Agranulocytosis under biotherapy in rheumatoid arthritis: three cases hypothesis of parvovirus B19 involvement in agranulocytosis observed under tocilizumab and rituximab for the treatment of rheumatoid arthritis.

Leukopenia is a considerably common complication of tocilizumab [TCZ] and rituximab [RTX] therapy. RTX-induced leukopenia typically exhibits delayed onset. While agranulocytosis has been reported linked to RTX treatment of lymphoma, this complication rarely occurs in rheumatoid arthritis (RA) treatment and, to our knowledge, has never been reported in association with TCZ therapy. We herein report four agranulocytosis cases in three patients, with the first two cases suspected to be secondary to human parvovirus B19 (PVB19) infection. Agranulocytosis manifested in the first patient 2 months following a third RTX course. Bone marrow (BM) polymerase chain reaction (PCR) was positive for PVB19. The patient relapsed after three TCZ courses, with her PCR again positive for PVB19. Both episodes resolved under granulocyte-macrophage colony-stimulating factor (GM-CSF). In the second patient, agranulocytosis manifested after the 74th TCZ course. Bone marrow PCR was positive for PVB19, and the evolution was favorable under intravenous immunoglobulin administration. The third case was a 53-year-old female patient with seropositive RA who presented agranulocytosis after the first infusion of her fourth RTX course. Unfortunately, no PCR PVB19 was made on myelogram. Evolution was favorable after 5 days of GM-CSF. PVB19 infection should be investigated in patients suffering from agranulocytosis manifesting during biotherapy. In cases manifesting from the 15th day of RTX treatment onwards, hemogram must be conducted before readministering the infusion.

Identification of a novel HLA allele, HLA-B*53:39, in a Guadaloupean individual.

The novel HLA-B*53:39 allele differs from HLA-B*53:01 by a single nucleotide substitution at codon 45 (ACG>AAG).

Can we identify response markers to antihypertensive drugs? First results from the IDEAL Trial.

Current antihypertensive strategies do not take into account that individual characteristics may influence the magnitude of blood pressure (BP) reduction. Guidelines promote trial-and-error approaches with many different drugs. We conducted the Identification of the Determinants of the Efficacy of Arterial blood pressure Lowering drugs (IDEAL) Trial to identify factors associated with BP responses to perindopril and indapamide. IDEAL was a cross-over, double-blind, placebo-controlled trial, involving four 4-week periods: indapamide, perindopril and two placebo. Eligible patients were untreated, hypertensive and aged 25-70 years. The main outcome was systolic BP (SBP) response to drugs. The 112 participants with good compliance had a mean age of 52. One in every three participants was a woman. In middle-aged women, the SBP reduction from drugs was -11.5 mm Hg (indapamide) and -8.3 mm Hg (perindopril). In men, the response was significantly smaller: -4.8 mm Hg (indapamide) and -4.3 (perindopril) (P for sex differences 0.001 and 0.015, respectively). SBP response to perindopril decreased by 2 mm Hg every 10 years of age in both sexes (P=0.01). The response to indapamide increased by 3 mm Hg every 10 years of age gradient in women (P=0.02). Age and sex were important determinants of BP response for antihypertensive drugs in the IDEAL population. This should be taken into account when choosing drugs a priori.

[Conservation and destruction of autologous and allogeneic cryopreserved cellular products: recommendations from the SFGM-TC]. / Modalités de conservation et de destruction des produits cellulaires cryopréservés : recommandations de la SFGM-TC.

Thousands of autologous and at less extent allogeneic hematopoietic stem cells (HSC) bags are cryopreserved in France. The majority of autologous HSC grafts are used within a year after collection. However, many bags are still unused and cryopreserved for many years. In France and on a European scale, the ever-growing number of cryopreserved bags represents a real economic health concern. Indeed, the cost of storage is about 100€ per bag and per year. In addition, quality and therapeutic value of these long-term cryopreserved grafts needs to be evaluated. In the attempt to harmonize clinical practices between different French transplantation centers, the French Society of Bone Marrow Transplantation and Cell Therapies (SFGM-TC) set up its fourth annual series of workshops which brought together practitioners from its member centers across France. These workshops took place in September 2013 in Lille. In this article, we addressed the issue of the destruction of long-term cryopreserved grafts be them autologous or allogeneic and provide recommendations regarding their destruction.

[Suicide attempts of 48 children aged 6-12 years]. / Tentatives de suicide chez 48 enfants âgés de 6 à 12ans.

AIM: Research is limited on suicide attempts in children under 13 years of age. The objective of this study was to provide an in-depth description of this population. MATERIALS AND METHODS: The present study is both retrospective and descriptive. Data were collected retrospectively from a file containing the causes for hospitalization of each child admitted into the Department of Child Psychiatry at the hôpital Femme-Mère-Enfant (hospices civils de Lyon). We included all patients under 13 years of age who were hospitalized for a suicide attempt between 2008 and 2011. The methods used to collect the medical records consisted in using a form made up of four major parts: suicide attempts, social environment, medical history, and therapy. RESULTS: The 26 girls and 22 boys included had a mean age of 11.52 years. The boys were younger than the girls (P=0.047) and their parents were usually separated (P=0.034). The boys used more violent means to commit suicide in comparison to the girls (P=0.048). On average, children using violent means were younger (P=0.013). Boys underwent more psychotherapy (P=0.027) and were prescribed more psychotropic medication in comparison to girls (P=0.051). Adjustment disorders (37.5%) and depression (27%) were the two main diagnoses for hospitalization. They were hospitalized on average (±standard deviation) 9.6 days (±10 days). Psychotherapy was organized when leaving the hospital (98%) with legal measures (8.3%), change of residence (12.5%), and prescription of psychotropic drugs (37.5%). None had physical complications. DISCUSSION: In children under 13 years of age, attempted suicide was more frequent in girls than boys. However, the sample included 18 girls and nine boys who were 12 years old (sex ratio of 12-year-olds, 0.5). There were more boys (16 boys/eight girls) in the children under 12 (sex ratio of 8- to 11-year-olds, 1.6). Children under 11 used more violent means (P=0.01). The literature also reports that more violent means lead to a greater risk of death by suicide. Consequently, suicidal behavior in children under 11 years of age is closer to a behavior of a person who has committed suicide than an adolescent attempting suicide. As a result of the sex ratio and non-violent means, 12-year-old children's behavior can be considered like that of adolescents. One factor that could explain children's attempted suicide is family cohesion. The children in this study were most often from broken families and had a difficult relationship with their parents. From 1981 to 1985, more than 50% of children who consulted for their first suicide attempt were not hospitalized. Now hospitalization is recommended for all children who consult for attempted suicide. They are hospitalized on average 8.9-9 days. Individual psychotherapy is systematic. The main difference between the treatments for adolescents and children is the importance of the social worker who will require legal measures or changing residences when necessary. CONCLUSION: The sex ratio in 6- to 12-year-olds attempting suicide is higher than the sex ratio in adolescents attempting suicides. Insecure attachment was found in all families in this sample. This population is particularly at risk knowing that in adulthood, the risk of death by suicide is higher when there is a background of attempted suicide by violent methods. These children should always be hospitalized for a psychological and socioenvironmental evaluation.

[How to handle unexpected biological abnormalities observed in the pre-donation workup for hematopoietic stem cell transplantation: an SFGM-TC report on pre-transplant cytomegalovirus, Epstein-Barr virus, Toxoplasma gondii, or syphilis IgM positive serology test]. / Conduite à tenir devant une anomalie biologique découverte lors du bilan pré-don cellules souches hématopoïétiques: sérologie IgM positive pour le cytomégalovirus, le virus d'Epstein-Barr, la toxoplasmose, ou la syphilis.

In the attempt to harmonize clinical practices between different French transplantation centers, the French Society of Bone Marrow Transplantation and Cell Therapy (SFGM-TC) set up the third annual series of workshops which brought together practitioners from all member centers and took place in October 2012 in Lille. Here we report our results and recommendations regarding the management of pre-transplant donor's cytomegalovirus, Epstein-Barr virus, Toxoplasma gondii, or syphilis IgM positive serology test.

[How to handle unexpected biological abnormalities observed in the pre-donation workup for hematopoietic stem cell transplantation: an SFGM-TC report on pre-transplant positive pregnancy test and monoclonal gammapathy]. / Conduite à tenir devant un problème du donneur de cellules souches hématopoïétiques: test de grossesse positif et gammapathie monoclonale.

In the attempt to harmonize clinical practices between different French transplantation centers, the French Society of Bone Marrow Transplantation and Cell Therapy (SFGM-TC) set up the third annual series of workshops which brought together practitioners from all member centers and took place in October 2012 in Lille. Here we report our results and recommendations regarding the management of common issues related to the donor: pre-transplant pregnancy and monoclonal gammopathy.

Long-term follow-up of corticosteroid refractory acute GVHD treated with an Inolimomab-based algorithm: a single center experience.

Acute corticosteroid refractory GVHD (aGVHD) remains a challenging problem after allogeneic hematopoietic SCT. Even though immunosuppressive therapies may achieve a response, unsatisfactory aGVHD control and toxicity of high cumulative doses of corticosteroids are frequent, notably with an increased infection rate. We report long-term follow-up of 33 consecutive patients who developed corticosteroid refractory aGVHD in our institution, treated homogeneously according to a unique algorithm combining an induction treatment (Inolimomab, 0.3 mg/kg per day), an associated immunosuppression (Mycophenolate Mofetil) and a predefined management of partial responses (PR) by the switch from Cyclosporin to Tacrolimus, together with an intensive infectious monitoring and supportive care. In this cohort, 17 patients (52%) achieved a complete response (CR) and 14 patients (42%) a PR, which converted to CR for 12 patients after Tacrolimus introduction. Transplant related mortality (TRM) was 15.5% and 29.7% at 1 and 3 years, respectively. OS was 54.5% at 3 years. Multivariate analysis identified CR after Inolimomab therapy as the unique prognostic factor on OS. Among the 30 evaluable patients, 19 (63%) developed extensive chronic GVHD. This Inolimomab-based algorithm allows for an efficient control of corticosteroid refractory aGVHD in a high proportion of patients with low toxicity, and deserves further investigation.

Placental transfer of maraviroc in an ex vivo human cotyledon perfusion model and influence of ABC transporter expression.

Nowadays, antiretroviral therapy is recommended during pregnancy to prevent mother-to-child transmission of HIV. However, for many antiretroviral drugs, including maraviroc, a CCR5 antagonist, very little data exist regarding placental transfer. Besides, various factors may modulate this transfer, including efflux transporters belonging to the ATP-binding cassette (ABC) transporter superfamily. We investigated maraviroc placental transfer and the influence of ABC transporter expression on this transfer using the human cotyledon perfusion model. Term placentas were perfused ex vivo for 90 min with maraviroc (600 ng/ml) either in the maternal-to-fetal (n = 10 placentas) or fetal-to-maternal (n = 6 placentas) direction. Plasma concentrations were determined by ultra performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS). Fetal transfer rates (FTR) and clearance indexes (CLI) were calculated as ratios of fetal to maternal concentrations at steady state (mean values between 30 and 90 min) and ratios of FTR of maraviroc to that of antipyrine, respectively. ABC transporter gene expression levels were determined by quantitative reverse transcription (RT)-PCR and ABCB1 protein expression by Western blotting. For the maternal-to-fetal direction, the mean FTR and CLI were 8.0% ± 3.0 and 0.26 ± 0.07, respectively, whereas the mean CLI was 0.52 ± 0.23 for the fetal-to-maternal direction. We showed a significant inverse correlation between maraviroc CLI and ABCC2, ABCC10, and ABCC11 placental gene expression levels (P < 0.05). To conclude, we report a low maraviroc placental transfer probably involving ABC efflux transporters and thus in all likelihood associated with a limited fetal exposition. Nevertheless, these results would need to be supported by in vivo data obtained from paired maternal and cord blood samples.

ABC drug transporter and nuclear receptor expression in human cytotrophoblasts: influence of spontaneous syncytialization and induction by glucocorticoids.

OBJECTIVES: ABC transporters in the human placenta play a major role in protecting the fetus against potential toxic drugs. The glucocorticoid dexamethasone has been shown to induce ABCB1 expression in enterocytes and hepatocytes. However, in placental cells, little data exists either for dexamethasone, betamethasone or prednisone while these three glucocorticoids may be used during pregnancy. We investigated the modulation of placental ABC transporter and nuclear receptor expression by these drugs. METHODS: Cytotrophoblasts were isolated from normal full-term placentas. We first assessed the influence of spontaneous syncytialization on transporter and nuclear receptor gene expression by taking samples of cytotrophoblasts after 24, 48 and 72 h of cell culture (n = 7 placentas). Incubations were then conducted with dexamethasone (50 nM-1 µM), betamethasone (20-400 nM) and prednisone (50 nM-1 µM) versus no drug for 24 h (n = 6). mRNA expression was determined by qRT-PCR. RESULTS: Influence of syncytialization was observed only for ABCB1, ABCC2 and ABCC5 gene expression between t = 24 and 48 h (p < 0.05). Therefore, the following induction studies were conducted between t = 48 h and 72 h. Dexamethasone and betamethasone significantly induced ABCB1 gene expression by around 4-fold (p < 0.01 and 0.001, respectively). In parallel, 100 nM betamethasone decreased the glucocorticoid receptor gene expression by 22% (p < 0.01). Prednisone showed no effect on transporter or receptor expression. CONCLUSIONS: These results suggest that dexamethasone or betamethasone administration may decrease the maternal-fetal transfer of an associated treatment being ABCB1 substrate, which may be either protective or deleterious for the fetus depending on the treatment's therapeutic aim.

[Quality and safety about hemapheresis practicing: from a self-evaluation of medical practices to a systematic risk management process]. / Qualité et sécurité des pratiques en hémaphérèse : d'un référentiel d'évaluation des pratiques professionnelles vers une démarche systématique de gestion des risques.

Hemapheresis is made up of a set of essential methods for modern medical practices. Applicated to donors, they allow production of labile blood products as well as raw material for plasma products industrial manufacturing; applicated to patients, they are indispensable for the treatment of many pathologies. Conditions for their implementation are very variable according to the teams who have few tools at their disposal to access, standardize and make reliable their activity. Therefore, an approach has been initiated by the Institut National de la Transfusion Sanguine and the Hemapheresis French Society to develop a guide which primary goal is to help teams evaluate themselves in different areas of their responsibilities. The generalization of the evaluation of professional practices in all activities and specialties required recently by the third iteration of the certification procedure of health institutions and a new law about French health system reinforce us in our approach. Finally, hemapheresis is not without risks, neither for donors nor for patients, which now encourages us to analyze sharply our procedures, to identify risk situations and develop means of prevention. This second step is already a current news for us.

[Quality control of defrosted cord blood units: results from an inter-laboratory study]. / Contrôle de qualité des greffons de sang placentaire décongelés : résultats d'une étude multicentrique.

PURPOSE: Today, haematopoietic stem cell graft from placental blood concerns more than 15 % of allogeneic grafts. An inter-laboratory study of the quality control of defrosted cord blood units has been coordinated by the French society for cell and tissue bioengineering (SFBCT), with the cord blood bank of Bourgogne Franche-Comté and controlled by the French health products safety agency (Afssaps). The aim of this study is to ensure the inter-laboratory reproducibility of the quality controls practised by the banks during defrosting. The cellular outputs were analyzed according to the defrosting techniques, according to the method used in flow cytometry: single-platform (SP) versus double-platform (DP), or the product nature, i.e. in total blood or miniaturized. METHODS: Forty-two units of placental blood (USP), which were out of range were provided for defrosting to 14 participating sites. USP were defrosted and controlled according to the procedures of each bank. Once the USP is defrosted, a part of the product was controlled by the site and the other part by Afssaps. Following controls were carried out: numeration of the total nucleated cells (TNC) and of CD34+ cells (made by a SP method in Afssaps) and functional assay. RESULTS: Concerning TNC, the defrosting sites obtained a cellular output of 94 %+/-28 in day 0 compared with an output of 72 %+/-24 in Afssaps showing a rather good stability of the USP transmitted with an average deviation of 23 %+/-22. The freezing process with or without reduction of volume does not affect this variation. Concerning the numeration of CD34+ cells, the average deviation between the participating sites and Afssaps was 29 %+/-23 compared with 21 %+/-16 for the sites using a SP method against 47 %+/-25 for those using a DP method. The CD34+ outputs are equal to 82 % +/- 60 in day 0 for the participating sites against 52 %+/-20 for Afssaps. For the sites using a DP method, it is stressed that this output is particularly high with a rate of 126 %+/-90 (n=15) whereas it is 62 %+/-20 (n=32) for the sites using a SP method. CONCLUSION: These results underline a good stability of viable CD34+ cells and a greater reliability of the SP methods for the CD34+ cell numeration for these defrosted USP. Lastly, the results of the functional assay regarding the average clonogenicities (equal to 15 %) reinforce the conclusions on the quality of the defrosted products.

Lack of P-glycoprotein induction by rifampicin and phenobarbital in human lymphocytes.

The efficacy of drugs acting within lymphocytes depends on their intracellular concentrations, which could be modulated by membrane efflux transporters including P-glycoprotein (P-gp), encoded by the MDR1 gene. In particular, P-gp induction may compromise the efficacy of its substrates. Rifampicin and phenobarbital have been shown to induce P-gp in hepatic and intestinal cells through the activation of the nuclear receptors PXR and CAR. However, controversial data exist in human lymphocytes. We investigated the effect of these drugs on P-gp activity and expression in lymphocytes in vitro and ex vivo. CCRF-CEM cells and peripheral blood mononuclear cells (PBMCs) from healthy volunteers were incubated in the presence of rifampicin, phenobarbital, or without any drug. P-gp activity was measured by flow cytometry using DiOC(6) efflux. MDR1, PXR and CAR mRNA expression were measured by quantitative RT-PCR. Neither P-gp activity nor MDR1 mRNA expression were modified by rifampicin or phenobarbital both in CCRF-CEM cells and PBMCs. Moreover, P-gp protein expression at the membrane was neither detectable nor induced. The very weak PXR and CAR mRNA expression levels in these cells could partly explain these results. Therefore, P-gp induction by rifampicin and phenobarbital may play a negligible role in drug interactions occurring within lymphocytes.

Length of the treatment and number of doses per day as major determinants of child adherence to acute treatment.

OBJECTIVE: To determine the rate of aftercare adherence to prescriptions from a paediatric emergency department and to identify predictors for nonadherence. METHODS: Patients discharged from a French paediatric emergency department with at least one oral drug prescription were included. A telephone interview questionnaire was used to determine whether the child had received the treatments according to the prescription. Adherence was assessed according to three items: frequency of drug administration, length of treatment and drug administering method. Complete adherence was defined as adherence to the three items mentioned above, and nonadherent as nonadherent to at least one of the items. Influence of age, sex, pathology, language spoken at home, type of medical insurance, type of medication prescribed, diagnosis, dissatisfaction with the explanation of the medical problem, number of prescribed medications, length of the treatment and number of doses per day was assessed. RESULTS: One hundred and five telephone interviews were exploited. The children were 60 boys (57%) and 45 girls (43%). The ages of these 105 children were between 0.2 and 12 years. The most common diagnoses were asthma and pulmonary infection. Complete adherence with the prescription was 36.2%. Three factors were significantly associated with nonadherence (p < 0.05): length of treatment, number of doses per day and male sex. CONCLUSION: This study suggests that simplifying treatment schedules is an effective strategy for improving compliance in paediatric emergency departments.

[Treatment of homozygous familial hypercholesterolemia with LDL-apheresis on a 4-year-old child]. / Traitement d'une hypercholestérolémie familiale homozygote par LDL-aphérèse chez un enfant de 4 ans.

Homozygous familial hypercholesterolemia (HFH) is a rare genetic disease associated with increased atherosclerosis, resulting in premature death near the age of 20 years. Treatment requires the LDL-apheresis system. M, born from a consanguineous union, suffers from HFH (total-cholesterol=12.29 g/l, LDL-cholesterol=9.65 g/l). Diet and drug treatment was not associated with decreased LDL-cholesterol. At the age of 4.5 years (body weight: 16.7 kg), M began treatment with LDL-apheresis. Apheresis treatment was given every 2 weeks using the Direct Adsorption of LIpoprotein (DALI system, a process that involves total-blood filtration. During the first 26 sessions, the mean reduction in LDL-cholesterol was 67+/-12%, while HDL-cholesterol decreased by only 17+/-11%. Mean LDL-cholesterol concentration decreased from 6.54+/-0.93 g/l (before apheresis) to 2.21+/-0.95 g/l (after apheresis). Apart from iron deficiency anemia, no major side effects were observed. LDL-apheresis using the DALI system is associated with significant reductions in LDL-cholesterol (similar to reports from the literature) without major side effects, even in a child weighing less than 20 kg. A long term, multinational (European) study is needed to confirm these results.

[Three cases of IgM monoclonal gammopathy with haemostasis dysfunction]. / Trois cas de gammapathies monoclonales IgM avec anomalies de l'hémostase.

We relate three cases of IgM gammopathy with haemostatic dysfunction in the perioperative period. The acquired von Willebrand syndrome due to IgM gammopathy is rare and sometimes serious. Its different treatments and their efficiency are discussed: desmopressin, intravenous gammaglobulin, chemotherapy and plasmapheresis.

High levels of P-glycoprotein activity in human lymphocytes in the first 6 months of life.

P-glycoprotein (P-gp) is an efflux transporter that controls the intracellular concentrations of drugs. Human development may modulate P-gp function. We investigated the effect of age on P-gp activity and MDR1 gene expression in lymphocytes. We also assessed the influence of human immunodeficiency virus (HIV) infection. We used 3,3'-diethyloxacarbocyanin iodide (DiOC(6)) efflux, estimated by flow cytometry, to quantify P-gp activity in 94 children (age range, 0-18 years) and 25 adults. MDR1 gene expression was quantified using reverse transcription-PCR (RT-PCR). In T and natural killer (NK) cell populations, P-gp activity peaked at birth, decreased between the ages of 0 and 6 months, and stabilized between the ages of 6 months and 2 years (P < 10(-6)). These maturation profiles were also strongly correlated (r = 0.67, P < 10(-6)). HIV infection did not affect P-gp activity in the lymphocytes of children. MDR1 gene expression was not influenced by age, nor was it correlated with P-gp activity. The high levels of P-gp activity observed in the lymphocytes of children ~6 months of age may affect the efficacy of intracellular drugs.

A Botrytis cinerea emopamil binding domain protein, required for full virulence, belongs to a eukaryotic superfamily which has expanded in euascomycetes.

A previous transcriptomic analysis of 3,032 fungal genes identified the Botrytis cinerea PIE3 (BcPIE3) gene to be up-regulated early in planta (A. Gioti, A. Simon, P. Le Pêcheur, C. Giraud, J. M. Pradier, M. Viaud, and C. Levis, J. Mol. Biol. 358:372-386, 2006). In the present study, BcPIE3 was disrupted in order to determine its implication in pathogenicity. BcPIE3 was shown to be a virulence factor, since the DeltaBcPIE3 mutant was blocked during the colonization of tomato and bean leaves, giving lesions reduced in size by at least 74%. Within the emopamil binding domain (EBD), BcPIE3 shows significant structural similarities to mammalian emopamil binding proteins (EBPs). Mammalian EBPs function as sterol isomerases, but an analysis of the sterol content and the results of growth inhibition experiments with the DeltaBcPIE3 strain indicated that BcPIE3 is dispensable for ergosterol biosynthesis. The systematic identification of EBD-containing proteins included in public databases showed that these proteins constitute a protein superfamily present only in eukaryotes. Phylogenetic analysis showed that the ancestral EBD-encoding gene was duplicated in the common ancestor of animals and fungi after the split from plants. Finally, we present evidence that the EBP phylogenetic clade of this superfamily has further expanded exclusively in euascomycetes, especially in B. cinerea, which contains three copies of the EBP gene.

Detection of human bocavirus in children with Kawasaki disease.

Human bocavirus (HboV) is an emerging virus that has been implicated as a cause of acute upper and lower respiratory tract infection in children. As no serological assay is available, PCR was used to screen nasopharyngeal, serum or stool samples from 16 patients with Kawasaki disease for HBoV nucleic acid. HBoV was identified by PCR in five (31.2%) patients, suggesting that this emerging virus may also play a pathogenic role in some cases of Kawasaki disease.