesia values, whereas phenylephrine does not. Conclusion: Administratión of phenylephrine and ephedrine has opposite effects on skin blood flow and sympathetic blockade does not modify this response. These findings could be explained by the distribution of the alpha-adrenoréceptor subtypes and their relative predominance among veins and arteries of different size perfusing the subcutaneous tissue and the skin. Ephedrine, due to its better efficacy to correct hypotensive episodes following SA, should be preferred, to phenylephrine, their opposite effects on SBF being not relevant for clinical practice.

"Marxisme et philosophie du langage" (1929) de V. Volochinov et sa réception chez R. Chor : deux voies du développement de la science du langage «marxiste» dans les années 1920 en Russie

Malgré le succès de  « Marxisme et philosophie du langage » (par la suite MPL) de V. Volochinov, ce texte n'a suscité que trois comptes rendus. Le quatrième est ébauché par A. Romm. Resté à l'état de notes, ce compte rendu est bien étudié par les chercheurs contemporains. En revanche, le compte rendu écrit par R. Chor n'a jamais été analysé de façon détaillée. Notre article a pour but de combler cette lacune et porte sur le contenu de l'analyse critique de MPL faite par Chor. Son examen révèle que le compte rendu écrit par cette dernière et MPL de Volochinov représentent la formulation de principes de base diamétralement opposés de la science du langage «marxiste». Cela se manifeste dans la réception des idées de F. de Saussure. Si Chor considère la conception de ce dernier comme fondement de la linguistique «théorique» portant sur la langue en tant que système de signes (ou de «signifiances» dans les termes de Chor), Volochinov rejette la théorie de Saussure. Il la considère comme «abstraite», d'où le nom «objectivisme abstrait» qu'il donne au saussurisme. En élaborant la philosophie du langage «marxiste», il s'appuie sur la néo-philologie idéaliste (les idées de Vossler et de son Ecole) définie dans MPL comme «subjectivisme individualiste». Volochinov propose d'étudier la «langue» comme fait réel, accessible à l'observation immédiate et non pas comme objet d'étude «théorique». Ce fait constitue une cible de la critique adressée par Chor à Volochinov

The V-region disease hypothesis: evidence from autoimmune encephalomyelitis.

Experimental allergic encephalomyelitis has been shown to have an immunological basis. In fact, the disease can be induced by T cells specific for myelin basic protein, a molecule found in abundance in the central nervous system. In this article, Ellen Heber-Katz and Hans Acha-Orbea discuss the T-cell receptor (TCR) repertoire of the encephalitogenic T-cell response, and show that a limited V gene pool, in fact a single V beta and two V alpha families, are being used by the PL/J and B10.PL mice and by every rat strain examined, even though the antigenic determinants and the major histocompatibility complex (MHC) molecules are different in all cases. This extraordinary finding suggests that the TCR is involved in encephalitogenicity in a way that not only involves the recognition of antigen in association with MHC, but also as an effector molecule that results in encephalitis. If this is true, it implies that TCRs, in general, play more than one role in mammalian physiology.

Manganese-induced integrin affinity maturation promotes recruitment of alpha V beta 3 integrin to focal adhesions in endothelial cells: evidence for a role of phosphatidylinositol 3-kinase and Src.

Integrin activity is controlled by changes in affinity (i.e. ligand binding) and avidity (i.e. receptor clustering). Little is known, however, about the effect of affinity maturation on integrin avidity and on the associated signaling pathways. To study the effect of affinity maturation on integrin avidity, we stimulated human umbilical vein endothelial cells (HUVEC) with MnCl(2) to increase integrin affinity and monitored clustering of beta 1 and beta 3 integrins. In unstimulated HUVEC, beta 1 integrins were present in fibrillar adhesions, while alpha V beta 3 was detected in peripheral focal adhesions. Clustered beta 1 and beta 3 integrins expressed high affinity/ligand-induced binding site (LIBS) epitopes. MnCl(2)-stimulation promoted focal adhesion and actin stress fiber formation at the basal surface of the cells, and strongly enhanced mAb LM609 staining and expression of beta 3 high affinity/LIBS epitopes at focal adhesions. MnCl(2)-induced alpha V beta 3 clustering was blocked by a soluble RGD peptide, by wortmannin and LY294002, two pharmacological inhibitors of phosphatidylinositol 3-kinase (PI 3-K), and by over-expressing a dominant negative PI 3-K mutant protein. Conversely, over-expression of active PI 3-K and pharmacological inhibiton of Src with PP2 and CGP77675, enhanced basal and manganese-induced alpha V beta 3 clustering. Transient increased phosphorylation of protein kinase B/Akt, a direct target of PI 3K, occurred upon manganese stimulation. MnCl(2) did not alter beta 1 integrin distribution or beta1 high-affinity/LIBS epitope expression. Based on these results, we conclude that MnCl(2)-induced alpha V beta 3 integrin affinity maturation stimulates focal adhesion and actin stress fiber formation, and promotes recruitment of high affinity alpha V beta 3 to focal adhesions. Affinity-modulated alpha V beta 3 clustering requires PI3-K signaling and is negatively regulate by Src.

Molecular basis of coagulation factor V deficiency caused by the R1698W inter-domain mutation.

Coagulation factor V (FV) deficiency is characterised by variable bleeding phenotypes and heterogeneous mutations. To add new insights into the FV genotype-phenotype relationship, we characterised the R1698W change in the A3 domain, at the poorly investigated interface with the A2 domain. The FV R1698W mutation was responsible for a markedly reduced expression level (10% of FV-WT) and specific activity in thrombin generation (0.39). Interestingly, the FVa1698W showed rapid activity decay upon activation due to increased dissociation rate between the heavy and light chains. The importance of the size and charge of the residue at position 1698 was investigated by three additional recombinant mutants, FVR1698A, FVR1698Q, and FVR1698E. FVR1698A and FVR1698Q expression (30 and 45% of FV-WT), specific activity (both 0.57) and stability were all reduced. Noticeably, FVR1698E showed normal activity and stability despite poor expression (10% of FV-WT). These data indicate the essential role of R1698 for normal biosynthetic process and support local flexibility for positively or negatively charged residues to produce stable and functional A3-A2 domain interactions. Their experimental alteration produces a gradient of FV defects, which help to interpret the wide spectrum of phenotypes in FV-deficient patients.