Protocol for preparing dynamic covalent macrocycles for co-delivering genes and drugs to cancer cell lines

Combination therapy using effective drug molecules and functional genes such as small interfering RNA (siRNA) has been suggested as a powerful strategy against multiple drug resistance. Here, we present a protocol for preparing a delivery system by developing dynamic covalent macrocycles using a dithiol monomer to co-deliver doxorubicin and siRNA. We describe steps for preparing the dithiol monomer, followed by co-delivery to form nanoparticles. We then detail procedures for cell uptake and assessing enhanced anti-cancer efficacy in vitro. For complete details on the use and execution of this protocol, please refer to Lyu et al.1 [Display omitted] •Synthetic procedures of a dithiol monomer that can generate ring-like molecules•Prepare drug and gene co-delivery systems using the above macrocycles•Experimental details for cell uptake and in vitro anti-cancer efficacy Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. Combination therapy using effective drug molecules and functional genes such as small interfering RNA (siRNA) has been suggested as a powerful strategy against multiple drug resistance. Here, we present a protocol for preparing a delivery system by developing dynamic covalent macrocycles using a dithiol monomer to co-deliver doxorubicin and siRNA. We describe steps for preparing the dithiol monomer, followed by co-delivery to form nanoparticles. We then detail procedures for cell uptake and assessing enhanced anti-cancer efficacy in vitro.