Benoît Grellier

A unique anti-CD115 monoclonal antibody which inhibits osteolysis and skews human monocyte differentiation from M2-polarized macrophages toward dendritic cells

Cancer progression has been associated with the presence of tumor-associated M2-macrophages (M2-TAMs) able to inhibit anti-tumor immune responses. It is also often associated with metastasis-induced bone destruction mediated by osteoclasts. Both cell types are controlled by the CD115 (CSF-1R)/colony-stimulating factor-1 (CSF-1, M-CSF) pathway, making CD115 a promising target for cancer therapy. Anti-human CD115 monoclonal antibodies (mAbs) that inhibit the receptor function have been generated in a number of laboratories. These mAbs compete with CSF-1 binding to CD115, dramatically affecting monocyte survival and preventing osteoclast and macrophage differentiation, but they also block CD115/CSF-1 internalization and degradation, which could lead to potent rebound CSF-1 effects in patients after mAb treatment has ended. We thus generated and selected a non-ligand competitive anti-CD115 mAb that exerts only partial inhibitory effects on CD115 signaling without blocking the internalization or the degradation of the CD115/CSF-1 complex. This mAb, H27K15, affects monocyte survival only minimally, but downregulates osteoclast differentiation and activity. Importantly, it inhibits monocyte differentiation to CD163+CD64+ M2-polarized suppressor macrophages, skewing their differentiation toward CD14-CD1a+ dendritic cells (DCs). In line with this observation, H27K15 also drastically inhibits monocyte chemotactic protein-1 secretion and reduces interleukin-6 production; these two molecules are known to be involved in M2-macrophage recruitment. Thus, the non-depleting mAb H27K15 is a promising anti-tumor candidate, able to inhibit osteoclast differentiation, likely decreasing metastasis-induced osteolysis, and able to prevent M2 polarization of TAMs while inducing DCs, hence contributing to the creation of more efficient anti-tumor immune responses.

3D modeling and characterization of the human CD115 monoclonal antibody H27K15 epitope and design of a chimeric CD115 target

The humanized monoclonal antibody H27K15 specifically targets human CD115, a type III tyrosine kinase receptor involved in multiple cancers and inflammatory diseases. Binding of H27K15 to hCD115 expressing cells inhibits the functional effect of colony-stimulating factor-1 (CSF-1), in a non-competitive manner. Both homology modeling and docking programs were used here to model the human CD115 extracellular domains, the H27K15 variable region and their interaction. The resulting predicted H27K15 epitope includes mainly the D1 domain in the N-terminal extracellular region of CD115 and some residues of the D2 domain. Sequence alignment with the non-binding murine CD115, enzyme-linked immunosorbent assay, nuclear magnetic resonance spectroscopy and affinity measurements by quartz crystal microbalance revealed critical residues of this epitope that are essential for H27K15 binding. A combination of computational simulations and biochemical experiments led to the design of a chimeric CD115 carrying the human epitope of H27K15 in a murine CD115 backbone that is able to bind both H27K15 as well as the murine ligands CSF-1 and IL-34. These results provide new possibilities to minutely study the functional effects of H27K15 in a transgenic mouse that would express this chimeric molecule.

Viral based vaccine TG4010 induces broadening of specific immune response and improves outcome in advanced NSCLC

BackgroundAdvanced non-small cell lung cancer patients receiving TG4010, a therapeutic viral vaccine encoding human Mucin 1 and interleukin-2 in addition to standard chemotherapy, displayed longer overall survival in comparison to that of patients treated with standard chemotherapy alone. Our study intended to establish the association between overall survival and vaccine-induced T cell responses against tumor associated antigens (TAA) targeted by the vaccine.MethodThe TIME trial was a placebo-controlled, randomized phase II study aimed at assessing efficacy of TG4010 with chemotherapy in NSCLC. 78 patients from the TIME study carrying the HLA-A02*01 haplotype were analyzed using combinatorial encoding of MHC multimers to detect low frequencies of cellular immune responses to TG4010 and other unrelated TAA.ResultsWe report that improvement of survival under TG4010 treatment correlated with development of T cell responses against MUC1. Interestingly, responses against MUC1 were associated with broadening of CD8 responses against non-targeted TAA, thus demonstrating induction of epitope spreading.ConclusionOur results support the causality of specific T-cell response in improved survival in NSCLC. Additionally, vaccine induced epitope spreading to other TAA participates to the enrichment of the diversity of the anti-tumor response. Hence, TG4010 appears as a useful therapeutic option to maximize response rate and clinical benefit in association with other targeted immuno-modulators.Trial registrationRegistered on ClinicalTrials.gov under identifier NCT01383148 on June 23rd, 2011.

Vaccinia Virus Shuffling: deVV5, a Novel Chimeric Poxvirus with Improved Oncolytic Potency

Oncolytic virus (OV) therapy has emerged as a promising approach for cancer treatment with the potential to be less toxic and more efficient than classic cancer therapies. Various types of OVs in clinical development, including Vaccinia virus (VACV)-derived OVs, have shown good safety profiles, but limited therapeutic efficacy as monotherapy in some cancer models. Many different methods have been employed to improve the oncolytic potency of OVs. In this study, we used a directed evolution process, pooling different strains of VACV, including Copenhagen, Western Reserve and Wyeth strains and the attenuated modified vaccinia virus Ankara (MVA), to generate a new recombinant poxvirus with increased oncolytic properties. Through selective pressure, a chimeric VACV, deVV5, with increased cancer cell killing capacity and tumor selectivity in vitro was derived. The chimeric viral genome contains sequences of all parental strains. To further improve the tumor selectivity and anti-tumor activity of deVV5, we generated a thymidine kinase (TK)-deleted chimeric virus armed with the suicide gene FCU1. This TK-deleted virus, deVV5-fcu1 replicated efficiently in human tumor cells, and was notably attenuated in normal primary cells. These studies demonstrate the potential of directed evolution as an efficient way to generate recombinant poxviruses with increased oncolytic potency, and with high therapeutic index to improve cancer therapy.

Abstract 288: TG3003, an immunomodulatory anti-CD115 mAb targeting M2-macrophage polarization in the tumor microenvironment

Cancer progression has been associated with the presence of tumor-associated M2-type macrophages (M2-TAMs) able to inhibit anti-tumor immune responses, stimulate neo-angiogenesis and facilitate metastasis. Colony-stimulating factor-1 (CSF-1, M-CSF) is a cytokine required for the survival and differentiation of myeloid cell lineages, and CSF-1 signaling is known to polarize macrophages towards the M2-type. M2-TAMs can represent the most abundant immunosuppressive cell population in the tumor microenvironment, notably recruited by CSF-1 and MCP-1/CCL2. Transgene has developed a monoclonal antibody (mAb), TG3003, directed against the CSF-1 cell-surface receptor, CD115 (CSF-1R, M-CSFR). This mAb does not block the binding of CSF-1 to its receptor, but down-modulates CD115 signaling. In contrast to other anti-CD115 mAbs currently in development, whose modes of action rely on the blockade of ligand binding, TG3003 is not cytotoxic to normal myeloid cells that require CD115-mediated signaling for their survival. In vitro, TG3003 skews monocyte differentiation from M2-type macrophages towards dendritic cells, most potent antigen-presenting cells capable of stimulating efficacious T cell responses. It inhibits the secretion of MCP-1/CCL2 by differentiating macrophages and decreases their IL-6 production. Through this inhibition of M2-TAMs, TG3003 may potentiate immune responses in patients and impact on tumor progression. Moreover, due to its unique non-competitive mode of action, TG3003 does not block the physiological pathway for CSF-1 clearing from the circulation, thus avoiding the issue of toxic or rebound effects in treated patients. To investigate the properties of TG3003 in vivo, we have generated a transgenic mouse strain where the mAb epitope has been inserted into murine CD115 without affecting murine CSF-1 binding nor signaling. We will present the results of preclinical proof-of-concept experiments validating the mechanism of action and the immunomodulatory properties of mAb TG3003. Citation Format: Helene Haegel, Christelle Ziller-Remy, Luc Barraud, Jean-Yves Bonnefoy, Sandrine Cochin, Vanessa Duong, Michel Geist, Benoit Grellier, Remy Hallet, Jean-Baptiste Marchand, Thierry Menguy, Ronald Rooke, Christine Thioudellet, Carine Reymann, Xavier Preville. TG3003, an immunomodulatory anti-CD115 mAb targeting M2-macrophage polarization in the tumor microenvironment. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 288. doi:10.1158/1538-7445.AM2015-288