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. 2015 Apr 3;11(3):650–656. doi: 10.1080/21645515.2015.1009345

Biodegradable polymeric microsphere-based vaccines and their applications in infectious diseases

Chi-Ying Lin 1, Shih-Jie Lin 1, Yi-Chen Yang 1, Der-Yuan Wang 1, Hwei-Fang Cheng 1, Ming-Kung Yeh 1,2,*
PMCID: PMC4514183  PMID: 25839217

Abstract

Vaccination, which provides effective, safe infectious disease protection, is among the most important recent public health and immunological achievements. However, infectious disease remains the leading cause of death in developing countries because several vaccines require repeated administrations and children are often incompletely immunized. Microsphere-based systems, providing controlled release delivery, can obviate the need for repeat immunizations. Here, we review the function of sustained and pulsatile release of biodegradable polymeric microspheres in parenteral and mucosal single-dose vaccine administration. We also review the active-targeting function of polymeric particles. With their shield and co-delivery functions, polymeric particles are applied to develop single-dose and mucosally administered vaccines as well as to improve subunit vaccines. Because polymeric particles are easily surface-modified, they have been recently used in vaccine development for cancers and many infectious diseases without effective vaccines (e.g., human immunodeficiency virus infection). These polymeric particle functions yield important vaccine carriers and multiple benefits.

Keywords: biodegradable, infectious diseases, immunization, polymeric microspheres, vaccines

Abbreviations and acronyms

APC

antigen-presenting cell

DEN-1–DEN-4

dengue virus serotypes 1–4

DC

dendritic cell

DT

diphtheria toxoid

DT or TD

diphtheria + tetanus vaccine

DTP

diphtheria + tetanus + pertussis vaccine

NS1

nonstructural protein 1

PEG

poly (ethylene glycol)

PLA

poly (lactide)

PLGA

Poly (lactic-co-glycolic acid)

TT

tetanus-toxoid

VC

Vibrio cholera

WHO

World Health Organization

Introduction

Infectious diseases, caused by pathogenic microorganisms, are among leading global health problems. Millions of people die of infectious diseases annually. One approach toward preventing infectious diseases is vaccination, which helps an individual develop resistance to an infectious disease.

The first vaccine targeted smallpox and was developed by Edward Jenner in 1796. Jenner initially inoculated a boy with cowpox virus and later reinoculated the boy again with smallpox virus after the boy had suffered from cowpox. The boy exhibited no symptoms of smallpox after the second inoculation. Thus, Jenner concluded that inoculation with the cowpox virus had protected the boy from smallpox. In 1980, after the achievement of global smallpox vaccination, the World Health Assembly endorsed the worldwide eradication of smallpox.

The earliest vaccines, such as smallpox vaccines, comprised inactivated or live attenuated viruses or bacteria. Bacterial toxoids, virus-like particles, and purified viral proteins and their subunits were introduced as knowledge about pathogens increased. These toxoid and subunit vaccines are considered safer and are not infectious.1

Today, several vaccines against infectious diseases are recommended for children. The recommended routine immunization schedule for children in Taiwan is listed in Table 1. Although vaccine immunization has succeeded in controlling several vaccine-preventable diseases in the developed countries, many children in developing countries continue to contract such diseases.2 A complete immunization schedule for infants and children typically includes repeated vaccine administrations over the course of several years.3 However, many rural children in developing countries have poor living conditions and medical care and therefore do not receive complete immunization.4 To improve patient accessibility and maintain long-lasting protection, single-dose vaccines that mimic repeated injections administered via vaccination schedules may provide a promising solution.5-10

Table 1.

Immunization schedule for children in Taiwan

Immunization schedule Vaccine <24 h after birth ≥24 h after birth 1 month 2 months 4 months 6 months 12 months 15 months 18 months 27 months >5 years old
Hepatitis B First dose Boost 1 Boost 2
BCG (Bacillus Calmette-Guérin) One dose
DTaP–Hib–IPV* First dose Boost 1 Boost 2 Boost 3
Varicella One dose
MMR (Measles, mumps and rubella) First dose Boost 1
JE (Japanese encephalitis) First dose Boost 1** Boost 2 Boost 3
Influenza First dose and boost 1 with an interval of 1 month and one boost every year after boost 1
Hepatitis A First dose Boost 1
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*DTaP-Hib-IPV: Diphtheria, Tetanus, acellular Pertussis, Haemophilus influenzae type b conjugate and poliovirus vaccine.

**An interval of 2 weeks between first dose and boost 1.

To mimic repeated injections of conventional vaccinations, single-dose vaccines have been used to release entrapped antigens over periods lasting weeks or months.10 Several controlled release technology materials, including liposomes, polymers, and virus-like particles, have been tested to improve the efficacy of conventional vaccination.6,11 The different types of polymers that can be used in controlled release delivery systems to encapsulate antigens and thus protect and control antigen release are listed in Table 2.12-14

Table 2.

Polymer microsphere materials used in microencapsulation

Types Materials
Natural Polymers Carbohydrates Agarose
Alginate
Chitosan
Polydextran
Polystarch
Starch
Proteins Albumin
Collagen
Gelatin
Others Calcium carbonate
Lipids
Tricalcium phosphate
Synthetic Polymers Non biodegradable Acrolein
Glycidyl methacrylate
Lactides
Polyanhydride
Polymethylmetharylate
Polyiminocarbonates
Biodegradable Glycolides
Epoxy polymers
Hydrogels
Paraffin
Pegylated poly(lactide)
Poly(lactide-co-glycolide)
Polyacrylates
Polyacrylonitrile
Polyamide
Polyamino acids
Polycaprolactones
Polyelectrolytes
Polyester
Polyethylene glycol
Polyphosphazenes
Polyurea
Polyurethane
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Compared with conventional vaccines, a benefit of particle-based controlled release systems is the ability to simultaneously co-deliver antigens and adjuvants to the same antigen-presenting cells (APCs). Another advantage is the ability to protect antigens and adjuvants from degradation before reaching the target cells. Other advantages include site-directed delivery and the ability to induce cell-mediated immune responses.15 Further, of the different materials used in particle-based controlled release systems, liposome delivery systems are less stable than polymer particle systems.16 Although virus-like particles are stable as polymeric particles, they introduced the issue of capsid component immunogenicity.17

Biodegradable polymers are superior to non-degradable polymers because the later may require additional removal procedures. For reliability and reproducibility, synthetic biodegradable polymers are the best choice for antigen encapsulation in single-dose vaccine production. Poly (lactic-co-glycolic acid) (PLGA) is among the most widely used synthetic biodegradable polymers.18 Biodegradable PLGA microspheres have been widely used because of their safety and ability to provide long-term controlled vaccine antigen release.5,19 The microspheres have not only been used for controlled release vaccines but also in delivery systems for other drugs, such as cancer therapies and birth control.20-22

To mimic repeated immunizations, 2 types of vaccine antigen release are possible with biodegradable PLGA microspheres: sustained release and pulsatile release. Sustained release, or continued vaccine antigen diffusion after the initial release, mimics the administration of several small boosters. Pulsatile release, or a second vaccine antigen diffusion distinct from the first release, mimics the current immunization schedule.6 In previous studies, microsphere-based vaccines were developed using sustained antigen releases; however, this method could not mimic clinical vaccine administration. Therefore, pulsatile antigen release was developed to improve the antigen release pattern.6,23

Here, we review the use of biodegradable polymeric microspheres for single-dose vaccines with parenteral and mucosal administrations. In addition to the function of sustained and pulsatile release of antigens, we also review the active-targeting function of polymeric particles. With their shield function and their ability to co-deliver antigens and adjuvants to the same target cells (e.g., dendritic cells), polymeric particles are applied to develop various vaccines. For examples, because they can prevent antigens from degradation and control release of antigens, they were applied to single-dose and mucosal administered vaccines. Furthermore, their ability to co-deliver antigens and adjuvants to the same target cells make them suitable to improve subunit vaccines, which are safer but sometimes fail to induce potent immune responses. Recently, polymeric particles have been used in the development of vaccines for cancers and many infectious diseases for which there are not currently effective vaccines (e.g., human immunodeficiency virus infection, malaria, and tuberculosis) because they are easy to be surface-modified. These functions of polymeric particles maintain their importance in the development of various vaccines and bring multiple benefits into vaccination.

Vaccines Encapsulated in Biodegradable Polymeric Microspheres—Parenteral Routes

To date, most vaccines are parenterally administered via intramuscular, subcutaneous, and intradermal injections. Several parenteral vaccines have been encapsulated in biodegradable polymeric microspheres, including the tetanus and diphtheria vaccine.

Tetanus vaccine

Tetanus, also known as lockjaw, is caused by the bacterium Clostridium tetani and is characterized by severe muscle spasms that initially occur in the jaw muscles. The administration of tetanus toxoid (TT) containing vaccines can prevent tetanus. Furthermore, a global initiative to eliminate neonatal tetanus was launched in 1989, and the World Health Organization (WHO) selected TT as the first single-dose vaccine to be administered via biodegradable polymeric microspheres.24,25

The sustained release of TT from biodegradable PLGA microspheres has been widely studied.19,26 A pattern of constant release with a decreasing release rate after the initial burst of TT has been identified. Small-sized TT-PLA microspheres with rapid release kinetics induced an earlier release compared with larger TT-PLGA 50:50 microspheres with slow release kinetics.26 A continuously increasing release rate after the initial burst was observed with low-molecular-weight TT-PLGA microspheres.19

A pulsatile release pattern that mimics the current vaccine regimen has also been investigated. The time between the first and second pulsed TT release is determined by the degradation rate of biodegradable polymers. The degradation rates of these polymers depend on the composition and molecular weight.27 Moreover, the time between the first and second TT release increases from 21 d to 52 d as the lactic acid ratio increases from 50:50 to 75:25 and the molecular weight increases from 0.33 dl/g to 0.80 dl/g. The pulsatile release pattern was achieved with a combination of different TT-biodegradable PLGA microspheres. A single administration of a combination of PLGA 50:50 and PLGA 75:25 microspheres yielded a pulsed release pattern with second and third releases after the initial release.28 The second release occurred between 3 and 5 weeks and the third release occurred between 9 and 11 weeks after the initial release. The antibody responses induced by single administrations of mixtures of TT-biodegradable PLGA microspheres with different particle sizes were similar to those obtained following 3 administrations of TT- aluminum.

In a mouse model, the in vivo induction of tetanus-specific antibodies following a single administration of TT-biodegradable PLGA microspheres with different compositions (PLGA 50:50 and PLGA 75:25) was compared with that following the conventional multiple administration of aluminum-adsorbed TT.29 The abilities of the TT-biodegradable PLGA microsphere combination (PLGA 50:50 and PLGA 75:25) and of aluminum-adsorbed TT to elicit antibody titers were similar. In addition, protection against a subcutaneous TT challenge following immunization with this TT-biodegradable PLGA microsphere combination (PLGA 50:50 and PLGA 75:25) or aluminum-adsorbed TT was compared in this same mouse model. Both preparations protected the preimmunized mice from a TT challenge.29 These results demonstrated that a single administration of TT-biodegradable PLGA microspheres provided similar protective immunity against TT as did conventionally administered aluminum-adsorbed TT. A comparison between a single dose of TT-biodegradable PLGA microspheres and multiple doses of conventional aluminum-adsorbed TT was also studied in a rat model.30 The antibody responses induced by a single dose of TT-biodegradable PLGA microspheres were similar to those induced by multiple doses of aluminum-adsorbed TT.30 These results revealed that antigen encapsulation by microspheres reduced the number of required vaccinations while yielding a performance similar to that of conventional vaccines administered in multiple shots.

Diphtheria vaccine

Diphtheria is caused by the bacterium Corynebacterium diphtheriae and is characterized by the presence of pseudomembranes (adherent membranes) in the upper respiratory tract. Diphtheria can be prevented by the administration of diphtheria vaccines, which are based on diphtheria toxoid (DT). Diphtheria vaccines are usually combined with tetanus and pertussis vaccines to yield combination vaccines against diphtheria, tetanus, and pertussis (DTP vaccines). Combination vaccines against diphtheria and tetanus are also available as DT or TD vaccines.

Studies of the in vitro release and in vivo induction of DT-specific antibodies following a single administration of DT-biodegradable PLGA microspheres have been reported.31 Sustained DT-specific antibodies were elicited in guinea pigs immunized with DT-biodegradable PLGA 50:50 microspheres. These antibody responses were comparable with those elicited by DT plus aluminum adjuvant.

Combination vaccines based on biodegradable PLGA microspheres have also been studied for tetanus and diphtheria.32 Guinea pigs were immunized with a single subcutaneous injection of a combination of TT-biodegradable PLGA microspheres and DT-biodegradable PLGA microspheres. The specific antibody titers following that immunization were comparable with those obtained from guinea pigs immunized with the licensed divalent vaccine. The protective immunity provided by immunization from the combined of TT- and DT-biodegradable PLGA microspheres was comparable with that induced by the licensed vaccine. Guinea pigs preimmunized with the combined of TT- and DT-biodegradable PLGA microspheres were also protected from tetanus and diphtheria toxins challenges 6 weeks after immunization.32

Vaccines Encapsulated in Biodegradable Polymeric Microspheres—Mucosal Routes

Although mucosal administration routes have the advantage of being needle-free, vaccine antigens are easily degraded during delivery through a mucosal route. The protection of vaccine antigens delivered via mucosal administration may be provided by encapsulation in biodegradable polymer microspheres.33

Mucosal immunity provides major protection against pathogenic microorganisms. Mucosal surfaces include the respiratory, alimentary, and urogenital tracts. Pathogenic microorganisms caused several infectious diseases adhering to mucosal surfaces. The induction of mucosal immunization through respiratory tracts by pathogenic antigens promotes the secretion of antigen-specific antibodies in various human body fluids.34-38 This finding suggests that the mucosal route may be a good option for vaccine inoculation.39

Biodegradable polymeric microspheres are suitable for mucosal administration because the spheres can prevent antigens from low pH, bile salts, and digestive enzymes present in the gastrointestinal tracts.40 PLGA and poly (lactide) (PLA) have been developed for vaccine delivery through the gastrointestinal tract and nasal cavity.41,42 The delivery routes are Peyer's patches in the intestinal cavity and mucosa-associated lymphoid tissues in the nasal cavity.43,44 The biodegradable polymeric microspheres have been used to encapsulate oral vaccines (e.g., cholera vaccine).

Cholera vaccine

Cholera, caused by the food- and water-borne bacterium Vibrio cholera (VC), is an acute intestinal infection. The main symptoms are copious, painless, watery diarrhea, and vomiting. Severe diarrhea and vomiting can lead to acute dehydration, and occasionally death. Cholera can be prevented by the administration of cholera vaccines.

In vivo induction of Vibrio-specific serum antibody titers following the oral immunization of VC-loaded PLG microspheres has been studied.45 Compared with the administration of a VC solution, significantly higher serum titers of Vibrio-specific immunoglobulin (Ig) G and IgM antibodies were elicited in mice immunized with VC-loaded microspheres. These VC-loaded microspheres were also prepared in different combinations: 50:50 PLG, 75:25 PLG, and PLA/poly (ethylene glycol) (PEG) blended microspheres.46 The serum titers of Vibrio-specific antibodies were examined in mice immunized with VC-loaded 50:50 PLG, 75:25 PLG, and PLA/PEG-blended microspheres. Higher antibody responses were elicited in mice immunized with VC-loaded 75:25 PLG microspheres, and the highest antibody responses were obtained in mice immunized with VC-loaded PLA/PEG-blended microspheres. The administration of VC-loaded PLA/PEG-blended microspheres protected preimmunized mice from a VC challenge with a survival rate of 92%.46

Vaccines Encapsulated in Biodegradable Polymeric Microspheres—Nonvaccine-Preventable Diseases

Vaccine-preventable diseases, such as tetanus and diphtheria, are infectious diseases, for which effective vaccines are available. In addition to these vaccine-preventable diseases, biodegradable polymeric microspheres have been studied in terms of encapsulation of antigens from diseases without effective vaccines.47,48

Dengue vaccine

Dengue, caused by dengue virus, is a mosquito-borne tropical disease. The main symptoms are mild or high fever, headache, muscle and joint pains, pain behind the eyes, and rash. No licensed dengue vaccine is currently available.

There are 4 distinct serotypes of the dengue virus, DEN-1–DEN-4. A study of the effectiveness of nonstructural protein 1 (NS1) protein-loaded microspheres against dengue 2 virus (DEN-2) has been reported.48 The NS1 protein of DEN-2 was encapsulated in PLGA/PEG microspheres. Strong antibody responses were elicited in mice immunized with these NS1 protein-loaded PLGA/PEG microspheres. In a dengue virus challenge test in mice, an increased survival was observed in mice immunized with NS1 protein-loaded PLGA/PEG microspheres compared with mice immunized with NS1 protein plus an aluminum adjuvant or PBS solution.48

Active-Targeting Polymeric System

Most vaccines are believed to block the spread of infection primarily through the induction of protective antibodies.49 Despite the importance of antibodies, in many infectious diseases for which no effective vaccines exist, such as human immunodeficiency virus infection, malaria, and tuberculosis, T-cell responses are believed to be required for protection.50-52 In addition, to induce both cellular and humoral immunity, dendritic cells (DCs), the APCs that initiate adaptive immunity, have become a key target of vaccine design.53

The first attempt to use DC as a vaccination target initiated from ex vivo antigen-loaded DC.54 In addition to cell-based immunotherapy involving in vitro-cultured, antigen-loaded DCs, another promising approach for designing DC-targeted vaccines is the selective targeting of DC-specific receptors by coupling the desired antigen to an antibody or ligand.55 Several studies have revealed that these direct-conjugate approaches efficiently induce antigen-specific CD4+ and CD8+ T-cell responses.56,57 However, direct antigen-antibody conjugation may alter the antigen conformation. Furthermore, besides antigens, additional stimulating signals, usually provided by adjuvants, are often required to induce an effective immune response, particularly for subunit vaccines.58 In the absence of an adjuvant, T cell tolerance may be induced instead of T-cell immunity.59 Therefore, particulate carriers such as polymeric particles have been studied because of their abilities to co-deliver antigens and adjuvants to target cells (e.g. DCs).60-62 In addition to protecting antigens and adjuvants from degradation, the selection of flexible combinations of target cells and adjuvants, which is important for inducing appropriate immune responses, is another advantage of the co-delivery of antigens and adjuvants by polymeric particles.

The direct incorporation of ligands, such as avidin–fatty acid conjugates, in the polymeric matrices of biodegradable PLGA polymeric particles has been reported.63 However, PLGA polymeric particles are often PEGylated by anchoring a layer of PEG chains to which the targeting ligands or antibodies are attached.64,65 The advantages of active-targeting polymeric particles are the protection of antigens from degradation; delivery of antigens to APCs, such as DCs, in a targeted and prolonged manner; prevention of antigen spread to the systemic circulation; co-delivery of antigens and adjuvants to DCs; lower required doses of antigens and adjuvants; and stability. However, antigen and adjuvant destabilization may occur during the preparation of polymeric particles.17

Conclusions

Biodegradable polymeric particles have been shown to be effective for the development of single-dose and mucosal vaccines and have been applied to the development of active-targeting delivery systems. These polymeric particles have been shown to release antigens in either a sustained or pulsatile pattern to provide long-lasting protection without repeated immunizations. The particles are also capable of delivering antigens via mucosal administration routes. Furthermore, the targeting functions of polymeric particles and their ability to co-deliver antigens and adjuvants make them important carriers in vaccine development. These functions permit the use of biodegradable polymeric particle-based vaccines not only for infectious diseases but also for cancers and chronic diseases. The development of such vaccines is expected to greatly improve protection against infectious diseases and cancers.

Particle-based delivery systems are used for mucosal vaccinations because they can protect antigens during gastrointestinal tract delivery. However, the uptake of particles by cells of the mucosal system is highly size-dependent.66 Other challenges of polymeric microspheres include the problem of production scale-up and remnants of unacceptable solvents in the final products.67 Moreover, the requirement of the microencapsulated polymeric microsphere solubility under harsh conditions, such as organic solvents or high temperatures, is problematic. Vaccine antigens and adjuvants may be degraded in such harsh conditions. Regarding the aspect of size, nanoparticles, which feature smaller sizes in the nanoscale range, can better penetrate mucosal barriers. Nanoparticles have gained considerable attention in recent years because of their broad applications for several uses, including industry, agriculture, medicine, cosmetics, and clothing.68 Although there have been some concerns about their safety and the toxicities of various nanomaterials have been reported,69-75 nanoparticle application for vaccines and immunotherapies remains very attractive. For example, vaccine delivery using nanoparticles has been recently reviewed.67 Nanoparticles have also been discussed in the context of cancer immunotherapy.76 Because the 2 size classes of polymeric particles have different properties,77 different applications of microspheres and nanoparticles should be considered according to their advantages and disadvantages. For example, DCs preferentially take up nanoparticles over microparticles.60 Nanoparticles also induce stronger humoral immune responses than microparticles.78 Smaller nanoparticles traffic to lymph nodes because they can penetrate tissue barriers, whereas larger particles are usually retained at the site of injection.79 In addition to size, surface charge is another factor that may affect the performance of polymeric particles. Positively charged particles induce stronger humoral immune responses than do negatively charged particles.80 In another study, immunization with positively charged liposomes induced stronger antibody responses than did antigen alone.81

In this review, we have summarized the functions of biodegradable polymeric particles, including their shield, controlled-release, targeting, and co-delivery functions, as well as their applications in vaccines for several infectious diseases. These functions promote the importance of these particles as carriers in vaccine development. As infectious diseases still cause the deaths of many children in developing countries, the shield and controlled release functions of biodegradable polymeric particles make them useful in developing single-shot vaccines. Considering the discovery of DCs and further great achievements in this field, recently, the targeting and co-delivery functions of polymeric particles maintain their importance in the development of vaccines for cancers and many infectious diseases for which there are not currently effective vaccines, such as human immunodeficiency virus infection, malaria, and tuberculosis. In addition to the functions listed herein, the development of new materials; improvements in encapsulation procedures; understanding of the immune responses elicited by these polymeric particles; and improvements in the co-encapsulation of adjuvants, additives, and stabilizers in these polymeric particles will further improve and extend the applications of these particles to additional vaccines and therapies, resulting in the improvement of human health.

Disclosure of Potential Conflicts of Interest

No potential conflicts of interest were disclosed.

References

  • 1. Pulendran B, Ahmed R. Immunological mechanisms of vaccination. Nat Immunol 2011; 12:509–17; PMID:21739679; http://dx.doi.org/ 10.1038/ni.2039 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 2. Wilson-Welder JH, Torres MP, Kipper MJ, Mallapragada SK, Wannemuehler MJ, Narasimhan B. Vaccine adjuvants: current challenges and future approaches. J Pharm Sci 2009; 98:1278-316; PMID:18704954; http://dx.doi.org/ 10.1002/jps.21523 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3. CDC. Immunization Schedules for Children in Easy-to-read Formats. CDC: Atlanta, GA; 2014 Available from http://www.cdc.gov/vaccines/schedules/easy-to-read/child.html. [Google Scholar]
  • 4. Maina LC, Karanja S, Kombich J. Immunization coverage and its determinants among children aged 12 - 23 months in a peri-urban area of Kenya. Pan Afr Med J 2013; 14:3; PMID:23504493; http://dx.doi.org/ 10.11604/pamj.2013.14.3.2181 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5. Jiang W, Gupta RK, Deshpande MC, Schwendeman SP. Biodegradable poly(lactic-co-glycolic acid) microparticles for injectable delivery of vaccine antigens. Adv Drug Del Rev 2005; 57:391-410; PMID:15560948; http://dx.doi.org/ 10.1016/j.addr.2004.09.003 [DOI] [PubMed] [Google Scholar]
  • 6. Cleland JL. Single-administration vaccines: controlled-release technology to mimic repeated immunizations. Trends Biotechnol 1999; 17:25-9; PMID:10098275; http://dx.doi.org/ 10.1016/S0167-7799(98)01272-4 [DOI] [PubMed] [Google Scholar]
  • 7. Aguado MT. Future approaches to vaccine development: single-dose vaccines using controlled-release delivery systems. Vaccine 1993; 11:596-7; PMID:8488720; http://dx.doi.org/ 10.1016/0264-410X(93)90241-O [DOI] [PubMed] [Google Scholar]
  • 8. Coombes AG, Lavelle EC, Jenkins PG, Davis SS. Single dose, polymeric, microparticle-based vaccines: the influence of formulation conditions on the magnitude and duration of the immune response to a protein antigen. Vaccine 1996; 14:1429-38; PMID:8994318; http://dx.doi.org/ 10.1016/S0264-410X(96)00077-1 [DOI] [PubMed] [Google Scholar]
  • 9. Langer R, Cleland JL, Hanes J. New advances in microsphere-based single-dose vaccines. Adv Drug Del Rev 1997; 28:97-119; PMID:10837567; http://dx.doi.org/ 10.1016/S0169-409X(97)00053-7 [DOI] [PubMed] [Google Scholar]
  • 10. Gupta RK, Singh M, O'Hagan DT. Poly(lactide-co-glycolide) microparticles for the development of single-dose controlled-release vaccines. Adv Drug Del Rev 1998; 32:225-46; PMID:10837646; http://dx.doi.org/ 10.1016/S0169-409X(98)00008-8 [DOI] [PubMed] [Google Scholar]
  • 11. Zhao Z, Leong KW. Controlled delivery of antigens and adjuvants in vaccine development. J Pharm Sci 1996; 85:1261-70; PMID:8961136; http://dx.doi.org/ 10.1021/js9602812 [DOI] [PubMed] [Google Scholar]
  • 12. Singh M, Singh O, Singh A, Talwar GP. Immunogenicity studies on diphtheria toxoid loaded biodegradable microspheres. Int J Pharm 1992; 85:R5-R8; http://dx.doi.org/ 10.1016/0378-5173(92)90157-W [DOI] [Google Scholar]
  • 13. Thomasin C, Corradin G, Men Y, Merkle HP, Gander B. Tetanus toxoid and synthetic malaria antigen containing poly(lactide)/poly(lactide-co-glycolide) microspheres: importance of polymer degradation and antigen release for immune response. J Control Release 1996; 41:131-45; http://dx.doi.org/ 10.1016/0168-3659(96)01363-6 [DOI] [Google Scholar]
  • 14. Lee HK, Park JH, Kwon KC. Double-walled microparticles for single shot vaccine. J Control Release 1997; 44:283-93; http://dx.doi.org/ 10.1016/S0168-3659(96)01534-9 [DOI] [Google Scholar]
  • 15. Cruz LJ, Tacken PJ, Rueda F, Domingo JC, Albericio F, Figdor CG. Targeting nanoparticles to dendritic cells for immunotherapy. Methods Enzymol 2012; 509:143-63.; PMID:22568905; http://dx.doi.org/ 10.1016/B978-0-12-391858-1.00008-3 [DOI] [PubMed] [Google Scholar]
  • 16. Soppimath KS, Aminabhavi TM, Kulkarni AR, Rudzinski WE. Biodegradable polymeric nanoparticles as drug delivery devices. J Control Release: Off J Control Release Soc 2001; 70:1-20; PMID:11166403; http://dx.doi.org/ 10.1016/S0168-3659(00)00339-4 [DOI] [PubMed] [Google Scholar]
  • 17. Kreutz M, Tacken PJ, Figdor CG. Targeting dendritic cells–why bother? Blood 2013; 121:2836-44; PMID:23390195; http://dx.doi.org/ 10.1182/blood-2012-09-452078 [DOI] [PubMed] [Google Scholar]
  • 18. Desai KG, Schwendeman SP. Active self-healing encapsulation of vaccine antigens in PLGA microspheres. J Control Release: Off J Control Release Soc 2013; 165:62-74; PMID:23103983; http://dx.doi.org/ 10.1016/j.jconrel.2012.10.012 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19. Alonso MJ, Gupta RK, Min C, Siber GR, Langer R. Biodegradable microspheres as controlled-release tetanus toxoid delivery systems. Vaccine 1994; 12:299-306; PMID:8178550; http://dx.doi.org/ 10.1016/0264-410X(94)90092-2 [DOI] [PubMed] [Google Scholar]
  • 20. Sanders LM, McRae GI, Vitale KM, Kell BA. Controlled delivery of an LHRH analogue from biodegradable injectable microspheres. J Control Release 1985; 2:187-95; http://dx.doi.org/ 10.1016/0168-3659(85)90044-6 [DOI] [Google Scholar]
  • 21. Cowsar DR, Tice TR, Gilley RM, English JP. Poly(lactide-co-glycolide) microcapsules for controlled release of steroids. Methods Enzymol 1985; 112:101-16; PMID:4046844; http://dx.doi.org/; http://dx.doi.org/ 10.1016/S0076-6879(85)12010-0 [DOI] [PubMed] [Google Scholar]
  • 22. Singh M, Singh O, Talwar GP. Biodegradable delivery system for a birth control vaccine: immunogenicity studies in rats and monkeys. Pharm Res 1995; 12:1796-800; PMID:8592689; http://dx.doi.org/ 10.1023/A:1016294512292 [DOI] [PubMed] [Google Scholar]
  • 23. Johansen P, Estevez F, Zurbriggen R, Merkle HP, Gluck R, Corradin G, Gander B. Towards clinical testing of a single-administration tetanus vaccine based on PLA/PLGA microspheres. Vaccine 2000; 19:1047-54; PMID:11137238; http://dx.doi.org/ 10.1016/S0264-410X(00)00343-1 [DOI] [PubMed] [Google Scholar]
  • 24. Bloom BR. Vaccines for the Third World. Nature 1989; 342:115-20; PMID:2812009; http://dx.doi.org/ 10.1038/342115a0 [DOI] [PubMed] [Google Scholar]
  • 25. Aguado MT, Lambert PH. Controlled-release vaccines–biodegradable polylactide/polyglycolide (PL/PG) microspheres as antigen vehicles. Immunobiology 1992; 184:113-25; PMID:1587538; http://dx.doi.org/ 10.1016/S0171-2985(11)80470-5 [DOI] [PubMed] [Google Scholar]
  • 26. Alonso MJ, Cohen S, Park TG, Gupta RK, Siber GR, Langer R. Determinants of release rate of tetanus vaccine from polyester microspheres. Pharm Res 1993; 10:945-53; PMID:8378256; http://dx.doi.org/ 10.1023/A:1018942118148 [DOI] [PubMed] [Google Scholar]
  • 27. Sanchez A, Gupta RK, Alonso MJ, Siber GR, Langer R. Pulsed controlled-released system for potential use in vaccine delivery. J Pharm Sci 1996; 85:547-52; PMID:8773947; http://dx.doi.org/ 10.1021/js960069y [DOI] [PubMed] [Google Scholar]
  • 28. Men Y, Thomasin C, Merkle HP, Gander B, Corradin G. A single administration of tetanus toxoid in biodegradable microspheres elicits T cell and antibody responses similar or superior to those obtained with aluminum hydroxide. Vaccine 1995; 13:683-9; PMID:7668038; http://dx.doi.org/ 10.1016/0264-410X(94)00046-P [DOI] [PubMed] [Google Scholar]
  • 29. Jung T, Koneberg R, Hungerer KD, Kissel T. Tetanus toxoid microspheres consisting of biodegradable poly(lactide-co-glycolide)- and ABA-triblock-copolymers: immune response in mice. Int J Pharm 2002; 234:75-90; PMID:11839439; http://dx.doi.org/ 10.1016/S0378-5173(01)00957-7 [DOI] [PubMed] [Google Scholar]
  • 30. Singh M, Li XM, Wang H, McGee JP, Zamb T, Koff W, Wang CY, O'Hagan DT. Immunogenicity and protection in small-animal models with controlled-release tetanus toxoid microparticles as a single-dose vaccine. Infect Immun 1997; 65:1716-21; PMID:9125552 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31. Johansen P, Moon L, Tamber H, Merkle HP, Gander B, Sesardic D. Immunogenicity of single-dose diphtheria vaccines based on PLA/PLGA microspheres in guinea pigs. Vaccine 1999; 18:209-15; PMID:10506644; http://dx.doi.org/ 10.1016/S0264-410X(99)00191-7 [DOI] [PubMed] [Google Scholar]
  • 32. Peyre M, Sesardic D, Merkle HP, Gander B, Johansen P. An experimental divalent vaccine based on biodegradable microspheres induces protective immunity against tetanus and diphtheria. J Pharm Sci 2003; 92:957-66; PMID:12712415; http://dx.doi.org/ 10.1002/jps.10361 [DOI] [PubMed] [Google Scholar]
  • 33. Josefsberg JO, Buckland B. Vaccine process technology. Biotechnol Bioeng 2012; 109:1443-60; PMID:22407777; http://dx.doi.org/ 10.1002/bit.24493 [DOI] [PubMed] [Google Scholar]
  • 34. Moldoveanu Z, Clements ML, Prince SJ, Murphy BR, Mestecky J. Human immune responses to influenza virus vaccines administered by systemic or mucosal routes. Vaccine 1995; 13:1006-12; PMID:8525683; http://dx.doi.org/ 10.1016/0264-410X(95)00016-T [DOI] [PubMed] [Google Scholar]
  • 35. Johansson EL, Wassen L, Holmgren J, Jertborn M, Rudin A. Nasal and vaginal vaccinations have differential effects on antibody responses in vaginal and cervical secretions in humans. Infect Immun 2001; 69:7481-6; PMID:11705923; http://dx.doi.org/ 10.1128/IAI.69.12.7481-7486.2001 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 36. Rudin A, Riise GC, Holmgren J. Antibody responses in the lower respiratory tract and male urogenital tract in humans after nasal and oral vaccination with cholera toxin B subunit. Infect Immun 1999; 67:2884-90; PMID:10338495 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37. Bergquist C, Johansson EL, Lagergard T, Holmgren J, Rudin A. Intranasal vaccination of humans with recombinant cholera toxin B subunit induces systemic and local antibody responses in the upper respiratory tract and the vagina. Infect Immun 1997; 65:2676-84; PMID:9199436 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 38. Kozlowski PA, Williams SB, Lynch RM, Flanigan TP, Patterson RR, Cu-Uvin S, Neutra MR. Differential induction of mucosal and systemic antibody responses in women after nasal, rectal, or vaginal immunization: influence of the menstrual cycle. J Immunol 2002; 169:566-74; PMID:12077289; http://dx.doi.org/ 10.4049/jimmunol.169.1.566 [DOI] [PubMed] [Google Scholar]
  • 39. Liang B, Hyland L, Hou S. Nasal-associated lymphoid tissue is a site of long-term virus-specific antibody production following respiratory virus infection of mice. J Virol 2001; 75:5416-20; PMID:11333927; http://dx.doi.org/ 10.1128/JVI.75.11.5416-5420.2001 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 40. Hanes J, Chiba M, Langer R. Polymer microspheres for vaccine delivery. Pharm Biotechnol 1995; 6:389-412; PMID:7551227; http://dx.doi.org/ 10.1007/978-1-4615-1823-5_16 [DOI] [PubMed] [Google Scholar]
  • 41. Eldridge JH, Meulbroek JA, Staas JK, Tice TR, Gilley RM. Vaccine-containing biodegradable microspheres specifically enter the gut-associated lymphoid tissue following oral administration and induce a disseminated mucosal immune response. Adv Exp Med Biol 1989; 251:191-202; PMID:2610110 [DOI] [PubMed] [Google Scholar]
  • 42. Eyles JE, Sharp GJ, Williamson ED, Spiers ID, Alpar HO. Intra nasal administration of poly-lactic acid microsphere co-encapsulated Yersinia pestis subunits confers protection from pneumonic plague in the mouse. Vaccine 1998; 16:698-707; PMID:9562689; http://dx.doi.org/ 10.1016/S0264-410X(97)00249-1 [DOI] [PubMed] [Google Scholar]
  • 43. Almeida AJ, Alpar HO. Nasal delivery of vaccines. J Drug Target 1996; 3:455-67; PMID:8863138; http://dx.doi.org/ 10.3109/10611869609015965 [DOI] [PubMed] [Google Scholar]
  • 44. Beier R, Gebert A. Kinetics of particle uptake in the domes of Peyer's patches. Am J Physiol 1998; 275:G130-7; PMID:9655693 [DOI] [PubMed] [Google Scholar]
  • 45. Yeh M-K, Liu Y-T, Chen J-L, Chiang C-H. Oral immunogenicity of the inactivated Vibrio cholerae whole-cell vaccine encapsulated in biodegradable microparticles. J Control Release 2002; 82:237-47; PMID:12175740; http://dx.doi.org/ 10.1016/S0168-3659(02)00140-2 [DOI] [PubMed] [Google Scholar]
  • 46. Yeh M, Chiang C. Inactive Vibrio cholerae whole-cell vaccine-loaded biodegradable microparticles: in vitro release and oral vaccination. J Microencapsul 2004; 21:91-106; PMID:14718189; http://dx.doi.org/ 10.1080/02652040310001619794 [DOI] [PubMed] [Google Scholar]
  • 47. Huang SS, Li IH, Hong PD, Yeh MK. Development of Yersinia pestis F1 antigen-loaded microspheres vaccine against plague. Int J Nanomed 2014; 9:813-22; PMID:24550673 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 48. Huang SS, Li IH, Hong PD, Yeh MK. Evaluation of protective efficacy using a nonstructural protein NS1 in DNA vaccine-loaded microspheres against dengue 2 virus. Int J Nanomed 2013; 8:3161-9; PMID:23990724 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 49. Plotkin SA. Vaccines: correlates of vaccine-induced immunity. Clin Infect Dis: Off Pub Infect Dis Soc Am 2008; 47:401-9; PMID:18558875; http://dx.doi.org/ 10.1086/589862 [DOI] [PubMed] [Google Scholar]
  • 50. Pantaleo G, Koup RA. Correlates of immune protection in HIV-1 infection: what we know, what we don't know, what we should know. Nat Med 2004; 10:806-10; PMID:15286782; http://dx.doi.org/ 10.1038/nm0804-806] [DOI] [PubMed] [Google Scholar]
  • 51. Hoft DF. Tuberculosis vaccine development: goals, immunological design, and evaluation. Lancet 2008; 372:164-75; PMID:18620952; http://dx.doi.org/ 10.1016/S0140-6736(08)61036-3 [DOI] [PubMed] [Google Scholar]
  • 52. Reyes-Sandoval A, Pearson FE, Todryk S, Ewer K. Potency assays for novel T-cell-inducing vaccines against malaria. Curr Opin Mol Therap 2009; 11:72-80; PMID:19169962 [PubMed] [Google Scholar]
  • 53. Steinman RM. Dendritic cells in vivo: a key target for a new vaccine science. Immunity 2008; 29:319-24; PMID:18799140; http://dx.doi.org/ 10.1016/j.immuni.2008.08.001 [DOI] [PubMed] [Google Scholar]
  • 54. Figdor CG, de Vries IJ, Lesterhuis WJ, Melief CJ. Dendritic cell immunotherapy: mapping the way. Nat Med 2004; 10:475-80; PMID:15122249; http://dx.doi.org/ 10.1038/nm1039 [DOI] [PubMed] [Google Scholar]
  • 55. Palucka K, Banchereau J. Cancer immunotherapy via dendritic cells. Nat Rev Cancer 2012; 12:265-77; PMID:22437871; http://dx.doi.org/ 10.1038/nrc3258 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 56. Idoyaga J, Cheong C, Suda K, Suda N, Kim JY, Lee H, Park CG, Steinman RM. Cutting edge: langerin/CD207 receptor on dendritic cells mediates efficient antigen presentation on MHC I and II products in vivo. J Immunol 2008; 180:3647-50; PMID:18322168; http://dx.doi.org/ 10.4049/jimmunol.180.6.3647 [DOI] [PubMed] [Google Scholar]
  • 57. Idoyaga J, Fiorese C, Zbytnuik L, Lubkin A, Miller J, Malissen B, Mucida D, Merad M, Steinman RM. Specialized role of migratory dendritic cells in peripheral tolerance induction. J Clin Invest 2013; 123:844-54; PMID:23298832 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 58. Gallucci S, Lolkema M, Matzinger P. Natural adjuvants: endogenous activators of dendritic cells. Nat Med 1999; 5:1249-55; PMID:10545990; http://dx.doi.org/ 10.1038/15200 [DOI] [PubMed] [Google Scholar]
  • 59. Kastenmuller W, Kastenmuller K, Kurts C, Seder RA. Dendritic cell-targeted vaccines - hope or hype? Nat Rev Immunol 2014; 14:705-11; PMID:25190285; http://dx.doi.org/ 10.1038/nri3727 [DOI] [PubMed] [Google Scholar]
  • 60. Cruz LJ, Tacken PJ, Fokkink R, Joosten B, Stuart MC, Albericio F, Torensma R, Figdor CG. Targeted PLGA nano- but not microparticles specifically deliver antigen to human dendritic cells via DC-SIGN in vitro. J Control Release: Off J Control Release Soc 2010; 144:118-26; PMID:20156497; http://dx.doi.org/ 10.1016/j.jconrel.2010.02.013 [DOI] [PubMed] [Google Scholar]
  • 61. Zhang Z, Tongchusak S, Mizukami Y, Kang YJ, Ioji T, Touma M, Reinhold B, Keskin DB, Reinherz EL, Sasada T. Induction of anti-tumor cytotoxic T cell responses through PLGA-nanoparticle mediated antigen delivery. Biomaterials 2011; 32:3666-78; PMID:21345488; http://dx.doi.org/ 10.1016/j.biomaterials.2011.01.067 [DOI] [PubMed] [Google Scholar]
  • 62. Schlosser E, Mueller M, Fischer S, Basta S, Busch DH, Gander B, Groettrup M. TLR ligands and antigen need to be coencapsulated into the same biodegradable microsphere for the generation of potent cytotoxic T lymphocyte responses. Vaccine 2008; 26:1626-37; PMID:18295941; http://dx.doi.org/ 10.1016/j.vaccine.2008.01.030 [DOI] [PubMed] [Google Scholar]
  • 63. Fahmy TM, Samstein RM, Harness CC, Mark Saltzman W. Surface modification of biodegradable polyesters with fatty acid conjugates for improved drug targeting. Biomaterials 2005; 26:5727-36; PMID:15878378; http://dx.doi.org/ 10.1016/j.biomaterials.2005.02.025 [DOI] [PubMed] [Google Scholar]
  • 64. Duncanson WJ, Figa MA, Hallock K, Zalipsky S, Hamilton JA, Wong JY. Targeted binding of PLA microparticles with lipid-PEG-tethered ligands. Biomaterials 2007; 28:4991-9; PMID:17707503; http://dx.doi.org/ 10.1016/j.biomaterials.2007.05.044 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 65. Cruz LJ, Tacken PJ, Fokkink R, Figdor CG. The influence of PEG chain length and targeting moiety on antibody-mediated delivery of nanoparticle vaccines to human dendritic cells. Biomaterials 2011; 32:6791-803; PMID:21724247; http://dx.doi.org/ 10.1016/j.biomaterials.2011.04.082 [DOI] [PubMed] [Google Scholar]
  • 66. Woodrow KA, Bennett KM, Lo DD. Mucosal vaccine design and delivery. Annu Rev Biomed Eng 2012; 14:17-46; PMID:22524387; 23532930http://dx.doi.org/10.1146/annurev-bioeng-071811-150054 [DOI] [PubMed] [Google Scholar]
  • 67. Gregory AE, Titball R, Williamson D. Vaccine delivery using nanoparticles. Front Cell Infect Microbiol 2013; 3:13; PMID:23532930; http://dx.doi.org/ 10.3389/fcimb.2013.00013 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 68. Fu PP. Introduction to the special issue: nanomaterials– toxicology and medical applications. J Food Drug Anal 2014; 22:1-2; PMID:24673899; http://dx.doi.org/ 10.1016/j.jfda.2014.01.013 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 69. Karmakar A, Zhang Q, Zhang Y. Neurotoxicity of nanoscale materials. J Food Drug Anal 2014; 22:147-60; PMID:24673911; http://dx.doi.org/ 10.1016/j.jfda.2014.01.012 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 70. McShan D, Ray PC, Yu H. Molecular toxicity mechanism of nanosilver. J Food Drug Anal 2014; 22:116-27; PMID:24673909; http://dx.doi.org/ 10.1016/j.jfda.2014.01.010 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 71. Guo X, Mei N. Assessment of the toxic potential of graphene family nanomaterials. J Food Drug Anal 2014; 22:105-15; PMID:24673908; http://dx.doi.org/ 10.1016/j.jfda.2014.01.009 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 72. Chen T, Yan J, Li Y. Genotoxicity of titanium dioxide nanoparticles. J Food Drug Anal 2014; 22:95-104; PMID:24673907; http://dx.doi.org/ 10.1016/j.jfda.2014.01.008 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 73. Wu H, Yin JJ, Wamer WG, Zeng M, Lo YM. Reactive oxygen species-related activities of nano-iron metal and nano-iron oxides. J Food Drug Anal 2014; 22:86-94; PMID:24673906; http://dx.doi.org/ 10.1016/j.jfda.2014.01.007 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 74. Li M, Yin JJ, Wamer WG, Lo YM. Mechanistic characterization of titanium dioxide nanoparticle-induced toxicity using electron spin resonance. J Food Drug Anal 2014; 22:76-85; PMID:24673905; http://dx.doi.org/ 10.1016/j.jfda.2014.01.006 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 75. Fu PP, Xia Q, Hwang HM, Ray PC, Yu H. Mechanisms of nanotoxicity: generation of reactive oxygen species. J Food Drug Anal 2014; 22:64-75; PMID:24673904; http://dx.doi.org/ 10.1016/j.jfda.2014.01.005 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 76. Serda RE. Particle platforms for cancer immunotherapy. Int J Nanomed 2013; 8:1683-96; PMID:23761969; http://dx.doi.org/ 10.2147/IJN.S31756 [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 77. Kohane DS. Microparticles and nanoparticles for drug delivery. Biotechnol Bioeng 2007; 96:203-9; PMID:17191251; http://dx.doi.org/ 10.1002/bit.21301 [DOI] [PubMed] [Google Scholar]
  • 78. Fifis T, Gamvrellis A, Crimeen-Irwin B, Pietersz GA, Li J, Mottram PL, McKenzie IF, Plebanski M. Size-dependent immunogenicity: therapeutic and protective properties of nano-vaccines against tumors. J Immunol 2004; 173:3148-54; PMID:15322175; http://dx.doi.org/ 10.4049/jimmunol.173.5.3148 [DOI] [PubMed] [Google Scholar]
  • 79. Manolova V, Flace A, Bauer M, Schwarz K, Saudan P, Bachmann MF. Nanoparticles target distinct dendritic cell populations according to their size. Eur J Immunol 2008; 38:1404-13; PMID:18389478; http://dx.doi.org/ 10.1002/eji.200737984 [DOI] [PubMed] [Google Scholar]
  • 80. Slutter B, Bal S, Keijzer C, Mallants R, Hagenaars N, Que I, Kaijzel E, van Eden W, Augustijns P, Löwik C, et al. . Nasal vaccination with N-trimethyl chitosan and PLGA based nanoparticles: nanoparticle characteristics determine quality and strength of the antibody response in mice against the encapsulated antigen. Vaccine 2010; 28:6282-91; PMID:20638455; http://dx.doi.org/ 10.1016/j.vaccine.2010.06.121 [DOI] [PubMed] [Google Scholar]
  • 81. Baca-Estrada ME, Foldvari M, Snider M. Induction of mucosal immune responses by administration of liposome-antigen formulations and interleukin-12. J Interf Cytok Res: Off J Int Soc Interf Cytok Res 1999; 19:455-62; PMID:10386857; http://dx.doi.org/ 10.1089/107999099313893 [DOI] [PubMed] [Google Scholar]

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