[Show abstract][Hide abstract] ABSTRACT: La surexpression de GALIG, un gène humain emboité dans celui de la Galectine-3, induit la mort cellulaire : au cours de l’apoptose spontanée des polynucléaires, son expression augmente de plus de 100 fois. Par ailleurs, l’expression de ce gène semble maximale dans les leucocytes et moindre dans la moelle osseuse (MO).
L’expression de GALIG augmente fortement lors de la différenciation in vitro en monocytes/macrophages de plusieurs lignées myéloïdes immatures (HL60, U937 et THP-1). A noter que ces différenciations ont été validées par des tests de phagocytose.
L’expression de GALIG a été dosée dans des prélèvements de Leucémie Aiguë Myéloïde (LAM), pathologie caractérisée par un arrêt de la différenciation cellulaire. Il existe différents types de LAM, classées en fonction du stade de différenciation auquel sont bloquées les cellules. Le taux de GALIG est inférieur dans les prélèvements de LAM-M2 (MO et cellules mononucléées-PBMC), comparé aux prélèvements sains. De plus, le taux de GALIG est plus important dans les LAM-M3 par rapport aux LAM-M2 : le niveau d’expression de GALIG semble donc corrélé au niveau de différenciation de la cellule myéloïde. Ce travail se poursuit actuellement avec le CHR d’Orléans : l’expression de GALIG sera dosée sur des prélèvements de tous les types de LAM (MO et PBMC), au diagnostic et à chaque retour d’aplasie post-cure.
Nous avons aussi étudié l’expression de GALIG au cours de la différenciation terminale du monocyte en macrophage. In vitro, ces macrophages acquièrent la capacité de phagocyter. Dans ce cas, l’expression du gène GALIG est augmentée d’un facteur 10.
Le gène GALIG voit son expression augmenter tout au long des processus de différenciation touchant les cellules myéloïdes, mais sa fonction précise reste encore à déterminer.
Prix du meilleur poster.
9èmes Journées du Cancéropole Grand Ouest (18-19 Juin 2015); 06/2015
[Show abstract][Hide abstract] ABSTRACT: Cancer-specific splice variants gain significant interest as they generate neo-antigens that could be targeted by immune cells. CD20, a membrane antigen broadly expressed in mature B cells and in B cell lymphomas, is subject to an alternative splicing named D393-CD20 leading to loss of membrane expression of the spliced isoform. D393-CD20 expression is detectable in transformed B cells and upregulated in various lymphoma B cells. In this study, we show that D393-CD20 is translated in malignant B cells and that D393-CD20 specific CD4 T cells producing IFN-γ are present in B-cell lymphoma patients. Then we have investigated whether the 20mer D393-CD20 peptide spanning the splicing site might be targeted by the immune system and we have shown that D393-CD20-specific CD4 Th1 clones could directly recognize malignant B cell lines and kill autologous lymphoma B cells indicating that D393-CD20-derived epitopes are naturally processed and presented on tumor cells. Finally, D393-CD20 peptide-based vaccination induced specific CD8 and CD4 T cell responses in HLA-humanized transgenic mice suggesting the presentation of D393-CD20 derived peptides on both HLA class-I and -II. These findings support further investigations on the potential use of D393-CD20 directed specific immunotherapy in B cell malignancies. This article is protected by copyright. All rights reserved.
International Journal of Cancer 11/2014; 137(1). DOI:10.1002/ijc.29366 · 5.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Background
There are several reports demonstrating the role of CD8 T cells against Leishmania species. Therefore peptide vaccine might represent an effective approach to control the infection. We developed a rational polytope-DNA construct encoding immunogenic HLA-A2 restricted peptides and validated the processing and presentation of encoded epitopes in a preclinical mouse model humanized for the MHC-class-I and II.
Methods and Findings
HLA-A*0201 restricted epitopes from LPG-3, LmSTI-1, CPB and CPC along with H-2Kd restricted peptides, were lined-up together as a polytope string in a DNA construct. Polytope string was rationally designed by harnessing advantages of ubiquitin, spacers and HLA-DR restricted Th1 epitope. Endotoxin free pcDNA plasmid expressing the polytope was inoculated into humanized HLA-DRB1*0101/HLA-A*0201 transgenic mice intramuscularly 4 days after Cardiotoxin priming followed by 2 boosters at one week interval. Mice were sacrificed 10 days after the last booster, and splenocytes were subjected to ex-vivo and in-vitro evaluation of specific IFN-γ production and in-vitro cytotoxicity against individual peptides by ELISpot and standard chromium-51(51Cr) release assay respectively. 4 H-2Kd and 5 HLA-A*0201 restricted peptides were able to induce specific CD8 T cell responses in BALB/C and HLA-A2/DR1 mice respectively. IFN-γ and cytolytic activity together discriminated LPG-3-P1 as dominant, LmSTI-1-P3 and LmSTI-1-P6 as subdominant with both cytolytic activity and IFN-γ production, LmSTI-1-P4 and LPG-3-P5 as subdominant with only IFN-γ production potential.
Here we described a new DNA-polytope construct for Leishmania vaccination encompassing immunogenic HLA-A2 restricted peptides. Immunogenicity evaluation in HLA-transgenic model confirmed CD8 T cell induction with expected affinities and avidities showing almost efficient processing and presentation of the peptides in relevant preclinical model. Further evaluation will determine the efficacy of this polytope construct protecting against infectious challenge of Leishmania. Fortunately HLA transgenic mice are promising preclinical models helping to speed up immunogenicity analysis in a human related mouse model.
PLoS ONE 10/2014; 9(10):e108848. DOI:10.1371/journal.pone.0108848 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: L’allogreffe de cellules souches hématopoïétiques (ACSH) est un traitement curatif des hémopathies. Cette thérapeutique est associée à un profond déficit immunitaire et un risque important d’infections opportunistes. La nocardiose est une infection bactérienne rare qui survient préférentiellement chez les patients ayant un déficit de l’immunité cellulaire comme les patients atteints de sida ou les greffés. Le diagnostic est difficile en raison de signes cliniques et biologiques peu spécifique de l’infection. La prophylaxie systématique par cotrimoxazole n’assure pas une protection totale. Cinq cas de nocardioses ont été diagnostiqués entre mai 2001 et décembre 2012 dans notre centre parmi 475 greffes d’ACSH. Quatre cas sur 5 sont des cas de nocardioses disséminées survenant dans les deux premières années de l’allogreffe. Le cinquième cas est une nocardiose localisée cutanée. Parmi les cas de nocardioses disséminées, le taux médian de lymphocytes T CD4+ était inférieur à 300/mm3. Les lymphocytes T CD4+ à phénotype naïf CD45RA+/RO– étaient quasiment indétectables. Les lymphocytes T CD8+ et les cellules Natural Killer étaient majoritairement inférieures aux normes et la reconstitution des lymphocytes B CD19+ complètement inexistante. Concernant le cas de nocardiose localisée, nous avons observé également un profond déficit de la reconstitution des lymphocytes T CD4+ naïfs issus récemment du thymus de phénotype CD45RA+/RO–.
[Show abstract][Hide abstract] ABSTRACT: GALIG gene expression induces apoptosis in cultured cells through a pathway still under investigation. It is highly expressed in leukocytes but weakly detectable in bone marrow, suggesting a role in the myeloid lineage homeostasis. We show here that GALIG-induced cell death is counteracted by the overexpression of MCL-1, a pro-survival member of the Bcl2 family. Moreover, during spontaneous neutrophil apoptosis, a substantial increase in GALIG gene expression is observed: GALIG still opposes MCL-1. Finally, in bone marrow and peripheral blood cells from patients with Acute Myeloid Leukemia type 2, the level of GALIG transcripts is massively down-regulated when compared to their normal counterparts, while MCL-1 is expressed to the same extent. These data suggest that GALIG could be a key player in the cell death pathway involved in leukocytes homeostasis and myeloid malignancies.
[Show abstract][Hide abstract] ABSTRACT: The aim of our study was to analyze the factors contributing to heterogeneity of prognosis in patients with hyperdiploidy>50 chromosomes (HD>50), a group of B-cell precursor acute lymphoblastic leukemia with favorable outcome. The 541 HD>50 patients registered prospectively in the 58951 CLG-EORTC trial, identified by karyotype (446 patients) and by DNA index (DI) (490 patients), had a 6-year EFS of 89.0% (SE=1.5%) and a 6-year overall survival (OS) of 95.9% (SE=0.9%). The strongest prognostic factor was the modal number of chromosomes (MNC): the 6-year EFS of 51-53, 54-57 and 58-66 MNC groups were 80%, 89% and 99% respectively (P<0.0001). Ploidy assessed by DNA index (DI) was also a favorable factor, the higher the DI the better the outcome: the 6-year EFS of the three subgroups of DI <1.16/≥1.16-<1.24/≥1.24 were 83%, 90% and 95% respectively (P=0.009). All usual combinations of trisomies (chromosomes 4,10,17,18) were significant favorable factors but had lower EFS when MNC was lower than 58. In multivariate analysis, MNC remained the strongest factor. Consequently, the best indicator for excellent outcome was ploidy assessed by karyotype because patients with 58-66 chromosomes stood every chance of being cured (OS of 100% at 6-year follow-up) with less intensive therapy. NCT00003728, Registered: http://www.eortc.org/, http://clinicaltrials.gov/show/NCT00003728.
[Show abstract][Hide abstract] ABSTRACT: Clofarabine alone or in combination with cyclophosphamide and etoposide has shown a good efficacy and a tolerable toxicity profile in previous studies of children with relapsed or refractory leukaemia. This report describes a retrospective study of 38 French patients who received clofarabine as a monotherapy or in combination for relapsed or refractory acute lymphoblastic leukaemia (ALL) outside of clinical trials after marketing authorization.
We retrospectively analysed data for 38 patients, up to 21 years old, attending 17 French centres. Thirty patients received clofarabine alone or in combination for a bone marrow relapse of acute lymphoblastic leukaemia (ALL) or refractory disease and eight patients for a high level of minimal residual disease (MRD). Survival and response durations were estimated by the Kaplan-Meier method.
For the 30 patients who received clofarabine for a bone marrow relapse of ALL (number of relapse, 1-3; median, 1), the overall remission rate (ORR) was 37%: eight complete remission (CR) and three complete remission without platelet recovery (CRp). Ten of the 11 responding patients subsequently underwent haematopoietic stem cell transplantation (HSCT).
Only four of the eight patients who received clofarabine while in remission for a high level of MRD, showed a moderate improvement of MRD. Seven of these eight patients received HSCT and six of them were alive at the end of the study. One other patient was alive without receiving HSCT.
However, clofarabine treatment was associated with a high risk of infection and hepatotoxicity. Febrile neutropenia grade ≥ 3 was reported in 79% of patients and documented infections grade ≥ 3 occurred in nine patients (24%). Hepatotoxicity grade 3 was reported in nine patients (24%). We observed four deaths related to treatment.
In our experience, the efficacy of clofarabine is poorer than previously reported. Its toxicity is high and can be life threatening. Prospective studies on clofarabine used during earlier phases of the disease may help to define how best this new drug can be exploited for childhood and adolescent ALL.
[Show abstract][Hide abstract] ABSTRACT: Congenital neutropenia is a group of genetic disorders that associates chronic neutropenia and susceptibility to infections. These neutropenias are either isolated, associated with immunologic defects or extra-hematopoietic manifestations. Complications may occur as infectious diseases but also less frequently as myelodysplastic syndromes (MDS) or acute myeloid leukemia (AML). Recently, GATA2, a transcription factor, has been identified as a new predisposing gene for familial AML/MDS. Here, we described the initial identification by exome sequencing of a GATA2 R396Q mutation in a family with a history of chronic mild neutropenia evolving to AML and/or MDS. The subsequent analysis of the French national registry of chronic neutropenia allowed the identification of 6 additional pedigrees and 10 patients with 6 different and not previously reported GATA2 mutations (R204X, E224X, R330X, A372T, M388V and a complete deletion of the GATA2 locus). The frequent evolution to MDS and AML of these patients reveals the importance of screening GATA2 in chronic neutropenia associated with monocytopenia due to their frequent hematopoietic transformation, their variable clinical expression at onset and the need of aggressive strategy therapy in patients with poor clinical outcome.
[Show abstract][Hide abstract] ABSTRACT: WHIM syndrome (WS), a rare congenital neutropenia due to mutations of the CXCR4 chemokine receptor, is associated with Human Papillomavirus (HPV)-induced Warts, Hypogammaglobulinemia, bacterial Infections and Myelokathexis. The long term follow up of eight patients highlights the clinical heterogeneity of this disease as well as the main therapeutic approaches and remaining challenges in the light of the recent development of new CXCR4 inhibitors.
This study aims to describe the natural history of WS based on a French cohort of 8 patients.
We have reviewed the clinical, biological and immunological features of patients with WS enrolled into the French Severe Chronic Neutropenia Registry.
We identified four pedigrees with WS comprised of eight patients and one foetus. Estimated incidence for WS was of 0.23 per million births. Median age at the last visit was 29 years. Three pedigrees encompassing seven patients and the fetus displayed autosomal dominant heterozygous mutations of the CXCR4 gene, while one patient presented a wild-type CXCR4 gene. Two subjects exhibited congenital conotruncal heart malformations. In addition to neutropenia and myelokathexis, all patients presented deep monocytopenia and lymphopenia. Seven patients presented repeated bacterial Ears Nose Throat as well as severe bacterial infections that were curable with antibiotics. Four patients with late onset prophylaxis developed chronic obstructive pulmonary disease (COPD). Two patients reported atypical mycobacteria infections which in one case may have been responsible for one patient’s death due to liver failure at the age of 40.6 years. HPV-related disease manifested in five subjects and progressed as invasive vulvar carcinoma with a fatal course in one patient at the age of 39.5 years. In addition, two patients developed T cell lymphoma skin cancer and basal cell carcinoma at the age of 38 and 65 years.
Continuous prophylactic anti-infective measures, when started in early childhood, seem to effectively prevent further bacterial infections and the consequent development of COPD. Long-term follow up is needed to evaluate the effect of early anti-HPV targeted prophylaxis on the development of skin and genital warts.
[Show abstract][Hide abstract] ABSTRACT: Chronic HIV infection leads to increased risk of non-Hodgkin B-cell lymphoma. However, only few recent data are available about their current management and prognosis in HIV-infected children since the advent highly active antiretroviral therapy (HAART). This multicenter retrospective study describes the 12 cases of B-cell non-Hodgkin lymphoma diagnosed in HIV-infected children in France between 1996 and 2009. All children had moderate to severe immunosuppression and high viral load at the time of diagnosis. Nine children had extracerebral primary sites and 3 had a primary central nervous system lymphoma. Eight patients had Burkitt lymphoma; 4 had diffuse large B-cell lymphoma. Concomitantly with HAART, all children with extracerebral lymphoma received intensive chemotherapy according to LMB protocol, those with primary central nervous system lymphoma received high-dose methotrexate. No toxicity-related deaths occurred. Ten patients entered complete remission (CR), 2 died of tumor progression despite a second line of therapy. No relapses occurred after CR (median follow-up, 72 mo). Thus, prognosis of patients unresponsive to first-line lymphoma treatment remains poor, but relapse seems to be rare when CR is achieved. Children without severe comorbidities can tolerate intensive chemotherapy with a mandatory HAART treatment, taking into account drug interactions.
[Show abstract][Hide abstract] ABSTRACT: HFE, an MHC class Ib molecule that controls iron metabolism, can be directly targeted by cytotoxic TCR αβ T lymphocytes. Transgenic DBA/2 mice expressing, in a Rag 2 KO context, an αβ TCR that directly recognizes mouse HFE (mHFE) were created to further explore the interface of HFE with the immune system. TCR-transgenic mHfe WT mice deleted mHFE-reactive T cells in the thymus, but a fraction of reprogrammed cells were able to escape deletion. In contrast, TCR-transgenic mice deprived of mHFE molecules (mHfe KO mice) or expressing a C282→Y mutated mHFE molecule - the most frequent mutation associated with human hereditary hemochromatosis - positively selected mHFE-reactive CD8(+) T lymphocytes and were not tolerant toward mHFE. By engrafting these mice with DBA/2 WT (mHFE(+)) skin, it was established, as suspected on the basis of similar engraftments performed on DBA/2 mHfe KO mice, that mHFE behaves as an autonomous skin-associated histocompatibility antigen, even for mHFE-C282→Y mutated mice. By contrast, infusion of DBA/2 mHFE(+) mice with naïve mHFE-reactive transgenic CD8(+) T lymphocytes did not induce GVHD. Thus, tolerance toward HFE in mHfe WT mice can be acquired at either thymic or peripheral levels but is disrupted in mice reproducing human familial hemochromatosis.
European Journal of Immunology 04/2012; 42(4):851-62. DOI:10.1002/eji.201141664 · 4.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The EORTC Children's Leukemia Group (CLG) is a spin-off from the EORTC Hemopathies Working Party (adults and children). After a decade of collaboration in the adult-pediatric group it became clear that there was not only a large difference in cure rates between children and adults, but many chemotherapeutic-toxic borders were substantially different. During the following decade the CLG was not only a witness of a very exciting battle against childhood leukemia, but it also contributed substantially to better cure rates for these diseases.The main activity of the CLG was concentrated on the field of acute lymphoblastic leukemia (ALL). Fine tuning of treatment elements using the BFM design as a backbone has been done very successfully over the past decades. The CLG has many achievements, and the major ones include: the 58831 trial showing the superfluity of the prophylactic Central Nervous System (CNS) radiotherapy in ALL patients when a adequate systemic and CNS directed chemotherapy is ascertained. The 58881 trial demonstrated that the assessment of minimal residual disease (MRD) at completion of induction in ALL is a key step in the process to categorizing and allocating patients into different risk groups, and that MRD is a powerful and independent prognostic factor. This same 58881 trial showed the clear difference in efficacy of different asparaginases resulting in an optimization of the use these drugs and a revival of interest for the asparaginases in the treatment of ALL.In the near future the CLG will focus mainly on translational research projects and innovative biologically targeted treatment approaches, on collaborations with other children's leukemia groups in intergroup studies, and on the evaluation of the long-term outcome of childhood cancer survivors.
[Show abstract][Hide abstract] ABSTRACT: A primary HCMV infection or virus reactivation may cause severe disease in hosts with a deficient immune system. The virus can disturb both innate and adaptive immunity by targeting dendritic cell (DC) functions. Monocytes, the precursors of DCs in vivo (MoDCs), are the primary targets of HCMV; they can also harbor latent virus. The DCs generated from infected monocytes (CMV-MoDCs) have an altered phenotype and functional defects. We have shown that CMV-MoDCs do not secrete IL-12 in response to lipopolysaccharide stimulation, cannot ingest dead cells, induce T(H)1 differentiation, or the proliferation of naive allogeneic CD4(+) T cells. We found that the GM-CSF signaling in an entire population of CMV-MoDCs was impaired, although only half of the cells were productively infected, and that IL-6 secretion and suppressors of cytokine signaling 3 induction contributed to this bystander effect. We also showed that MoDCs derived ex vivo from monocytes of viremic patients had the same altered phenotype as CMV-MoDCs, including decreased STAT5 phosphorylation, indicating defective GM-CSF signaling. We have thus described a new mechanism of HCMV-induced immunosupression, indicated how infection may disturb both GM-CSF-dependent physiologic processes and proposed GM-CSF-based therapeutic approaches.
[Show abstract][Hide abstract] ABSTRACT: The current screening for eligibility of unrelated volunteer marrow donors comprises a complete clinical check-up, a blood CBC and serum protein immunoelectrophoresis. This allows to eliminate acute leukemias, myeloproliferative and myelodysplastic disorders, myelomas and MGUS. To date, the risk of transmission of chronic lymphocytic leukemia (CLL) disease is only evaluated by the clinical evaluation and CBC. We report here the case of a CLL-type MBL disease occurring in a 12-year-old boy after unrelated BMT. Deep biological investigations, as Immunophenotyping, cytogenetic and molecular biology allow us to determine the donor origin of the CLL clone. In 2010, 14.2% donor (105/737) for unrelated hematopoietic stem cell transplantation were over 45y. It is currently estimated (USA) that 1 in 210 men and women will be diagnosed with CLL during their lifetime. Given the long asymptomatic phase of CLL, this raises the case for a detection strategy analog to that used for MGUS and myeloma through serum protein electrophoresis. This case-report, to our knowledge, of a CLL-type MBL unrelated donor-to-recipient transmission through BMT raises ethical and practical questions, such as the proper information about disease transmission risk. The cost-effectiveness of a systematic peripheral blood Immunophenotyping in donors elder than 40y at time of stem cell donation should be evaluated.
European Journal Of Haematology 12/2011; 88(3):269-72. DOI:10.1111/j.1600-0609.2011.01741.x · 2.41 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Digestive cryptosporidiosis (DC) can mimic GVHD after allogeneic haematopoietic stem cell transplantation (HSCT), thus requiring a reduction of immunosuppressive drugs and a specific therapy, whereas GVHD requires an intensification of immunosuppression. We systematically searched for cryptosporidiosis by light microscopy, immunochromatography and PCR in HSCT recipients who presented with at least one episode of diarrhoea. Of 115 consecutive patients allografted between July 2006 and November 2008, we analysed stools in 52 of 56 patients meeting these criteria. We identified Cryptosporidium parvum in 5 of the 52 patients (9.6%) at a median of 503 days (range 20-790) after HSCT. In those five patients, the median CD4+ cell and B lymphocyte counts were 60/mm3 (0-234) and 0/mm3 (0-96), respectively. Two patients died of invasive fungal infections. In the other three patients, diarrhoea disappeared after a median of 5 weeks following onset of bitherapy with azithromycine and nitazoxanide; they were still alive 433, 380 and 1179 days after the DC diagnosis. DC is probably under diagnosed after HSCT because it is difficult to detect during the asymptomatic phase. Early bitherapy and reduction of immunosuppression seem efficacious. In our series, DC has a seasonal pattern and is promoted by profound T lymphopenia.
Bone marrow transplantation 06/2011; 46(6):858-62. DOI:10.1038/bmt.2010.200 · 3.47 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Autoimmune hemolytic anemia is a rare condition in children. Little is known about its initial presentation and the subsequent progression of the disease.
Since 2004, a national observational study has been aiming to thoroughly describe cases and identify prognostic factors. Patients from all French hematologic pediatric units have been included if they had a hemoglobin concentration less than 11 g/dL, a positive direct antiglobulin test and hemolysis. Evans' syndrome was defined by the association of autoimmune hemolytic anemia and immunological thrombocytopenic purpura. Data from patients' medical records were registered from birth to last follow-up. Autoimmune hemolytic anemia was classified as primary or secondary. Remission criteria, qualifying the status of anemia at last follow-up, were used with the aim of identifying a subgroup with a favorable prognosis in continuous complete remission.
The first 265 patients had a median age of 3.8 years at diagnosis. In 74% of cases the direct antiglobulin test was IgG/IgG+C3d. Consanguinity was reported in 8% of cases and first degree familial immunological diseases in 15% of cases. Evans' syndrome was diagnosed in 37% of cases. Autoimmune hemolytic anemia was post-infectious in 10%, immunological in 53% and primary in 37% of cases. After a median follow-up of 3 years, 4% of children had died, 28% were still treatment-dependent and 39% were in continuous complete remission. In multivariate analysis, IgG and IgG+C3d direct antiglobulin tests were associated with a lower rate of survival with continuous complete remission (adjusted hazard ratio, 0.43; 95% confidence interval, 0.21-0.86).
This nationwide French cohort is the largest reported study of childhood autoimmune hemolytic anemia. The rarity of this condition is confirmed. Subgroups with genetic predisposition and underlying immune disorders were identified.