Critical size limit of biodegradable nanoparticles for enhanced lymph node trafficking and paracortex penetration


Lymph node (LN) targeting through interstitial drainage of nanoparticles (NPs) is an attractive strategy to stimulate a potent immune response, as LNs are the primary site for lymphocyte priming by antigen presenting cells (APCs) and triggering of an adaptive immune response. NP size has been shown to influence the efficiency of LN-targeting and retention after subcutaneous injection. For clinical translation, biodegradable NPs are preferred as carrier for vaccine delivery. However, the selective “size gate” for effective LN-drainage, particularly the kinetics of LN trafficking, is less well defined. This is partly due to the challenge in generating size-controlled NPs from biodegradable polymers in the sub-100-nm range. Here, we report the preparation of three sets of poly(lactic-co-glycolic)-b-poly(ethylene-glycol) (PLGA-b-PEG) NPs with number average diameters of 20-, 40-, and 100-nm and narrow size distributions using flash nanoprecipitation. Using NPs labeled with a near-infrared dye, we showed that 20-nm NPs drain rapidly across proximal and distal LNs following subcutaneous inoculation in mice and are retained in LNs more effectively than NPs with a number average diameter of 40-nm. The drainage of 100-nm NPs was negligible. Furthermore, the 20-nm NPs showed the highest degree of penetration around the paracortex region and had enhanced access to dendritic cells in the LNs. Together, these data confirmed that small, size-controlled PLGA-b-PEG NPs at the lower threshold of about 30-nm are most effective for LN trafficking, retention, and APC uptake after s.c. administration. This report could inform the design of LN-targeted NP carrier for the delivery of therapeutic or prophylactic vaccines.

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  1. [1]

    Trevaskis, N. L.; Kaminskas, L. M.; Porter, C. J. H. From sewer to saviour-targeting the lymphatic system to promote drug exposure and activity. Nat. Rev. Drug Discov. 2015, 14, 781–803.

    Article  Google Scholar 

  2. [2]

    Willard-Mack, C. L. Normal structure, function, and histology of lymph nodes. Toxicol. Pathol. 2006, 34, 409–424.

    Article  Google Scholar 

  3. [3]

    Wilson, N. S.; El-Sukkari, D.; Belz, G. T.; Smith, C. M.; Steptoe, R. J.; Heath, W. R.; Shortman, K.; Villadangos, J. A. Most lymphoid organ dendritic cell types are phenotypically and functionally immature. Blood 2003, 102, 2187–2194.

    Article  Google Scholar 

  4. [4]

    Swartz, M. A.; Hubbell, J. A.; Reddy, S. T. Lymphatic drainage function and its immunological implications: From dendritic cell homing to vaccine design. Semin. Immunol. 2008, 20, 147–156.

    Article  Google Scholar 

  5. [5]

    Tostanoski, L. H.; Chiu, Y.-C.; Gammon, J. M.; Simon, T.; Andorko, J. I.; Bromberg, J. S.; Jewell, C. M. Reprogramming the local lymph node microenvironment promotes tolerance that is systemic and antigen specific. Cell Rep. 2016, 16, 2940–2952.

    Article  Google Scholar 

  6. [6]

    Supsersaxo, A.; Hein, W. R.; Steffen, H. Effect of molecular weight on the lymphatic absorption of water-soluble compounds following subcutaneous administration. Pharm. Res. 1990, 7, 167–169.

    Article  Google Scholar 

  7. [7]

    Oussoren, C.; Storm, G. Liposomes to target the lymphatics by subcutaneous administration. Adv. Drug Deliv. Rev. 2001, 50, 143–156.

    Article  Google Scholar 

  8. [8]

    Kaminskas, L. M.; Porter, C. J. H. Targeting the lymphatics using dendritic polymers (dendrimers). Adv. Drug Deliv. Rev. 2011, 63, 890–900.

    Article  Google Scholar 

  9. [9]

    Reddy, S. T.; Rehor, A.; Schmoekel, H. G.; Hubbell, J. A.; Swartz, M. A. In vivo targeting of dendritic cells in lymph nodes with poly(propylene sulfide) nanoparticles. J. Control. Release 2006, 112, 26–34.

    Article  Google Scholar 

  10. [10]

    Reddy, S. T.; van der Vlies, A. J.; Simeoni, E.; Angeli, V.; Randolph, G. J.; O’Neil, C. P.; Lee, L. K.; Swartz, M. A.; Hubbell, J. A. Exploiting lymphatic transport and complement activation in nanoparticle vaccines. Nat. Biotechnol. 2007, 25, 1159–1164.

    Article  Google Scholar 

  11. [11]

    Mottram, P. L.; Leong, D.; Crimeen-Irwin, B.; Gloster, S.; Xiang, S. D.; Meanger, J.; Ghildyal, R.; Vardaxis, N.; Plebanski, M. Type 1 and 2 immunity following vaccination is influenced by nanoparticle size: Formulation of a model vaccine for respiratory syncytial virus. Mol. Pharm. 2007, 4, 73–84.

    Article  Google Scholar 

  12. [12]

    Fifis, T.; Gamvrellis, A.; Crimeen-Irwin, B.; Pietersz, G. A.; Li, J.; Mottram, P. L.; McKenzie, I. F.; Plebanski, M. Size-dependent immunogenicity: Therapeutic and protective properties of nano-vaccines against tumors. J. Immunol. 2004, 173, 3148–3154.

    Article  Google Scholar 

  13. [13]

    Kumar, S.; Anselmo, A. C.; Banerjee, A.; Zakrewsky, M.; Mitragotri, S. Shape and size-dependent immune response to antigen-carrying nanoparticles. J. Control. Release 2015, 220, 141–148.

    Article  Google Scholar 

  14. [14]

    Manolova, V.; Flace, A.; Bauer, M.; Schwarz, K.; Saudan, P.; Bachmann, M. F. Nanoparticles target distinct dendritic cell populations according to their size. Eur. J. Immunol. 2008, 38, 1404–1413.

    Article  Google Scholar 

  15. [15]

    Randolph, G. J.; Angeli, V.; Swartz, M. A. Dendritic-cell trafficking to lymph nodes through lymphatic vessels. Nat. Rev. Immunol. 2005, 5, 617–628.

    Article  Google Scholar 

  16. [16]

    Li, X. R.; Sloat, B. R.; Yanasarn, N.; Cui, Z. R. Relationship between the size of nanoparticles and their adjuvant activity: Data from a study with an improved experimental design. Eur. J. Pharm. Biopharm. 2011, 78, 107–116.

    Article  Google Scholar 

  17. [17]

    Chaney, E. J.; Tang, L.; Tong, R.; Cheng, J. J.; Boppart, S. A. Lymphatic biodistribution of polylactide nanoparticles. Mol. Imaging 2010, 9, 153–162.

    Article  Google Scholar 

  18. [18]

    Rao, D. A.; Forrest, M. L.; Alani, A. W. G.; Kwon, G. S.; Robinson, J. R. Biodegradable PLGA based nanoparticles for sustained regional lymphatic drug delivery. J. Pharm. Sci. 2010, 99, 2018–2031.

    Article  Google Scholar 

  19. [19]

    Zheng, S. S.; Qin, T.; Lu, Y.; Huang, Y. F.; Luo, L.; Liu, Z. G.; Bo, R. N.; Hu, Y. L.; Liu, J. G.; Wang, D. Y. Maturation of dendritic cells in vitro and immunological enhancement of mice in vivo by pachyman-and/or OVA-encapsulated poly(D, L-lactic acid) nanospheres. Int. J. Nanomedicine 2018, 13, 569–583.

    Article  Google Scholar 

  20. [20]

    Zhang, W. F.; Wang, L. Y.; Liu, Y.; Chen, X. M.; Liu, Q.; Jia, J. L.; Yang, T. Y.; Qiu, S. H.; Ma, G. H. Immune responses to vaccines involving a combined antigen-nanoparticle mixture and nanoparticle-encapsulated antigen formulation. Biomaterials 2014, 35, 6086–6097.

    Article  Google Scholar 

  21. [21]

    Maldonado, R. A.; LaMothe, R. A.; Ferrari, J. D.; Zhang, A. H.; Rossi, R. J.; Kolte, P. N.; Griset, A. P.; O’Neil, C.; Altreuter, D. H.; Browning, E. et al. Polymeric synthetic nanoparticles for the induction of antigen-specific immunological tolerance. Proc. Natl. Acad. Sci. USA 2015, 112, E156–E165.

    Article  Google Scholar 

  22. [22]

    Johnson, B. K.; Prud’homme, R. K. Mechanism for rapid self-assembly of block copolymer nanoparticles. Phys. Rev. Lett. 2003, 91, 118302.

    Article  Google Scholar 

  23. [23]

    Saad, W. S.; Prud’homme, R. K. Principles of nanoparticle formation by flash nanoprecipitation. Nano Today 2016, 11, 212–227.

    Article  Google Scholar 

  24. [24]

    Xiang, S. D.; Scholzen, A.; Minigo, G.; David, C.; Apostolopoulos, V.; Mottram, P. L.; Plebanski, M. Pathogen recognition and development of particulate vaccines: Does size matter?. Methods 2006, 40, 1–9.

    Article  Google Scholar 

  25. [25]

    Harrell, M. I.; Iritani, B. M.; Ruddell, A. Lymph node mapping in the mouse. J. Immunol. Methods 2008, 332, 170–174.

    Article  Google Scholar 

  26. [26]

    Reddy, S. T.; Berk, D. A.; Jain, R. K.; Swartz, M. A. A sensitive in vivo model for quantifying interstitial convective transport of injected macromolecules and nanoparticles. J. Appl. Physiol. 2006, 101, 1162–1169.

    Article  Google Scholar 

  27. [27]

    Tomura, M.; Hata, A.; Matsuoka, S.; Shand, F. H. W.; Nakanishi, Y.; Ikebuchi, R.; Ueha, S.; Tsutsui, H.; Inaba, K.; Matsushima, K. et al. Tracking and quantification of dendritic cell migration and antigen trafficking between the skin and lymph nodes. Sci. Rep. 2014, 4, 6030.

    Article  Google Scholar 

  28. [28]

    Thomas, S. N.; Schudel, A. Overcoming transport barriers for interstitial-, lymphatic-, and lymph node-targeted drug delivery. Curr. Opin. Chem. Eng. 2015, 7, 65–74.

    Article  Google Scholar 

  29. [29]

    Sixt, M.; Kanazawa, N.; Selg, M.; Samson, T.; Roos, G.; Reinhardt, D. P.; Pabst, R.; Lutz, M. B.; Sorokin, L. The conduit system transports soluble antigens from the afferent lymph to resident dendritic cells in the T cell area of the lymph node. Immunity 2005, 22, 19–29.

    Article  Google Scholar 

  30. [30]

    Roozendaal, R.; Mempel, T. R.; Pitcher, L. A.; Gonzalez, S. F.; Verschoor, A.; Mebius, R. E.; von Andrian, U. H.; Carroll, M. C. Conduits mediate transport of low-molecular-weight antigen to lymph node follicles. Immunity 2009, 30, 264–276.

    Article  Google Scholar 

  31. [31]

    Phan, T. G.; Grigorova, I.; Okada, T.; Cyster, J. G. Subcapsular encounter and complement-dependent transport of immune complexes by lymph node B cells. Nat. Immunol. 2007, 8, 992–1000.

    Article  Google Scholar 

  32. [32]

    Kang, S.; Ahn, S.; Lee, J.; Kim, J. Y.; Choi, M.; Gujrati, V.; Kim, H.; Kim, J.; Shin, E. C.; Jon, S. Effects of gold nanoparticle-based vaccine size on lymph node delivery and cytotoxic T-lymphocyte responses. J. Control. Release 2017, 256, 56–67.

    Article  Google Scholar 

  33. [33]

    Kim, H.; Uto, T.; Akagi, T.; Baba, M.; Akashi, M. Amphiphilic poly(amino acid) nanoparticles induce size-dependent dendritic cell maturation. Adv. Funct. Mater. 2010, 20, 3925–3931.

    Article  Google Scholar 

  34. [34]

    Hirosue, S.; Kourtis, I. C.; van der Vlies, A. J.; Hubbell, J. A.; Swartz, M. A. Antigen delivery to dendritic cells by poly(propylene sulfide) nanoparticles with disulfide conjugated peptides: Cross-presentation and T cell activation. Vaccine 2010, 28, 7897–7906.

    Article  Google Scholar 

  35. [35]

    Jiang, H.; Wang, Q.; Sun, X. Lymph node targeting strategies to improve vaccination efficacy. J. Control. Release 2017, 267, 47–56.

    Article  Google Scholar 

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This work was funded by support from the National Institutes of Health (Nos. R01-AI114609 and T32-OD11089) and NSF GRFP (No. DGE1746891). Partial support was received from the University of Florida Emerging Pathogens Institute.

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Correspondence to Rhoel R. Dinglasan or Hai-Quan Mao.

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Howard, G.P., Verma, G., Ke, X. et al. Critical size limit of biodegradable nanoparticles for enhanced lymph node trafficking and paracortex penetration. Nano Res. 12, 837–844 (2019).

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  • biodegradable nanoparticle
  • lymph node trafficking
  • vaccine delivery
  • nanoparticle size
  • antigen presenting cells
  • in vivo imaging