RESUMO
Tamoxifen (Tam) has long been a top treatment option for breast cancer patients, but the challenge of eliminating cancer recurrence remains. Here, we identify a signalling pathway involving ELOVL2, ELOVL2-AS1, and miR-1233-3p, which contributes to drug resistance in Tam-resistant (TamR) breast cancer. ELOVL2-AS1, a long noncoding RNA, was significantly upregulated by its antisense gene, ELOVL2, which is known to be downregulated in TamR cells. Additionally, ELOVL2-AS1 underwent the most hypermethylation in MCF-7/TamR cells. Furthermore, patients with breast cancer who developed TamR during chemotherapy had significantly lower expression of ELOVL2-AS1 compared to those who responded to Tam. Ectopic downregulation of ELOVL2-AS1 by siRNA both stimulated cancer cell growth and deteriorated TamR. We also found that ELOVL2-AS1 sponges miR-1233-3p, which has pro-proliferative activity and elevates TamR, leading to the activation of potential target genes, such as MYEF2, NDST1, and PIK3R1. These findings suggest that ELOVL2-AS1, in association with ELOVL2, may contribute to the suppression of drug resistance by sponging miR-1233-3p in breast cancer.
Assuntos
Neoplasias da Mama , MicroRNAs , RNA Longo não Codificante , Feminino , Humanos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Metilação de DNA , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , MicroRNAs/metabolismo , Recidiva Local de Neoplasia/tratamento farmacológico , Recidiva Local de Neoplasia/genética , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Tamoxifeno/farmacologia , Tamoxifeno/uso terapêutico , RNA AntissensoRESUMO
While a number of coding genes have explained the anticancer activity of ginsenoside Rh2, little is known about noncoding RNAs. This study was performed to elucidate the regulatory activity of long noncoding RNA (lncRNA) CFAP20DC-AS1, which is known to be downregulated by Rh2. MiR-3614-3p, which potentially binds CFAP20DC-AS1, was screened using the LncBase Predicted program, and the binding was verified by assaying the luciferase activity of a luciferase/lncRNA recombinant plasmid construct. The competitive endogenous RNA (ceRNA) relationship of the two RNAs was further validated by quantitative PCR after deregulation of each RNA using siRNA. The effect of miRNA and target genes on the MCF-7 cancer cell growth was determined by monitoring proliferation and apoptosis in the presence of Rh2 after deregulating the corresponding gene. The miRNA decreased the luciferase activity of the luciferase/CFAP20DC-AS1 fusion vector, confirming the binding. SiRNA-based deregulation of CFAP20DC-AS1 attenuated the expression of miR-3614-3p and vice versa. In contrast to CFAP20DC-AS1, miR-3614-3p was upregulated by Rh2, inhibiting proliferation but stimulating apoptosis of the MCF-7 cells. Target genes of miR-3614-3p, BBX and TNFAIP3, were downregulated by Rh2 and the miRNA but upregulated by the lncRNA. Rh2 inhibits CFAP20DC-AS1, which obscures the association of the lncRNA with miR-3614-3p, resulting in the suppression of oncogenic BBX and TNFAIP3. Taken together, the Rh2/CFAP20DC-AS1/miR-3614-3p/target gene axis contributes to the antiproliferation activity of Rh2 in cancer cells.
Assuntos
Neoplasias da Mama , MicroRNAs , RNA Longo não Codificante , Apoptose/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Ginsenosídeos , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Interferente Pequeno , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/genética , Proteína 3 Induzida por Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND: Tamoxifen (tam) is widely used to treat estrogen-positive breast cancer. However, cancer recurrence after chemotherapy remains a major obstacle to achieve good patient prognoses. In this study, we aimed to identify genes responsible for epigenetic regulation of tam resistance in breast cancer. METHODS: Methylation microarray data were analyzed to screen highly hypomethylated genes in tam resistant (tamR) breast cancer cells. Quantitative RT-PCR, Western blot analysis, and immunohistochemical staining were used to quantify expression levels of genes in cultured cells and cancer tissues. Effects of matrix metalloproteinase-1 (MMP1) expression on cancer cell growth and drug resistance were examined through colony formation assays and flow cytometry. Xenografted mice were generated to investigate the effects of MMP1 on drug resistance in vivo. RESULTS: MMP1 was found to be hypomethylated and overexpressed in tamR MCF-7 (MCF-7/tamR) cells and in tamR breast cancer tissues. Methylation was found to be inversely associated with MMP1 expression level in breast cancer tissues, and patients with lower MMP1 expression exhibited a better prognosis for survival. Downregulating MMP1 using shRNA induced tam sensitivity in MCF-7/tamR cells along with increased apoptosis. The xenografted MCF-7/tamR cells that stably expressed short hairpin RNA (shRNA) against MMP1 exhibited retarded tumor growth compared to that in cells expressing the control shRNA, which was further suppressed by tam. CONCLUSIONS: MMP1 can be upregulated through promoter hypomethylation in tamR breast cancer, functioning as a resistance driver gene. MMP1 can be a potential target to suppress tamR to achieve better prognoses of breast cancer patients.