Septins是具有形成细丝能力的细胞骨架蛋白，其在细胞分裂、细胞极性、形态发生和膜转运过程中发挥多重作用。反对septin-5的自身抗体与非肿瘤性小脑失调有关，而反对septin-7的自身抗体与突出的神经精神症状的脑病有关。在这里，我们报告了在患有旁肿瘤性小脑失调的患者中新识别的反对septin-3的自身抗体。我们还提出了一种反septin自身抗体测定的策略。来自三名在小脑和海马区切片上产生相似免疫荧光染色模式的患者的血清经免疫沉淀后进行质谱分析。所鉴定的候选抗原均为septin，在HEK293细胞中单独、复合或缺少单个septin的组合中进行了重组细胞间接免疫荧光试验（RC-IIFA）。通过组织IIFA中和实验证实了对septin-3的特异性。最后，肿瘤组织切片通过免疫组化分析septin-3的表达。用大鼠小脑溶液进行免疫沉淀表明，septin-3、-5、-6、-7和-11是候选的靶抗原。所有三名患者的血清与共表达septin-3/5/6/7/11的重组细胞反应，而149份健康对照血清则没有类似的反应。在RC-IIFAs中，患者血清仅识别单独表达septin-3和复合体中的septin-3的细胞。用五种不同的缺少其中一个septin的septin组合物对患者血清进行孵育，证实了自身抗体对septin-3的特异性。患者血清的组织IIFA反应通过预先孵育septin-3/5/6/7/11复合体或单独的septin-3重组细胞溶液消除，但未在过表达septin-5的HEK293细胞溶液中消除。所有三名患者均有癌症（2个黑色素瘤，1个小细胞肺癌），表现为进行性小脑综合征，并对免疫治疗反应不良。从一名患者的切除肿瘤组织中证实了septin-3的表达。Septin-3是旁肿瘤性小脑综合征患者中的一种新型自身抗体靶标。根据我们的发现，HEK293重组细胞表达septin-3/5/6/7/11复合体的RC-IIFA可以作为筛查工具，用于研究在神经元组织切片上具有特征性染色模式的血清中的抗septin自身抗体。然后可以通过表达单个septin的RC-IIFA来确认反对单个septin的自身抗体。 © 2023. The Author(s)。

Septins are cytoskeletal proteins with filament forming capabilities, which have multiple roles during cell division, cellular polarization, morphogenesis, and membrane trafficking. Autoantibodies against septin-5 are associated with non-paraneoplastic cerebellar ataxia, and autoantibodies against septin-7 with encephalopathy with prominent neuropsychiatric features. Here, we report on newly identified autoantibodies against septin-3 in patients with paraneoplastic cerebellar ataxia. We also propose a strategy for anti-septin autoantibody determination.Sera from three patients producing similar immunofluorescence staining patterns on cerebellar and hippocampal sections were subjected to immunoprecipitation followed by mass spectrometry. The identified candidate antigens, all of which were septins, were expressed recombinantly in HEK293 cells either individually, as complexes, or combinations missing individual septins, for use in recombinant cell-based indirect immunofluorescence assays (RC-IIFA). Specificity for septin-3 was further confirmed by tissue IIFA neutralization experiments. Finally, tumor tissue sections were analyzed immunohistochemically for septin-3 expression.Immunoprecipitation with rat cerebellum lysate revealed septin-3, -5, -6, -7, and -11 as candidate target antigens. Sera of all three patients reacted with recombinant cells co-expressing septin-3/5/6/7/11, while none of 149 healthy control sera was similarly reactive. In RC-IIFAs the patient sera recognized only cells expressing septin-3, individually and in complexes. Incubation of patient sera with five different septin combinations, each missing one of the five septins, confirmed the autoantibodies' specificity for septin-3. The tissue IIFA reactivity of patient serum was abolished by pre-incubation with HEK293 cell lysates overexpressing the septin-3/5/6/7/11 complex or septin-3 alone, but not with HEK293 cell lysates overexpressing septin-5 as control. All three patients had cancers (2 × melanoma, 1 × small cell lung cancer), presented with progressive cerebellar syndromes, and responded poorly to immunotherapy. Expression of septin-3 was demonstrated in resected tumor tissue available from one patient.Septin-3 is a novel autoantibody target in patients with paraneoplastic cerebellar syndromes. Based on our findings, RC-IIFA with HEK293 cells expressing the septin-3/5/6/7/11 complex may serve as a screening tool to investigate anti-septin autoantibodies in serological samples with a characteristic staining pattern on neuronal tissue sections. Autoantibodies against individual septins can then be confirmed by RC-IIFA expressing single septins.© 2023. The Author(s).