Phase Ⅰ Study In Patients With Colorectal Cancer Of A Vaccine With Antigen Pulsed Human Dendritic Cells

Although multiple options including surgery, radiotherapy and chemotherapy for the treatment of malignant tumors have often been the mainstream, the traditional treatments have the disadvantage of inhibiting the immunity system. With several years' researches on cell biology and immunology, it was evident that tumor-specific immunity presented by themselves might be an effective method to control tumor metastasis and prevent recurrence.Dendritc cells(DCs) are the known unique strongest antigen presenting cells which can activate naive T cells. They are the initiators of immune response [1]. It was the basis for DC vaccine to treat malignant tumors which dendritic cells can induce tumor-specifi longlasting immune response.Autologous monocytes isolated from the peripheral blood can develop to and function as dendritic cells co-cultured in GM-CSF and IL-4 ex vivo. As a result, a good many DCs as vaccine have been acquired by this mean. Using surgery specimens as tumor antigen stimulates dendritic cells. The advantage of the approach is easy to gain the most antigens of the specimens, to search the tumor-specific antigens and to operate.Current international clinical trials showed antigen-pulsed dendritic cell(APDC) activated tumor-specific CTLs that recognize the naturally processed target antigens on tumor cells and set up longlasting anti-tumor specific immune response. The patients had no obvious acute and chronic toxitic response and a good tolerance.A large scale clinical studies on DC vaccine have been mainly aimed to malignant melanoma and prostate cancer and gained a good effect. There has been an increasing incidence rate, a high recurrence rate and metastatic rate of colon cancer in China. No standardized DC vaccine clinical trial has been set up for cancer patients in China up to now. Therefore, we applied autologous peripheral monocyte-derived DCs to induce anti-caner immunotherapy for colon cancerpatients.The object of the study was to preliminarily assess safety and tolerance and to test immune response of human APDC. We hoped that the results of our study could provide a safe and effective regimen for phase II clinical trial of APDC vaccination. APDC Generation and measurementThe surgical resection samples were tested for sterility test, the concentration of protein and tumorogenesis. And then the samples were stored frozen. 10ml blood were isolated from patients and l-5><109 monocytes centrifuged were cultured for 5 days in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4 . At the end of this period, DCs were collected and co-cultured with colon cancer cells. On day 6 of their culture, APDCs were obtained. The assessment of APDC activity and tumorogenesis were performed.Results: The concentration of generated specific-tumor antigen was kept between 544 and 1544ug/ml. No tumorogenetic clone was formed. There is no significance between the control group and APDC vaccine group by means of mixed leukocyte reaction (student-t test, P<0.05). The percentages of DCs expressing CD86, those expressing HLA-DR were both above 80%. The number of generated APDCs was more than 5><107/Lfor each patient, and the survival rate of those also no less than 80%. There is a negative result in sterility test. The residual calf serum was no more than 25ng/ml and the residual IL-4 less than lOpg/ml.Protocol and rerults of APDC vaccine clinical trial PatientsColon carcinoma specimens were obtained from surgical resections confirmed by histopathologic and immunohistochemical staining. Patients were required to meet the following criteria: above the age of 18;KPS at least 70;with Approximately normal renal, hepatic and bone marrow functions;with the access to obtain adequate amounts of tumor samples. However, such patients should be excluded: with severe hepatic or renal or heart impairment;with autoimmunedisease history;pregnancy or lactation;with prior allergic reactions to certain drugs;with previous chemotherapy, radiotherapy, immunosuppressive treatment or any other immune therapy within less than 4 weeks. TherapyAPDCs were diluted with 50ml Sodium Chloride and intravenous injected within 15 minutes. All the patients were divided into three groups. The three groups were vaccinated as followed: A group, lxl07/vaccination;B group, 3xl07/ vaccination;C group, 6><107/ vaccination. 12.5mg Promethazine was intramuscular injected 30 minutes before APDC vaccination.Adverse events were observed and recorded and the end-points of the study were evaluated according National Cancer Institute definitions of grades of adverse events. If grade 3 of adverse events were observed, the therapy was terminated. If there was no grade 3 of adverse events, the therapy was completed after C group APDC vaccination. ELISPOT, DTH AND OT assays were tested before and after the APDC vaccination. Results General data21 colon cancer patients were divided into three groups and 7 cases for each. There was no significant difference for ages, sex and KPS. There was neither no significant difference for vital signs between before and after vaccination. No grade 3 or more adverse events related to APDC were observed. The adverse events related APDC vaccination included fever, algor, rhinocleisis, nasal discharge, weakness and myalgia which were grade 1 -2 and could be relieved after symptomatic treatment. And the incidence rates of adverse events for the three groups were separately 14.29%, 14.29% and 57.14%. No abnormal results were found in blood test, urine test, hepatic function and renal function test and ECG. Immune responseThe OT value of DTH for the three groups were separately 4.29±3.82, 3.43±1.90, 4.29±6.45. The OT value of DTH 7 days were separately 4.57±3.78, 4.29±3.15, 5.14±6.87 7 days after the last vaccination. There was no significant difference between three groups. However, a significant difference was observed in DTH between before and after APDC vaccination (P=0.0475).Autologous skin DTHThere was no significant difference for autologous skin DTH between before and after APDC vaccination. Tumor-specific immune responseThe difference of tumor-specific immune response between before and after APDC vaccination was tested by ELOSPOT assay. It was shown that the level of v-IFN was significantly elevated in 3 cases of A group, 2 cases of B group and 2 cases of C group. v-IFN elevation represented enhancement of tumor-specific immune response. The study illustrated that APDC vaccination could enhance tumor-specific immune response.Conclusion:APDC vaccine was safe and no severe adverse effect. It had a good tolerance at the dose of 3><107/vaccination which could induce tumor-specific immune response.