In Vitro and in Vivo Study of Individualized Cancer Immunotherapy and a Pilot Trail on Patients with Gastric Cancer

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Introduction:

Cancer immunotherapy has been used in clinical trials and shows a powerful life force. But if it is benefit for the treatment of gastric cancer is still not well studied. Present study discussed the antitumor efficiency of individualized immunotherapy induced by dendritic cells transfected with total antigen of autologouse gastric cancer cells and reported the initial results of our pilot clinical trail.

Methods:

Autologous DCs were isolated from PBMC and proliferated in supplement with GM-CSF and IL-4. Resulting immature DCs were loaded with autologous tumor antigen (tumor lysate or total RNA) and matured in supplement with TNF-α (tu-DC). Autologous T lymphocytes were isolated from PBMC and cocultured with tu-DC to prepare antigen-specific CTLs. Autologous primary cancer cells were obtained from resected specimens of the patients with gastric cancer and cultured as target cells. Phenotypic characters of DCs were detected and evaluated by FCM. The efficiency of cytotoxicity and cytokine release was measured and estimated by using assays of Cytotoxicity and ELISA.

Humanized immune system was reconstructed on 4-week-old nude mice by intraperitoneal (i.p.) injection of human PBMC with cell number of 3×10⁷ each. Prophylactic assay and therapeutic assay were performed by subcutaneously challenge of primary cancer cells after or before autologous DC vaccination, respectively, and the tumor burden were observed.

Nine patients with gastric cancer were subjected to a pilot clinical trail. Two patients with stage Ib-IIIa underwent a radical D2 resection, 5 patients with stage IIIb-IV underwent palliative resection, and 2 patients with stage IV underwent an open-and-close operation. Autologous tumor lysate were prepared from fresh specimens of the patients. DC vaccines and tumor specific CTLs were prepared by using the methods above mentioned. To suppress Treg and to promote calreticulin (CRT) exposure, CTX 1000 mg and ADM 60 mg were intravenously infused a day before immunotherapy. Tumor-specific CTLs were therapied by intravenous infusing with one day interval, and DC vaccines were inoculated to the axillary lymph nodes with two therapies interval. Thymosin alpha 1 (Tα1) was injected once a day subcutaneously during the therapy. The levels of Treg were measured before and after chemotherapy by FCM, and the Incidence of recurrence, Karnofsky performance scale, tumor burden, and side effects were observed and evaluated.

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