Tamoxifen resistance remains a clinical challenge for hormone receptor-positive breast cancer. Recently, dysregulations in autophagy have been suggested as a potential mechanism for tamoxifen resistance. Although the long noncoding RNA H19 is involved in various stages of tumorigenesis, its role in tamoxifen resistance remains unknown. Here, we assessed the role of H19 in the development of tamoxifen-resistant breast cancer. Quantitative real-time PCR analyzed expression of H19 in tamoxifen-resistant breast cancer tissues. Knockdown of H19 was used to assess the sensitivity to tamoxifen in vitro and in vivo. Both knockdown and overexpression of H19 were used to analyze the status of autophagy. Real-time quantitative methylation-specific polymerase chain reaction, chromatin immunoprecipitation, immunofluorescence, and Western blot were used to explore the tamoxifen resistance mechanism of H19. In this study, we observed that the expression of H19 was substantially upregulated in tamoxifen-resistant breast cancer cell line and tumor tissues, and knockdown of H19 enhanced the sensitivity to tamoxifen both in vitro and in vivo. Furthermore, knockdown of H19 significantly inhibited autophagy in MCF7 tamoxifen-resistant (MCF7/TAMR) cells. Conversely, overexpression of H19 promoted autophagy. Interestingly, overexpression of H19 in MCF7 tamoxifen-sensitive cells could recapitulate tamoxifen resistance. Moreover, an increase in methylation in the promoter region of Beclin1 was observed in MCF7/TAMR-shH19 cells. In the double knockdown groups, both shH19+shSAHH and shH19+shDNMT3B rescued the Beclin1 promoter region methylation levels and reactivated autophagy functions. A chromatin immunoprecipitation assay further validated that DNMT3B binds to the Beclin1 promoter region and the knockdown of H19 increases this binding. Our findings demonstrate that H19 induces autophagy activation via the H19/SAHH/DNMT3B axis, which could contribute to tamoxifen resistance in breast cancer.

中文翻译：

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长的非编码RNA H19通过自噬促进乳腺癌中他莫昔芬的抗性。

他莫昔芬的耐药性仍然是激素受体阳性乳腺癌的临床挑战。最近，自噬失调被认为是他莫昔芬耐药的潜在机制。尽管长的非编码RNA H19参与肿瘤发生的各个阶段，但其在他莫昔芬抗药性中的作用仍然未知。在这里，我们评估了H19在他莫昔芬耐药性乳腺癌的发生中的作用。实时定量PCR分析了他莫昔芬耐药性乳腺癌组织中H19的表达。击倒H19用于评估他莫昔芬在体外和体内的敏感性。击倒和过表达H19都用于分析自噬的状态。实时定量甲基化特异性聚合酶链反应，染色质免疫沉淀，免疫荧光，免疫印迹法和Western blot研究了他莫昔芬对H19的耐药机制。在这项研究中，我们观察到H19的表达在耐他莫昔芬的乳腺癌细胞系和肿瘤组织中显着上调，而敲除H19则增强了他莫昔芬在体外和体内的敏感性。此外，H19的敲低显着抑制了耐MCF7他莫昔芬（MCF7 / TAMR）细胞的自噬。相反，H19的过表达促进自噬。有趣的是，MCF7他莫昔芬敏感细胞中H19的过表达可以概括他莫昔芬的耐药性。此外，在MCF7 / TAMR-shH19细胞中观察到Beclin1启动子区域甲基化的增加。在双重击倒组中，shH19 + shSAHH和shH19 + shDNMT3B均可挽救Beclin1启动子区域的甲基化水平并重新激活自噬功能。染色质免疫沉淀试验进一步验证了DNMT3B与Beclin1启动子区域结合，而敲除H19则增加了这种结合。我们的发现表明，H19通过H19 / SAHH / DNMT3B轴诱导自噬激活，这可能有助于乳腺癌中他莫昔芬的耐药性。