CIBLAGE DU GANGLIOSIDE GD2 O-ACÉTYLÉ COMME NOUVELLE STRATÉGIE THÉRAPEUTIQUE ET DIAGNOSTIQUE DANS LE CANCER À CELLULES SOUCHES TUMORALES

Type de la demande A1 N° et date de dépôt EP14723706

UTILISATION D'UN ANTICORPS CONTRE LE GANGLIOSIDE GD2 O-ACÉTYLÉ DANS LE BUT D'AMÉLIORER LE POTENTIEL THÉRAPEUTIQUE DE MÉDICAMENTS

Type de la demande A1 N° et date de dépôt EP1782959

Identification of 9-O-acetyl-N-acetylneuraminic acid (Neu5,9Ac2) as main O-acetylated sialic acid species of GD2 in breast cancer cells

International audienc

O-acetylated gangliosides: Structure, biosynthesis, immunogenicity, functions and their potential for cancer immunotherapy

International audienceSialic acid O-acetylation is a developmentally regulated modification of gangliosides implicated in ontogeny and tumor progression. Their existence has been underestimated in the past because of their alkali-labile nature and their transient expression. New data indicates, however, that O-acetylated gangliosides perform important function in tumor malignancy. Best studied O-acetyl-GD3 blocks the pro-apoptotic activity of GD3 and promotes survival of cancer cells. In acute lymphoblastic leukaemia cells, O-acetyl-GD3 expression level also correlates with survival and drug resistance. The recent identification of the enigmatic O-acetyltransferase opens new experimental approaches for designing novel effective therapeutics targeting drug-resistant cancer cells in acute lymphoblastic leukaemia. In addition, O-acetylated gangliosides expressed at the tumor cell surface are accessible for specific monoclonal antibodies to inhibit cell growth, to induce apoptosis, and to inhibit tumor metastasis formation. Thus, passive immunotherapy using murine or murine/human chimeric monoclonal anti-O-acetylated ganglioside antibodies are currently being investigated. Particularly, targeting of O-acetyl-GD2 could reduce the acute toxicities currently associated with anti-GD2 therapeutic antibodies. This review summarizes the molecular mechanisms involved in the biosynthesis and the expression of O-acetylated gangliosides and presents the new experimental approaches that allow the characterization of their importance in tumor progression. The different strategies used by different teams to develop specific monoclonal antibodies against these poorly immunogenic glycolipids for therapeutic application are also discussed

Targeting and killing glioblastoma with monoclonal antibody toO-acetyl GD2 ganglioside

International audienceThere are still unmet medical needs in the treatment of glioblastoma, themost common and the most aggressive glioma of all brain tumors. Here, we foundthat O-acetyl GD2 is expressed in surgically resected human glioblastoma tissue.In addition, we demonstrated that 8B6 monoclonal antibody specific for O-acetylatGD2 could effectively inhibit glioblastoma cell proliferation in vitro and in vivo. Takentogether, these results indicate that O-acetylated GD2 represents a novel antigen forimmunotherapeutic-based treatment of high-grade gliomas

Targeting and killing glioblastoma with monoclonal antibody toO-acetyl GD2 ganglioside

International audienceThere are still unmet medical needs in the treatment of glioblastoma, themost common and the most aggressive glioma of all brain tumors. Here, we foundthat O-acetyl GD2 is expressed in surgically resected human glioblastoma tissue.In addition, we demonstrated that 8B6 monoclonal antibody specific for O-acetylatGD2 could effectively inhibit glioblastoma cell proliferation in vitro and in vivo. Takentogether, these results indicate that O-acetylated GD2 represents a novel antigen forimmunotherapeutic-based treatment of high-grade gliomas

Chimeric Antibody c.8B6 to O-Acetyl-GD2 Mediates the Same Efficient Anti-Neuroblastoma Effects as Therapeutic ch14.18 Antibody to GD2 without Antibody Induced Allodynia

<div><p>Background</p><p>Anti-GD2 antibody is a proven therapy for GD2-postive neuroblastoma. Monoclonal antibodies against GD2, such as chimeric mAb ch14.18, have become benchmarks for neuroblastoma therapies. Pain, however, can limit immunotherapy with anti-GD2 therapeutic antibodies like ch14.18. This adverse effect is attributed to acute inflammation via complement activation on GD2-expressing nerves. Thus, new strategies are needed for the development of treatment intensification strategies to improve the outcome of these patients.</p><p>Methodology/Principal Findings</p><p>We established the mouse-human chimeric antibody c.8B6 specific to OAcGD2 in order to reduce potential immunogenicity in patients and to fill the need for a selective agent that can kill neuroblastoma cells without inducing adverse neurological side effects caused by anti-GD2 antibody immunotherapy. We further analyzed some of its functional properties compared with anti-GD2 ch14.18 therapeutic antibody. With the exception of allodynic activity, we found that antibody c.8B6 shares the same anti-neuroblastoma attributes as therapeutic ch14.18 anti-GD2 mAb when tested in cell-based assay and <i>in vivo</i> in an animal model.</p><p>Conclusion/Significance</p><p>The absence of OAcGD2 expression on nerve fibers and the lack of allodynic properties of c.8B6–which are believed to play a major role in mediating anti-GD2 mAb dose-limiting side effects–provide an important rationale for the clinical application of c.8B6 in patients with high-risk neuroblastoma.</p></div

Intravenous injection of mAb c.8B6 does not induce allodynia.

<p>(A) Intraveinous injection of either 1 mg/kg (•) or 3 mg/kg (♦)of mAb c.8B6 in Sprague-Dawley rats did not result in a decrease in mechanical thresholds. In contrast, ch14.18 injected at 1 mg/kg (▪) resulted in a prolonged decrease in threshold. Threshold of animal injected with anti-CD20 chimeric control antibody rituximab (1 mg/kg) (□) are shown for comparison. (B) Results shown in panel B were recalculated as area under the curve and then normalized such that 100 would be equal to a theoretical maximal allodynia. Animals administered ch14.18 displayed more allodynia than any of the other treatment groups (p≤ 0.0001).</p

Binding specificity chimeric mAbs c.8B6 and ch14.18 measured by flow-cytometric analysis (A), by TLC-immunostaining (B) and by Immunochemistry on NXS2 tumor (C).

<p>(A) NXS2 and the OAcGD2/GD2⁻ Neuro2A cells were stained as described in Materials and Methods. Representative flow cytometry histogram of OAcGD2 expression. Antibody c.8B6 or mAb ch14.18 (<i>grey</i>) and control antibody (<i>black</i>). The same staining pattern was observed for mAb c.8B6 or ch14.18 on OAcGD2/GD2⁺ NXS2 cells whereas neither mAb c.8B6 nor mAb ch14.18 bind OAcGD2/GD2-negative Neuro-2a cells. These experiments were independently replicated 3 times. (B) TLC of gangliosides extracted from rat brain (Lane 0) and NXS2 cells with (Lane 1) or without (Lane 2) alkaline treatment and stained with resorcinol-HCl (Panel a) or immunostained with mAbs c.8B6 (Panel b), or ch14.18 (Panel c). Chimeric mAb 8B6 reacted with the alkali-labile <i>O</i>AcGD2 ganglioside with no cross-reactivity against GD2 ganglioside (Panel b) whereas ch14.18 reacted with GD2 and OAcGD2 (Panel c). The same results were obtained in 3 independent experiments. (C) An immunoperoxidase assay was performed on NX2 neuroblastoma tumor sections as described in Material and Methods. Strong immunostaining was detected on neuroblastoma cells with either mAb c.8B6 (c) or mAb ch14.18 (d). The anti-CD20 chimeric antibody ritximab was used as a negative control (b). No antibody (a). NXS2 neuroblastoma tumors from 6 different mice were tested with the same results. Scale bar = 20 µm.</p

Immune effector functions of both mAbs c.8B6 and ch14.18.

<p>(A) Complement-dependent specific lysis was determined for the NXS2 cells as described in Materials and Methods. (B) The NK-92-RFcγIII+ ADCC activity with NXS2 target cells at E/T ratio 12 to 1 as described in Materials and Methods. Horizontally hatched columns, mAb c.8B6; white columns, mAb ch14.18; black columns, irrelevant antibody. Data are presented as the mean ± SEM for three independent experiments, each in triplicate.</p