205 AgenT-797, a native allogeneic ‘off-the-shelf’ iNKT cell therapy product shows anti-tumor activity in preclinical xenograft models

BackgroundagenT-797 is an allogeneic, native invariant natural killer T (iNKT) cell therapy product currently in phase I clinical trials for cancer (heme and solid). iNKT cells are a distinct population of T cells that can recognize tumors via direct recognition of CD1d (an MHC-I like molecule presenting glycolipids) through the TCR or recognition of NK cell receptor ligands via various NK receptors. We developed agenT-797 from isolated and ex-vivo expanded peripheral blood iNKT cells. Here we describe in vivo xenograft models to demonstrate the overall tissue distribution, tumor infiltration and efficacy of agenT-797 in liquid as well as solid tumors.MethodsWe utilized NOG-hIL15 (human IL-15) transgenic mice to ensure persistence/maintenance of ex-vivo expanded human iNKT cells throughout the studies. For studying efficacy in liquid tumors, we used NALM6, an acute lymphoblastic leukemia (ALL) cell line and for solid tumors selected A375, a melanoma cell line. Both cell lines were engineered to overexpress CD1d. Upon injection of iNKT cells, tumor growth and iNKT cell tissue/tumor infiltration as well as phenotype were studied.ResultsInjection of iNKT cells in NALM6- engrafted NOG-hIL15 mice resulted in an overall reduction in leukemic burden as measured by luminescence-based imaging. Flow cytometric analysis revealed infiltration of iNKT cells at the site of leukemic expansion, namely blood, spleen, bone marrow and liver. Cells were activated when reaching the site of the tumor. In addition, iNKT cells produced IFNγ and TNFα and low levels of IL-13/IL-4, consistent with a Th1 response. When iNKT cells were injected into A375 engrafted mice we observed infiltration of iNKT cells into the tumor, where they become activated and proliferate overtime. We observed an overall reduction in tumor size when iNKT cells were injected compared to control group, demonstrating the impact of iNKT cells on tumor growth.ConclusionsWe established xenograft mouse models to address various biological questions around human iNKT cells as a cell therapy product. We have demonstrated homing and infiltration of iNKT cells at the site of tumor and relative proliferation and expansion. These models provide a suitable platform for in-vivo preclinical studies on agenT -797 in cancer.Ethics ApprovalAll procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.

Selecting a Cell Engineering Methodology During Cell Therapy Product Development

When considering the development pathway for a genetically modified cell therapy product, it is critically important that the product is engineered consistent with its intended human use. For scientists looking to develop and commercialize a new technology, the decision to select a genetic modification method depends on several practical considerations. Whichever path is chosen, the developer must understand the key risks and potential mitigations of the cell engineering approach. The developer should also understand the clinical implications: permanent/memory establishment versus transient expression, and clinical manufacturing considerations when dealing with transplantation of genetically engineered cells. This review covers important topics for mapping out a strategy for developers of new cell-based therapeutics. Biological, technological, manufacturing, and clinical considerations are all presented to map out development lanes for the initiation and risk management of new gene-based cell therapeutic products for human use.

772 A potent and off-the-shelf oNK cell therapy product targets HER2+ cancer cells and resists suppressive tumor microenvironment

BackgroundAutologous or allogeneic natural killer (NK) cells possess efficient cytotoxicity against tumor cells without severe side effects such as CRS or graft-versus-host disease (GvHD). In addition to chimeric antigen receptor (CAR) strategy, antibody-cell conjugates (ACC) platform provides more efficient way to arm NK cells with binding specificity and enhanced potency against target cells. In this work, we develop a NK cell therapy product ACE1702, a novel NK cell line oNK conjugated with trastuzumab, and assess its potency against HER2+ solid tumors.Methods oNK cells were covalently conjugated with monoclonal antibody Trastuzumab after sublethal irradiation by our patented antibody-cell conjugates (ACC) platform to become our cryopreserved final product ACE1702 compliant with current good manufacturing practice (cGMP). Function of ACE1702 was validated by real-time xCELLigence analyzer and MTT assay in vitro. Efficacy of intraperitoneally (ip.) delivered ACE1702 was evaluated in tumor-bearing female immune compromised NSG mice. Characterization of ACE1702 was analyzed by flow cytometry.ResultsWe demonstrated that the trastuzumab-armed oNK cells, ACE1702, exerted human epidermal growth factor 2 (HER2) binding specificity and enhanced cytotoxicity against various types of cancer cells with different grade of HER2 expressions compared to control oNK cells in vitro. In vivo results in human ovarian cancer cell line SK-OV-3-bearing xenograft mouse model further supported the in vitro observations. Of note, ACE1702 also displayed a better cytotoxicity against HER2+ cancer cells than trastuzumab and its derived antibody-drug conjugate. ACE1702 also remained cytotoxicity against cancer cells in the suppressive tumor microenvironment. Characterization revealed a preferential expression of NK activation receptors, and conjugation of trastuzumab with cell membrane proteins responsible for NK activity capacitated ACE1702 with enhanced cytotoxicity. These results underscore the potency of ACE1702 in eradication of cancer cells.ConclusionsHere we introduced a novel trastuzumab-modified oNK cell product with enhanced specificity against myriad types of HER2+ cancers. Selective conjugation of trastuzumab with membrane proteins contributing to NK activation conferred ACE1702 with enhanced cytotoxicity even in the suppressive tumor microenvironment.AcknowledgementsNoneTrial RegistrationNoneEthics ApprovalThe animal study was conducted according to protocols approved by the Institutional Animal Care and Use Committee of Muragenics.ConsentNone

From protocol to product: ventral midbrain dopaminergic neuron differentiation for the treatment of Parkinson's disease

Current cell therapy product limitations include the need for in-depth product understanding to ensure product potency, safety and purity. New technologies require development and validation to address issues of production scale-up to meet clinical need; assays are required for process control, validation and release. Prior to clinical realization, an understanding of production processes is required to implement process changes that are essential for process control. Identification of key parameters forms the basis of process tolerances, allowing for validated, adaptive manufacturing processes. This enables greater process control and yield while withstanding regulatory scrutiny. This report summaries key milestones in specifically for ventral midbrain dopaminergic neuroprogenitor differentiation and key translational considerations and recommendations to enable successful, robust and reproducible current cell therapy product-manufacturing.