Investigation of Dendriplexes by Ion Mobility-Mass Spectrometry

Abstract

Abstract:Highly branched polyamidoamine (PAMAM) dendrimers presenting biological activities have been envisaged as non-viral gene delivery vectors. They are known to associate with nucleic acid (DNA) in non-covalent complexes via electrostatic interactions. Although their transfection efficiency has been proved, PAMAMs present a significant cytotoxicity due to their cationic surface. To overcome such a drawback, different chemical modifications of the PAMAM surface have been reported such as the attachment of hydrophobic residues. In the present work, we studied the complexation of DNA duplexes with different low-generation PAMAM; ammonia-cored G0(N) and G1(N) PAMAM, native or chemically modified with aromatic residues, i.e., phenyl-modified-PAMAM G0(N) and phenylalanine-modified-PAMAM G1(N). To investigate the interactions involved in the PAMAM/DNA complexes, also called dendriplexes, we used electrospray ionization (ESI) coupled to ion mobility spectrometry-mass-spectrometry (IM-MS). ESI is known to allow the study of non-covalent complexes in native conditions while IM-MS is a bidimensional separation technique particularly useful for the characterization of complex mixtures. IM-MS allows the separation of the expected complexes, possible additional non-specific complexes and the free ligands. Tandem mass spectrometry (MS/MS) was also used for the structural characterization. This work highlights the contribution of IM-MS and MS/MS for the study of small dendriplexes. The stoichiometries of the complexes and the

Perspective: Dendrimer drugs for infection and inflammation

“…It is now important for a consensus to emerge on this matter; a lack of clarity amongst those working with dendrimer drugs means that the hurdle for obtaining FDA regulatory approval for clinical trial testing is very likely to remain too high for others to achieve. In this specific context, the recent analytical work of French groups to reliably and reproducibly define the precise MWt of their dendrimers (with all of the challenges that this involves) using mass spec based analytical techniques is to be highly commended [14,15,36].…”

Structural Characterization of Dendriplexes In Vacuo: A Joint Ion Mobility/Molecular Dynamics Investigation

“…Mass spectrometry analysis of intact polyanionic nucleic acids has nowadays acquired sufficient maturity to be considered as a method of choice for the structural investigation of nucleic acids, including nucleic acids complexed to dendrimers . In particular, ion mobility spectrometry (IMS) experiments represent an invaluable analytical method to generate structural data on gas-phase ions .…”

“…In 2014, Leriche et al studied for the first time dendriplexes by mass spectrometry methods, including collision-induced dissociation (CID) and ion mobility mass spectrometry experiments . Based on their CID data, the PAMAM/ds-DNA interaction was proposed to be only electrostatically driven, while for a phenyl-modified PAMAM, they suggested that intercalation also contributes to the dendrimer/ds-DNA binding . In that report, even if IMS experiments were conducted, no collision cross sections were provided and no MD simulations were presented to support the experimental-derived conclusions …”

“…18 Mass spectrometry analysis of intact polyanionic nucleic acids has nowadays acquired sufficient maturity to be considered as a method of choice for the structural investigation of nucleic acids, 19 including nucleic acids complexed to dendrimers. 20 In particular, ion mobility spectrometry (IMS) experiments represent an invaluable analytical method to generate structural data on gas-phase ions. 21 By comparing the measured collision cross sections (CCS) with theoretical CCS obtained on candidate ion structures generated upon MD simulations, description of the gas phase ion structures may be afforded at the molecular level.…”

“…28 In 2014, Leriche et al studied for the first time dendriplexes by mass spectrometry methods, including collision-induced dissociation (CID) and ion mobility mass spectrometry experiments. 20 Based on their CID data, the PAMAM/ds-DNA interaction was proposed to be only electrostatically driven, while for a phenyl-modified PAMAM, they suggested that intercalation also contributes to the dendrimer/ds-DNA binding. 20 In that report, even if IMS experiments were conducted, no collision cross sections were provided and no MD simulations were presented to support the experimentalderived conclusions.…”