Vaccine delivery system

Contents
IntroductionWhat are vaccine?
History of vaccineIdeal properties of vaccine.
Mechanism of vaccine
Types of vaccineUptake of antigen
Single shot vaccine
Mucosal delivery vaccine
Transdermal delivery vaccineReferences

1. Vaccine Delivery System. DEPARTMENT OF PHARMACEUTICS . GUIDED BY: Dr MOHD.WAIS PRESENTED BY SIDDIQUE ADNAN REHMATULLAH ARIFA 1

2. CONTENT Introduction What are vaccine? History of vaccine Ideal properties of vaccine. Mechanism of vaccine Types of vaccine Uptake of antigen Single shot vaccine Mucosal delivery vaccine Transdermal delivery vaccine References 2

3. Introduction to vaccine delivery system The concept of vaccine delivery include a range of devices and physical delivery systems that are designed to allow immunization using non invasive routes and to achieve different routes of vaccine administration. 3

4. What are Vaccine ? Vaccines are biological preparation that provides Active acquired Immunity to a particular disease. A vaccine typically contains an agent that resembles a disease causing microorganism. It is often made from weakened or killed forms of the causative microbes or its toxin or one of its surface proteins . The agent stimulate the body's immune system to recognize foreign agent, destroy it, and keep a record of it, so that the immune system can more easily recognize and destroy any of these microorganisms that it later encounters. 4

5. Brief History of vaccine The term VACCINE and VACCINATION are derived from a Latin word variolae vaccinae . F irst vaccine was developed in 1978 for small pox by E dward jenner. His innovations begun with successful use of cowpox material to create immunity against smallpox. L ater louis pasteur invented rabies vaccine in 1985. 5 Figure 1 Figure 2

6. Mechanism of vaccine V accines contains antigenic material which after administration into body, stimulates recipient's immune system to recognize it as a threat and to remember and destroy it. V accines can be prophylactic (e.g. to prevent or ameliorate the effects of a future infection by a natural or wild pathogen), or therapeutic (e.g., vaccines against cancer are being investigated) 6 Figure 3 Mechansim of Vaccine

7. Properties of ideal vaccine Should provide long lasting immunity . Should induce both humoral and cellular immunity . Should not induce autoimmunity or hypersensitivity . Should be inexpensive to produce, easy to store and administer It should be easy to administer preferably orally It should be stable under various condition (temperature ,light , transportation) It should be economically cheap. 7

8. Types of vaccine Vaccines are Dead or inactivated micro-organism or purified product derived from them . 1 . Traditional vaccine :-Traditional vaccines use a dead or a weakened pathogenic microbe or a toxin from a pathogen. The introduction of an attenuated or dead pathogen into a healthy individual generates an immune response. 2 . Innovative Vaccine :- Example conjugate vaccine , recombinant vector vaccine , T cell receptor peptide vaccine , valent (monovalent, multivalent) 8

9. Inactivated - Virulent factor of microbes inactivated by chemicals, heat, irradiation, etc. (E.g.: Influenza, Cholera ) Live attenuated vaccine - Contain live, weakened microbes that are cultivated under specific conditions. ( E.g.Measles , Yellow fever , BCG) Toxoid : Prepared from inactivated toxic compounds causing illness. ( E.g.Tetanus , Diptheria ) Protein Sub-unit : Fragment of inactivated/attenuated microbe used to elicit immune response. ( Eg : Sub-unit vaccine of Hepatitis B) Conjugate : Poorly immunogenic outer coats of certain bacteria linked to proteins/ toxins. ( eg : Haemophilus influenza Type B) 9

10. Recombinant Vector : Combination of physiology of one microbe with DNA of another.(Hepatitis B) DNA vaccine : Insertion of viral or bacterial DNA into human cells. The immune system recognizes against this proteins and the cells expressing them . T-cell receptor peptide vaccine - They show thermodulation of cytokine production and improve cell mediated immunity and are under development . Valence:- a Monovalent- Use to immunize against single antigen b . Multivalent- Used to immunize against two or more microorganism Heterotypic - Vaccines that are pathogens of other animals that either do not cause disease or cause mild disease in the organism being treated 10

11. Uptake of antigen. Antigens generated by endogenous and exogenous antigen processing activate different effector functions EXOGENOUS PATHOGENS ENDOGENOUS PATHOGENS Eliminated by : Antibodies and phagocyte activation by T helper cells that use antigens generated by EXOGENOUS PROCESSING Eliminated by : Killing of infected cells by CTL that use antigens generated by ENDOGENOUS PROCESSING 11 T able1

12. Stages of exogenous antigen uptake 12

13. 13 Figure 4 Exogenous uptake

14. Stages of endogenous antigen uptake 14

15. 15 Figure 5 E ndogenous uptake

16. Delivery system Used to promote Uptake of antigens in Vaccine Absorption enhancers. Lipid carrier systems. Oral immunization Controlled release micro particles for vaccine development Single dose vaccine delivery systems using bio degradable polymers. Peptide based vaccines Nucleic acid based vaccines I DNA vaccines II RNA vaccines 16

17. SINGLE SHOT VACCINE 17

18. Introduction Single shot vaccines are given for preventing 4 to 6 diseases with only one injection . To provide effective patient protection, many traditional vaccines requires multiple injections, which results in a costly and inconvenient regimen . These disadvantages have spurred the development of single-shot vaccines that can provide protection against infection with only one injection . DEFINATION: The single shot vaccine is a Combination Product of a Prime Component Antigen with an Microsphere Component and appropriate Adjuvants and an encapsulated antigen which will provide the booster immunizations by delayed release of the antigen . 18

19. Components of single shot vaccine Involves following things:- Prime component antigen : Responsible for the production of antibody. Adjuvant : added in formulation to increase the efficacy and to improve the perfomance of the single shot vaccine Microsphere component : Involves use of different biopolymers which forms microsphere encpasulating the antigen which released over a long period of time to avoid need of booster dose. 19

20. Adjuvant of Single shot vaccine V ACCINE ADJUVANTS :- Adjuvants are the substances added to vaccines to help them work better . Adding an adjuvant triggers the immune system to become more sensitive to the vaccine. NEED FOR ADJUVANTS :- • To increase the therapeutic efficiency. • They form depot of antigen at the site of inoculation with slow release of antigens. • It can improves the performance of vaccines by targeting the antigen to APC . 20

21. Type of adjuvants 1 Gel types eg . :- aluminum hydroxide, calcium phosphate. 2 Oil emulsion and emulsifier Particulate based type eg :- liposomes , biodegradable microspheres . 3 Virosomes :- T hey resemble to viruses but non-infectious that are incorporated in these vaccines have antigens and other proteins on their surface, but they can not cause infection because it does not contain any genetic material. M echanism : some immune cells recognize these particulate cells and engulf them and present them to immune system and mount protective response. 21

22. Biodegradable polymer • Biodegradable polymers are defined as polymers comprised of monomers linked to one another through functional groups and have unstable links in the backbone. • Broken down into biologically acceptable molecules that are metabolized and removed from the body via normal metabolic pathways 22

23. Types of biodegradable polymer • There are two types of biodegradable polymers They are : 1) Natural biodegradable polymers :- eg : Albumin, Collagen, Gelatin etc.,. 2) Synthetic biodegradable polymers :- eg : Aliphatic poly(esters), Pseudo poly aminoacid , etc., 23

24. 24 Figure 6 :Classification of biodegradable polymers based on production

25. Important determinants for single shot vaccine development 25

26. Microsphere componenet of single shot vaccine • The microsphere component uses OctoVAX microsphere technology, which is based on cross-linked modified dextran polymers. • Dextrans are ideal polymers to form biocompatible hydrogels, Two major advantages of dextran microspheres as protein delivery systems , that the particles are prepared in the absence of organic solvents , and that degradation of the microspheres does not result in a pH drop. • Several different dextrans have been developed for hydrogel formation . One of these dextran-based polymers is derivatized with hydroxy -ethyl methacrylate ( DexHEMA ). • Which introduces hydrolytically sensitive carbonate ester groups that ensure biodegradation under physiological conditions. 26

27. Procedure for the formation of Microsphere suspension To ensure consistently high encapsulation efficiencies, the protein to be encapsulated is added to the dex -HEMA solution before adding the PEG solution. Subsequently , microspheres are obtained by polymerizing the HEMA groups using potassium persulfate (KPS) as initiator and N,N,N',N'- tetramethylethylenediamine (TEMED) as the catalyst Several factors are critical parameters for the formulation of consistent microspheres. First, the size distribution of the microspheres can be controlled by the shear force applied during the emulsification step in the bioreactor vessel . 27

28. 28 Figure 7 Formulation of Microsphere suspension

29. • Dextran hydroxyethyl methacyrylate ( Dex -HEMA) is very much suitable for the formation of the hydrogel that facilitates controlled release of encapsulated proteins . 29 Figure 8 comparision graph between Conventional vaccine and s i n g l e s h o t l vaccine

30. Controlling Particle Size During Process Scale-up The dextran microsphere preparation method is described by Stenekes was initially performed on a 5-g scale ( containing 120 mg of microspheres), and used vortexing to emulsify the dex -HEMA phase in the continuous PEG phase . 30 Figure 9 Microscopic pictures of microspheres produced at a high stirring speed of 700 rpm

31. • A direct correlation was observed between stirring speed and mean particle diameter of the microspheres, thus confirming that the particle size of the dextran emulsion is dependent on the energy input during emulsification. • It is important to note that, despite the larger mean diameter of microspheres prepared at the 500-g scale, more than 90% of the resulting particles had a size below 90 μm , a size suitable for subcutaneous injection. 31

32. Delayed antigen release from dex -HEMA microspheres Once the freeze-dried microsphere product is rehydrated by reconstitution in an aqueous solution, hydrolysis of the carbonate ester groups in the dex -HEMA will be initiated. This will increase the mesh size in the hydrogel network. The encapsulated protein will be released when the mesh size exceeds the hydrodynamic diameter of the protein. 32

33. Factor affecting antigen release 1. Polymer nature 2 . Crystallinity 3. Method of preparation 4. Molecular weight of drug 5. Carrier size and morphology Adverse effects 1 Fever 2 Pain around injection site 3 Muscle aches 33

34. Advantages Disadvantages Economic. With one Injection 4 to 6 Infections can be prevented. Patient compliance is Improved because, they would replace the need for a prime boost regimen, consequently eliminating the repeated visits to the doctor for mother and their children. The primary risk associated with vaccines, especially vaccines that utilize live organisms, so that the vaccine itself causes illness. The vaccine may behave as a super antigen and over stimulate the immune system. Some are not as effective as Multi-dose vaccines, because infection can occur due to micro organisms. 34 Table No-2

35. Example of Single shot vaccine Single-shot Ad26 vaccine protects against SARS-CoV-2 in rhesus macaques(under development ) Single-shot hepatitis B vaccine formulated with PLGA microspheres single-shot subunit vaccine for HIV-1 . 35

36. Mucosal delivery of vaccine 36

37. INTRODUCTION Mucosal surfaces area is Major portal of entry for many human pathogens that are the cause of infectious disease worldwide . Efforts have focused on efficient delivery of vaccine antigens to mucosal sites that facilitate uptake by local antigen-presenting cells to generate protective mucosal immune responses . The adult human mucosa lines the surfaces of the Digestive, Respiratory and genitourinary tracts, covering an immense surface area (400 m2 ) that is~200 times greater than that of the skin. Mucosal surfaces are typically categorized as Type-1 and Type-2 mucosa . Type-1 mucosa include surface of the lung and gut, where as Type-2 mucosa include surfaces of the mouth, esophagus and cornea 37

38. Most mucosal sites have organized lymphoid follicles, such as NALT ( Nasal associated lymphoid tissue ), and GALT (Gastric associated lymphoid tissue) , which have assembly of scattered antigen-reactive cells of immune system, such as B cells, T cells, and professional antigen presenting cells such as dendritic cells(APCs ). These cells are responsible for the induction and maintenance of immune responses against mucosally delivered antigen . It is widely accepted that mucosal vaccination can induce immune responses at both systemic and mucosal sites and prevent the invasion and colonization of pathogens at mucosal surfaces. 38

39. 39 Figure 10 Figure 11

40. WHY? Mucosal delivery of vaccines. Because injectable vaccines pose several problems such as- • High production cost • Low/poor compliance • Fear of needle borne infection • Lack of mucosal immune response • Injection site pain • Local side effects 40

41. Challenges in Mucosal vaccine delivery system. 41

42. Advantages and limitations Advantages Low cost as compared to injectable vaccines. Easily adaptable for the purpose of mass vaccination ( especially desirable in pandemic situation) The mucosal surface has an area of > 400m2 in an adult and produces more antibodies than all other types of antibodies in the blood combined, around 5gm are secreted at the mucosal surface each day. L imitations Hostile nature of gastrointestinal tract in case of oral vaccine . Impermeability of bio macromolecule through the mucosal epithelial barrier . presence of degrading enzyme and fear of the induction of tolerance . 42

43. POLYMERS USED FOR MUCOSAL VACCINES The concept of polymeric carrier system(s) offers advantage of delivering drugs/antigens to a specific target site, where it has to be released from the carrier. 43 Figure 12-Classification of polymer

44. DESGIN AND STRATEGIES FOR MUCOSAL DELIVERY Emulsion type delivery Liposome based delivery Polymeric nano particle Virosomes Melt in mouth strips 44

45. EMULSION TYPE DELIVERY Emulsions are heterogeneous liquid systems may be w/o or o/w Antigens are dissolved in a water phase and emulsified in the oil in the presence of an appropriate emulsifier. The controlled release characteristics of an emulsion are determined by factors such as :- Viscosity of oil phase. Oil to water phase ratio. Emulsion droplet size. H uang et developed a novel emulsion type vaccine delivery systems of the amphiphilic bioresorbable polymer poly (ethylene glycol)- block-poly( lactide -co-epsilon- caprolactone ) (peg-b- placl ) using ovalbumin as model antigen. 45

46. LIPOSOME BASED DELIVERY Liposomes are spherical shape vesicles containing an aqueous core which is enclosed by a lipid bilayer . They are most often composed of phospholipids, especially phosphatidylcholine , but may also include other lipids, like Ethanolamine . Preparation of Liposome Vaccine Delivery System:- Depending on the chemical properties, water-soluble antigens ( proteins, peptide, nucleic acids, carbohydrates, haptens ) are entrapped within the aqueous inner space of liposomes Lipophilic compounds ( lipopeptides , antigens, adjuvants, linker molecules ) are intercalated into the lipid bilayer Antigens or adjuvants can be attached to the liposome surface either by adsorption or stable chemical linking 46

47. Polymeric nano particles Polymeric nano particles are submicron-sized colloidal particles. Polymeric nanoparticles because of their size are preferentially taken up by the mucosa associated lymphoid Tissue. Limited doses of antigen are sufficient to induce effective immunization . Hence, the use of nanoparticles for oral delivery of antigens is suitable because of their ability to release proteins and to protect them from enzymatic degradation in the GIT. 47

48. B iodegradable poly(alkyl cyano -acrylate) ( paca ) N anoparticles have been shown to enhance the secretory immune response after their oral administration in association with ovalbumin in rats. B iodegradable poly(methyl metha acrylate)( pmma ) N anoparticles being very slowly degradable appear to be particularly suitable for vaccine purposes because P rolonged contact between antigen and immunocompetent cells favors persistent immunity. 48

49. VIROSOMES • A Virosome is a a drug or vacine delivery mechanism consisting of unilamellar phospholipid membrane (150mm ) vesicle incorporating virus derived proteins to allow the virosomes to fuse with target cells . • These proteins enable the virosome membranes to fuse with cells of the immune system and thus deliver the specific antigens directly to their target cells . • They elicit a specific immune response even with weak-immunogenic antigens. • Once they have delivered the antigens, the virosomes are completely degraded within the cells. 49 Figure 13-components of virosome Figure 14-Virosome

50. MELT IN MOUTH STRIPS • Quick dissolving films containing I mmunogens . • Melts into liquid that children and infants will swallow easily . • These strips stick and dissolves on the tongue in less than a minute. (useful for newborns who sometimes spit out the liquid) • EXAMPLE: ROTAVIRUS is a common cause of severe diarrhea and vomiting in children. ROTAVIRUS VACCINE at present is available in a liquid or freeze-dried form that must be chilled for transport and storage, making it very expensive for use in impoverished areas 50

51. 51 Figure 15 Figure 16 Figure 17

52. Advantages of mucosal vaccine over conventional vaccine C onventional Administrated by needle Require professional skills patient compliance is less Pain at injecton site Induce systemic immunity Defence occur after the invasion Mucosal Administrated orally No requirement of professional skill Patient compliance is more No pain Induce both systemic and mucosal immunity Defence at both front of invasion and after the invasion 52

53. Example of mucosal vaccine delivery system 53 Table No-3

54. Transdermal vaccine delivery system. 54

55. Introduction The skin is the largest and most accessible organ of the body. V accine administration to the skin offers many advantages including ease of access, a potential for generation of both systemic and mucosal immune response. F ormulation approaches such as liposomes, physical penetration enhancers such as electroporation, and technologies that create micron-sized pores in the skin, such as micro-needles have been used for transdermal delivery of vaccine. 55

56. Skin as a site of vaccine delivery T he skin has multiple barrier properties to minimize water loss from the body and prevent the permeation of environmental contaminants into the body . T hese barriers can be considered as physical, enzymatic and immunological. 1) Physical barriers T he outermost layer, stratum corneum presents an effective physical barrier to the permeation of large molecules such as vaccines. this is the first barrier property that must be overcome to provide effective transdermal vaccine delivery. 56

57. 2 ) Enzymatic barrier T he skin possesses many enzymes that are capable of hydrolyzing peptides and proteins their potential to degrade topically applied vaccine antigens should be considered. 3) Immunological barriers When the skin is damaged, environmental contaminants can access the epidermis to initiate an immunological response . 57

58. M any approaches have been investigated to overcome the skins barrier properties in order to deliver antigens via the skin . A ll methods aim to overcome the stratum corneum barrier and target vaccine to immune responsive cells such as langerhans cells . 58 Figure 18-Mechanism of antigen release

59. Immunization by Transdermal route includes approaches like: 1 Liquid jet injector 2 Epidermal powder immunization 3 Colloidal carrier A)Nano particles and nano carriers B)Liposomes and elastic vesicles 4 Energy Based Approaches A) Electroporation B)Ultra sound or Sonophoresis C)Micro needles 59

60. Liquid jet injectors L iquid jet injectors use a high-velocity jet () to deliver molecules through the skin region typically into the subcutaneous or intramuscular at rate of 100 to 200 m/s.- J et injectors can be broadly classified into:- 1) multi-use nozzle jet injectors (MUNJIS) 2 ) disposable cartridge jet injectors (DCJIS) Commercially available liquid jet injectors consists of a)power source (compressed gas or spring) b) Piston c)drug or vaccine-loaded compartment d)application nozzle, with typical orifice size in the range of 150 to 300 um 60

61. Working of Jet injector 1)Upon actuation the power source pushes the piston & rapidly increases the pressure within the drug-loaded compartment 2) Thereby forcing the drug solution through the orifice as a high velocity liquid jet. 3) When the jet impacts on the skin it creates a hole &allows the liquid to enter the skin. 4)The process of hole formation and liquid jet deposition occurs within microseconds. 5)The deposited liquid can then disperse within the tissues to illicit an immune response. 61

62. Commercially available jet injector. 1)VISION® AND CHOICE® which delivers a variable dose of insulin 2)V-GO MINI-JECT system for insulin 3 ) BIOJECTOR 2000 and PENJET for smallpox vaccination 4 ) INJEX for administration of insulin 5 ) ZENEO for administration of human growth hormone 62

63. Epidermal Powder Immunization P owder injectors were first used for DNA and RNA transfection into plants. T he technique has subsequently been investigated for transdermal protein delivery, gene therapy and vaccination. T he device design principles are similar to liquid injectors, with a powder compartment and compressed carrier gas, such as helium. upon actuation, the particles are carried by the gas, to impact the skin surface at high velocity, puncturing micron-sized holes in the epidermis to facilitate skin deposition 63

64. Advantages 1)Powder injectors offer advantages over liquids in terms of formulation and stability issues. 2)Initial safety studies suggest that the powder injectors are reasonably well tolerated, and the particle bombardment offers advantages with regard to langerhans cell targeting and immune system activation. 64

65. Colloidal Carriers T he rationale for the use of colloidal carriers is that compounds with unfavourable permeation characteristics can be packaged within carriers that will permeate the skin. 65

66. A) Nano particles and N ano c arriers Nanoparticles and micro particles are polymeric particles in the nanometer and micro metere size range respectively. Compounds can be incorporated into the particles in form of a solid dispersion or a solid solution, or bound to the particle surface by physical adsorption and chemical binding Thus allowing the particles to act as carriers or as adjuvants for the vaccine. Problems associated The general problem is that nanoparticles administered to the skin do not permeate the intact stratum corneum , but may accumulate in hair follicles. So their potential utility for passive transdermal vaccine delivery is limited. 66

67. B)Liposomes and elastic vessicles Liposomes consist of multiple bilayers of phospholipids capable of solubilising both lipophilic and hydrophilic compounds within their structure. They could act as skin permeation carriers. Evidence of their permeation across the stratum corneum intact has not emerged. But alteration of the composition including incorporation of surfactants will give elastic or deformable liposomes which are capable of deforming in shape. So they could be "squeeze through“ the stratum corneum 67

68. IV) Energy Based Approaches Exposure of the skin to energy in the form of electrical pulses or ultrasonic waves can disrupt the stratum corneum barrier and increases its permeability. This approach has been extensively investigated for drugs, macromolecules and for vaccine delivery. 68

69. A) ELECTROPORATION Electroporation involves the administration of electrical pulses to create transient pores in the skin and thus increase the skin permeability to drugs and macromolecules. Inovio biomedical corporation has developed a series of hand-held, cordless electroporation devices that have been used in vaccine delivery studies. Delivery of DNA vaccines into muscle or skin tissue with electroporation systems generated robust immune responses. 69

70. 70 Figure 19-Electroporation device

71. B)ULTRASOUND OR SONOPHORESIS Low frequency sonophoresis invo lves application of ultrasound waves at frequencies between 20 to 100khz to the skin surface to reduce the stratum corneum barrier and thereby increasing skin permeability. Pretreatment is given prior to the application of a drug solution or patch. Low frequency ultrasound (20 khz ) was used to deliver a tetanus toxoid, eliciting a robust immune response in mice to mice. A commercial ultrasound device, sonoprep , for administration of local anesthetic, was launched in2004 but withdrawn in 2007. 71

72. 72 Figure 20-Ultrasound

73. C)Micro needles Micro needles consist of pointed micro-sized projections , fabricated into arrays with up to a hundred needles, that penetrate through the stratum cornea to create microscopic holes, thus providing delivery pathways for vaccines and drugs. A number of different micro needle systems have been investigated including:- A ) Solid micro needles for permeabilizing skin via formation of micron-sized holes-solid or insoluble micro needles are generally composed of metal such as titanium or silicone . 73

74. 74 Figure 21-Microneedle

75. The micro needle array is then removed and a drug/vaccine containing patch is applied. This approach is termed poke & patch B)Solid micro needles coated with dry drug or vaccine. C) Polymeric micro needles with encapsulated drug or vaccine. Coated micro needles have an insoluble core coated with drug that dissolves off within the skin The so called "coat & poke" approach 75

76. Example of transdermal vaccine 76 Table No-4

77. References. N.K. Jain TEXTBOOK OF PHARMACEUTICAL MICROBIOLOGY. KIM A. WOODROW , 1 KAILA M. BENNETT,2,3 AND DAVID D. LO2 MUCOSAL VACCINE DESIGN AND DELIVERY 3. SARIKA NAMJOSHI AND HEATHER A.E. BENSON CURTIN HEALTH TRANSDERMAL DELIVERY OF VACCINES . Garg Neeraj , Mangal Sharad , Khambete Hemant ."Recent patents on drug delivery & formulation"; Mucosal Delivery of Vaccines: Role of Mucoadhesive /Biodegradable Polymers 2010;4,114-128 . Saroja CH, Lakshmi PK"Recent trends in vaccine delivery systems":A Review International J pharm investig 2011 apr-jun;1(2):64-74 77

78. Carino GP. Vaccine delivery. In: Mathiowitz E, editor. Encyclopedia of Controlled Drug Delivery. Vol.2. United States:Wiley Interscience ; 1999 (996 ). Controlled and novel drug delivary by N.K. Jain CBS publication. Advances In Vaccination: A Review By Swarnali Dasa ', Rohitas Deshmukh,International Journal Applied Pharmaceutics Vol 1 Issue 1, 2009 Neutra MR. Kozlowski PA. Mucosal vaccines: the promise and the challenge. NatRev Immunol 2006;6:148-58 . Mucosal delivery of vaccines: a review zara sheikh", nishat jahan , rejaul karim Zara Sheikh*et al. International Journal of Pharmacy & Technology 78

79. Vaccines Wim Jiskoot Division of Drug Delivery Technology Leiden/Amsterdam Center for Drug Research (LACDR) Leiden University The Netherlands.Mucosal Vaccine Design and Delivery Kim A. Woodrow, 1 Kaila M. Bennett,2,3 and David D. Lo2 The Annual Review of Biomedical Engineering is online at bioeng.annualreviews.org Recent advances in vaccine delivery by Soni Khyati J., Patel Rakesh P., Asari Vaishnavi M. and Prajapati Bhupendra G Journal of Applied Pharmaceutical Science 01 (01); 2011: 30-37 Bernard KW., Mallonee J., Wright JC. Pre exposure immunization with intraderma human diploid cell rabies vaccine, Risks and benefits of primary and booster vaccination. The Journal of the American Medi Asso.2005; 257(8): 1059-1063.A 79

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