Phase I clinical trial of autologous ascites-derived exosomes combined with GM-CSF for colorectal cancer - PubMed (original) (raw)

Clinical Trial

Phase I clinical trial of autologous ascites-derived exosomes combined with GM-CSF for colorectal cancer

Shengming Dai et al. Mol Ther. 2008 Apr.

Abstract

Exosomes are small membrane vesicles that are secreted by a multitude of cell types. The exosomes derived from dendritic cells (Dex), tumor cells (Tex), and malignant effusions demonstrate immunomodulatory functions, and are even under clinical trial for cancer treatments. In this study we report the phase I clinical trial of the ascites-derived exosomes (Aex) in combination with the granulocyte-macrophage colony-stimulating factor (GM-CSF) in the immunotherapy of colorectal cancer (CRC). The Aex isolated by sucrose/D(2)O density gradient ultracentrifugation are 60-90-nm vesicles that contain the diverse immunomodulatory markers of exosomes and tumor-associated carcinoembryonic antigen (CEA). Totally 40 patients (HLA-A0201(+)CEA(+)) with advanced CRC were enrolled in the study, and randomly assigned to treatments with Aex alone or Aex plus GM-CSF. Patients in both groups received a total of four subcutaneous immunizations at weekly intervals. We found that both therapies were safe and well tolerated, and that Aex plus GM-CSF but not Aex alone can induce beneficial tumor-specific antitumor cytotoxic T lymphocyte (CTL) response. Therefore, our study suggests that the immunotherapy of CRC with Aex in combination with GM-CSF is feasible and safe, and thus can serve as an alternative choice in the immunotherapy of advanced CRC.

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Figures

Figure 1

Figure 1

Characterization of Aex. (a) Electron microscopy assay of the isolated Aex. The data were representative of Aex derived from patient H5. Bar = 100 nm. (b) Western blot assay of the protein markers in Aex derived from patient H5. Thirty microgram of cell lysates derived from the ascites (lane 1) or Aex (lane 2) were separated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gel and examined by Western blot. (c) Western blot assay of major histocompatibility complex class I (MHC-I), HSC70, and CD71 contained in Aex derived from representative patients. Lanes 1–7 correspond to Aex (30 μg per lane) derived from patients A1, B1, C1, D1, E1, F1, and G1, respectively.

Figure 2

Figure 2

Induction of carcinoembryonic antigen (CEA)-specific cytotoxic T lymphocyte (CTL) by the treatments. (a) Tetramer tests. Representative results derived from triplicate samples were shown. Delayed-type hypersensitivity (DTH) sites–derived emigrated cells were stained with phycoerythrin-labeled HLA-A*0201 tetramers (CAP-1 or SSp-1-specific) and fluorescein isothiocyanate (FITC)-conjugated CD8 monoclonal antibody and finally analyzed by flow cytometry. Numbers indicated for percentages of cells with positive staining. (b) Cytotoxicity assay. DTH site–derived cultured cells were cocultured with 51Cr-labeled SW480 cells, LoVo cells, T2 cells pulsed with CAP-1 (T2/CAP-1), or SSp-1 (T2/SSp-1) peptide. Specific cytotoxicity was evaluated by 51Cr release assay. Results were presented as the mean percentage of specific lysis ± SD of triplicate samples. (c) Interferon-γ (IFN-γ) release assay. CD8+ T cells isolated from the DTH site–derived cultured cells were cocultured with native T2 cells or T2/CAP-1 or T2/SSp-1 for 24 hours. IFN-γ level in the culture supernatant was determined by enzyme-linked immunosorbent assay, and the results were presented as mean ± SD of triplicate samples. Data presented here were the results derived from patient G1 who was treated with 300 μg Aex plus 50 μg granulocyte–macrophage colony-stimulating factor.

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