Department of Hematology 54P4, Center for Cancer Immune Therapy, University Hospital Herlev, Herlev, Denmark.
Cancer Immunol Immunother. 2012 Aug;61(8):1221-31. doi: 10.1007/s00262-011-1199-8. Epub 2012 Jan 12.
Adoptive cell transfer (ACT) of in vitro expanded autologous tumor-infiltrating lymphocytes (TIL) has been shown to exert therapeutic efficacy in melanoma patients. We aimed to develop an ACT protocol based on tumor-specific T cells isolated from peripheral blood and in vitro expanded by Dynabeads® ClinExVivo™CD3/CD28. We show here that the addition of an in vitro restimulation step with relevant peptides prior to bead expansion dramatically increased the proportion of tumor-specific T cells in PBMC-cultures. Importantly, peptide-pulsed dendritic cells (DCs) as well as allogeneic tumor lysate-pulsed DCs from the DC vaccine preparation could be used with comparable efficiency to peptides for in vitro restimulation, to increase the tumor-specific T-cell response. Furthermore, we tested the use of different ratios and different types of Dynabeads® CD3/CD28 and CD3/CD28/CD137 T-cell expander, for optimized expansion of tumor-specific T cells. A ratio of 1:3 of Dynabeads® CD3/CD28 T-cell expander to T cells resulted in the maximum number of tumor-specific T cells. The addition of CD137 did not improve functionality or fold expansion. Both T-cell expansion systems could generate tumor-specific T cells that were both cytotoxic and effective cytokine producers upon antigen recognition. Dynabeads®-expanded T-cell cultures shows phenotypical characteristics of memory T cells with potential to migrate and expand in vivo. In addition, they possess longer telomeres compared to TIL cultures. Taken together, we demonstrate that in vitro restimulation of tumor-specific T cells prior to bead expansion is necessary to achieve high numbers of tumor-specific T cells. This is effective and easily applicable in combination with DC vaccination, by use of vaccine-generated DCs, either pulsed with peptide or tumor-lysate.
过继细胞转移（ACT）体外扩增的自体肿瘤浸润淋巴细胞（TIL）已被证明在黑色素瘤患者中具有治疗效果。我们旨在开发一种基于从外周血中分离并通过 Dynabeads®ClinExVivo™CD3/CD28 体外扩增的肿瘤特异性 T 细胞的 ACT 方案。我们在这里表明，在珠子扩增之前进行体外再刺激步骤，用相关肽脉冲，可大大增加 PBMC 培养物中肿瘤特异性 T 细胞的比例。重要的是，肽脉冲树突状细胞（DC）以及来自 DC 疫苗制备的同种异体肿瘤裂解物脉冲的 DC 可以与肽一样有效地用于体外再刺激，以增加肿瘤特异性 T 细胞反应。此外，我们测试了使用不同比例和不同类型的 Dynabeads®CD3/CD28 和 CD3/CD28/CD137 T 细胞扩增剂，以优化肿瘤特异性 T 细胞的扩增。Dynabeads®CD3/CD28 T 细胞扩增剂与 T 细胞的比例为 1:3 可获得最大数量的肿瘤特异性 T 细胞。添加 CD137 不会改善功能或折叠扩增。两种 T 细胞扩增系统都可以产生肿瘤特异性 T 细胞，这些细胞在抗原识别时具有细胞毒性和有效的细胞因子产生能力。Dynabeads®-扩增的 T 细胞培养物显示出记忆 T 细胞的表型特征，具有在体内迁移和扩增的潜力。此外，它们的端粒比 TIL 培养物更长。总之，我们证明在珠子扩增之前进行肿瘤特异性 T 细胞的体外再刺激是获得大量肿瘤特异性 T 细胞所必需的。这与 DC 疫苗接种有效且易于应用，通过使用疫苗生成的 DC，无论是用肽还是肿瘤裂解物脉冲。
