On-Chip Synthesis of Hyaluronic Acid-Based Nanoparticles for Selective Inhibition of CD44+ Human Mesenchymal Stem Cell Proliferation

Enrica Chiesa; Federica Riva; Rossella Dorati; Antonietta Greco; Stefania Ricci; Silvia Pisani; Maddalena Patrini; Tiziana Modena; Bice Conti; Ida Genta

doi:10.3390/pharmaceutics12030260

On-Chip Synthesis of Hyaluronic Acid-Based Nanoparticles for Selective Inhibition of CD44+ Human Mesenchymal Stem Cell Proliferation

Pharmaceutics. 2020 Mar 13;12(3):260. doi: 10.3390/pharmaceutics12030260.

Authors

Enrica Chiesa¹, Federica Riva², Rossella Dorati^{1

3}, Antonietta Greco¹, Stefania Ricci², Silvia Pisani⁴, Maddalena Patrini⁵, Tiziana Modena^{1

3}, Bice Conti^{1

3}, Ida Genta^{1

3}

Affiliations

¹ Department Drug Sciences, University of Pavia, V.le Taramelli 12, 27100 Pavia, Italy.
² Department of Public Health, Experimental and Forensic Medicine, Histology and Embryology Unit, University of Pavia, Via Forlanini 10, 27100 Pavia, Italy.
³ Polymerix srl, V.le Taramelli 24, 27100 Pavia, Italy.
⁴ Immunology and Transplantation Laboratory, Pediatric Hematology Oncology Unit, Department of Maternal and Children's Health, Fondazione IRCCS Policlinico S. Matteo, 27100 Pavia, Italy.
⁵ Department of Physics, University of Pavia, Via Bassi 6, 27100 Pavia, Italy.

Abstract

In this study, an innovative microfluidics-based method was developed for one-step synthesis of hyaluronic acid (HA)-based nanoparticles (NPs), by exploiting polyelectrolytic interactions between HA and chitosan (CS), in order to improve reliability, reproducibility and possible scale-up of the NPs preparation. The on-chip synthesis, using a staggered herringbone micromixer, allowed to produce HA/CS NPs with tailored-made size and suitable for both parenteral (117.50 ± 4.51 nm) and loco-regional (349.15 ± 38.09 nm) administration, mainly composed by HA (more than 85% wt) with high negative surface charge (< -20 mV). HA/CS NPs were successfully loaded with a challenging water-insoluble molecule, Everolimus (EVE), an FDA- and EMA-approved anticancer drug able to lead to cell cycle arrest, reduced angiogenesis and promotion of apoptosis. HA/CS NPs resulted to be massively internalized in CD44+ human mesenchymal stem cells via CD44 receptor-mediated endocytosis. HA/CS NPs selectiveness towards CD44 was highlighted by blocking CD44 receptor by anti-CD44 primary antibody and by comparison to CS-based NPs cellular uptake. Eventually, high effectiveness in inhibiting cell proliferation was demonstrated on-chip synthetized EVE loaded HA/CS NPs by tracking in vitro DNA synthesis.

Keywords: CD44 targeting; Everolimus; chitosan; human mesenchymal stem cells; hyaluronic acid-based nanocarriers; microfluidics.