Eradicating neuroblastomas by an RBM39 degrader in combination with anti-GD2 immunotherapy regardless of cancer cell state alterations [RNA-seq]

The lineage plasticity of neuroblastoma cells is defined by a mixture of two major cell states, adrenergic (ADRN) and mesenchymal (MES), which presumably contributes to therapy resistance. However, how the neuroblastoma cells switch their cell states during therapy remains largely unknown and how to eradicate neuroblastoma cells regardless of their cell state alterations is a clinical challenge. By using four distinct human and murine neuroblastoma models treated with indisulam, a molecular glue drug serving as a selective RBM39 degrader, we show that cancer cells not only can undergo bidirectional switch of ADRN and MES cell states, but also can acquire additional cell states of melanoma and Schwann cell progenitors, reminiscent of the cellular pliancy of neural crest cells. The lineage alterations of neuroblastoma cells are coupled with epigenetic changes and dependency switch of lineage-specific transcription factors. Nevertheless, indisulam treatment induces an inflamed tumor microenvironment leading to an immune cell infiltration. The combination of indisulam with anti-GD2 monoclonal immunotherapy results in a durable complete response in transgenic neuroblastoma models driven by MYCN and ALK mutant oncogenes, providing a therapeutic rationale to eradicate tumor cells even when they acquire alternative cell lineage(s) in response to treatment. Overall design: To determine the gene expression profile To determine the gene expression and splicing profile with and without GAC and KGA expression in of neuroblastoma cells, BE2C and SKNAS were transduced with Lenti virus control, GAC and KGA to overexpression GCA and KGA.of neuroblastoma cell with, line BE2C, SKNAS

神经母细胞瘤细胞的谱系可塑性由两种主要细胞状态——肾上腺素能（adrenergic, ADRN）与间质型（mesenchymal, MES）——的混合状态所定义，这被认为是其产生治疗抵抗的重要原因。然而，神经母细胞瘤细胞在治疗过程中如何切换细胞状态仍未被充分阐明，且无论细胞状态发生何种改变，如何根除神经母细胞瘤都是一项临床难题。本研究采用经吲哚西拉姆（indisulam）——一种作为选择性RNA结合基序蛋白39（RBM39）降解剂的分子胶类药物——处理的四种不同人源和鼠源神经母细胞瘤模型，结果显示，肿瘤细胞不仅可在ADRN与MES细胞状态间双向切换，还可获得黑色素细胞与雪旺细胞祖细胞的额外细胞状态，这与神经嵴细胞的细胞可塑性特征相似。神经母细胞瘤细胞的谱系改变与表观遗传变化及谱系特异性转录因子的依赖性切换密切相关。值得注意的是，吲哚西拉姆治疗可诱导肿瘤微环境发生炎症重塑，进而促进免疫细胞浸润。将吲哚西拉姆与抗GD2单克隆免疫疗法联合使用，可在由MYCN与ALK突变癌基因驱动的转基因神经母细胞瘤模型中实现持久完全缓解，这为即便肿瘤细胞在治疗压力下获得其他细胞谱系时仍能根除肿瘤细胞提供了治疗理论依据。整体实验设计：为明确神经母细胞瘤细胞的基因表达谱，以及在GAC与KGA表达存在或缺失条件下的基因表达与剪接谱，我们将BE2C与SKNAS细胞系分别以对照慢病毒（Lenti virus）、过表达GAC的慢病毒及过表达KGA的慢病毒进行转导，以实现GAC与KGA的过表达。