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. 2003 May 15;372(Pt 1):65–75. doi: 10.1042/BJ20021945

Poly(2-alkylacrylic acid) polymers deliver molecules to the cytosol by pH-sensitive disruption of endosomal vesicles.

Rachel A Jones 1, Charles Y Cheung 1, Fiona E Black 1, Jasmine K Zia 1, Patrick S Stayton 1, Allan S Hoffman 1, Mark R Wilson 1
PMCID: PMC1223370  PMID: 12583812

Abstract

The permeability barrier posed by cell membranes represents a challenge for the delivery of hydrophilic molecules into cells. We previously proposed that poly(2-alkylacrylic acid)s are endocytosed by cells into acidified vesicles and are there triggered by low pH to disrupt membranes and release the contents of endosomes/lysosomes to the cytosol. If this hypothesis is correct, these polymers could be valuable in drug-delivery applications. The present paper reports functional comparisons of a family of three poly(2-alkylacrylic acid)s. Poly(2-propylacrylic acid) (PPAA), poly(2-ethylacrylic acid) (PEAA) and poly(2-methylacrylic acid) (PMAA) were compared in red-blood-cell haemolysis assays and in a lipoplex (liposome-DNA complex) assay. We also directly examined the ability of these polymers to disrupt endosomes and lysosomes in cultured human cells. Our results show that: (i) unlike membrane-disruptive peptides, the endosomal-disruptive ability of poly(2-alkylacrylic acid)s cannot necessarily be predicted from their haemolytic activity at low pH, (ii) PPAA (but not PEAA or PMAA) potently facilitates gene transfection by cationic lipoplexes and (iii) endocytosed poly(2-alkylacrylic acid)s are triggered by luminal acidification to selectively disrupt endosomes (not lysosomes) and release their contents to the cytosol. These results will facilitate the rational design of future endosomal-disrupting polymers for drug delivery.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akasaki K., Michihara A., Mibuka K., Fujiwara Y., Tsuji H. Biosynthetic transport of a major lysosomal membrane glycoprotein, lamp-1: convergence of biosynthetic and endocytic pathways occurs at three distinctive points. Exp Cell Res. 1995 Oct;220(2):464–473. doi: 10.1006/excr.1995.1338. [DOI] [PubMed] [Google Scholar]
  2. Boussif O., Lezoualc'h F., Zanta M. A., Mergny M. D., Scherman D., Demeneix B., Behr J. P. A versatile vector for gene and oligonucleotide transfer into cells in culture and in vivo: polyethylenimine. Proc Natl Acad Sci U S A. 1995 Aug 1;92(16):7297–7301. doi: 10.1073/pnas.92.16.7297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cain C. C., Sipe D. M., Murphy R. F. Regulation of endocytic pH by the Na+,K+-ATPase in living cells. Proc Natl Acad Sci U S A. 1989 Jan;86(2):544–548. doi: 10.1073/pnas.86.2.544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cheung C. Y., Murthy N., Stayton P. S., Hoffman A. S. A pH-sensitive polymer that enhances cationic lipid-mediated gene transfer. Bioconjug Chem. 2001 Nov-Dec;12(6):906–910. doi: 10.1021/bc0100408. [DOI] [PubMed] [Google Scholar]
  5. Duncan R., Gac-Breton S., Keane R., Musila R., Sat Y. N., Satchi R., Searle F. Polymer-drug conjugates, PDEPT and PELT: basic principles for design and transfer from the laboratory to clinic. J Control Release. 2001 Jul 6;74(1-3):135–146. doi: 10.1016/s0168-3659(01)00328-5. [DOI] [PubMed] [Google Scholar]
  6. Eccleston M. E., Kuiper M., Gilchrist F. M., Slater N. K. pH-responsive pseudo-peptides for cell membrane disruption. J Control Release. 2000 Nov 3;69(2):297–307. doi: 10.1016/s0168-3659(00)00316-3. [DOI] [PubMed] [Google Scholar]
  7. Finsinger D., Remy J. S., Erbacher P., Koch C., Plank C. Protective copolymers for nonviral gene vectors: synthesis, vector characterization and application in gene delivery. Gene Ther. 2000 Jul;7(14):1183–1192. doi: 10.1038/sj.gt.3301227. [DOI] [PubMed] [Google Scholar]
  8. Godbey W. T., Wu K. K., Mikos A. G. Tracking the intracellular path of poly(ethylenimine)/DNA complexes for gene delivery. Proc Natl Acad Sci U S A. 1999 Apr 27;96(9):5177–5181. doi: 10.1073/pnas.96.9.5177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Granger B. L., Green S. A., Gabel C. A., Howe C. L., Mellman I., Helenius A. Characterization and cloning of lgp110, a lysosomal membrane glycoprotein from mouse and rat cells. J Biol Chem. 1990 Jul 15;265(20):12036–12043. [PubMed] [Google Scholar]
  10. Griffith K. J., Chan E. K., Lung C. C., Hamel J. C., Guo X., Miyachi K., Fritzler M. J. Molecular cloning of a novel 97-kd Golgi complex autoantigen associated with Sjögren's syndrome. Arthritis Rheum. 1997 Sep;40(9):1693–1702. doi: 10.1002/art.1780400920. [DOI] [PubMed] [Google Scholar]
  11. Guicciardi M. E., Deussing J., Miyoshi H., Bronk S. F., Svingen P. A., Peters C., Kaufmann S. H., Gores G. J. Cathepsin B contributes to TNF-alpha-mediated hepatocyte apoptosis by promoting mitochondrial release of cytochrome c. J Clin Invest. 2000 Nov;106(9):1127–1137. doi: 10.1172/JCI9914. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hughes J. A., Aronsohn A. I., Avrutskaya A. V., Juliano R. L. Evaluation of adjuvants that enhance the effectiveness of antisense oligodeoxynucleotides. Pharm Res. 1996 Mar;13(3):404–410. doi: 10.1023/a:1016044609972. [DOI] [PubMed] [Google Scholar]
  13. Kim J. H., Lingwood C. A., Williams D. B., Furuya W., Manolson M. F., Grinstein S. Dynamic measurement of the pH of the Golgi complex in living cells using retrograde transport of the verotoxin receptor. J Cell Biol. 1996 Sep;134(6):1387–1399. doi: 10.1083/jcb.134.6.1387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lackey C. A., Murthy N., Press O. W., Tirrell D. A., Hoffman A. S., Stayton P. S. Hemolytic activity of pH-responsive polymer-streptavidin bioconjugates. Bioconjug Chem. 1999 May-Jun;10(3):401–405. doi: 10.1021/bc980109k. [DOI] [PubMed] [Google Scholar]
  15. Lackey Chantal A., Press Oliver W., Hoffman Allan S., Stayton Patrick S. A biomimetic pH-responsive polymer directs endosomal release and intracellular delivery of an endocytosed antibody complex. Bioconjug Chem. 2002 Sep-Oct;13(5):996–1001. doi: 10.1021/bc010053l. [DOI] [PubMed] [Google Scholar]
  16. Li W., Yuan X., Nordgren G., Dalen H., Dubowchik G. M., Firestone R. A., Brunk U. T. Induction of cell death by the lysosomotropic detergent MSDH. FEBS Lett. 2000 Mar 17;470(1):35–39. doi: 10.1016/s0014-5793(00)01286-2. [DOI] [PubMed] [Google Scholar]
  17. Mellman I. Endocytosis and molecular sorting. Annu Rev Cell Dev Biol. 1996;12:575–625. doi: 10.1146/annurev.cellbio.12.1.575. [DOI] [PubMed] [Google Scholar]
  18. Mellman I., Fuchs R., Helenius A. Acidification of the endocytic and exocytic pathways. Annu Rev Biochem. 1986;55:663–700. doi: 10.1146/annurev.bi.55.070186.003311. [DOI] [PubMed] [Google Scholar]
  19. Merlin J. L., Dolivet G., Dubessy C., Festor E., Parache R. M., Verneuil L., Erbacher P., Behr J. P., Guillemin F. Improvement of nonviral p53 gene transfer in human carcinoma cells using glucosylated polyethylenimine derivatives. Cancer Gene Ther. 2001 Mar;8(3):203–210. doi: 10.1038/sj.cgt.7700289. [DOI] [PubMed] [Google Scholar]
  20. Mu F. T., Callaghan J. M., Steele-Mortimer O., Stenmark H., Parton R. G., Campbell P. L., McCluskey J., Yeo J. P., Tock E. P., Toh B. H. EEA1, an early endosome-associated protein. EEA1 is a conserved alpha-helical peripheral membrane protein flanked by cysteine "fingers" and contains a calmodulin-binding IQ motif. J Biol Chem. 1995 Jun 2;270(22):13503–13511. doi: 10.1074/jbc.270.22.13503. [DOI] [PubMed] [Google Scholar]
  21. Murthy N., Robichaud J. R., Tirrell D. A., Stayton P. S., Hoffman A. S. The design and synthesis of polymers for eukaryotic membrane disruption. J Control Release. 1999 Aug 27;61(1-2):137–143. doi: 10.1016/s0168-3659(99)00114-5. [DOI] [PubMed] [Google Scholar]
  22. Noguchi Y., Wu J., Duncan R., Strohalm J., Ulbrich K., Akaike T., Maeda H. Early phase tumor accumulation of macromolecules: a great difference in clearance rate between tumor and normal tissues. Jpn J Cancer Res. 1998 Mar;89(3):307–314. doi: 10.1111/j.1349-7006.1998.tb00563.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ohkuma S., Shimizu S., Noto M., Sai Y., Kinoshita K., Tamura H. Inhibition of cell growth by bafilomycin A1, a selective inhibitor of vacuolar H(+)-ATPase. In Vitro Cell Dev Biol Anim. 1993 Nov;29A(11):862–866. doi: 10.1007/BF02631364. [DOI] [PubMed] [Google Scholar]
  24. Plank C., Oberhauser B., Mechtler K., Koch C., Wagner E. The influence of endosome-disruptive peptides on gene transfer using synthetic virus-like gene transfer systems. J Biol Chem. 1994 Apr 29;269(17):12918–12924. [PubMed] [Google Scholar]
  25. Plank C, Zauner W, Wagner E. Application of membrane-active peptides for drug and gene delivery across cellular membranes. Adv Drug Deliv Rev. 1998 Oct 5;34(1):21–35. doi: 10.1016/s0169-409x(98)00005-2. [DOI] [PubMed] [Google Scholar]
  26. Simões S., Slepushkin V., Gaspar R., de Lima M. C., Düzgüneş N. Gene delivery by negatively charged ternary complexes of DNA, cationic liposomes and transferrin or fusigenic peptides. Gene Ther. 1998 Jul;5(7):955–964. doi: 10.1038/sj.gt.3300674. [DOI] [PubMed] [Google Scholar]
  27. Stayton P. S., Hoffman A. S., Murthy N., Lackey C., Cheung C., Tan P., Klumb L. A., Chilkoti A., Wilbur F. S., Press O. W. Molecular engineering of proteins and polymers for targeting and intracellular delivery of therapeutics. J Control Release. 2000 Mar 1;65(1-2):203–220. doi: 10.1016/s0168-3659(99)00236-9. [DOI] [PubMed] [Google Scholar]
  28. Sullivan P. C., Ferris A. L., Storrie B. Effects of temperature, pH elevators, and energy production inhibitors on horseradish peroxidase transport through endocytic vesicles. J Cell Physiol. 1987 Apr;131(1):58–63. doi: 10.1002/jcp.1041310110. [DOI] [PubMed] [Google Scholar]
  29. Thomas J. L., Barton S. W., Tirrell D. A. Membrane solubilization by a hydrophobic polyelectrolyte: surface activity and membrane binding. Biophys J. 1994 Sep;67(3):1101–1106. doi: 10.1016/S0006-3495(94)80575-2. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wagner E., Plank C., Zatloukal K., Cotten M., Birnstiel M. L. Influenza virus hemagglutinin HA-2 N-terminal fusogenic peptides augment gene transfer by transferrin-polylysine-DNA complexes: toward a synthetic virus-like gene-transfer vehicle. Proc Natl Acad Sci U S A. 1992 Sep 1;89(17):7934–7938. doi: 10.1073/pnas.89.17.7934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. van Deurs B., Holm P. K., Sandvig K. Inhibition of the vacuolar H(+)-ATPase with bafilomycin reduces delivery of internalized molecules from mature multivesicular endosomes to lysosomes in HEp-2 cells. Eur J Cell Biol. 1996 Apr;69(4):343–350. [PubMed] [Google Scholar]
  32. van de Wetering P., Moret E. E., Schuurmans-Nieuwenbroek N. M., van Steenbergen M. J., Hennink W. E. Structure-activity relationships of water-soluble cationic methacrylate/methacrylamide polymers for nonviral gene delivery. Bioconjug Chem. 1999 Jul-Aug;10(4):589–597. doi: 10.1021/bc980148w. [DOI] [PubMed] [Google Scholar]

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