OpenAIRE is about to release its new face with lots of new content and services.
During September, you may notice downtime in services, while some functionalities (e.g. user registration, login, validation, claiming) will be temporarily disabled.
We apologize for the inconvenience, please stay tuned!
For further information please contact helpdesk[at]

fbtwitterlinkedinvimeoflicker grey 14rssslideshare1
Khayat, Zeena.
Languages: English
Types: Doctoral thesis
Subjects: RS
Polyamidoamines PAAs are a family of synthetic, water-soluble, linear polymers that are synthesised by hydrogen-transfer polyaddition of aliphatic amines or bis-secondary amines to bisacrylamides (reviewed by Ferruti et al. 2002). Over the last 15 years, PAAs been developed as drug carriers and as pH-responsive polymers for protein and gene delivery (Richardson et al., 2001). The latter, called endosomolytic PAAs, can disrupt membranes at low pH, but the delivery of genes and proteins is still poorly efficient. The precise mode of action is still poorly understood, therefore the main aim of this work was to examine the physico-chemical properties of PAAs in order to better define their mechanism of membrane permeabilisation to allow design of more efficient chemical structures. First PAA ISA1 (23 k g/mol) and ISA23 (52 - 67 k g/mol) were synthesised and were characterised using H-NMR, GPC and acid-base titration. In agreement with past studies, ISA23.HC1 demonstrated a pH-dependant haemolytic activity. The solution conformation of ISA23.HC1 was then investigated using small-angle neutron scattering (SANS). ISA23.HC1 possessed a Gaussian coil like-shape in solution at all pHs (pH 2 to 14) and polymer concentrations examined (10 mg/ml to 50 mg/ml). However as the pH decreased the radius of gyration increased to a maximum (Rg ~8 nm) at ~pH 3. Surface tension, electron paramagnetic resonance (EPR) and SANS were then used to investigate the interaction of ISA23.HC1 with simple micelles as model surfaces. Both pH and micelle composition played an important role in the strength of interaction seen. For SDS micelles, three different responses were observed, strong interaction at pH < 4.5, weak interaction at pH 4.5 - 6.5, and no interaction at pH > 7. However, ISA23.HC1 did not appear to interact with more biologically relevant lyso-PC surfactants at pHs 7.4 - 5.5. EPR experiments showed that micelle fluidity was effected when a PAA-micelle interaction occurred, and SANS reinforced this conclusion. Finally, liposomes were used as a more complex model membrane. Liposomes were prepared to mimic three phospholipids composition of plasma, endosomal and lysosomal membranes. The liposomes were stable over 2 days at pH 7.4, 5.5 and 4, but not at pH 3. SANS was used to study polymer-liposomes interaction, and the Schultz polydisperse 3 shell sphere model was used to fit the liposomal (alone) SANS scattering data. The radius of the plasma, endosome and lysosome liposomes was ~ 50 nm. When liposomes were incubated in the presence of ISA23.HC1, it was difficult to interpret the scattering data. However, it was clear that at pH 4 and 5.5 scattering of model liposomes alters in the presence of PAA indicating possible interaction. Using a contrast approach the scattering data for ISA23.HC1 in the presence of model liposomes could be extracted. This was again fitted using Gaussian coil model. Generally, the polymer increased in size in the presence of liposomes, at pHs where there was an interaction. However, these are only preliminary experiments, and there is a need for further mathematical modelling. In conclusion, this project emphasise the importance of combining the different disciplines to fully understand a system that can be the first step in finding a way to cure devastating genetic diseases.
  • The results below are discovered through our pilot algorithms. Let us know how we are doing!

    • Aswal, V.K., 2003. Salt effect in ionic mixed micelles. Chem. Phys. Lett. 371, 371-377.
    • Barbucci, R., Casolar, M., Ferruti, P., Barone, V., Lelj, F.L., Oliva, L., 1981. Macroinorganics. 7. Property-structure relationships for polymeric bases whose monomeric units behave independently toward protonation. Macromolecules 14, 1203-1209.
    • Bergstrom, M.L.L., Kjellin, M.U.R., Claesson, P.M., 2004. Small-angle neutron scattering study of mixtures o f cationic polyelectrolyte and anionic surfactant: effect of polyelectrolyte charge density. J. Phys. Chem. B 108, 1874-1881.
    • Boussif, O., Lezoualch, F., Zanta, M.A., Mergnym, M.D., Scherman, D„ Demeneix, B., Behr, J.-P., 1995. A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: polyethylenimine. Proc. Natl. Acad. Sci. U.S.A. 92, 7297-7301.
    • Check, E., 2003. Harmful potential o f viral vectors fuels doubts over gene therapy. Nature 423, 573-574.
    • Debye, P., 1947. Molecular-weight determination by light scattering. J. Phys. Colloid Chem. 51, 18-32.
    • Duncan, R., Ferruti, P., Sgouras, D„ Tuboku-Metzger, A., Ranucci, E., Bignotti, F.A., 1994. Polymer-triton X-100 conjugate capable of pHdependent red blood cell lysis: a model system illustrating the possibility o f drug delivery within acidic intracellular compartments. J. Drug Target. 2, 341-347.
    • Duncan, R„ 2003. The dawning era o f polymer therapeutics. Nat. Rev. Drug Discov. 2, 347-360.
    • Edelstein, M.L., Abedi, M.R., Wixon, J., Edelstein, R.M., 2004. J. Gene Med. 6, 560-597.
    • Feng, J., Huang, R., 1997. Studies on the aqueous solution properties o f AMPS-DMAEMA polyampholytes. Gaofenzi Cailiao Kexue Yu Gongcheng 13, 109-113.
    • Ferruti, P., Marchisio, M.A., Barbucci, R., 1985. Synthesis, physico-chemical properties and biomedical applications of poly(amido-amine)s. Polymer 26, 1336-1348.
    • Ferruti, P., Manzoni, S., Richardson, S.C.W., Duncan, R., Pattrick, N.G., Mendichi, R., Casolaro, M„ 2000. Amphoteric linear poly(amidoamine)s as endosomolytic polymers: correlation between physicochemical and biological properties. Macromolecules 33, 7793-7800.
    • Ferruti, P., Marchisio, M.A., Duncan, R., 2002. Poly(amido-amine)s: biomedical applications. Macromol. Rapid Commun. 23, 332-355.
    • Fischer, D., Li, Y.X., Ahlemeyer, B„ Krieglstein, J., Kissel, T„ 2003. In vitro cytotoxicity testing of poly cations: influence o f polymer structure on cell viability and hemolysis. Biomaterials 24, 1121-1131.
    • Funhoff, A.M., van Nostrum, C.F., Koning, G.A., Schuurmans-Nieuwenbroek, N.M.E., Crommelin, D.J.A., Hennink, W.E., 2004. Endosomal escape of polymeric gene delivery complexes is not always enhanced by polymers buffering at low pH. Biomacromolecules 5, 32-39.
    • Griffiths, P.C., Paul, A., Khayat, Z., Wan, K.W., King, S.M., Grillo, I., Schweins, R., Ferruti, P., Franchini, J., Duncan, R., 2004a. Understanding the mechanism of action of poly(amidoamine)s as endosomolytic polymers: correlation of physicochemical and biological properties. Biomacromolecules 5, 1422-1427.
    • Griffiths, P.C., Paul, A., Heenan, R.K., Penfold, J., Ranganathan, R„ Bales, B.L., 2004b. Role of counterion concentration in determining micelle aggregation: evaluation of the combination of constraints from small-angle neutron scattering, electron paramagnetic resonance, and time-resolved fluorescence quenching. J. Phys. Chem. B 108, 3810-3816.
    • Guinier, A., Foumet, G., 1955. Small Angle Scattering of X-rays. Wiley, New York.
    • Hacein-Bey-Abina, S., von Kalle, C., Schmidt, M., Le Deist, F., Wulffraat, N., Mcintyre, E., Radford, I., Villeval, J.L., Fraser, C.C., Cavazzana-Calvo, M., Fischer, A., 2003. A serious adverse event after successful gene therapy for X-linked severe combined immunodeficiency. New Engl. J. Med. 348, 255-256.
    • Hedden, R.C., Bauer, B.J., 2003. Structure and dimensions of PAMAM/PEG dendrimer-star polymers. Macromolecules 36, 1829-1835.
    • Hickl, P., Ballauff, M., Scherf, U., Muellen, K., Lindner, P., 1997. Characterization of a ladder polymer by small-angle X-ray and neutron scattering. Macromolecules 30, 273-279.
    • Ilarduya, C.T.D., DuzgUnes, N., 2000. Efficient gene transfer by transferrin lipoplexes in the presence of serum. Biochim. Biophys. Acta 1463, 333-342.
    • Kholodenko, A.L., 1993a. Analytical calculation of the scattering function for polymers of arbitrary flexibility using the Dirac propagator. Macromolecules 26, 4179-4183.
    • Kholodenko, A.L., 1993b. Scattering function for semiflexible polymers: Dirac versus Kratky-Porod. Phys. Lett. A 178, 1-2.
    • King, S., Griffiths, P.C., Hone, J., Cosgrove, T., 2002. SANS from adsorbed polymer layers. Macromol. Symp. 190, 33-42.
    • Koetz, J., Hahn, M., Philipp, B., Bekturov, E.A., Kudaibergenov, S.E., 1993. Inter- and intramolecular interactions in polyelectrolyte complex formation with polyampholytes. Makromol. Chem. 194, 397-410.
    • Lavignac, N., Lazenby, M., Foka, P., Malgesini, B., Verpilio, I., Ferruti, P., Duncan, R., 2004. Synthesis and endosomolytic properties o f poly(amidoamine) block copolymers. Macromol. Biosci. 20, 922- 929.
    • Mendichi, R., Ferruti, P., Malgesini, B., 2005a. Evidence of aggregation in dilute solution of amphoteric poly(amido-amine)s by size exclusion chromatography. Biomed. Chromatogr. 19, 96-201.
    • Mendichi, R., Ferruit, P., Malgesini, B., 2005b. Evidence of aggregation in dilute solution of amphoteric poly(amido-amines)s by size exclusion chromatography. Biomed. Chromatogr. 19, 196-201.
    • Murthy, N., Robichaud, J., Tirrel, D., Stayton, P., Hoffman, A., 1999. The design and synthesis of polymers for eukaryotic membrane disruption. J. Contr. Release 61, 137-143.
    • Nakamura, K., Shikata, T., Takahashi, N., Kanaya, T., 2005. Highly extended conformation of polyelectrolytes incorporated into hybrid threadlike micelles studied by small angle neutron scattering. J. Am. Chem. Soc. 127, 4570-4571.
    • Nishida, K., Kaji, K., Kanaya, T., Shibano, T., 2002. Added salt effect on the intermolecular correlation in flexible polyelectrolyte solutions: smallangle scattering study. Macromolecules 35, 4084-4089.
    • Ohana, P., Gofrit, O., Ayesh, S., Al-Sharef, W„ Mizrahi, A., Birman, T„ Schneider, T., Matouk, I., de Groot, N„ Tavdy, E., Sidi, A.A., Hochberg, A., 2004. Regulatory sequences of the HI9 gene in DNA based therapy o f bladder. Gene Ther. Mol. Biol. 8, 181-192.
    • Pack, D.W., Hoffman, A.S., Pun, S., Stayton, P., 2005. Design and development of polymers for gene delivery. Nat. Rev. Drug Discov. 4, 581- 593.
    • Pattrick, N.G., Richardson, S.C.W., Casolaro, M„ Ferruti, P., Duncan, R„ 2001a. Poly(amidoamine) mediated intracytoplasmic delivery of ricin Achain and gelonin. J. Contr. Release 77, 225-232.
    • Pattrick, N.G., Ferruti, P., Duncan, R„ 2001b. Demonstration of poly(amidoamine)-mediated lysosomal membrane perturbation after administration to rats in vivo. Proc. Int. Symp. Contr. Release Bioact. Mater. 28, 864-865.
    • Pederson, J.S., 2002. Modelling of small-angle scattering data from colloids and polymers systems. In: Lindner, P., Zemb, Th. (Eds.), Neutrons, Xrays and Light: Scattering Methods Applied to Soft Condensed Matter. North-Holland.
    • Qin, L.H., Pahud, D.R., Ding, Y.Z., Beilinska, A.U., Kukowska-Latallo, J.F., Baker, J.R., Brombeig, J.S., 1998. Efficient transfer of genes into murine cardiac grafts by starburst polyamidoamine dendrimer. Hum. Gene Ther. 9, 553-560.
    • Ramzi, A., Scherrenberg, R., Joosten, J., Lemstra, P., Mortensen, K., 2000. Structure-property relations in dendritic polyelectrolyte solutions at different ionic strength. Macromolecules 35, 827-833.
  • No related research data.
  • No similar publications.

Share - Bookmark

Cite this article

Cookies make it easier for us to provide you with our services. With the usage of our services you permit us to use cookies.
More information Ok