Bioengineering Novel Chimeric microRNA-34a for Prodrug Cancer Therapy: High-Yield Expression and Purification, and Structural and Functional Characterization

• DOI: 10.1124/jpet.115.225631
• Publication Date: 2015-05-29T02:30:22Z
• Authors: Wei-Peng Wang; Pui Yan Ho; Qiu-Xia Chen; Balasubrahmanyam ...; Patrick A. Limbach; Mei-Mei Li; Wen-Juan Wu; Joseph L. Jilek; Jing-Xin Qiu; Hong-Jian Zhang; Tianhong Li; Theodore Wun; Ralph DeVere White; Kit S. Lam; Ai-Ming Yu
• Keywords: 0301 basic medicine; Cell Survival; Nude; 610; Mice, Nude; Antineoplastic Agents; Bioengineering; Cell Line; Dose-Response Relationship; Mice; Structure-Activity Relationship; 03 medical and health sciences; Cell Line, Tumor; Genetics; Animals; Humans; Prodrugs; Pharmacology & Pharmacy; Lung; Inbred BALB C; Cancer; Mice, Inbred BALB C; Tumor; Biomedical and Clinical Sciences; Dose-Response Relationship, Drug; Lung Cancer; Pharmacology and Pharmaceutical Sciences; Hep G2 Cells; Xenograft Model Antitumor Assays; 3. Good health; MicroRNAs; Pharmacology and pharmaceutical sciences; Drug; Biotechnology

Development of anticancer treatments based on microRNA (miRNA/miR) such as miR-34a replacement therapy is limited to the use synthetic RNAs with artificial modifications. Herein, we present a new approach high-yield and large-scale biosynthesis, in <i>Escherichia coli</i> using transfer RNA (tRNA) scaffold, chimeric agent, which may act prodrug for therapy. The recombinant tRNA fusion pre–miR-34a (tRNA/mir-34a) was quickly purified high degree homogeneity (&gt;98%) anion-exchange fast protein liquid chromatography, whose primary sequence post-transcriptional modifications were directly characterized by mass spectrometric analyses. Chimeric tRNA/mir-34a showed favorable cellular stability while it degradable several ribonucleases. Deep sequencing quantitative real-time polymerase chain reaction studies revealed that tRNA-carried precisely processed mature within human carcinoma cells, same fragments produced from control scaffold (tRNA/MSA). Consequently, inhibited proliferation various types cells dose-dependent manner much greater than tRNA/MSA, mechanistically attributable reduction target genes. Furthermore, significantly suppressed growth non–small-cell lung cancer A549 hepatocarcinoma HepG2 xenograft tumors mice, compared dose tRNA/MSA. In addition, had no or minimal effect blood chemistry interleukin-6 level mouse models, suggesting well tolerated. These findings provoke conversation producing biologic miRNAs perform miRNA actions, point toward direction developing miRNA-based therapies.