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Abstract：
BACKGROUND: Neuroendocrine prostate cancer (NEPC) is a lethal subset of prostate cancer which is characterized by neuroendocrine differentiation and loss of androgen receptor (AR) signaling. Growing evidence reveals that cell lineage plasticity is crucial in the failure of NEPC therapies. Although studies suggest the involvement of the neural transcription factor PAX6 in drug resistance, its specific role in NEPC remains unclear.
METHODS: The expression of PAX6 in NEPC was identified via bioinformatics and immunohistochemistry. CCK8 assay, colony formation assay, tumorsphere formation assay and apoptosis assay were used to illustrate the key role of PAX6 in the progression of in vitro. ChIP and Dual-luciferase reporter assays were conducted to confirm the binding sequences of AR in the promoter region of PAX6, as well as the binding sequences of PAX6 in the promoter regions of STAT5A and MET. For in vivo validation, the xenograft model representing NEPC subtype underwent pathological analysis to verify the significant role of PAX6 in disease progression. Complementary diagnoses were established through public clinical datasets and transcriptome sequencing of specific cell lines. ATAC-seq was used to detect the chromatin accessibility of specific cell lines.
RESULTS: PAX6 expression was significantly elevated in NEPC and negatively regulated by AR signaling. Activation of PAX6 in non-NEPC cells led to NE trans-differentiation, while knock-down of PAX6 in NEPC cells inhibited the development and progression of NEPC. Importantly, loss of AR resulted in an enhanced expression of PAX6, which reprogramed the lineage plasticity of prostate cancer cells to develop NE phenotypes through the MET/STAT5A signaling pathway. Through ATAC-seq, we found that a high expression level of PAX6 elicited enhanced chromatin accessibility, mainly through attenuation of H4K20me3, which typically causes chromatin silence in cancer cells.
CONCLUSIONS: This study reveals a novel neural transcription factor PAX6 could drive NEPC progression and suggest that it might serve as a potential therapeutic target for the management of NEPC.
METHODS: The expression of PAX6 in NEPC was identified via bioinformatics and immunohistochemistry. CCK8 assay, colony formation assay, tumorsphere formation assay and apoptosis assay were used to illustrate the key role of PAX6 in the progression of in vitro. ChIP and Dual-luciferase reporter assays were conducted to confirm the binding sequences of AR in the promoter region of PAX6, as well as the binding sequences of PAX6 in the promoter regions of STAT5A and MET. For in vivo validation, the xenograft model representing NEPC subtype underwent pathological analysis to verify the significant role of PAX6 in disease progression. Complementary diagnoses were established through public clinical datasets and transcriptome sequencing of specific cell lines. ATAC-seq was used to detect the chromatin accessibility of specific cell lines.
RESULTS: PAX6 expression was significantly elevated in NEPC and negatively regulated by AR signaling. Activation of PAX6 in non-NEPC cells led to NE trans-differentiation, while knock-down of PAX6 in NEPC cells inhibited the development and progression of NEPC. Importantly, loss of AR resulted in an enhanced expression of PAX6, which reprogramed the lineage plasticity of prostate cancer cells to develop NE phenotypes through the MET/STAT5A signaling pathway. Through ATAC-seq, we found that a high expression level of PAX6 elicited enhanced chromatin accessibility, mainly through attenuation of H4K20me3, which typically causes chromatin silence in cancer cells.
CONCLUSIONS: This study reveals a novel neural transcription factor PAX6 could drive NEPC progression and suggest that it might serve as a potential therapeutic target for the management of NEPC.
摘要：
背景：神经内分泌前列腺癌（NEPC）是前列腺癌的致死性亚群，其特征在于神经内分泌分化和雄激素受体（AR）信号传导丧失。越来越多的证据表明，细胞谱系可塑性对于NEPC疗法的失败至关重要。尽管研究表明神经转录因子PAX6参与了耐药性，其在NEPC中的具体作用尚不清楚。
方法：通过生物信息学和免疫组织化学鉴定PAX6在NEPC中的表达。CCK8测定，集落形成试验,肿瘤球形成试验和细胞凋亡试验用于说明PAX6在体外进展中的关键作用。进行ChIP和双荧光素酶报告基因测定以确认PAX6启动子区域中AR的结合序列,以及STAT5A和MET启动子区域中PAX6的结合序列。对于体内验证，对代表NEPC亚型的异种移植模型进行病理分析,以验证PAX6在疾病进展中的重要作用.通过公共临床数据集和特定细胞系的转录组测序建立补充诊断。ATAC-seq用于检测特定细胞系的染色质可及性。
结果：PAX6表达在NEPC中显著升高，并受AR信号的负调控。PAX6在非NEPC细胞中的激活导致NE转分化，而PAX6在NEPC细胞中的敲除抑制NEPC的发生和发展。重要的是,AR的缺失导致PAX6的表达增强,这通过MET/STAT5A信号通路重新编程前列腺癌细胞的谱系可塑性以发展NE表型.通过ATAC-seq,我们发现PAX6的高表达水平引起染色质可及性增强，主要通过H4K20me3的衰减，这通常会导致癌细胞染色质沉默。
结论：这项研究揭示了一种新的神经转录因子PAX6可以驱动NEPC进展，并表明它可能作为NEPC治疗的潜在治疗靶点。
方法：通过生物信息学和免疫组织化学鉴定PAX6在NEPC中的表达。CCK8测定，集落形成试验,肿瘤球形成试验和细胞凋亡试验用于说明PAX6在体外进展中的关键作用。进行ChIP和双荧光素酶报告基因测定以确认PAX6启动子区域中AR的结合序列,以及STAT5A和MET启动子区域中PAX6的结合序列。对于体内验证，对代表NEPC亚型的异种移植模型进行病理分析,以验证PAX6在疾病进展中的重要作用.通过公共临床数据集和特定细胞系的转录组测序建立补充诊断。ATAC-seq用于检测特定细胞系的染色质可及性。
结果：PAX6表达在NEPC中显著升高，并受AR信号的负调控。PAX6在非NEPC细胞中的激活导致NE转分化，而PAX6在NEPC细胞中的敲除抑制NEPC的发生和发展。重要的是,AR的缺失导致PAX6的表达增强,这通过MET/STAT5A信号通路重新编程前列腺癌细胞的谱系可塑性以发展NE表型.通过ATAC-seq,我们发现PAX6的高表达水平引起染色质可及性增强，主要通过H4K20me3的衰减，这通常会导致癌细胞染色质沉默。
结论：这项研究揭示了一种新的神经转录因子PAX6可以驱动NEPC进展，并表明它可能作为NEPC治疗的潜在治疗靶点。
