AAPS PharmSciTech

, 20:23 | Cite as

Tunable Properties of Poly-DL-Lactide-Monomethoxypolyethylene Glycol Porous Microparticles for Sustained Release of Polyethylenimine-DNA Polyplexes

  • Treniece L. Terry
  • Brittany E. Givens
  • Victor G. J. Rodgers
  • Aliasger K. SalemEmail author
Research Article Theme: Translational Multi-Disciplinary Approach for the Drug and Gene Delivery Systems
Part of the following topical collections:
  1. Theme: Translational Multi-Disciplinary Approach for the Drug and Gene Delivery Systems


Direct pulmonary delivery is a promising step in developing effective gene therapies for respiratory disease. Gene therapies can be used to treat the root cause of diseases, rather than just the symptoms. However, developing effective therapies that do not cause toxicity and that successfully reach the target site at therapeutic levels is challenging. We have developed a polymer-DNA complex utilizing polyethylene imine (PEI) and DNA, which was then encapsulated into poly(lactic acid)-co-monomethoxy poly(ethylene glycol) (PLA-mPEG) microparticles via double emulsion, solvent evaporation. Then, the resultant particle size, porosity, and encapsulation efficiency were measured as a function of altering preparation parameters. Microsphere formation was confirmed from scanning electron micrographs and the aerodynamic particle diameter was measured using an aerodynamic particle sizer. Several formulations produced particles with aerodynamic diameters in the 0–5 μm range despite having larger particle diameters which is indicative of porous particles. Furthermore, these aerodynamic diameters correspond to high deposition within the airways when inhaled and the measured DNA content indicated high encapsulation efficiency. Thus, this formulation provides promise for developing inhalable gene therapies.


PLA PEG polyplexes microspheres 



The authors would like to acknowledge Terra M. Kruger for help with the NMR spectra, and Prof. Kristan S. Worthington for assistance with the polymerization reaction. The SEM images were obtained using an SEM instrument in the Central Microscopy Research Facilities at The University of Iowa.


A.K.S acknowledges support from the NIH P30 CA086862 grant and the Lyle and Sharon Bighley Chair of Pharmaceutical Sciences. T.L.T. acknowledges support from the Department of Education GAANN Fellowship program. B.E.G. acknowledges funding support from the Alfred P. Sloan Foundation, the University of Iowa Graduate College, and the National GEM Consortium. V. G. J. R. acknowledges support from the Jacques S. Yeager, Sr. endowment.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12249_2018_1215_MOESM1_ESM.docx (779 kb)
ESM 1 (DOCX 778 kb)


  1. 1.
    Jacobson SG, Cideciyan AV, Roman AJ, Sumaroka A, Schwartz SB, Heon E, et al. Improvement and decline in vision with gene therapy in childhood blindness. N Engl J Med. 2015;372(20):1920–6.PubMedPubMedCentralGoogle Scholar
  2. 2.
    Finotti A, Breda L, Lederer CW, Bianchi N, Zuccato C, Kleanthous M, et al. Recent trends in the gene therapy of beta-thalassemia. J Blood Med. 2015;6:69–85.PubMedPubMedCentralGoogle Scholar
  3. 3.
    Villate-Beitia I, Zarate J, Puras G, Pedraz JL. Gene delivery to the lungs: pulmonary gene therapy for cystic fibrosis. Drug Dev Ind Pharm. 2017;43(7):1071–81.PubMedGoogle Scholar
  4. 4.
    Griesenbach U, Pytel KM, Alton EW. Cystic fibrosis gene therapy in the UK and elsewhere. Hum Gene Ther. 2015;26(5):266–75.PubMedPubMedCentralGoogle Scholar
  5. 5.
    Sabatel C, Radermecker C, Fievez L, Paulissen G, Chakarov S, Fernandes C, et al. Exposure to bacterial CpG DNA protects from airway allergic inflammation by expanding regulatory lung interstitial macrophages. Immunity. 2017;46(3):457–73.PubMedGoogle Scholar
  6. 6.
    Olin JT, Wechsler ME. Asthma: pathogenesis and novel drugs for treatment. BMJ. 2014;349(7):g5517.PubMedGoogle Scholar
  7. 7.
    Gomes Dos Reis L, Svolos M, Hartwig B, Windhab N, Young PM, Traini D. Inhaled gene delivery: a formulation and delivery approach. Expert Opin Drug Deliv. 2017;14(3):319–30.PubMedGoogle Scholar
  8. 8.
    Shalash AO, Khalafallah NM, Molokhia AM, Elsayed MMA. The relationship between the permeability and the performance of carrier-based dry powder inhalation mixtures: new insights and practical guidance. AAPS PharmSciTech. 2018;19(2):912–22.PubMedGoogle Scholar
  9. 9.
    Sheth P, Stein SW, Myrdal PB. Factors influencing aerodynamic particle size distribution of suspension pressurized metered dose inhalers. AAPS PharmSciTech. 2015;16(1):192–201.PubMedGoogle Scholar
  10. 10.
    Singh B, Bandopadhyay S, Kapil R, Singh R, Katare O. Self-emulsifying drug delivery systems (SEDDS): formulation development, characterization, and applications. Crit Rev Ther Drug Carrier Syst. 2009;26(5):427–521.PubMedGoogle Scholar
  11. 11.
    Morales JO, Fathe KR, Brunaugh A, Ferrati S, Li S, Montenegro-Nicolini M, et al. Challenges and future prospects for the delivery of biologics: oral mucosal, pulmonary, and transdermal routes. AAPS J. 2017;19(3):652–68.PubMedGoogle Scholar
  12. 12.
    Liechty WB, Kryscio DR, Slaughter BV, Peppas NA. Polymers for drug delivery systems. Annu Rev Chem Biomol Eng. 2010;1:149–73.PubMedPubMedCentralGoogle Scholar
  13. 13.
    Kleemann E, Jekel N, Dailey LA, Roesler S, Fink L, Weissmann N, et al. Enhanced gene expression and reduced toxicity in mice using polyplexes of low-molecular-weight poly(ethylene imine) for pulmonary gene delivery. J Drug Target. 2009;17(8):638–51.PubMedGoogle Scholar
  14. 14.
    Ebeid K, Meng X, Thiel KW, Do AV, Geary SM, Morris AS, et al. Synthetically lethal nanoparticles for treatment of endometrial cancer. Nat Nanotechnol. 2018;13(1):72–81.PubMedGoogle Scholar
  15. 15.
    Do AV, Geary SM, Seol D, Tobias P, Carlsen D, Leelakanok N, et al. Combining ultrasound and intratumoral administration of doxorubicin-loaded microspheres to enhance tumor cell killing. Int J Pharm. 2018;539(1–2):139–46.PubMedGoogle Scholar
  16. 16.
    Wongrakpanich A, Morris AS, Geary SM, Joiner MA, Salem AK. Surface-modified particles loaded with CaMKII inhibitor protect cardiac cells against mitochondrial injury. Int J Pharm. 2017;520(1–2):275–83.PubMedPubMedCentralGoogle Scholar
  17. 17.
    Wafa EI, Geary SM, Goodman JT, Narasimhan B, Salem AK. The effect of polyanhydride chemistry in particle-based cancer vaccines on the magnitude of the anti-tumor immune response. Acta Biomater. 2017;50:417–27.PubMedPubMedCentralGoogle Scholar
  18. 18.
    Morris AS, Sebag SC, Paschke JD, Wongrakpanich A, Ebeid K, Anderson ME, et al. Cationic CaMKII inhibiting nanoparticles prevent allergic asthma. Mol Pharm. 2017;14(6):2166–75.PubMedPubMedCentralGoogle Scholar
  19. 19.
    Ahmed KK, Geary SM, Salem AK. Surface engineering tumor cells with adjuvant-loaded particles for use as cancer vaccines. J Control Release. 2017;248:1–9.PubMedPubMedCentralGoogle Scholar
  20. 20.
    Uchida S, Itaka K, Chen Q, Osada K, Ishii T, Shibata MA, et al. PEGylated polyplex with optimized PEG shielding enhances gene introduction in lungs by minimizing inflammatory responses. Mol Ther. 2012;20(6):1196–203.PubMedPubMedCentralGoogle Scholar
  21. 21.
    Laube BL. The expanding role of aerosols in systemic drug delivery, gene therapy, and vaccination. Respir Care. 2005;50(9):1161–76.PubMedGoogle Scholar
  22. 22.
    Al-Dosari MS, Gao X. Nonviral gene delivery: principle, limitations, and recent progress. AAPS J. 2009;11(4):671–81.PubMedPubMedCentralGoogle Scholar
  23. 23.
    Intra J, Salem AK. Fabrication, characterization and in vitro evaluation of poly(D,L-lactide-co-glycolide) microparticles loaded with polyamidoamine-plasmid DNA dendriplexes for applications in nonviral gene delivery. J Pharm Sci. 2010;99(1):368–84.PubMedPubMedCentralGoogle Scholar
  24. 24.
    Abbas AO, Donovan MD, Salem AK. Formulating poly(lactide-co-glycolide) particles for plasmid DNA delivery. J Pharm Sci. 2008;97(7):2448–61.PubMedGoogle Scholar
  25. 25.
    Zhang XQ, Intra J, Salem AK. Conjugation of polyamidoamine dendrimers on biodegradable microparticles for nonviral gene delivery. Bioconjug Chem. 2007;18(6):2068–76.PubMedGoogle Scholar
  26. 26.
    Zhang XQ, Dahle CE, Baman NK, Rich N, Weiner GJ, Salem AK. Potent antigen-specific immune responses stimulated by codelivery of CpG ODN and antigens in degradable microparticles. J Immunother. 2007;30(5):469–78.PubMedGoogle Scholar
  27. 27.
    Intra J, Salem AK. Characterization of the transgene expression generated by branched and linear polyethylenimine-plasmid DNA nanoparticles in vitro and after intraperitoneal injection in vivo. J Control Release. 2008;130(2):129–38.PubMedPubMedCentralGoogle Scholar
  28. 28.
    Kim N, Jiang D, Jacobi AM, Lennox KA, Rose SD, Behlke MA, et al. Synthesis and characterization of mannosylated pegylated polyethylenimine as a carrier for siRNA. Int J Pharm. 2012;427(1):123–33.PubMedGoogle Scholar
  29. 29.
    Jiang D, Salem AK. Optimized dextran-polyethylenimine conjugates are efficient non-viral vectors with reduced cytotoxicity when used in serum containing environments. Int J Pharm. 2012;427(1):71–9.PubMedGoogle Scholar
  30. 30.
    Edwards DA, Hanes J, Caponetti G, Hrkach J, Ben-Jebria A, Eskew ML, et al. Large porous particles for pulmonary drug delivery. Science. 1997;276(5320):1868–71.PubMedGoogle Scholar
  31. 31.
    Klassen CD, Watkins JBI. Casarett & Doull’s essentials of toxicology. 2nd ed: McGraw-Hill Education / Medical; 2015. 528 pGoogle Scholar
  32. 32.
    Kamada Ltd. International study evaluating the safety and efficacy of inhaled, human, alpha-1 antitrypsin (AAT) in alpha-1 antitrypsin deficient patients with emphysema. 2014.Google Scholar
  33. 33.
    Duncan GA, Jung J, Hanes J, Suk JS. The mucus barrier to inhaled gene therapy. Mol Ther. 2016;24(12):2043–53.PubMedPubMedCentralGoogle Scholar
  34. 34.
    Kim N, Duncan GA, Hanes J, Suk JS. Barriers to inhaled gene therapy of obstructive lung diseases: a review. J Control Release. 2016;240:465–88.PubMedPubMedCentralGoogle Scholar
  35. 35.
    Abramoff MD, Magalhaes PJ, Ram SJ. Image processing with ImageJ. Biophoton Int. 2004;11(7):36–42.Google Scholar
  36. 36.
    Singh NA, Mandal AKA, Khan ZA. Fabrication of PLA-PEG nanoparticles as delivery systems for improved stability and controlled release of catechin. J Nanomater. 2017;2017:1–9.Google Scholar
  37. 37.
    Sakhalkar HS, Dalal MK, Salem AK, Ansari R, Fu J, Kiani MF, et al. Leukocyte-inspired biodegradable particles that selectively and avidly adhere to inflamed endothelium in vitro and in vivo. Proc Natl Acad Sci U S A. 2003;100(26):15895–900.PubMedPubMedCentralGoogle Scholar
  38. 38.
    Sinclair J, Salem AK. Rapid localized cell trapping on biodegradable polymers using cell surface derivatization and microfluidic networking. Biomaterials. 2006;27(9):2090–4.PubMedGoogle Scholar
  39. 39.
    Jain A, Kunduru KR, Basu A, Mizrahi B, Domb AJ, Khan W. Injectable formulations of poly(lactic acid) and its copolymers in clinical use. Adv Drug Deliv Rev. 2016;107:213–27.PubMedGoogle Scholar
  40. 40.
    Kulkarni RK, Moore EG, Hegyeli AF, Leonard F. Biodegradable poly(lactic acid) polymers. J Biomed Mater Res. 1971;5(3):169–81.PubMedGoogle Scholar
  41. 41.
    Woodland JH, Yolles S, Blake DA, Helrich M, Meyer FJ. Long-acting delivery systems for narcotic antagonists. 1. J Med Chem. 1973;16(8):897–901.PubMedGoogle Scholar
  42. 42.
    Yolles S, Leafe TD, Woodland JH, Meyer FJ. Long acting delivery systems for narcotic antagonists II: release rates of naltrexone from poly(lactic acid) composites. J Pharm Sci. 1975;64(2):348–9.PubMedGoogle Scholar
  43. 43.
    Vila A, Sanchez A, Evora C, Soriano I, McCallion O, Alonso MJ. PLA-PEG particles as nasal protein carriers: the influence of the particle size. Int J Pharm. 2005;292(1–2):43–52.PubMedGoogle Scholar
  44. 44.
    Lu S, Morris VB, Labhasetwar V. Codelivery of DNA and siRNA via arginine-rich PEI-based polyplexes. Mol Pharm. 2015;12(2):621–9.PubMedGoogle Scholar
  45. 45.
    Zhu KJ, Lin XZ, Yang SL. Preparation, characterization, and properties of polylactide (Pla) poly(ethylene glycol) (peg) copolymers - a potential-drug carrier. J Appl Polym Sci. 1990;39(1):1–9.Google Scholar
  46. 46.
    Dechy-Cabaret O, Martin-Vaca B, Bourissou D. Controlled ring-opening polymerization of lactide and glycolide. Chem Rev. 2004;104(12):6147–76.PubMedGoogle Scholar
  47. 47.
    Kricheldorf HR, Sumbel MV, Kreiser-Saunders I. Polylactones. 20. Polymerization of iε-caprolactone with tributyltin derivatives: a mechanistic study. Macromolecules. 1991;24(8):1944–9.Google Scholar
  48. 48.
    Kricheldorf HR, Kreiser-Saunders I, Stricker A. Polylactones 48. SnOct2-initiated polymerizations of lactide: a mechanistic study. Macromolecules. 2000;33(3):702–9.Google Scholar
  49. 49.
    Beletsi A, Leontiadis L, Klepetsanis P, Ithakissios DS, Avgoustakis K. Effect of preparative variables on the properties of poly(dl-lactide-co-glycolide)-methoxypoly(ethyleneglycol) copolymers related to their application in controlled drug delivery. Int J Pharm. 1999;182(2):187–97.PubMedGoogle Scholar
  50. 50.
    Zheng X, Kan B, Gou M, Fu S, Zhang J, Men K, et al. Preparation of MPEG-PLA nanoparticle for honokiol delivery in vitro. Int J Pharm. 2010;386(1–2):262–7.PubMedGoogle Scholar
  51. 51.
    Alibolandi M, Sadeghi F, Sazmand SH, Shahrokhi SM, Seifi M, Hadizadeh F. Synthesis and self-assembly of biodegradable polyethylene glycol-poly (lactic acid) diblock copolymers as polymersomes for preparation of sustained release system of doxorubicin. Int J Pharm Investig. 2015;5(3):134–41.PubMedPubMedCentralGoogle Scholar
  52. 52.
    Luo WJ, Li SM, Bei JZ, Wang SG. Synthesis and characterization of poly(L-lactide)- poly(ethylene glycol) multiblock copolymers. J Appl Polym Sci. 2002;84(9):1729–36.Google Scholar
  53. 53.
    Vert M. Degradable polymers in medicine: updating strategies and terminology. Int J Artif Organs. 2011;34(2):76–83.PubMedGoogle Scholar
  54. 54.
    Salem AK, Weiner GJ. CpG oligonucleotides as immunotherapeutic adjuvants: innovative applications and delivery strategies. Adv Drug Deliv Rev. 2009;61(3):193–4.PubMedGoogle Scholar
  55. 55.
    Krishnamachari Y, Salem AK. Innovative strategies for co-delivering antigens and CpG oligonucleotides. Adv Drug Deliv Rev. 2009;61(3):205–17.PubMedPubMedCentralGoogle Scholar
  56. 56.
    Joshi VB, Adamcakova-Dodd A, Jing X, Wongrakpanich A, Gibson-Corley KN, Thorne PS, et al. Development of a poly (lactic-co-glycolic acid) particle vaccine to protect against house dust mite induced allergy. AAPS J. 2014;16(5):975–85.PubMedPubMedCentralGoogle Scholar
  57. 57.
    Makkouk A, Joshi VB, Wongrakpanich A, Lemke CD, Gross BP, Salem AK, et al. Biodegradable microparticles loaded with doxorubicin and CpG ODN for in situ immunization against cancer. AAPS J. 2015;17(1):184–93.PubMedGoogle Scholar
  58. 58.
    Hsu YY, Hao T, Hedley ML. Comparison of process parameters for microencapsulation of plasmid DNA in poly(D,L-lactic-co-glycolic) acid microspheres. J Drug Target. 1999;7(4):313–23.PubMedGoogle Scholar
  59. 59.
    Bivas-Benita M, Romeijn S, Junginger HE, Borchard G. PLGA-PEI nanoparticles for gene delivery to pulmonary epithelium. Eur J Pharm Biopharm. 2004;58(1):1–6.PubMedGoogle Scholar
  60. 60.
    Zhang XQ, Intra J, Salem AK. Comparative study of poly (lactic-co-glycolic acid)-poly ethyleneimine-plasmid DNA microparticles prepared using double emulsion methods. J Microencapsul. 2008;25(1):1–12.PubMedGoogle Scholar
  61. 61.
    Bailey MM, Berkland CJ. Nanoparticle formulations in pulmonary drug delivery. Med Res Rev. 2009;29(1):196–212.PubMedGoogle Scholar
  62. 62.
    Biddiscombe MF, Usmani OS. Inhaler characteristics in asthma. EU Respir Pulm Dis. 2017;3:32–7.Google Scholar
  63. 63.
    Huang YY, Chung TW, Tzeng TW. Drug release from PLA/PEG microparticulates. Int J Pharm. 1997;156(1):9–15.Google Scholar

Copyright information

© American Association of Pharmaceutical Scientists 2019

Authors and Affiliations

  1. 1.Department of Pharmaceutical Sciences and Experimental Therapeutics, College of PharmacyUniversity of IowaIowa CityUSA
  2. 2.Department of Chemical and Biochemical Engineering, College of EngineeringUniversity of IowaIowa CityUSA
  3. 3.Department of BioengineeringUniversity of CaliforniaRiversideUSA

Personalised recommendations