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Abstract：
The pediatric liver cancers, hepatoblastoma and hepatocellular carcinoma, are dangerous cancers which often spread to the lungs. Although treatments with cisplatin significantly improve outcomes, cisplatin may not eliminate metastasis-initiating cells. Our group has recently shown that the metastatic microenvironments of hepatoblastoma contain Cancer Associated Fibroblasts (CAFs) and neuron-like cells, which initiate cancer spread from liver to lungs. In this study, we found that these cells express high levels of HDAC1; therefore, we examined if histone deacetylase inhibition improves cisplatin anti-proliferative effects and reduces the formation of tumor clusters in pediatric liver cancer metastatic microenvironments.
METHODS: New cell lines were generated from primary hepatoblastoma liver tumors (hbl) and lung metastases (LM) of HBL patients. In addition, cell lines were generated from hepatocellular neoplasm, not otherwise specified (HCN-NOS) tumor samples, and hcc cell lines. Hbl, LM and hcc cells were treated with cisplatin, SAHA or in combination. The effect of these drugs on the number of cells, formation of tumor clusters and HDAC1-Sp5-p21 axis were examined.
RESULTS: Both HBL and HCC tissue specimens have increased HDAC1-Sp5 pathway activation, recapitulated in cell lines generated from the tumors. HDAC inhibition with vorinostat (SAHA) increases cisplatin efficacy to eliminate CAFs in hbl and in hcc cell lines. Although the neuron-like cells survive the combined treatments, proliferation was inhibited. Notably, combining SAHA with cisplatin overcame cisplatin resistance in an LM cell line from an aggressive case with multiple metastases. Underlying mechanisms of this enhanced inhibition include suppression of the HDAC1-Sp5 pathway and elevation of an inhibitor of proliferation p21. Similar findings were found with gemcitabine treatments suggesting that elimination of proliferative CAFs cells is a key event in the inhibition of mitotic microenvironment.
CONCLUSIONS: Our studies demonstrate the synergistic benefits of HDAC inhibition and cisplatin to eliminate metastasis-initiating cells in pediatric liver cancers.
METHODS: New cell lines were generated from primary hepatoblastoma liver tumors (hbl) and lung metastases (LM) of HBL patients. In addition, cell lines were generated from hepatocellular neoplasm, not otherwise specified (HCN-NOS) tumor samples, and hcc cell lines. Hbl, LM and hcc cells were treated with cisplatin, SAHA or in combination. The effect of these drugs on the number of cells, formation of tumor clusters and HDAC1-Sp5-p21 axis were examined.
RESULTS: Both HBL and HCC tissue specimens have increased HDAC1-Sp5 pathway activation, recapitulated in cell lines generated from the tumors. HDAC inhibition with vorinostat (SAHA) increases cisplatin efficacy to eliminate CAFs in hbl and in hcc cell lines. Although the neuron-like cells survive the combined treatments, proliferation was inhibited. Notably, combining SAHA with cisplatin overcame cisplatin resistance in an LM cell line from an aggressive case with multiple metastases. Underlying mechanisms of this enhanced inhibition include suppression of the HDAC1-Sp5 pathway and elevation of an inhibitor of proliferation p21. Similar findings were found with gemcitabine treatments suggesting that elimination of proliferative CAFs cells is a key event in the inhibition of mitotic microenvironment.
CONCLUSIONS: Our studies demonstrate the synergistic benefits of HDAC inhibition and cisplatin to eliminate metastasis-initiating cells in pediatric liver cancers.
摘要：
小儿肝癌，肝母细胞瘤和肝细胞癌，是经常扩散到肺部的危险癌症。尽管顺铂治疗显著改善了预后，顺铂可能不能消除转移起始细胞。我们的研究小组最近表明，肝母细胞瘤的转移微环境包含癌症相关成纤维细胞（CAFs）和神经元样细胞，导致癌症从肝脏扩散到肺部。在这项研究中,我们发现这些细胞表达高水平的HDAC1；因此，我们研究了组蛋白去乙酰化酶抑制是否能改善顺铂的抗增殖作用并减少小儿肝癌转移微环境中肿瘤簇的形成.
方法：从HBL患者的原发性肝母细胞瘤肝肿瘤（hbl）和肺转移（LM）产生新的细胞系。此外,细胞系是从肝细胞肿瘤中产生的，未指定（HCN-NOS）肿瘤样本，和hcc细胞系。Hbl,用顺铂处理LM和hcc细胞，SAHA或组合。这些药物对细胞数量的影响，检查肿瘤簇的形成和HDAC1-Sp5-p21轴。
结果：HBL和HCC组织标本均增加了HDAC1-Sp5通路的激活，在由肿瘤产生的细胞系中重现。用伏立诺他(SAHA)抑制HDAC增加顺铂功效以消除hbl和hcc细胞系中的CAF。尽管神经元样细胞在联合治疗中幸存下来，增殖受到抑制。值得注意的是,SAHA与顺铂联合治疗克服了多发性转移侵袭性病例LM细胞系的顺铂耐药性。这种增强的抑制的潜在机制包括HDAC1-Sp5途径的抑制和增殖抑制剂p21的升高。在吉西他滨治疗中发现了类似的发现，表明消除增殖性CAF细胞是抑制有丝分裂微环境的关键事件。
结论：我们的研究证明了HDAC抑制和顺铂在消除小儿肝癌转移起始细胞方面的协同益处。
方法：从HBL患者的原发性肝母细胞瘤肝肿瘤（hbl）和肺转移（LM）产生新的细胞系。此外,细胞系是从肝细胞肿瘤中产生的，未指定（HCN-NOS）肿瘤样本，和hcc细胞系。Hbl,用顺铂处理LM和hcc细胞，SAHA或组合。这些药物对细胞数量的影响，检查肿瘤簇的形成和HDAC1-Sp5-p21轴。
结果：HBL和HCC组织标本均增加了HDAC1-Sp5通路的激活，在由肿瘤产生的细胞系中重现。用伏立诺他(SAHA)抑制HDAC增加顺铂功效以消除hbl和hcc细胞系中的CAF。尽管神经元样细胞在联合治疗中幸存下来，增殖受到抑制。值得注意的是,SAHA与顺铂联合治疗克服了多发性转移侵袭性病例LM细胞系的顺铂耐药性。这种增强的抑制的潜在机制包括HDAC1-Sp5途径的抑制和增殖抑制剂p21的升高。在吉西他滨治疗中发现了类似的发现，表明消除增殖性CAF细胞是抑制有丝分裂微环境的关键事件。
结论：我们的研究证明了HDAC抑制和顺铂在消除小儿肝癌转移起始细胞方面的协同益处。
