Manufacturing of a Secretoneurin Drug Delivery System with Self-Assembled Protamine Nanoparticles by Titration

Abstract

Since therapeutic peptides and oligonucleotides are gathering interests as active pharmaceutical ingredients (APIs), nanoparticulate drug delivery systems are becoming of great importance. Thereby, the possibility to design drug delivery systems according to the therapeutic needs of APIs enhances clinical implementation. Over the last years, the focus of our group was laid on protamine-oligonucleotide-nanoparticles (so called proticles), however, the possibility to modify the size, zeta potential or loading efficiencies was limited. Therefore, at the present study we integrated a stepwise addition of protamine (titration) into the formation process of proticles loaded with the angiogenic neuropeptide secretoneurin (SN). A particle size around 130 nm was determined when proticles were assembled by the commonly used protamine addition at once. Through application of the protamine titration process it was possible to modify and adjust the particle size between approx. 120 and 1200 nm (dependent on mass ratio) without influencing the SN loading capacity. Dynamic light scattering pointed out that the difference in particle size was most probably the result of a secondary aggregation. Initially-formed particles of early stages in the titration process aggregated towards bigger assemblies. Atomic-force-microscopy images also revealed differences in morphology along with different particle size. In contrast, the SN loading was only influenced by the applied mass ratio, where a slight saturation effect was observable. Up to 65% of deployed SN could be imbedded into the proticle matrix. An in-vivo biodistribution study (i.m.) showed a retarded distribution of SN from the site of injection after the application of a SN-proticle formulation. Further, it was demonstrated that SN loaded proticles can be successfully freeze-dried and resuspended afterwards. To conclude, the integration of the protamine titration process offers new possibilities for the formulation of proticles in order to address key parameters of drug delivery systems as size, API loading or modified drug release.

Protamine – A Review on an Oligonucleotide-Binding Peptide Applied in Nanopharmaceuticals including Vaccines

“…As one of the most remarkable findings, it is possible to use protamine as delivery system for biomolecules, such as CPPs for in vivo gene transport. The researchers mostly focus on protamine's cell penetrating and nucleus targeting properties (Sorgi, Bhattacharya, and Huang 1997;Junghans, Kreuter, and Zimmer 2000;Lochmann et al 2005;Scheicher et al 2016;Fresacher et al 2019). In addition, there are several working groups introducing protamine in different nanosized formulations to enhance cell penetration (Delgado et al 2012;Mayer et al 2005;Dinauer et al 2004).…”

“…The data imply that the nanoparticle manufacture by titration facilitates the modification of particle size, which is most probably connected to the second titration step. Only the applied mass ratio, but not the manufacturing method influenced the drug loading (Scheicher et al 2016).…”

“…In an in-vivo biodistribution study they could demonstrate a retarded distribution of secretoneurin after secretoneurin-proticle injection. The innovative nanoparticle production method also offers new possibilities for Proticle engineering and handling with respect to stability and storing (Scheicher et al 2016).…”

“…Protamines are non-invasive cell penetrating peptides, showing the ability to target drugs to specific molecules within the cells (Park et al 2005;Heitz, Morris, and Divita 2009a;Chugh, Eudes, and Shim 2010;Choi et al 2010;Bashyal et al 2016;Zhang, Wang, and Xu 2016). Their penetration and targeting effect can be further enhanced by creating innovative, nanosized drug delivery systems (Lochmann, Jauk, and Zimmer 2004;Delgado et al 2012;Yuan et al 2013;Scheicher, Schachner-Nedherer, and Zimmer 2015;Scheicher et al 2016;Silva, Almeida, and Vale 2019;Fresacher et al 2019). Thus, the first part of this review will especially highlight the ongoing research in the field of protamine-associated nanotechnology, giving details about the self-assembling manufacturing processes, the properties of the resulting nanoparticles and how they can be functionalized.…”

Use of Protamine in Nanopharmaceuticals—A Review

“…Protamines are noninvasive cell-penetrating peptides, showing the ability to target drugs to specific molecules within the cells [9,[16][17][18][19][20][21][22] . Their penetration and targeting effect can be further enhanced by creating innovative, nanosized drug delivery systems [23][24][25][26][27][28][29].…”