# Camptothecin-Bearing PEGylated Polypropylenimine Dendriplexes for Prostate Cancer Gene Therapy: Impact of Microfluidic Processing on Physicochemical Properties and Transfection

## TL;DR

This study compares microfluidic and traditional methods for making drug and gene delivery systems for prostate cancer treatment.

## Contribution

The study introduces microfluidic processing as a scalable and reproducible method for preparing camptothecin-bearing dendriplexes for prostate cancer gene therapy.

## Key findings

- Microfluidic processing produced stable, nanosized dendriplexes with efficient DNA condensation.
- Microfluidic dendriplexes showed enhanced cellular uptake in prostate cancer cells.
- Transfection efficiency in PC3-Luc cells was comparable to conventional methods.

## Abstract

Background/Objectives: Prostate cancer is the most commonly diagnosed cancer in men and a leading cause of cancer-related mortality, highlighting the need for delivery systems capable of efficiently transporting both chemotherapeutic drugs and therapeutic genes to tumor cells. Generation-3 diaminobutyric polypropylenimine (DAB) dendrimers display low toxicity, high drug loading capacity and efficient gene delivery, and can be engineered as camptothecin-bearing PEGylated carriers complexed with plasmid DNA. The aim of this study was to compare microfluidic processing with conventional hand mixing for the preparation of camptothecin-bearing PEGylated DAB dendriplexes and to evaluate the impact of formulation methods and microfluidic parameters on their physicochemical properties, cellular uptake and gene expression in prostate cancer cells. Methods: Camptothecin-bearing PEGylated DAB dendrimers were synthesized and complexed with plasmid DNA to form dendriplexes. Formulations were prepared either by microfluidics, using different total flow rates and aqueous: organic flow rate ratios, or by conventional hand mixing. The resulting dendriplexes were characterized for DNA condensation, particle size, polydispersity index and zeta potential. Morphology was assessed by transmission electron microscopy. Cellular uptake of fluorescein-labelled DNA and β-galactosidase reporter gene expression were evaluated in PC3-Luc and DU145 prostate cancer cells. Results: Both microfluidic and hand-mixed methods produced stable, nanosized, positively charged dendriplexes with efficient and sustained DNA condensation (more than 99% over 24 h). Microfluidic processing, particularly at an aqueous: organic flow rate ratio of 3:1, yielded dendriplexes with hydrodynamic diameters and zeta potentials comparable to or slightly improved over hand-mixed formulations. These microfluidic conditions significantly enhanced cellular uptake in both PC3-Luc and DU145 cells. In PC3-Luc cells, this translated into β-galactosidase expression levels comparable to hand-mixed dendriplexes and higher than naked DNA, whereas in DU145 cells, transfection efficiencies remained modest for all formulations despite increased uptake. Conclusions: Microfluidic processing enables the reproducible and scalable preparation of camptothecin-bearing PEGylated DAB dendriplexes with tunable physicochemical properties. Under selected conditions, in vitro cellular uptake and gene expression were comparable to conventional hand mixing, supporting microfluidics as a robust alternative platform for the manufacture of dendrimer-based systems for combined chemo–gene delivery in prostate cancer.