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International audience; Estrogen receptor-, progesterone receptor- and HER2-negative breast cancers, also known as triple-negative breast cancers (TNBCs), have poor prognoses and are refractory to current therapeutic agents, including epidermal growth factor receptor (EGFR) inhibitors. Resistance to anti-EGFR therapeutic agents is often associated with sustained kinase phosphorylation, which promotes EGFR activation and translocation to the nucleus and prevents these agents from acting on their targets. The mechanisms underlying this resistance have not been fully elucidated. In addition, the IL-17E receptor is overexpressed in TNBC tumors and is associated with a poor prognosis. We have previously reported that IL-17E promotes TNBC resistance to anti-mitotic therapies. Here, we investigated whether IL-17E promotes TNBC resistance to anti-EGFR therapeutic agents by exploring the link between the IL-17E/IL-17E receptor axis and EGF signaling. We found that IL-17E, similarly to EGF, activates the EGFR in TNBC cells that are resistant to EGFR inhibitors. It also activates the PYK-2, Src and STAT3 kinases, which are essential for EGFR activation and nuclear translocation. IL-17E binds its specific receptor, IL-17RA/IL17RB, on these TNBC cells and synergizes with the EGF signaling pathway, thereby inducing Src-dependent EGFR transactivation and pSTAT3 and pEGFR translocation to the nucleus. Collectively, our data indicate that the IL-17E/IL-17E receptor axis may underlie TNBC resistance to EGFR inhibitors and suggest that inhibiting IL-17E or its receptor in combination with EGFR inhibitor administration may improve TNBC management.

Background KIR3DL2, an inhibitory receptor expressed by natural killer cells and a subset of normal CD8(+) T cells, is aberrantly expressed in neoplastic cells in transformed mycosis fungoides and Sezary syndrome. Anti-KIR3DL2 targeted antibody therapy has shown potent activity in preclinical models for these diseases. Objectives To examine the expression of KIR3DL2 and its potential use as a therapeutic target in patients with primary cutaneous anaplastic large-cell lymphoma (pcALCL), the most aggressive cutaneous CD30(+) lymphoproliferative disease. Methods Samples from 11 patients with pcALCL and three CD30(+) lymphoproliferative disease cell lines - Mac1, Mac2a and Mac2b - were used in KIR3DL2 expression studies using immunohistochemistry, flow cytometry and reverse-transcriptase quantitative polymerase chain reaction. The effect of IPH4102, a monoclonal humanized IgG1 targeting KIR3DL2, was assessed by in vitro cytotoxicity assays against Mac1, Mac2a and Mac2b using allogeneic peripheral blood mononuclear cells as effectors. Results KIR3DL2 mRNA and protein were found in all human samples of pcALCL, and in the Mac2a and Mac2b cell lines. KIR3DL2 protein expression was present on 85·8 ± 14·0% of CD30(+) skin-infiltrating tumour cells. In vitro functional studies showed that KIR3DL2(+) Mac2a and Mac2b pcALCL lines are sensitive to antibody-derived cytotoxicity mediated by IPH4102, through activation of natural killer cells, in a concentration-dependent manner. Conclusions pcALCL tumour cells express KIR3DL2, and we provide preclinical proof of concept for the use of IPH4102, a humanized anti-KIR3DL2 antibody, to treat patients with primary cutaneous CD30(+) ALCL.

After allogeneic hematopoietic stem cell transplantation (AHSCT), acute GVHD is almost entirely mediated by donor T cells whereas chronic graft- versus -host disease (cGVHD) also involves donor B cells.[1][1] A breakdown in peripheral B cell homeostasis in cGVHD is in part due to high levels of B

KIT mutations are frequent in acral, mucosal and chronic sun-damage (CSD) melanoma, but little is known about the mechanisms driving the transformation of KIT-mutated melanocytes into melanoma cells. We showed that exposition of melanocytes harboring the (L576P)KIT mutation to a hypoxic environment induced their transformation into malignant cells. Transformed (L576P)KIT melanocytes showed downregulation of MITF expression characteristic of melanoma initiating cells (MICs). In agreement, these cells were able to form spheres in neural crest cell medium and low-adherence conditions, also a characteristic of MICs. Downregulation of MITF by RNA interference induced transformation of KIT-mutated melanocytes in normoxia and acquisition of a MIC phenotype by these cells. Hence, low level of MITF cooperates with oncogenic KIT to transform melanocytes. Activation of the cAMP pathway in transformed (L576P)KIT melanocytes stimulated MITF expression, and reduced cellular proliferation and sphere formation. These findings highlight the essential role of MITF in revealing the oncogenic activity of KIT in melanocytes and suggest that the cAMP pathway is a therapeutic target in KIT-mutated melanoma.

Psoriasis is a frequent inflammatory disease that involves mostly the skin and sometimes the joints. This chronic disease is rarely life-threatening but impairs significantly the patient's quality of life. It is characterized, in its typical form, by erythematous and squamous plaques with well-defined borders, associated with increased proliferation of the keratinocytes, inflammation and greater number of dilated blood vessels in the upper dermis. A role of Th1 CD4 T cells was initially suspected. More recently, Th17 CD4 T cells have been shown to play a major role in the disease. It has led to the development of Th17 inhibitors, such as anti-IL-23 (cytokine that induces Th17 CD4 T cell differentiation), anti-IL-17, anti-IL-17RA (IL-17 receptor) and anti-IL-22 (cytokines that are notably produced by Th17 CD4 T cells).

e20040 Background: Advanced stage cutaneous T-cell lymphomas (CTCLs) remain an unmet medical need. Immunomodulatory agents such as mogamulizumab, anti-KIR3DL2 and brentuximab vedotin (BV), an anti-CD30 antibody–drug conjugate (ADC) coupled to monomethyl-auristatin-E (MMAE), provided encouraging results but new targeted therapies are needed. Inducible Co-Stimulator (ICOS), a T-cell costimulatory receptor involved in the development of CTCLs, arouses interest. Methods: We used immunohistochemistry to study ICOS expression in skin biopsies of 23 patients with early-stage mycosis fungoides (MF), 12 with transformed MF (TMF) and 17 with Sézary Syndrome (SS), at diagnosis or in relapse. ICOS expression by circulating Sézary cells and regulatory T cells (Tregs) in patients with SS was evaluated using flow cytometry, and compared to healthy donors (HD) lymphocytes. In 5 patients with SS, we also analyzed concomitant biopsies from involved nodes. Then, we investigated the efficacy of anti-ICOS ADCs generated by coupling murine anti-ICOS monoclonal antibodies with MMAE, in comparison to BV. We used ICOS+ CTCL cell lines (MyLa and MJ), murine xenograft models with MyLa and ICOS+ Patient Derived Xenografts (PDXs) from patients with SS and angioimmunoblastic T-cell lymphoma (AITL). Results: ICOS was highly expressed by the cutaneous atypical lymphocytic infiltrates in respectively 61%, 75% and 88% of patients with early-stage MF, TMF and SS, such as in the 5 patients with node involvement. ICOS expression by circulating Sézary cells was strong: 69±7.3% versus 38.8±7.1% of non-tumoral CD4+ cells ( p< 0.009; CI95%: 8.7-51.6); and 31±3.2% of CD4+ cells in HD ( p< 0.0001; CI95%:20.3-46.3). Percentages of ICOS+ Tregs were significantly higher in patients with SS than in HD. In CTCL cell lines, we observed a significant dose-dependent decrease in cell viability in the presence of anti-ICOS ADCs. In a mouse xenograft model (MyLa), anti-ICOS ADCs provided a longer overall survival (OS) than BV (HR = 15.2;CI95%:3.2-71.1; p< 0.0006). Finally, in ICOS+ PDXs anti-ICOS ADCs significantly improved OS, and reduced the number of tumor cells in the blood, bone marrow and spleen. No evidence of ADC toxicity was observed in treated mice. Conclusions: All together our results show that ICOS is a therapeutic target of interest in CTCLs and provide the preliminary basis for a therapeutic trial

Introduction Dans cette etude, nous avons developpe une emulsion de pickering co-encapsulant un medicament immunosuppresseur (ciclosporine A ou tacrolimus) dans des nanoparticules de PLGA (NP) biodegradables et biocompatibles, qui stabilisent la phase huileuse contenant du calcitriol. L’objectif est de quantifier leurs effets biologiques in vitro sur les cellules cibles cutanees residentes (keratinocytes) ou infiltrant la peau (lymphocytes), et de determiner leur penetration sur des explants de peau humaine. Materiel et methodes Des cellules mononucleees fraichement isolees du sang de volontaires sains ont ete cultivees en presence de billes d’anticorps CD3*CD28 sur 6 jours. Les effets des emulsions sont quantifies sur leur production d’IL-2 par ELISA, sur leur proliferation apres coloration CFSE par cytometrie en flux et leur activation par analyse de l’expression membranaire du CD25. La translocation de la voie NFAT est analysee apres transfection d’un gene reporter NFAT-Luc dans la lignee lymphocytaire JURKAT. Les emulsions sont etudiees sur les keratinocytes de la lignee HaCaT par MTT pour leur cytotoxicite et par ELISA pour leurs effets sur la production de cytokines. La penetration des NP de PLGA marquees a la rhodamine est evaluee sur des explants cutanes humains par microscopie confocale. Resultats Une reduction significative de la translocation de la voie NFAT (p Discussion L’encapsulation d’agents actifs dans la formulation de nanoparticules peut offrir plusieurs avantages : protection des agents actifs, ciblage des couches cutanees et liberation prolongee du medicament. Ceci permettrait d’obtenir une meilleure observance du patient grâce a une administration topique non invasive et non agressive, en particulier en cas d’inflammation chronique, et une reduction des effets secondaires avec une amelioration du rapport benefice/risque. Conclusion Les donnees de notre etude suggerent que les emulsions de pickering sont efficaces sur l’activation des lymphocytes et la reponse pro-inflammatoire des keratinocytes. De plus, les nanoparticules penetrent dans les couches profondes de l’epiderme, apportant ainsi de nouvelles perspectives therapeutiques aux dermatoses inflammatoires, comme le psoriasis et la dermatite atopique.

Introduction Le mycosis fongoide (MF) et son variant leucemique, le syndrome de Sezary (SS), sont les formes les plus frequentes des lymphomes T cutanes (LTC). Leur diagnostic clinique et histologique peut etre difficile. Notre equipe a decrit quatre marqueurs sanguins prometteurs du SS : Twist, T-plastine, NKp46 et KIR3DL2. De plus Tox a ete decrit par d’autres equipes comme etant specifique du MF et du SS. Le but de notre etude est de decrire l’utilite de ces marqueurs dans une nouvelle cohorte de validation pour le diagnostic de SS, MF erythrodermique (MFe) et MF debutant. Materiel et methodes Nous avons inclus apres consentement ecrit des patients ayant un SS, un MF ou une dermatose inflammatoire. A partir du sang peripherique, les ARNm ont ete extraits des cellules mononucleees peripheriques et des cellules CD4+ enrichies. L’expression des ARNm de Twist, T-plastine, NKp46, KIR3DL2 et Tox a ete quantifiee par RTqPCR. Les resultats ont ete compares entre les differents stades de LTC et les autres maladies cutanees par le t-test de Welch. La sensibilite et la specificite ont ete etudiees statistiquement avec des courbes ROC. Resultats Nous avons inclus 156 patients atteints de SS, 221 ayant un MF et 70 patients ayant une dermatose inflammatoire. Des differences significatives d’expression de T-plastine, Twist, KIR3DL2 et Tox ont ete observees entre les patients SS ou MFe et ceux atteints de dermatoses inflammatoires (pour SS p ≤ 0,001, pour MFe p ≤ 0,05). L’expression combinee de Twist et T-plastine permettait d’obtenir un diagnostic de SS ou MFe avec 95,2 % de specificite et 84 % de sensibilite. L’ajout de l’expression de KIR3DL2 permettait d’obtenir un diagnostic avec 90 % de specificite et 94 % de sensibilite. A noter que les patients exprimant KIR3DL2 rechutaient plus rapidement que les patients ne l’exprimant pas. Discussion La combinaison de l’expression de Twist et T-plastine est validee pour distinguer les MFe et SS des autres maladies cutanees dans cette nouvelle cohorte. L’ajout de KIR3DL2 a cette combinaison permet d’augmenter la sensibilite du diagnostic et l’expression de ce marqueur est associee a une rechute plus rapide. Conclusion Ces donnees affirment la validite de la combinaison des marqueurs Twist, T-plastine et KIR3DL2 comme nouvel outil rapide et efficace pour le diagnostic des LTC. Elles offrent de nouvelles perspectives therapeutiques, notamment en ciblant le marqueur KIR3DL2 a la surface des cellules tumorales.

Introduction L’alopecie androgenique affecte la zone frontale et le vertex. Les cellules du derme papillaire (DPCs) perdent leurs proprietes regeneratives in vitro en culture bidimensionnelle (2D). Il a ete recemment demontre que la structure tridimensionnelle (3D) des DPCs peut restaurer 22 % de la signature genetique moleculaire du derme papillaire intact. Cette etude compare la difference d’expression genetique du DP entre les cuirs chevelus alopecique et non alopecique et leurs variations en culture 2D, 3D et co-culture avec les cellules souches derivees des adipocytes (ASCs). Materiel et methodes Deux lignees de cellules immortalisees du cuir chevelu alopecique (B-DPC) et du cuir chevelu non alopecique (NB-DPC) ont ete etablies par notre equipe. Elles presentent respectivement 75 % et 65 % de signatures moleculaires de DP. Nous avons etudie si la culture 3D et la co-culture avec ASCs peuvent augmenter l’expression des marqueurs de folliculogenese. La culture 2D et les spheres ont ete creees de chaque lignee, suivies de l’analyse genetique par sequencage ARN (alignement 98 %, 14 millions lde ectures) et l’analyse par logiciel Panther. Resultats Les deux lignees immortalisees ont montre une augmentation de certains marqueurs de folliculogenese en 3D compares a 2D, comme WNT5A et FGF7. Cependant, l’augmentation de ces marqueurs est plus notable en co-culture avec ASCs qu’en 3D seule. L’expression de DKK1 (inhibiteur de la voie de signalisation Wnt) est fortement diminuee en 3D et co-culture comparee a 2D. Il a ete precedemment demontre que l’expression de DKK1 diminue dans les spheroides et nos resultats confirment ce fait. Contrairement aux DPC en culture 2D, le DPC intact est trichogenique. Certains genes presents en DP intact sont seulement sont surexprimes en co-culture avec ASCs et non pas en 3D comme COL23A1, PTGS1, MEGF10 ans RASD1. Les genes surexprimes, communs et differentiellement exprimes en B-DPC versus NB-DPC sont impliques dans les voies de signalisation inflammatoire. Alors que ceux surexprimes, communs et differentiellement exprimes en NB-DPC versus B-DPC, sont impliques dans les processus d’adhesion cellulaire. Conclusion En resume, les spheres de DPC et leur co-culture avec ASCs semblent avoir un profil plus inductif de folliculogenese que les DPC en culture 2D. Le modele de co-culture semble restaurer certains traits moleculaires propres au derme papillaire intact dans les lignees immortalisees mieux que la culture 3D.

Normal ageing is associated with an impaired systemic immune response contributing to an increased susceptibility to infectious diseases. The aim of this study was to compare the lymphocyte phenotype in human skin from old and young healthy subjects. Skin samples from donors were used for explant cultures before flow cytometric analysis. Our results depicted a higher proportion of CD4

Purpose Anti-angiogenic agents stand first in the treatment of neovascular diseases of the retina. CD160 appeared in several experimental studies as a marker of activated endothelial cells, suggesting it could represent a promising target for novel anti-angiogenic therapies. The aim of the present study was to assess the distribution of CD160 in the human eye, and to search for a possible correlation with retinal neovascular diseases. Methods The physiological distribution of CD160 in the normal eye was assessed with immunolabeling in 10 human donor eyes. Then, in a retrospective cohort of 75 surgical retinal specimens, the density of CD160+ microvessels was evaluated, along with immunolabeling on serial sections against ERG (pan-endothelial cell marker), CD105 (activated endothelial cell marker), and α-SMA (pericyte cell marker). The cohort was divided into two groups: 29 patients with neovascular disease (NV+) and 46 control patients (NV-). Results CD160 was physiologically expressed by several cell types: endothelial cells of retinal blood vessels, ganglion cells, macrophages, epithelial cells of the conjunctiva, ciliary body, and retinal pigment epithelium. In the patient cohort, the percentage of CD160+ vessels in the retina was significantly and independently higher in patients suffering from neovascular diseases (P = 0.04). On the contrary, the expression of CD105 was correlated neither with retinal neovascular diseases, nor with CD160 expression. Conclusions CD160 was expressed in some retinal vessels in both normal and pathologic eyes. CD160 expression by endothelial cells of retinal vessels was correlated with ocular neovascular diseases. CD160 could therefore represent an interesting target for novel anti-angiogenic therapies.

International audience; No abstract available

Introduction Corticotherapy is the leading medication worldwide for patients with history of repeated implantation failures (RIF) after IVF/ICSI. Nevertheless, we still do not know its local mechanism of action, hence its precise indication. Our objective is to document the impact of prednisone on the endometrial expression of immune biomarkers (CD56 cells count, IL-18/TWEAK, IL-15/Fn-14 mRNA ratio) at the time of uterine receptivity in a RIF population. Materials and method An endometrial biopsy was realized in the mid luteal phase for immune profiling: IL-15/Fn-14 and IL-18/TWEAK mRNA ratios were determined by quantitative RT-PCR and CD56 mobilization per IHC. Fifty-five patients with a RIF history were diagnosed to have local over-immune activation [high IL-18/TWEAK mRNA ratio, and/or high IL-15/Fn-14 mRNA ratio] likely to impair the implantation process. They underwent a second immune profiling with supplementation of prednisone. A paired comparison of the immune profile before and under prednisone was performed in the subset of patients subsequently pregnant under prednisone. Finding In 54.5% of the cases, both immune biomarkers were normalized and in 16.5%, only one was normalized under prednisone. In 29% we observed a paradoxical increase of both immune biomarkers. The IL-18/TWEAK mRNA ratio reflecting the Th-1/Th-2 local equilibrium was significantly reduced (0.29 versus 0.10, p = .004), through very significant increase of TWEAK expression, in patients who were subsequently pregnant under prednisone. Conclusion Testing the response to prednisone in a RIF context may be very useful. Less than half of RIF patients with immune deregulation may be prednisone responders and would benefit from its administration.

International audience; Interleukin 17B (IL-17B) is a pro-inflammatory cytokine that belongs to the IL-17 cytokines family and binds to IL-17 receptor B (IL-17RB). Here we found that high expression of IL-17B and IL-17RB is associated with poor prognosis in patients with breast cancer and that IL-17B expression upregulation is specifically associated with poorer survival in patients with basal-like breast cancer. We thus focused on IL-17B role in breast cancer by using luminal and triple negative (TN)/basal-like tumor cell lines. We found that IL-17B induces resistance to conventional chemotherapeutic agents. In vivo, IL-17B induced resistance to paclitaxel and treatment with an anti-IL-17RB neutralizing antibody completely restored breast tumor chemosensitivity, leading to tumor shrinkage. We next focused on the signaling pathways activated in human breast cancer cell lines upon incubation with IL-17B. We observed that IL-17B induces ERK1/2 pathway activation, leading to upregulation of anti-apoptotic proteins of the BCL-2 family. IL-17B-induced chemoresistance was completely abolished by incubation with PD98059, an inhibitor of the MAPK/ERK pathway, indicating that the ERK pathway plays a crucial role. Altogether our results emphasize the role of the IL-17B/IL-17RB signaling pathway in breast tumors and identify IL-17B and its receptor as attractive therapeutic targets for potentiating breast cancer chemotherapy.

Introduction KIR3DL2 est un recepteur NK exprime par 85 % des lymphomes T cutanes (LTC). IPH4102 est un anticorps (Ac) monoclonal « first-in-class » anti-KIR3DL2 qui deplete les cellules par des mecanismes d’ADCC et d’ADCP (cytotoxicite et phagocytose dependantes de l’anticorps). Il a ete teste pour la premiere fois chez l’homme dans une etude de phase I afin d’evaluer sa tolerance et de determiner la dose maximale toleree (DMT) et la dose recommandee pour une etude de phase 2 (RP2D). Les objectifs secondaires comportaient des etudes pharmacocinetiques et l’evaluation de l’efficacite clinique et biologique. Patients et methodes Les malades inclus etaient des LTC de stade avance refractaires a au moins 2 lignes de traitement systemique, dont les cellules tumorales exprimaient KIR3DL2 dans la peau et/ou le sang. L’escalade de dose comportait dix niveaux (0,0001 a 10 mg/kg). L’Ac etait administre par voie IV toutes les semaines pendant 4 semaines, toutes les 2 semaines jusqu’a la semaine 26 puis toutes les 4 semaines. Les malades pouvaient recevoir des doses croissantes et etaient traites jusqu’a progression ou survenue d’une toxicite. L’efficacite clinique etait evaluee selon les criteres internationaux. Les lymphocytes KIR3DL2 etaient monitores dans la peau par immunohistochimie et dans le sang par cytometrie de flux. La maladie residuelle etait evaluee par « deep sequencing » du recepteur T pour l’antigene. Resultats Vingt-cinq patients ont ete inclus : 20 syndromes de Sezary (SS), 4 mycosis fongoides (MF) et un LTC CD4+ NOS. L’âge median etait de 71 ans (42–90) et le nombre median de traitements precedents de 4 (2–14). Dix-neuf malades ont recu des doses croissantes d’Ac. Aucune toxicite limitante n’a ete observee et la DMT n’a pas ete atteinte. Six patients ont presente des evenements indesirables (EI) de grade 3, dont 2 lies au traitement et 2 des EI de grade 4 non lies au traitement. Les EI etaient pour la majorite des evenements de bas grade frequents lors de perfusions d’Ac. Le taux de reponses globales etait de 50 % pour les SS, dont 65 % de reponses dans le sang incluant 25 % de reponses completes. Les scores de prurit etaient tres ameliores chez tous les malades repondeurs et stables. Les lymphocytes KIR3DL2 ont ete progressivement et durablement depletes dans la peau et le sang. L’evaluation de la maladie residuelle a montre la disparition totale et durable du clone tumoral dans le sang et la peau. Une posologie de 10 mg/kg a ete choisie pour la phase d’extension. Conclusion Ces resultats montrent une excellente tolerance et une efficacite clinique tres prometteuse d’IPH4102 chez des malades ayant un LTC refractaire a plusieurs traitements systemiques. Une phase d’expansion de 15 malades ayant un syndrome de Sezary est en cours, les malades recevant une « flat dose » de 750 mg.

Reproductive immunology applies general immunology principles to specialised targets, reproduction and development. The involvement of colony-stimulating factors (CSFs) in reproduction illustrates this. The CSF family includes CSF-1 or macrophage CSF (M-CSF), CSF-2 or granulocyte macrophage CSF (GM-CSF), and CSF-3 or granulocyte CSF (G-CSF). Each member has a specific localisation and timed expression in the reproductive tract with specific functions involving them in ovulation, embryo implantation, placentation and further embryonic development. They are used in reproductive medicine, either as biomarkers of oocyte quality and competence (follicular G-CSF), or to supplement embryo culture media with human recombinant GM-CSF, or they are used as an innovative therapy by using human recombinant G-CSF for infertile patients. Given fundamental considerations on CSFs and their strong implication in reproduction, this review aimed to detail the current knowledge for each member of the family to improve our understanding of their implication in the maternal–foetal cytokinic dialogue and in possibly preventing reproductive disorders.

Interleukin 10 (IL-10)-producing B cells (regulatory B cells [Bregs]) regulate autoimmunity in mice and humans, and a regulatory role of IL-10-producing plasma cells has been described in mice. Dysfunction of B cells that maintain homeostasis may play a role in the pathogenesis of chronic graft-versus-host disease (cGVHD) after allogeneic stem cell transplantation. Here, we found a relation between decreased Breg frequencies and cGVHD severity. An impaired ability of B cells to produce IL-10, possibly linked to poor signal transducer and activator of transcription 3 and extracellular signal-regulated kinase phosphorylation, was found in patients with active cGVHD. IL-10 production was not confined to a single B-cell subset, but enriched in both the CD24(hi)CD27(+) and CD27(hi)CD38(hi) plasmablast B-cell compartments. In vitro plasmablast differentiation increased the frequency of IL-10-producing B cells. We confirmed that allogeneic transplant recipients had an impaired reconstitution of the memory B-cell pool. cGVHD patients had less CD24(hi)CD27(+) B cells and IL-10-producing CD24(hi)CD27(+) B cells. Patients with cGVHD had increased plasmablast frequencies but decreased IL-10-producing plasmablasts. These results suggest a role of CD24(hi)CD27(+) B-cell and plasmablast-derived IL-10 in the regulation of human cGVHD.

The ectonucleotidases CD39 and CD73 hydrolyze extracellular adenosine triphosphate (ATP) and adenosine diphosphate (ADP) to generate adenosine, which binds to adenosine receptors and inhibits T-cell and natural killer (NK)–cell responses, thereby suppressing the immune system. The generation of adenosine via the CD39/CD73 pathway is recognized as a major mechanism of regulatory T cell (Treg) immunosuppressive function. The number of CD39+ Tregs is increased in some human cancers, and the importance of CD39+ Tregs in promoting tumor growth and metastasis has been demonstrated using several in vivo models. Here, we addressed whether CD39 is expressed by tumor cells and whether CD39+ tumor cells mediate immunosuppression via the adenosine pathway. Immunohistochemical staining of normal and tumor tissues revealed that CD39 expression is significantly higher in several types of human cancer than in normal tissues. In cancer specimens, CD39 is expressed by infiltrating lymphocytes, the tumor stroma, and tumor cells. Furthermore, the expression of CD39 at the cell surface of tumor cells was directly demonstrated via flow cytometry of human cancer cell lines. CD39 in cancer cells displays ATPase activity and, together with CD73, generates adenosine. CD39+CD73+ cancer cells inhibited the proliferation of CD4 and CD8 T cells and the generation of cytotoxic effector CD8 T cells (CTL) in a CD39- and adenosine-dependent manner. Treatment with a CD39 inhibitor or blocking antibody alleviated the tumor-induced inhibition of CD4 and CD8 T-cell proliferation and increased CTL- and NK cell–mediated cytotoxicity. In conclusion, interfering with the CD39–adenosine pathway may represent a novel immunotherapeutic strategy for inhibiting tumor cell–mediated immunosuppression. Cancer Immunol Res; 3(3); 254–65. ©2014 AACR.

Objective: Neoadjuvant chemotherapy (NACT) allows for a more frequent use of breast-conservative surgery; it is also an in vivo model of individual tumor sensitivity which permits to determine new prognostic factors to personalize the therapeutic approach. Methods: Between 2000 and 2012, 318 patients with primary invasive breast cancer were treated with a median of 6 cycles of NACT; they received either an anthracycline-based FEC 100 protocol (31.1%), or anthracyclines + taxanes (53.5%), with trastuzumab if indicated (15.4%). Results: After a median follow-up of 44.2 months, the pathological complete response rate according to the classification of Chevallier et al. [Am J Clin Oncol 1993;16:223-228] was 19.3%, and overall (OS) and disease-free survival (DFS) at 10 years were 60.2 and 69.6%, respectively. Univariate analyses demonstrated that the Residual Disease in Breast and Nodes (RDBN) index was the most significant prognostic factor for OS (p = 0.0082) and DFS (p = 0.0022), and multivariate analyses mainly revealed that the residual tumor size, residual involved node number and post-chemotherapy Scarff-Bloom-Richardson (SBR) grading were the most significant prognostic factors. Conclusions: In a cohort of patients who were all homogeneously treated with some of the most common drugs for breast cancer, we demonstrate that NACT may provide additional prognostic factors and confirm the RDBN index. As this index allows for the prediction of survival with different breast cancer subtypes, we suggest that it should be calculated routinely to help clinicians to select patients who need adjuvant treatments.

We previously identified the NK cell receptor KIR3DL2 as a valuable diagnostic and prognostic marker for the detection of the tumoral T cell burden of Sezary syndrome (SS) patients. However, the function of this receptor on the malignant T lymphocyte population remained unexplored. We here demonstrate that engagement of KIR3DL2 by its recently identified ligand CpG oligodeoxynucleotide (ODN) induces the internalization of the receptor and leads to a caspase-dependent apoptosis of malignant T cells. This process of cellular death is correlated to a dephosphorylation of the transcription factor STAT3 (signal transducer and activator of transcription 3), which is found constitutively phosphorylated and activated in Sezary cells. Our results indicate that KIR3DL2 can directly promote SS malignant cell death through the use of CpG ODN.

To the editor: KIR3DL2, a killer immunoglobulin-like receptor normally expressed by a subset of natural killer (NK) cells, is aberrantly expressed in Sezary syndrome (SS), an aggressive cutaneous T-cell lymphoma (CTCL). In SS patients, the detection of KIR3DL2 expression on circulating tumor cells

BACKGROUND: Male androgenetic alopecia (AGA) is the most common form of hair loss in men and is characterized by a distinct pattern of progressive hair loss starting from the frontal area and the vertex of the scalp. Although several genetic risk loci have been identified, relevant genes for AGA remain to be defined. OBJECTIVES: Herein, molecular biomarkers associated with premature AGA were identified through gene expression analysis using cDNA generated from scalp vertex biopsies of hairless/bald men with premature AGA and healthy volunteers. RESULTS: This monocentric study reveals that genes encoding mast cell granule enzymes, inflammatory and immunoglobulin-associated immune mediators were significantly over-expressed in AGA. In contrast, under-expressed genes appear to be associated with the Wnt/β-catenin and BMP/TGF-β signaling pathways. Although involvement of these pathways in hair follicle regeneration is well-described, functional interpretation of the transcriptomic data highlights different events that account for their inhibition. In particular, one of these events depends on the dysregulated expression of proopiomelanocortin (POMC), as confirmed by RT-qPCR and immunohistochemistry. In addition, lower expression of CYP27B1 in AGA subjects supports the notion that changes in vitamin D metabolism contributes to hair loss. CONCLUSION: This study provides compelling evidence for distinct molecular events contributing to alopecia that may pave way for new therapeutic approaches. This article is protected by copyright. All rights reserved.

Autophagy is one of the chemotherapy resistance mechanisms in breast cancer. The aim of this study was to determine the level of recruitment of the autophagy pathway in the triple-negative breast cancer (TNBC) cell line MDA-MB231 compared with that in the control luminal breast cancer cell line MCF7 before and after treatment with chemotherapy drugs. Furthermore, we investigated the relationship between autophagy and EGFR, MUC1 and IL17-receptors as activators of autophagy. Immunohistochemistry was performed in cell culture blocks using LC3b, MUC1-C, EGFR, IL17A, IL17-RA and IL17-RB antibodies. We found that the basal autophagy level in MDA-MB231 was high, whereas it was low in MCF7. However, in contrast to MDA-MB231, the autophagy level was increased in MCF7 upon treatment with chemotherapy agents. Interestingly, we observed that the expression levels of MUC1-C, EGFR, IL17-RA, and IL17-RB were not modified by the same treatments. Furthermore, the chemotherapy treatments did not increase autophagy in TNBC cells without affecting the expression levels of MUC1-C, EGFR, IL17-RA or IL17-RB.

Summary Introduction Adult T-cell leukemia/lymphoma (ATLL) is a hematological malignancy associated with chronic HTLV-1 infection. Aim To describe skin lesions in ATLL. Methods A descriptive, retrospective study between 1996 and 2016, including all patients diagnosed with ATLL at Saint-Louis Hospital (Paris, France). Results Thirty-seven ATLL patients were included. Fifteen patients (41%) had a cutaneous localization of the disease, which was present from the beginning of the disease for two thirds of them. ATLL types in patients with cutaneous localization of the disease were as follows: lymphoma, n = 5, chronic, n = 4, smoldering, n = 4, acute, n = 2. Half the patients had 2 or more cutaneous manifestations. The cutaneous localizations observed were as follows: nodulotumoral (n = 8), plaques (n = 7), multipapular (n = 6), macular (n = 4), purpuric (n = 2). Among the 15 patients with cutaneous localization, median overall survival was significantly shorter in the acute and lymphoma types compared to the smoldering and chronic types (8.7 months vs. 79 months, P = 0.003). Discussion ATLL is a hematologic malignancy with variable expression that is diagnosed only very rarely in metropolitan France, but that should be sought in patients from countries with high HTLV-1 prevalence in the event of a chronic eruption with patches, papules, plaques and/or tumors. The chronic and smoldering types are relatively indolent, whereas the acute and lymphoma forms have a poor prognosis.

Allogeneic human cardiac-derived stem/progenitor cells (hCPC) are currently under clinical investigation for cardiac repair. While cellular immune response against allogeneic hCPC could be part of their beneficial-paracrine effects, their humoral immune response remains largely unexplored. Donor-specific HLA antibodies (DSA-HLA-I/DSA-HLA-II), primary elements of antibody-mediated allograft injury, might present an unidentified risk to allogeneic hCPC therapy. Here we established that the binding strength of anti-HLA monoclonal antibodies delineates hCPC proneness to antibody-mediated injury. In vitro modeling of clinical setting demonstrated that specific DSA-HLA-I of high/intermediate binding strength are harmful for hCPC whereas DSA-HLA-II are benign. Furthermore, the Luminex-based solid-phase assays are suitable to predict the DSA-HLA risk to therapeutic hCPC. Our data indicate that screening patient sera for the presence of HLA antibodies is important to provide an immune-educated choice of allogeneic therapeutic cells, minimize the risk of precipitous elimination and promote the allogeneic reparative effects.

Extranodal natural killer–T-cell lymphoma (NKTCL) of nasal type is a malignant disorder of cytotoxic lymphocytes of natural killer or more rarely T cells, associated with clonal Epstein–Barr virus infection. NKTCL is an aggressive neoplasm with very poor prognosis. Although the pathogenesis of NKTCL is little understood, some insight has been gained in the recent years, especially from genome-wide studies, which revealed a deletion on chromosome 6q21 in more than 50% of patients. Of interest, this deleted region contains four candidate tumor suppressor genes whose decreased expression has been confirmed at the mRNA level: PRDM1 , ATG5 , AIM1 , and HACE1 . Mutations and methylation in PRDM1 , ATG5 , and AIM1 have been reported in NKTCL cell lines. We investigated the involvement of HACE1 in NKTCL pathophysiology. Even though the hypermethylation of CpG-177 island located directly upstream of HACE1 locus led to down-regulation of HACE1 mRNA, the protein product was expressed at nearly normal levels and was functional in the NKTCL cell lines regardless of 6q21 deletion (and indeed no double deletion of 6q21 and no nonfunctional mutations have been reported). Furthermore, contrary to previous report, overexpression of HACE1 by transduction of recombinant protein did not affect proliferation or survival of NKTCL cell lines. We therefore conclude that HACE1 is not directly involved in NKTCL pathophysiology.

Advanced cutaneous T-cell lymphoma (CTCL) remains an unmet medical need, which lacks effective targeted therapies. In this study, we report the development of IPH4102, a humanized monoclonal antibody that targets the immune receptor KIR3DL2, which is widely expressed on CTCL cells but few normal immune cells. Potent antitumor properties of IPH4102 were documented in allogeneic human CTCL cells and a mouse model of KIR3DL2+ disease. IPH4102 antitumor activity was mediated by antibody-dependent cell cytotoxicity and phagocytosis. IPH4102 improved survival and reduced tumor growth in mice inoculated with KIR3DL2+ tumors. Ex vivo efficacy was further evaluated in primary Sézary patient cells, sorted natural killer–based autologous assays, and direct spiking into Sézary patient peripheral blood mononuclear cells. In these settings, IPH4102 selectively and efficiently killed primary Sézary cells, including at unfavorable effector-to-target ratios characteristic of unsorted PBMC. Together, our results offer preclinical proof of concept for the clinical development of IPH4102 to treat patients with advanced CTCL. Cancer Res; 74(21); 6060–70. ©2014 AACR.

Multifactorial predisposition to melanoma includes genes involved in pigmentation, immunity and DNA repair. Nonetheless, missing heritability in melanoma is still important. We studied the role of 335 candidate SNPs in melanoma susceptibility by using a dedicated chip and investigating 110 genes involved in different pathways. A discovery set was comprised of 1069 melanoma patients and 925 controls from France. Data were replicated using validation phases II (1085 cases and 801 controls from Spain) and III (1808 cases and 1894 controls from Germany and a second set of Spanish samples). In addition, an exome sequencing study was performed in three high-risk French melanoma families. Nineteen SNPs in 17 genes were initially associated with melanoma in the French population. Six SNPs were replicated in phase II, including two new SNPs in the WNT3 (rs199524) and VPS41 (rs11773094) genes. The role of VPS41 and WNT3 was confirmed in a meta-analysis (3940 melanoma cases and 3620 controls) with two-side p values of 0.002, (OR 50.86) and 4.07 310 210 (OR 50.80), respectively. Exome sequencing revealed a non-synonymous VPS41 variant in one family that was shown to be strongly associated with familial melanoma (OR 54.46, p 50.001) in an independent sample of 178 melanoma families. WNT3 belongs to WNT pathway known to play a crucial role in melanoma, whereas VPS41 regulates vesicular trafficking and is thought to play a role in pigmentation. Our work identified two new pathways involved in melanoma predisposition. These results may be useful in the future for identifying individuals highly predisposed to melanoma. The knowledge of the genetic predisposition to melanoma has significantly increased during the last several years. Linkage studies, candidate gene approaches, genome-wide association studies (GWAS) and whole-exome sequencing studies have identified several loci that influence susceptibility to cutaneous malignant melanoma (MM). 1–3 Mutations in the highly penetrant CDKN2A (MIM#60160), CDK4 (MIM#609048), TERT (MIM#187270) and POT1 (MIM#606478) genes are well

> Les lymphocytes B sont consideres comme des acteurs effecteurs de la reponse immunitaire : ils donnent naissance, au cours de leur differenciation terminale, aux plasmocytes qui produisent les anticorps ; ils produisent des cytokines, presentent l’antigene aux lymphocytes T effecteurs pour les activer, et regulent la fonction des cellules dendritiques [1] (Figure 1). La tolerance immunologique definit la capacite du systeme immunitaire a ne pas developper de reaction immunitaire forte visa-vis de certains antigenes et a reguler negativement les reponses immunitaires, afin, par exemple en cas d’infection, d’eteindre la reaction inflammatoire une fois le pathogene neutralise. Plusieurs populations de lymphocytes T contribuent au maintien de la tolerance immunitaire, parmi lesquelles les lymphocytes T regulateurs naturels (nTreg) CD4+CD25+FoxP3+ et les lymphocytes T regulateurs de type 1 (Tr1) qui produisent de l’interleukine 10 (IL-10), une cytokine anti-inflammatoire [2]. Parallelement, de nombreux travaux chez la souris ont montre que les lymphocytes B pouvaient avoir des fonctions regulatrices, celles-ci etant pour l’instant essentiellement attribuees a la capacite de ces lymphocytes B a produire de l’IL-10 [3]. Bien que la capacite des lymphocytes B humains a produire de l’IL-10 soit connue depuis longtemps [4], l’existence de lymphocytes B regulateurs (Breg) humains n’a ete demontree que recemment, et leur ontogenie et leurs mecanismes d’action chez l’homme sont encore tres mal compris.

CD160 is a GPI-anchored Ig-like receptor identified by the BY55 mAb on human circulating CD56dim+ NK cells and TCRγδ lymphocytes. In addition, while most intestinal T lymphocytes express it, only a minor circulating CD4+ or CD8+ T lymphocyte subset is CD160+. Here we describe a population of CD4+CD160+ human blood T lymphocytes of circulating cutaneous T cells. These rare T lymphocytes represent 2.1 ± 1.9% of the circulating CD3+CD4+ T cells, coexpress CD8αα, CD244, and perforin but lack CD28 expression, a phenotype corresponding to effector memory cytotoxic T-lymphocytes. Functional studies further confirmed their cytotoxic potential. These cells lack αEβ7 integrin and CCR7 expression but do express skin-addressing molecules CLA, and CCR4. In normal human skin, CD4+CD160+ cells represent 34.6 ± 14.7% of the CD4+ T lymphocytes extracted by collagenase treatment. These T cells coexpress CLA (81 ± 13.6%), CCR4 (62.3 ± 15.9%), and some CD8αα (19.6 ± 13%) or CCR7 (24.4 ± 11.7%) expression. Cutaneous T-cell lymphoma cells express the natural killer receptor KIR3DL2 (CD158k) used as a tumor marker. Not only we confirmed the expression of this marker in the blood and/or skin of mycosis fungoides patients but we also show for the first time CD158k expression (often associated with CD160) on cutaneous CD4+ T cells from healthy individuals (25.3 ± 15%). Therefore, CD4+CD160+ T cells expressing CD158k might represent specialized cutaneous lymphocytes devoted to immune surveillance, from which could originate cutaneous T-cell lymphomas such as mycosis fungoides. © 2014 International Society for Advancement of Cytometry

Summary Background Alemtuzumab has been proposed as salvage therapy for refractory cuta-neous T-cell lymphomas (CTCLs). Long-term follow-up data are scarce.Objectives To assess the efficacy and safety of alemtuzumab in the treatment ofadvanced CTCL.Methods A multicentre retrospective analysis was carried out of 39 patients withadvanced CTCL treated with alemtuzumab between 2003 and 2013.Results Thirty-nine patients (median age 62 years, range 20–83) with Sezary syn-drome (SS, n = 23) or advanced mycosis fungoides (MF, n = 16) received ale-mtuzumab 30 mg two to three times per week for a median duration of12 weeks (range 1–35). Fifteen patients received maintenance therapy for a med-ian duration of 24 weeks (range 6–277). Eleven patients (28%) had transformeddisease (MF, n = 10; SS, n = 1). After a median follow-up of 24 months (range0 3–124), eight patients (21%) were still alive. The overall response rate was51% in the whole study group (partial response, n = 13; complete response,n = 7); 70% in patients with SS and 25% in patients with MF (P = 0 009). Themedian time to progression was 3 4 months (range 0 4–42). Six patients (15%;SS, n = 5; MF, n = 1) remained progression free for > 2 years (median56 months, range 28–117). Five patients experienced cutaneous large T-celltransformation during alemtuzumab treatment and one patient developedprimary cutaneous large B-cell lymphoma. Twenty-four patients (62%) had agrade three or higher infectious adverse event and 10 (26%) a haematologicaltoxicity, which led to treatment discontinuation in 17 cases (44%) and deathin two (5%).Conclusions Alemtuzumab may induce long-term remission in SS but seems inef-fective in MF and transformed CTCL.

Primary mucosal melanomas represent a minority of melanomas, but have a significantly worse prognosis than cutaneous melanomas. A better characterization of the molecular pathogenesis of this melanoma subtype could help us understand the risk factors associated with the development of mucosal melanomas and highlight therapeutic targets. Because the Mitogen-Activated Protein Kinase (MAPK) pathway plays such a significant role in melanoma development, we explore v-raf murine sarcoma viral oncogene homolog B (BRAF) and neuroblastoma RAS viral oncogene homolog (NRAS) mutations in mucosal melanoma and compare them to the mutation profiles in cutaneous melanoma and other tumors with BRAF and NRAS mutations. We show that in addition to being less frequent, BRAF and NRAS mutations are different in mucosal melanoma compared to cutaneous melanomas. Strikingly, the BRAF and NRAS mutation profiles in mucosal melanoma are closer to those found in cancers such as lung cancer, suggesting that mutations in mucosal melanoma could be linked to some genotoxic agents that remain to be identified. We also show that the atypical BRAF and NRAS mutations found in mucosal melanomas have particular effects on protein activities, which could be essential for the transformation of mucosal melanocytes.

Introduction La maladie de Still de l’adulte (MSA) est une maladie rare associee a une activation macrophagique, neutrophilique et a une secretion incontrolee de cytokines pro-inflammatoires. La presentation dermatologique de la MSA est caracterisee par une eruption cutanee typique evanescente (ECTE), mais de nombreux cas d’eruption cutanee atypique persistante (ECAP) associee a un moins bon pronostic ont ete rapportes. Nous avons etudie l’expression des cytokines et chimiokines inflammatoires dans les lesions cutanees de MSA et avons recherche une eventuelle difference d’expression dans les ECTE et les ECAP. Materiel et methodes Sept prelevements cutanes de MSA (4 ECTE et 3 ECAP) et 3 prelevements cutanes de peau saine etaient recueillis et inclus en paraffine. Apres extraction et dosage de l’ARN (ratio A260/A280 : 1,6–2,0) a partir de 5 coupes de 20 μm d’epaisseur, une RT-PCR quantitative etait realisee pour les genes suivants : IL-1β, IL-18, IL-6, IL-8, IFNγR, TNFα, MCP1, CXCL10, CXCL13 et β2 m. Resultats La comparaison des lesions cutanees des patients avec une MSA et des peaux saines de sujets temoins mettait en evidence une augmentation des taux d’IL-1β (p = 0,01), de CXCL13 (p = 0,001), d’IFNγR (p = 0,02) et une tendance a l’augmentation d’IL-6 (p = 0,06) dans les lesions cutanees de MSA. Par contre aucune modification significative des taux d’IL-18, d’IL-8, de TNFα, de MCP1 et de CXCL10 n’a ete trouvee. Lorsqu’on comparait les ECAP aux ECTE on pouvait observer une tendance a l’augmentation des taux d’IL-1β (p = 0,08) et d’IL-6 (p = 0,1). Concernant l’IL-18, l’IL-8, le MCP1, le CXCL10, le CXCL13, le TNFα et l’IFNγR, aucune variation significative n’etait obtenue entre les deux types de lesions cutanees. Discussion Les taux seriques d’IL-1β, d’IL-18, d’IL-6, d’IL-8, de TNFα et d’IFNγ sont augmentes chez les patients ayant une MSA, certaines de ces cytokines etant correlees a l’activite de la maladie. Nous montrons ici une augmentation de l’IL-1β, de l’IFNγR, de CXCL13, ainsi que de l’IL-6 (tendance) dans les lesions cutanees de ces patients. Par ailleurs, une tendance a l’augmentation d’expression d’IL-1β et d’IL-6 dans les ECAP comparees aux ECTE pourrait etre en accord avec une maladie plus severe chez les patients presentant ce type de lesions. Conclusion Les cytokines inflammatoires sont augmentees dans les lesions cutanees de la MSA, certaines semblent davantage exprimees dans les eruptions cutanees atypiques que dans le rash evanescent classique, ces resultats devant etre confirmes sur une population plus large de patients.