Hayden Hubert, Friedl Josef, Dettke Markus, Sachet Monika, Hassler Michaela, Dubsky Peter, Bachleitner-Hofmann Thomas, Gnant Michael, Stift Anton
Department of Surgery, Medical University of Vienna, Vienna, Austria.
J Immunother. 2009 Jul-Aug;32(6):638-54. doi: 10.1097/CJI.0b013e3181a5bc13.
Cryopreservation of immature or mature dendritic cells (DC) has been proposed as a suitable method to gain large numbers of DC for immunotherapeutic trials against cancer. However, clinical studies using cryopreserved DC have demonstrated only limited success so far. The aim of this study was to investigate whether cryopreservation of monocytes elicits more potent DC and whether these DC are comparable to freshly generated DC preparations. Monocytes, either separated immunomagnetically or by means of leukapheresis and elutriation, were differentiated into DC and cryopreserved at various developmental stages. DC preparations were analyzed regarding recovery, viability, phenotype, and functional properties. In contrast to DC frozen at their immature or semi-mature state, generation of DC from cryopreserved monocytes elicited viability values comparable with freshly generated DC. Furthermore, using frozen monocytes for DC differentiation revealed improved expression of DC surface markers and interleukin-12p70 secretion as compared with DC generated from frozen immature or frozen semi-mature DC. Impaired phenotypical appearance of the latter DC variants was further substantiated by functional analysis. T-cells cocultured with these DC showed decreased expression of interferon-gamma and granzyme B, and lowered proliferation when compared with T-cells cocultured with DC generated from frozen monocytes or DC generated from freshly isolated monocytes. Induction of regulatory T-cell populations was negligible among all investigated DC preparations. These findings may further improve DC-based immunotherapeutical protocols. Cryopreservation of unchallenged monocytes enables targeted therapy by loading DC with varying antigenic compositions in case of tumor escape during treatment.
将未成熟或成熟的树突状细胞（DC）进行冷冻保存，已被提议作为一种获取大量DC用于癌症免疫治疗试验的合适方法。然而，迄今为止，使用冷冻保存的DC进行的临床研究仅取得了有限的成功。本研究的目的是调查单核细胞的冷冻保存是否能产生更强效的DC，以及这些DC是否与新鲜生成的DC制剂相当。通过免疫磁珠分离或白细胞单采及淘洗法分离得到的单核细胞，在不同发育阶段分化为DC并进行冷冻保存。对DC制剂的回收率、活力、表型和功能特性进行了分析。与在未成熟或半成熟状态下冷冻的DC不同，由冷冻保存的单核细胞生成的DC的活力值与新鲜生成的DC相当。此外，与从未成熟或半成熟冷冻DC生成的DC相比，使用冷冻单核细胞进行DC分化显示DC表面标志物的表达有所改善，白细胞介素-12p70的分泌也有所增加。通过功能分析进一步证实了后一种DC变体的表型外观受损。与与从冷冻单核细胞生成的DC或从新鲜分离的单核细胞生成的DC共培养的T细胞相比，与这些DC共培养的T细胞显示干扰素-γ和颗粒酶B的表达降低，增殖也降低。在所有研究的DC制剂中，调节性T细胞群体的诱导可以忽略不计。这些发现可能会进一步改进基于DC的免疫治疗方案。在治疗过程中肿瘤逃逸的情况下，对未受刺激的单核细胞进行冷冻保存能够通过用不同抗原组成加载DC来实现靶向治疗。
