Milano Francesca, Rygiel Agnieszka M, Buttar Navtej, Bergman Jacques J G H M, Sondermeijer Carine, van Baal Jantine W P M, ten Brinke Anja, Kapsenberg Martien, van Ham S Marieke, Peppelenbosch Maikel P, Krishnadath Kausilia K
Department of Experimental Internal Medicine, Academic Medical Center, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands.
Cancer Immunol Immunother. 2007 Dec;56(12):1967-77. doi: 10.1007/s00262-007-0341-0. Epub 2007 Jun 13.
Esophageal cancer is a highly malignant disease that despite surgery and adjuvant therapies has an extremely poor outcome. Dendritic cell (DC) immunotherapy as a novel promising strategy could be an alternative for treating this malignancy. Effective DC-mediated immune responses can be achieved by raising cytotoxic T lymphocyte (CTL) response against multiple antigens through loading DCs with total tumor RNA. However, the efficacy of this strategy first needs to be evaluated in a pre-clinical setting. The aim of the study was to set up an ex vivo autologous human readout assay for assessing the effects of DC-mediated cytotoxic responses, using total tumor RNA as an antigen load. Biopsy specimens of seven esophageal cancer patients were used to establish primary cultures of normal and cancer cells and to obtain autologous RNA for loading DCs. Mature DCs loaded with either normal or tumor RNA were obtained and subsequently used to raise various lymphocytes populations. Apoptosis levels of the autologous cultures were measured before and after incubating the cultures with the different lymphocytes populations. The mean apoptosis levels in the tumor cell cultures, induced by lymphocytes instructed by DCs loaded with tumor RNA, significantly increased with 15.6% +/-2.9 SEM (range 3.4-24.5%, t-test, P < 0.05). Incubation of the normal cultures with the lymphocytes populations showed a mean non-significant increase in apoptosis of 0.4% +/-3.4 SEM (range -13.9 to 9.8%, t-test, P = 0.7). Here, we introduce a practical, patient-specific autologous readout assay for pre-clinical testing of DC-mediated cytotoxic responses. Additionally, we demonstrated that the use of autologous tumor RNA as a strategy for raising cytotoxic responses against multiple tumor antigens could be effective for treating esophageal cancer.
食管癌是一种高度恶性的疾病，尽管进行了手术和辅助治疗，但其预后极差。树突状细胞（DC）免疫疗法作为一种新的有前景的策略，可能是治疗这种恶性肿瘤的一种替代方法。通过用肿瘤总RNA负载DCs，引发针对多种抗原的细胞毒性T淋巴细胞（CTL）反应，可以实现有效的DC介导的免疫反应。然而，这一策略的疗效首先需要在临床前环境中进行评估。本研究的目的是建立一种体外自体人读数分析方法，以评估DC介导的细胞毒性反应的效果，使用肿瘤总RNA作为抗原负载。使用7例食管癌患者的活检标本建立正常细胞和癌细胞的原代培养物，并获得自体RNA用于负载DCs。获得负载正常或肿瘤RNA的成熟DCs，随后用于培养各种淋巴细胞群体。在用不同淋巴细胞群体孵育培养物之前和之后，测量自体培养物的凋亡水平。由负载肿瘤RNA的DCs指导的淋巴细胞诱导的肿瘤细胞培养物中的平均凋亡水平显著增加，为15.6%±2.9 SEM（范围3.4 - 24.5%，t检验，P < 0.05）。用淋巴细胞群体孵育正常培养物显示凋亡平均无显著增加，为0.4%±3.4 SEM（范围 - 13.9至9.8%，t检验，P = 0.7）。在此，我们介绍一种实用的、针对患者的自体读数分析方法，用于DC介导的细胞毒性反应的临床前测试。此外，我们证明使用自体肿瘤RNA作为引发针对多种肿瘤抗原的细胞毒性反应的策略，可能对治疗食管癌有效。
