Guangdong Pharmaceutical University, Guangzhou, Guangdong, China.
Missouri State University, Springfield, MO, USA.
Technol Cancer Res Treat. 2019 Jan 1;18:1533033819873636. doi: 10.1177/1533033819873636.
Unmethylated cytosine-phosphorothioate-guanine oligodeoxynucleotide, a synthetic oligodeoxynucleotide, has been used as an adjuvant in clinic and in the antitumor activity. However, the antitumor mechanism of cytosine-phosphorothioate-guanine oligodeoxynucleotide against human bladder cancer is unknown. The purpose of this study is to evaluate the cytotoxicity and molecular mechanism of anticancer effect of cytosine-phosphorothioate-guanine oligodeoxynucleotide on T24 cells (a human bladder cancer cell line).
The cytotoxic activity of cytosine-phosphorothioate-guanine oligodeoxynucleotide was examined by cell viability assay in the presence and absence of 5-fluorouracil, respectively. Apoptosis and cell-cycle phase distribution were detected by flow cytometry analysis. To investigate the molecular mechanisms of cytosine-phosphorothioate-guanine oligodeoxynucleotide cytotoxicity, the expression of antiapoptotic factors (B-cell lymphoma-2 and Survivin, β-actin as control) in RNA, and protein level was assayed by quantitative real-time polymerase chain reaction and automated capillary Western blot.
The inhibition ratio of T24 cells treated with both cytosine-phosphorothioate-guanine oligodeoxynucleotide and 5-fluorouracil was higher than those treated with either cytosine-phosphorothioate-guanine oligodeoxynucleotide or 5-fluorouracil alone. In the combination group (cytosine-phosphorothioate-guanine oligodeoxynucleotide and 5-fluorouracil), the apoptosis rate was significantly increased, and more cells were arrested at "S" and "G2/M" phases compared to those in cytosine-phosphorothioate-guanine oligodeoxynucleotide or 5-fluorouracil alone. Furthermore, the expression of antiapoptotic factors was decreased by cytosine-phosphorothioate-guanine oligodeoxynucleotide alone or combined with 5-fluorouracil.
Cytosine-phosphorothioate-guanine oligodeoxynucleotide promoted apoptosis and enhanced the chemosensitivity of 5-fluorouracil in T24 cells. Cytosine-phosphorothioate-guanine oligodeoxynucleotide downregulated the expression of antiapoptotic factors and inhibited cell-cycle phase by arresting more cells at "S" and "G2/M" phases. This study indicated the potential ability of cytosine-phosphorothioate-guanine oligodeoxynucleotide as a candidate drug for human bladder cancer.
非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸是一种合成的寡脱氧核苷酸，已被用作临床和抗肿瘤活性的佐剂。然而，非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸对人膀胱癌的抗肿瘤机制尚不清楚。本研究旨在评估非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸对 T24 细胞（人膀胱癌细胞系）的细胞毒性和抗癌作用的分子机制。
分别在存在和不存在 5-氟尿嘧啶的情况下，通过细胞活力测定法检测非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸的细胞毒性。通过流式细胞术分析检测细胞凋亡和细胞周期时相分布。为了研究非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸细胞毒性的分子机制，通过定量实时聚合酶链反应和自动毛细管 Western blot 测定 RNA 和蛋白质水平的抗凋亡因子（B 细胞淋巴瘤-2 和 Survivin，β-肌动蛋白作为对照）的表达。
同时用非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸和 5-氟尿嘧啶处理的 T24 细胞的抑制率高于单独用非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸或 5-氟尿嘧啶处理的细胞。在联合组（非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸和 5-氟尿嘧啶）中，凋亡率显著增加，与单独用非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸或 5-氟尿嘧啶处理的细胞相比，更多的细胞停滞在“S”和“G2/M”期。此外，非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸单独或联合 5-氟尿嘧啶处理均可降低抗凋亡因子的表达。
非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸促进 T24 细胞凋亡并增强 5-氟尿嘧啶的化疗敏感性。非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸通过将更多细胞阻滞在“S”和“G2/M”期来下调抗凋亡因子的表达并抑制细胞周期时相。本研究表明非甲基化胞嘧啶-硫代磷酸鸟嘌呤寡脱氧核苷酸作为人膀胱癌候选药物的潜力。
