title: Understanding the mechanism of action of poly(amidoamine)s as endosomolytic polymers: correlation of physicochemical and biological properties.
abstract: Bioresponsive poly(amidoamine)s (PAA)s are currently under development as endosomolytic polymers for intracellular delivery of proteins and genes. Here for the first time, small-angle neutron scattering (SANS) is used to systematically investigate the pH-dependent conformational change of an endosomolytic polymer, the PAA ISA 23. The radius of gyration of the ISA23 was determined as a function of pH and counterion, the aim being to correlate changes in polymer conformation with membrane activity assessed using a rat red blood cell haemolysis assay. With decreasing pH, the ISA23 radius of gyration increased to a maximum (R(g) approximately 80 A) around pH = 3, before subsequently decreasing once more. At high pH and therefore high ionic strengths, the polymer is negatively charged and adopts a rather compact structure (R(g) approximately 20 A), presumably with the dissociated carboxylic groups on the exterior of the polymer coil. At low pH, the coil again collapses (R(g) < 20 A), presumably due to the effects of the high ionic strength. It is concluded that the nature of the salt form has no direct bearing on the size of the polymer coil, but it does indirectly determine the prevailing pH and, hence, polymer conformation. Pulsed-gradient spin-echo NMR measurements were in good agreement with the SANS estimates of the radius of gyration, although ISA23 polydispersity does complicate the data interpretation/comparison. These results support the proposed mode of action of PAAs, namely a coil expansion on passing from a neutral pH (extracellular) to an acidic pH (endosomal and lysosomal) environments. The results do, however, suggest that the charge on the polymer shows a closer correlation with the haemolysis activity rather than the polymer conformation.
authors.affiliation: School of Chemistry, Cardiff University, PO Box 912, Cardiff CF10 3TB, United Kingdom. griffithspc@cardiff.ac.uk
authors.forename: P C
authors.initials: PC
authors.lastname: Griffiths
authors.forename: A
authors.initials: A
authors.lastname: Paul
authors.forename: Z
authors.initials: Z
authors.lastname: Khayat
authors.forename: Ka-Wai
authors.initials: KW
authors.lastname: Wan
authors.forename: S M
authors.initials: SM
authors.lastname: King
authors.forename: I
authors.initials: I
authors.lastname: Grillo
authors.forename: R
authors.initials: R
authors.lastname: Schweins
authors.forename: P
authors.initials: P
authors.lastname: Ferruti
authors.forename: J
authors.initials: J
authors.lastname: Franchini
authors.forename: R
authors.initials: R
authors.lastname: Duncan
mesh_terms: D000818:Animals; D001672:Biocompatible Materials / Q000737:chemistry / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D011992:Endosomes / Q000187:drug effects*; D004912:Erythrocytes / Q000187:drug effects; D006461:Hemolysis / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D008422:Materials Testing; D008855:Microscopy, Electron, Scanning / Q000379:methods; D008968:Molecular Conformation; D008970:Molecular Weight; D033363:Neutron Diffraction; D009994:Osmolar Concentration; D010879:Piperazines / Q000737:chemistry*; D011073:Polyamines / Q000737:chemistry* / Q000494:pharmacology; D011108:Polymers / Q000737:chemistry* / Q000494:pharmacology; D051381:Rats
chemical_list: D001672:Biocompatible Materials; C496056:ISA23 polymer; D010879:Piperazines; D011073:Polyamines; D011108:Polymers