Injectable cell transplantation system for cytokine-based cancer immunotherapy

• Main Author: Gerard Nathanael Adrianus.
• Other Authors: Wang Dongan
• Format: Final Year Project
• Language: English
• Published: 2009
• Subjects: DRNTU::Engineering::Chemical engineering::Biotechnology
• Online Access: http://hdl.handle.net/10356/16593
• Institution: Nanyang Technological University

Cancer is a major killing disease, causing 6.7 million deaths globally in 2002. Standard cancer treatments, such as chemotherapy and radiation, are either not effective or result in temporary tumour regression while causing excessive damage to normal tissues. Immunotherapy is a promising therapy that attempts to use components of the immune system to bolster host immune rejection against tumour tissues. Cytokines, the regulatory peptides that control modulation of cell proliferation, differentiation, and survival, have been widely studied for cancer immunotherapy. Systemic injection of most cytokines has only shown modest clinical benefits, while associated with significant side-effects akin to chronic inflammation. Local, targeted delivery of cytokines in tumour sites is beneficial to achieve high therapeutic dose within tumour while limiting systemic dose. We have designed a system to deliver immunocytes to tumour in situ, which will release cytokines favouring tumour regression in the tumour microenvironment. Immunocytes are microencapsulated in hydrogel, suspended in gel solution, and injected to tumour in situ, where the solution gelify and put the system in place. In this study, we assessed the design using an in vitro model. Murine macrophages were encapsulated in agarose spheres using water-in-oil emulsification technique, activated with lipopolysaccaride (LPS), suspended in another gel solution, and moulded into constructs. The constructs were co-cultured with several malignant and normal cell lines. The material was characterized; viability and proliferation of the macrophages in microcapsules were assayed using cell counting, Live/Dead, and WST-1 assay; target cells were assayed with Annexin, Live/Dead, and WST-1 assay; finally the gene and protein expression of the activated macrophages were studied using PCR array, quantitative PCR, and ELISA. The fabricated microcapsules were in the range of 50-200 µm in diameter, within injectable size. The microcapsules were able to contain macrophages without leak for at least a month. LPS was able to freely diffuse in and out of the agarose microcapsules. The macrophages did not proliferate in the microcapsules, but remained viable for one month. The construct was able to exert toxicity to two malignant cell lines, MCF-7 and HepG2, but no effect was observed in HeLa and H1229 cells. No toxicity was observed to the normal cells lines used, MSC and Fibroblast (Fb). Genetic analysis confirmed up-regulation of several anti-tumour cytokines, TNF-α, IFN-β1, IL-10, and M-CSF by the macrophages. ELISA suggested the TNF-α was released in the first 24-hour post LPS stimulation, and then rapidly subsided. These results confirmed the proofs-of-principle and feasibility of the proposed system.