Cloning cyclodipeptide synthases of Nocardiopsis prasina in eukaryotic expression vector for evaluation of antitumor activity

• Grant number: 24/02802-1
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: March 01, 2024
• End date: February 28, 2025
• Field of knowledge: Biological Sciences - Biochemistry - Molecular Biology
• Principal Investigator: Rodrigo Esaki Tamura
• Grantee: Laura Maria Pinto Dias
• Host Institution: Instituto de Ciências Ambientais, Químicas e Farmacêuticas (ICAQF). Universidade Federal de São Paulo (UNIFESP). Campus Diadema. Diadema , SP, Brazil
• Associated research grant: 19/15619-2 - Development of high capacity adenoviral system expressing antitumoral genes, AP.JP

Abstract

The main molecule found in the supernatant of Bacillus paralicheniforms bacteria, which showed antitumor activity in prostate cancer cells, was Cycle (Phe-Tyr). In canine kidney cells (MDCK- Madin-Darby canine kidney) cycle (Phe-tyr) showed cytotoxicity that kills 50% of cells (CC50) at concentrations of 90 µM (Zhang et al. 2018). In cells of cervical carcinoma (HeLa), colon carcinoma (HT-29) and breast carcinoma (MCF-7) Phe-tyr cycle) also showed cytotoxic activity (Kilian et al. 2005). Dicyclopeptides such as cycle (Phe-tyr) are mostly found (90%) in bacteria and exhibit a wide range of biological activities (Giessen et al. 2014). These molecules are generally formed by the action of cyclodipeptide synthases (CDPs) (Gondry et al. 2009). A mutation in the cyclodipeptide synthase of Nocardiopsis prasina (CDPS-Np) favors the synthesis of cycle (phe-tyr) instead of cycle (tyr-tyr) and, therefore, is an interesting gene to be used as a gene with antitumor activity (Brockmeyer et al. 2017). Therefore, we intend to optimize the CDPS-Np gene for the human codon usage and clone this gene into a eukaryotic expression vector pcDNA3 fused to a flag peptide. Next, the ability of overexpression of this gene to induce the production of cyclo(phe-tyr) detected by nuclear magnetic resonance and the ability to reduce the viability of PC3 and DU145 prostate cancer cells will be evaluated.