Applied Virology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
Applied Biotechnology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran.
Arch Iran Med. 2020 Sep 1;23(9):593-599. doi: 10.34172/aim.2020.69.
Selective therapy has always been the main challenge in cancer treatments. Various non-replicative oncolytic viral systems have revealed the safety and efficacy of using viruses and these products. The aim of this paper is to examine the impact of recombinant apoptin on the proliferation of lung cancer and breast cancer cell lines.
The present study consisted of two steps of expression of recombinant apoptin and its anti-proliferative effects on normal and cancer cells. In the first step, following bioinformatics and optimizing apoptin gene sequencing and synthesis, it was expressed using vector PET28a and E. coli BL21 (DE3). The expressed recombinant apoptin was confirmed by analytical SDSPAGE and then purified using Ni affinity chromatography. In the second step, the antiproliferative effects of recombinant apoptin on lung cancer, breast cancer and primary cell lines were determined using MTT assay.
According to the results of SDS-PAGE gel assay, recombinant apoptin was visible in the 14 kDa band. Also, the MTT assay results indicated that the antiproliferative effects of recombinant apoptin in cancer cell lines was different compared with the primary cell line, and followed a dose-dependent manner in both cell lines. The highest cytotoxicity (lowest cell viability) groups were 0.2 mg/mL in lung cancer (0.32 ± 0.015) (<0.001), and in breast cancer (0.33 ± 0.031) (<0.001) and 0.032 mg/mL in primary cells (0.17 ± 0.004) (<0.01), as compared to the control groups.
Our results confirmed that recombinant apoptin can induce antiproliferative effects in lung cancer and breast cancer cell lines, but not in normal monkey kidney cell line Vero; thus, it can be introduced as a promising novel specific antitumor agent after further evaluation in clinical trials.
选择性治疗一直是癌症治疗的主要挑战。各种非复制性溶瘤病毒系统已经揭示了使用病毒和这些产品的安全性和有效性。本文旨在研究重组凋亡素对肺癌和乳腺癌细胞系增殖的影响。
本研究包括表达重组凋亡素及其对正常和癌细胞的抗增殖作用的两个步骤。在第一步中,通过生物信息学和优化凋亡素基因测序和合成,使用载体 PET28a 和大肠杆菌 BL21(DE3)进行表达。通过分析 SDS-PAGE 确认表达的重组凋亡素,然后使用 Ni 亲和层析进行纯化。在第二步中,通过 MTT 测定法测定重组凋亡素对肺癌、乳腺癌和原代细胞系的抗增殖作用。
根据 SDS-PAGE 凝胶分析结果,在 14 kDa 带中可见重组凋亡素。此外,MTT 测定结果表明,与原代细胞系相比,重组凋亡素在癌细胞系中的抗增殖作用不同,并且在两种细胞系中均呈剂量依赖性。肺癌(0.32±0.015)(<0.001)和乳腺癌(0.33±0.031)(<0.001)中最高细胞毒性(最低细胞活力)组分别为 0.2mg/mL,而原代细胞中为 0.032mg/mL(0.17±0.004)(<0.01),与对照组相比。
我们的结果证实,重组凋亡素可以诱导肺癌和乳腺癌细胞系的抗增殖作用,但不能诱导正常猴肾细胞系 Vero 的抗增殖作用;因此,在临床试验中进一步评估后,它可以作为一种有前途的新型特异性抗肿瘤药物引入。