Klauer Lara Kristina, Schutti Olga, Ugur Selda, Doraneh-Gard Fatemeh, Amberger Daniel Christoph, Rogers Nicole, Krämer Doris, Rank Andreas, Schmid Christoph, Eiz-Vesper Britta, Schmetzer Helga Maria
Department of Medicine III, University Hospital of Munich, Munich, Germany.
Department of Haematology and Oncology, St.-Josefs-Hospital, Hagen, Germany.
Transfus Med Hemother. 2021 Jul 7;49(1):44-61. doi: 10.1159/000516886. eCollection 2022 Feb.
Myeloid leukaemic blasts can be converted into leukaemia-derived dendritic cells (DC), characterised by the simultaneous expression of dendritic- and leukaemia-associated antigens, which have the competence to prime and enhance (leukaemia-specific) immune responses with the whole leukaemic antigen repertoire. To display and further specify dendritic cell (DC)- and DC-mediated immune responses, we analysed the interferon gamma (IFNy) secretion of innate and adaptive immune cells.
DC/DC were generated from leukaemic whole blood (WB) with (blast)modulatory Kit-I (granulocyte-macrophage colony-stimulating factor [GM-CSF] + Picibanil [OK-432]) and Kit-M (GM-CSF + prostaglandin E1) and were used to stimulate T cell-enriched immunoreactive cells. Initiated anti-leukaemic cytotoxicity was investigated with a cytotoxicity fluorolysis assay. Initiated IFNy secretion of T, NK, CIK, and iNKT cells was investigated with a cytokine secretion assay (CSA). IFNy positivity was additionally evaluated with an intracellular cytokine assay (ICA). Recent activation of leukaemia-specific cells was verified through addition of leukaemia-associated antigens (LAA; WT-1 and Prame).
We found Kit-I and Kit-M competent to generate mature DC and DC from leukaemic WB without induction of blast proliferation. Stimulation of immunoreactive cells with DC/DC regularly resulted in an increased anti-leukaemic cytotoxicity and increased IFNy secretion of T, NK, and CIK cells, pointing to the significant role of DC/DC in leukaemia-specific alongside anti-leukaemic reactions. Interestingly, an addition of LAA did not further increase IFNy secretion, suggesting an efficient activation of leukaemia-specific cells. Here, both the CSA and ICA yielded comparable frequencies of IFNy-positive cells. Remarkably, the anti-leukaemic cytotoxicity positively correlated with the IFNy secretion in T, T, T, and NK cells.
Ultimately, the IFNy secretion of innate and adaptive immune cells appeared to be a suitable parameter to assess and monitor the efficacy of in vitro and potentially in vivo acute myeloid leukaemia immunotherapy. The CSA in this regard proved to be a convenient and reproducible technique to detect and phenotypically characterise IFNy-secreting cells. In respect to our studies on DC-based immunomodulation, we were able to display the potential of DC/DC to induce or improve leukaemia-specific and anti-leukaemic activity.
髓系白血病原始细胞可转化为白血病衍生的树突状细胞（DC），其特征是同时表达树突状细胞相关抗原和白血病相关抗原，这些细胞有能力利用整个白血病抗原库启动并增强（白血病特异性）免疫反应。为了展示并进一步明确树突状细胞（DC）及DC介导的免疫反应，我们分析了天然免疫细胞和适应性免疫细胞的γ干扰素（IFNγ）分泌情况。
使用（原始细胞）调节试剂盒-I（粒细胞-巨噬细胞集落刺激因子[GM-CSF]+沙培林[OK-432]）和试剂盒-M（GM-CSF+前列腺素E1）从白血病全血（WB）中生成DC/DC，并用于刺激富含T细胞的免疫反应性细胞。采用细胞毒性荧光溶解试验研究启动的抗白血病细胞毒性。采用细胞因子分泌试验（CSA）研究T细胞、自然杀伤细胞（NK）、细胞因子诱导的杀伤细胞（CIK）和不变自然杀伤T细胞（iNKT）启动的IFNγ分泌情况。还通过细胞内细胞因子试验（ICA）评估IFNγ阳性情况。通过添加白血病相关抗原（LAA；WT-1和Prame）验证白血病特异性细胞的近期激活情况。
我们发现试剂盒-I和试剂盒-M能够从白血病WB中生成成熟的DC和DC，而不会诱导原始细胞增殖。用DC/DC刺激免疫反应性细胞通常会导致抗白血病细胞毒性增加，以及T细胞、NK细胞和CIK细胞的IFNγ分泌增加，这表明DC/DC在白血病特异性反应以及抗白血病反应中发挥着重要作用。有趣的是，添加LAA并没有进一步增加IFNγ分泌，这表明白血病特异性细胞已被有效激活。在此，CSA和ICA产生的IFNγ阳性细胞频率相当。值得注意的是，抗白血病细胞毒性与T细胞、T细胞、T细胞和NK细胞中的IFNγ分泌呈正相关。
最终，天然免疫细胞和适应性免疫细胞的IFNγ分泌似乎是评估和监测体外以及潜在体内急性髓系白血病免疫治疗疗效的合适参数。在这方面，CSA被证明是一种方便且可重复的技术，用于检测和表型鉴定分泌IFNγ的细胞。关于我们基于DC的免疫调节研究，我们能够展示DC/DC诱导或改善白血病特异性和抗白血病活性的潜力。
