Authors: Nina Erwin, Umasankar De, Yufeng Xiao, Lei Wang, Chandra Maharjan, Xiaoshu Pan, Nikee Awasthee, Guangrong Zheng, Daiqing Liao, Weizhou Zhang, and Mei He
Abstract
Proteolysis targeting chimeras (PROTACs) are an emerging targeted cancer therapy approach, but wide-spread clinical use of PROTAC is limited due to poor cell targeting and penetration, and instability in vivo. To overcome such issues and enhance the in vivo efficacy of PROTAC drugs, microfluidic droplet-based electroporation (μDES) was developed as a novel extracellular vesicle (EVs) transfection system, which enables the high-efficient PROTAC loading and effective delivery in vivo. Our previously developed YX968 PROTAC drug had shown the selectively degradation of HDAC3 and 8, which effectively suppresses the growth of breast tumor cell lines, including MDA-MB-231 triple negative breast cancer (TNBC) line, via dual degradation without provoking a global histone hyperacetylation. In this study, we demonstrated that μDESbased PROTAC loading in EVs significantly enhanced therapeutic function of PROTAC drug in vivo in the TNBC breast tumor mouse model. NSG mice with pre-established MDA-MB-231 tumors and treated with intraperitoneal injection of EVs for tumor inhibition study, which showed significantly higher HDAC 3 and 8 degradation efficiency and tumor inhibition than PROTAC only group. The liver, spleen, kidney, lung, heart, and brain were collected for safety testing, which exhibited improved toxicity. The EV delivery of PROTAC drug enhances drug stability and bioavailability in vivo, transportability, and drug targeting ability, which fills an important gap in current development of PROTAC therapeutic functionality in vivo and clinical translation. This novel EV-based drug transfection and delivery strategy could be applicable to various therapeutics for enhancing in vivo delivery, efficacy, and safety.
Fig 1. (A) Schematic of the novel microfluidic droplet-based electroporation (μDES) device for PROTAC loading into EVs. (B) Uniform size and morphology of the droplets produced following μDES. Insert is the bright field image of produced droplets. The transfected PROTAC EVs were characterized in terms of (C) size, (D) concentration, (E) zeta potential, and (F) morphology, compared to conventional electroporation EVs and native EVs as the control group.
Fig 2. (A) Transfection efficiency of PROTAC loaded into EVs. (B) Transfection efficiency following storage at various conditions, representative of PROTAC loading stability. Effect of storage conditions on EV (C) size and (D) concentration.
Selected Figures
Keywords: PROTACs; microfluidics; breast cancer; HDAC; droplet generation; electroporation
bioRxiv 2024