Abstract
Background:Telomerase activity is up regulated in most breast cancer subtypes but not in the adjacent normal tissues. Thus, it is a promising target for anticancer therapy. The present work investigated the effects of telomerase inhibition by siRNA on breast cancer cell lines and studied the feasibility of whether the combined effect of doxorubicin with siRNA treatment on breast cancer cells potentiates a rapid cellular response to the cytotoxic effect of chemotherapy. Methods:This study was performed on Luminal A (MCF-7), triple negative (MDA-MB-468), and HER-2/neu (SKBR-3) human breast cancer cell lines, wherein telomerase activity inhibition by hTERT siRNA and doxorubicin was detected by measuring telomerase activity using Telomeric Repeat Amplification Protocol (TRAP assay), assessing cell viability through MTT assay, and evaluating apoptosis through scanning electron microscopy (SEM) and through estimating caspase-3 and -8 activities using enzyme-linked immunosorbent assay (ELISA). Results:In the present study, hTERT siRNA effectively reduced telomerase activity and cell viability to more than 90% and 60%, respectively, in most breast cancer cell lines within 72 hours after transfection. The combination of hTERT siRNA and doxorubicin showed a cumulative effect compared with either treatment alone (P < 0.05). Meanwhile, SEM demonstrated apoptotic morphologic cell changes. Conclusion:Telomerase inhibition is a promising strategy for the effective treatment of breast cancer. When used in combination with doxorubicin, it could potentiate the cytotoxic effect of the drug on breast cancer cells.
Highlights
Telomeres are repetitive, non-coding DNA sequences located at the ends of each chromosome (Louzon et al, 2019)
< 0.01), compared with the control group, as shown in Effects of different treatments on telomerase activity In Figure 2, the MCF-7 Breast cancer (BC) cell line (Figure 2a) showed a significant inhibition of telomerase activity (P < 0.001) in the combined treated group, and the small interference RNA (siRNA)-transfected group showed a significant reduction in telomerase activity (P < 0.001), in which 0.53% ± 0% of telomerase activity was only detected, the doxorubicin-treated group revealed a significant reduction in telomerase activity to 74.83% ± 3.8% (P < 0.001), compared with the control group
In the MDA-MB-468 BC cell line (Figure 2b), telomerase activity was significantly reduced to 2.0% ± 0.10% and 6.20% ± 0.96% (P < 0.001) in the siRNA-transfected and combined treated groups, respectively, and it was significantly reduced to only 52.13% ± 8.46% (P < 0.001) in the doxorubicintreated group, compared with the control group
Summary
Non-coding DNA sequences located at the ends of each chromosome (Louzon et al, 2019). HTERT has other evident biological activities beside its function in telomere lengthening It protects cancer cells from apoptosis induced by chemotherapeutic drugs. The present work investigated the effects of telomerase inhibition by siRNA on breast cancer cell lines and studied the feasibility of whether the combined effect of doxorubicin with siRNA treatment on breast cancer cells potentiates a rapid cellular response to the cytotoxic effect of chemotherapy. Methods: This study was performed on Luminal A (MCF-7), triple negative (MDA-MB-468), and HER-2/neu (SKBR-3) human breast cancer cell lines, wherein telomerase activity inhibition by hTERT siRNA and doxorubicin was detected by measuring telomerase activity using Telomeric Repeat Amplification Protocol (TRAP assay), assessing cell viability through MTT assay, and evaluating apoptosis through scanning electron microscopy (SEM) and through estimating caspase-3 and -8 activities using enzyme-linked immunosorbent assay (ELISA). When used in combination with doxorubicin, it could potentiate the cytotoxic effect of the drug on breast cancer cells
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