PDF(Pubmed)
Abstract：
BACKGROUND: Increasing evidence suggests that immunotherapy, especially immune checkpoint inhibitors (ICIs), has the potential to facilitate long-term survival in various cancer besides prostate cancer. Emerging evidence indicated that pyroptosis, an immunogenic form of cell death, could trigger an anti-tumor immune microenvironment and enhance the effectiveness of immunotherapy. Nevertheless, the mechanism underlying the regulation of pyroptosis signaling in prostate cancer remains unclear.
METHODS: The differential expression of human E3 ligases in prostate cancer was integratedly analyzed from five independent public datasets. Moreover, the immunohistochemistry analysis of a tissue microarray derived from prostate cancer patients confirmed the results from the bioinformatic analysis. Furthermore, prostate cancer cell lines were evaluated via the next-generation RNA sequencing to assess transcriptomic profile upon CDC20 depletion. Next, qRT-PCR, Western blotting, cycloheximide assay, immunoprecipitation, and ubiquitination assay were employed to explore the correlation and interaction between CDC20 and GSDME. Both immune-deficient and immune-competent murine models were utilized to examine the anti-tumor efficacy of CDC20 inhibition with or without the anti-PD1 antibodies, respectively. To analyze the immune microenvironment of the xenografts, the tumor tissues were examined by immunohistochemistry and flow cytometry.
RESULTS: The analysis of multiple prostate cancer cohorts suggested that CDC20 was the most significantly over-expressed E3 ligase. In addition, CDC20 exerted a negative regulatory effect on the pyroptosis pathway by targeting GSDME for ubiquitination-mediated proteolysis in a degron-dependent manner. Knockdown of CDC20 leads to increased GSDME abundance and a transition from apoptosis to pyroptosis in response to death signals. Furthermore, in our syngeneic murine models, we found that depletion of CDC20 significantly enhances the anti-tumor immunity by promoting the infiltration of CD8+ T lymphocytes dependent on the existence of GSDME, as well as reducing myeloid immune cells. More importantly, Apcin, a small molecular inhibitor that targets CDC20, exhibited synergistic effects with anti-PD1-based immunotherapy in murine models of prostate cancer.
CONCLUSIONS: Overall, these findings provide new insights into the upstream regulation of GSDME-mediated pyroptosis by CDC20, which specifically interacts with GSDME and facilitates its ubiquitination in a degron-dependent manner. Importantly, our data highlight novel molecular pathways for targeting cellular pyroptosis and enhancing the effectiveness of anti-PD1-based immunotherapy.
METHODS: The differential expression of human E3 ligases in prostate cancer was integratedly analyzed from five independent public datasets. Moreover, the immunohistochemistry analysis of a tissue microarray derived from prostate cancer patients confirmed the results from the bioinformatic analysis. Furthermore, prostate cancer cell lines were evaluated via the next-generation RNA sequencing to assess transcriptomic profile upon CDC20 depletion. Next, qRT-PCR, Western blotting, cycloheximide assay, immunoprecipitation, and ubiquitination assay were employed to explore the correlation and interaction between CDC20 and GSDME. Both immune-deficient and immune-competent murine models were utilized to examine the anti-tumor efficacy of CDC20 inhibition with or without the anti-PD1 antibodies, respectively. To analyze the immune microenvironment of the xenografts, the tumor tissues were examined by immunohistochemistry and flow cytometry.
RESULTS: The analysis of multiple prostate cancer cohorts suggested that CDC20 was the most significantly over-expressed E3 ligase. In addition, CDC20 exerted a negative regulatory effect on the pyroptosis pathway by targeting GSDME for ubiquitination-mediated proteolysis in a degron-dependent manner. Knockdown of CDC20 leads to increased GSDME abundance and a transition from apoptosis to pyroptosis in response to death signals. Furthermore, in our syngeneic murine models, we found that depletion of CDC20 significantly enhances the anti-tumor immunity by promoting the infiltration of CD8+ T lymphocytes dependent on the existence of GSDME, as well as reducing myeloid immune cells. More importantly, Apcin, a small molecular inhibitor that targets CDC20, exhibited synergistic effects with anti-PD1-based immunotherapy in murine models of prostate cancer.
CONCLUSIONS: Overall, these findings provide new insights into the upstream regulation of GSDME-mediated pyroptosis by CDC20, which specifically interacts with GSDME and facilitates its ubiquitination in a degron-dependent manner. Importantly, our data highlight novel molecular pathways for targeting cellular pyroptosis and enhancing the effectiveness of anti-PD1-based immunotherapy.
摘要：
背景：越来越多的证据表明，尤其是免疫检查点抑制剂（ICIs），除前列腺癌外，有可能促进各种癌症的长期生存。新出现的证据表明焦亡，细胞死亡的免疫原性形式，可以触发抗肿瘤免疫微环境并增强免疫治疗的有效性。然而，前列腺癌中焦凋亡信号调节的潜在机制尚不清楚.
方法：从五个独立的公共数据集中综合分析前列腺癌中人类E3连接酶的差异表达。此外，来自前列腺癌患者的组织微阵列的免疫组织化学分析证实了生物信息学分析的结果.此外，通过下一代RNA测序评估前列腺癌细胞系,以评估CDC20缺失后的转录组概况.接下来,qRT-PCR,西方印迹,环己酰亚胺测定,免疫沉淀,采用泛素化检测方法探讨CDC20与GSDME的相关性和相互作用。免疫缺陷和免疫活性小鼠模型均用于检查有或没有抗PD1抗体的CDC20抑制的抗肿瘤功效。分别。为了分析异种移植物的免疫微环境，用免疫组织化学和流式细胞术检测肿瘤组织。
结果：对多个前列腺癌队列的分析提示CDC20是最显著的过表达E3连接酶。此外,CDC20通过靶向GSDME以依赖degron的方式进行泛素化介导的蛋白水解，对焦亡途径产生负调节作用。CDC20的敲低导致GSDME丰度增加以及响应于死亡信号从细胞凋亡到焦亡的转变。此外，在我们的同基因鼠模型中，我们发现CDC20的耗竭通过促进CD8+T淋巴细胞的浸润而显著增强抗肿瘤免疫,以及减少髓系免疫细胞。更重要的是,Apcin,一种靶向CDC20的小分子抑制剂,在小鼠前列腺癌模型中与基于抗PD1的免疫疗法表现出协同作用.
结论：总体而言，这些发现为CDC20对GSDME介导的焦亡的上游调节提供了新的见解,CDC20与GSDME特异性相互作用,并以依赖degron的方式促进其泛素化.重要的是,我们的数据突出了靶向细胞焦亡和增强基于抗PD1的免疫疗法有效性的新分子途径.
方法：从五个独立的公共数据集中综合分析前列腺癌中人类E3连接酶的差异表达。此外，来自前列腺癌患者的组织微阵列的免疫组织化学分析证实了生物信息学分析的结果.此外，通过下一代RNA测序评估前列腺癌细胞系,以评估CDC20缺失后的转录组概况.接下来,qRT-PCR,西方印迹,环己酰亚胺测定,免疫沉淀,采用泛素化检测方法探讨CDC20与GSDME的相关性和相互作用。免疫缺陷和免疫活性小鼠模型均用于检查有或没有抗PD1抗体的CDC20抑制的抗肿瘤功效。分别。为了分析异种移植物的免疫微环境，用免疫组织化学和流式细胞术检测肿瘤组织。
结果：对多个前列腺癌队列的分析提示CDC20是最显著的过表达E3连接酶。此外,CDC20通过靶向GSDME以依赖degron的方式进行泛素化介导的蛋白水解，对焦亡途径产生负调节作用。CDC20的敲低导致GSDME丰度增加以及响应于死亡信号从细胞凋亡到焦亡的转变。此外，在我们的同基因鼠模型中，我们发现CDC20的耗竭通过促进CD8+T淋巴细胞的浸润而显著增强抗肿瘤免疫,以及减少髓系免疫细胞。更重要的是,Apcin,一种靶向CDC20的小分子抑制剂,在小鼠前列腺癌模型中与基于抗PD1的免疫疗法表现出协同作用.
结论：总体而言，这些发现为CDC20对GSDME介导的焦亡的上游调节提供了新的见解,CDC20与GSDME特异性相互作用,并以依赖degron的方式促进其泛素化.重要的是,我们的数据突出了靶向细胞焦亡和增强基于抗PD1的免疫疗法有效性的新分子途径.
