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Safe and Reproducible Preparation of Functional Dendritic Cells for Immunotherapy in Glioblastoma Patients.

ABSTRACT: Cell therapy based on dendritic cells (DCs) pulsed with tumor lysate is a promising approach in addition to conventional therapy for the treatment of patients with glioblastoma (GB). The success of this approach strongly depends on the ability to generate high-quality, functionally mature DCs (mDCs), with a high level of standardization and in compliance with Good Manufacturing Practices. In the cell factory of the Carlo Besta Foundation, two phase I clinical trials on immunotherapy with tumor lysate-loaded DCs as treatment for GB are ongoing. From 2010 to 2014, 54 patients were enrolled in the studies and 54 batches of DCs were prepared. We retrospectively analyzed the results of the quality control tests carried out on each produced batch, evaluating yield of mDCs and their quality in terms of microbiological safety and immunological efficacy. The number of mDCs obtained allowed the treatment of all the enrolled patients. All 54 batches were sterile, conformed to acceptable endotoxin levels, and were free of Mycoplasma species and adventitious viruses. During culture, cells maintained a high percentage of viability (87%-98%), and all batches showed high viability after thawing (mean±SD: 94.6%±2.9%). Phenotype evaluation of mDCs showed an evident upregulation of markers typical of DC maturation; mixed lymphocyte reaction tests for the functional evaluation of DCs demonstrated that all batches were able to induce lymphocyte responses. These results demonstrated that our protocol for DC preparation is highly reproducible and permits generation of large numbers of safe and functional DCs for in vivo use in immunotherapy approaches.Cell therapy based on antigen-pulsed dendritic cells (DCs) is a promising approach for the treatment of glioblastoma patients. The success of this approach strongly depends on the ability to generate high-quality, functional DCs with a high level of standardization, ensuring reproducibility, efficacy, and safety of the final product. This article summarizes the results of the quality controls on 54 batches, to demonstrate the feasibility of producing a therapeutic cell-based vaccine via a well-controlled Good Manufacturing Practices (GMP)-compliant production process. The findings may be of scientific interest to those working in the field of preparation of GMP-compliant products for cell-therapy applications.

SUBMITTER: Nava S

PROVIDER: S-EPMC4572908 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

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Safe and Reproducible Preparation of Functional Dendritic Cells for Immunotherapy in Glioblastoma Patients.

Nava Sara S Lisini Daniela D Pogliani Simona S Dossena Marta M Bersano Anna A Pellegatta Serena S Parati Eugenio E Finocchiaro Gaetano G Frigerio Simona S

Stem cells translational medicine 20150813 10

<h4>Unlabelled</h4>Cell therapy based on dendritic cells (DCs) pulsed with tumor lysate is a promising approach in addition to conventional therapy for the treatment of patients with glioblastoma (GB). The success of this approach strongly depends on the ability to generate high-quality, functionally mature DCs (mDCs), with a high level of standardization and in compliance with Good Manufacturing Practices. In the cell factory of the Carlo Besta Foundation, two phase I clinical trials on immunot ...[more]

PMID: 26273063

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Project description:BACKGROUND:Immunotherapeutic approaches, such as dendritic cell (DC) vaccination, have emerged as promising strategies in the treatment of glioblastoma. Despite their promise, however, the absence of objective biomarkers and/or immunological monitoring techniques to assess the clinical efficacy of immunotherapy still remains a primary limitation. To address this, we sought to identify a functional biomarker for anti-tumor immune responsiveness associated with extended survival in glioblastoma patients undergoing DC vaccination. METHODS:28 patients were enrolled and treated in two different Phase 1 DC vaccination clinical trials at UCLA. To assess the anti-tumor immune response elicited by therapy, we studied the functional responsiveness of pre- and post-vaccination peripheral blood lymphocytes (PBLs) to the immunostimulatory cytokines interferon-gamma (IFN-?) and interleukin-2 (IL-2) in 21 of these patients for whom we had adequate material. Immune responsiveness was quantified by measuring downstream phosphorylation events of the transcription factors, STAT-1 and STAT-5, via phospho-specific flow cytometry. RESULTS:DC vaccination induced a significant decrease in the half-maximal concentration (EC-50) of IL-2 required to upregulate pSTAT-5 specifically in CD3(+)CD8(+) T lymphocytes (p?<?0.045). Extended survival was also associated with an increased per cell phosphorylation of STAT-5 in cytotoxic T-cells following IL-2 stimulation when the median post/pre pSTAT-5 ratio was used to dichotomize the patients (p?=?0.0015, log-rank survival; hazard ratio?=?0.1834, p?=?0.018). Patients whose survival was longer than two years had a significantly greater pSTAT-5 ratio (p?=?0.015), but, contrary to our expectations, a significantly lower pSTAT-1 ratio (p?=?0.038). CONCLUSIONS:Our results suggest that monitoring the pSTAT signaling changes in PBL may provide a functional immune monitoring measure predictive of clinical efficacy in DC-vaccinated patients.

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Safe and Reproducible Preparation of Functional Dendritic Cells for Immunotherapy in Glioblastoma Patients.

Publications

Safe and Reproducible Preparation of Functional Dendritic Cells for Immunotherapy in Glioblastoma Patients.

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