Single-Dose Vaccine Carrier for Modulation of Immune Response Mechanisms Matt J. Kipper 1, Jennifer...
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Transcript of Single-Dose Vaccine Carrier for Modulation of Immune Response Mechanisms Matt J. Kipper 1, Jennifer...
Single-Dose Vaccine Carrier for Modulation of Immune Response Mechanisms
Matt J. Kipper1, Jennifer Wilson2, Michael Wannemuehler2, and Balaji Narasimhan1
1Department of Chemical Engineering, Iowa State University2Department of Veterinary Microbiology and Preventive
Medicine, Iowa State University
68a – AIChE Annual Meeting, November 9, 2004
Controlled Release Improves on Conventional Administration Schedules
Conventional• Poor patient
compliance
• Poor concentration control
• Overdose potential
Therapeutic Range
Toxicity
No Activity
Time
Con
cen
trati
on
Controlled Release• Single dose
• Tailored release kinetics
• Targeted to specific organs tissues or cells
Many Controlled Release Formulations have been Marketed
Brand Name ApplicationRelease
timeDelivery
route
Anti-tumor Weeks Implant
Allergy 1 Day Oral
Contraceptive
1 Month Injection
Anti-depressant
1 Week Oral
Polyanhydrides are Excellent Candidates for Controlled Release
Poly(Sebacic anhydride) Poly(SA)
Poly[1,6-bis (p-carboxyphenoxy)hexane] Poly(CPH)
Hydrophobic Surface erosion
Mutually incompatible
Biocompatible degradation products
-COOH
Hydrolytically labile
Narasimhan and Kipper, Adv. Chem. Eng. (2004)
Polymer Chemistry and Microstructure Affects Erosion and Release Kinetics
TimeC
um
ula
tive M
ass
R
ele
ase
dShen, Kipper, Dziadul, Lim, Narasimhan, J. Controlled Release (2002)
Tailored release kinetics
Surface Erosion
Bulk Erosion
Combined
Current Vaccine Administration Schedules are Non-Ideal• >700,000 neonatal deaths world-wide
from tetanus• Conventional injection schedules
– Many injections– Patient compliance
• Controlled release technology– Single injection– Multiple formulations
• NIH Lists Single Dose Vaccines as #1 Grand Challenge in Global Health
http://www.grandchallengesgh.org/
Controlled Release Formulations Offer Several Advantages for Vaccines
• Polyester-based (PLGA) single-dose vaccines– Protective immunity
possible w/single-dose (Corradin, O’Hagan)
– Antibody titer and isotype/subclass similar to that in alum-based systems (Corradin)
– Acidic/aqueous microenvironment reduces antigenicity (Schwendeman, Langer)
• Polyanhydrides– Hydrophobic
microenvironment– Protein
stabilization– Modulated release
kinetics– Reduced acidity– No studies with
vaccine formulations
Two Immune Response Mechanisms Offer Different Protection
DC
Pathogen
Migration to draining
lymph node
MHC II
Th1 Th1 B Cell
Secreted Antibody
MHC I
TCR
Th2 Th2Cytotoxic T Cell MHC I
Infected Cell
Humoral (Th1) response
Cellular (Th2) response
Circulation
Circulation
TCR
Research Paradigm
Hydrophobic Hydrolyzable
Surface Erosion
Polymer/APC Interactions
Controlled Release
Antigen Stabilization
Cytokine Profile
Immune Activation
Novel Adjuvants for Single-Dose Vaccines
Goal
Engineer tetanus toxoid (TT)-loaded polyanhydride microspheres and study in vivo immune response
Microspheres Fabricated by W/O/O Double Emulsion
100mg polymer in 4ml MeCl25mg protein in 100l water
Emulsify, add 4ml of silicon oil (dropwise) saturated with MeCl2
Continue emulsifying, to form outer emulsionAdd to 300ml of n-heptane, stir three hours to allow MeCl2 to evaporate
Filter, rinse, dry
Non-Porous Microspheres Provide Extended Release Kinetics
20:80 • Microspheres
incubated in 0.1M phosphate buffer (pH 7.4)
• TT antigen concentration determined by BCA assay
50:50
• 3 C3He/OuJ mice per group
• IM injection (right quadriceps)
• Unencapsulated TT and blank 20:80 CPH:SA microspheres
• 50% cottonseed oil/saline emulsion
• ELISA for TT-specific IgG at 1:400 dilution
Polyanhydride Microspheres Induce Dose-Dependent Inhibition
Group
Polymer
TT
I 3mg 3g
II 1mg 3g
III 0.5mg 3g
IV 3mg 3g (day 3)
V 3mg 3g (opposite leg)
VI none 3g
0
0.5
1
1.5
2
2.5
1
Optical Denisty (A.U.)
I II III IV V VI
TT-Loaded Microspheres Provide Immunity
• 5 C3He/OuJ mice per group• Injected IM (right quadriceps) with 2%
loaded TT-loaded microspheres (0.5mg) and/or bolus of unencapsulated TT (0.5g)
• Bled weekly from saphenous vein• TT-specific IgG antibody titer
determined by ELISA
TT-Loaded Microspheres Provide Immunity
+ Bolus only
Blank microspheres
Blank plus bolus
TT microspheres
TT microspheres plus bolus
x Equivalent dose of TT
50:50 CPH:SA
20:80 CPH:SA
20:80TT Microspheres Provide High-Avidity Antibody
• 10–week serum samples tested by ELISA
• Sodium thiocyanate dissociates antibody from TT
• Avidity index is maximum molarity of NaSCN that results in <50% dissociation in 20-minute incubation
20:80 CPH:SA Microspheres Provide High-Avidity Antibody
20:80 CPH:SA provides high titer and high avidity protective immunity
0
1
2
3
VIII IX X XII XIII XIV XV
Avidity Index
VIII – 20:80 blank w/bolus
IX – 20:80TT
X – 20:80TT w/bolus
XII – 50:50 blank w/bolus
XIII – 50:50TT
XIV – 50:50TT w/bolus
XV – Equivalent TT dose
Immune Response Pathway can be Tuned by Microsphere Formulation
•IgG1 and IGg2a isotypes determined by ELISA
•Ratio of OD at 1:400 dilution
•IgG2a = Th1
•IgG1 = Th2
0
0.2
0.4
0.6
0.8
1
1.2
IX X XIII XIV Eq.Dose
Fraction of IgG1 + IgG2a titer
IgG2a
IgG1
20:80 50:50
How do 20:80 Microspheres Mediate Immune Response?
Immunogen-loaded
microspheres
DC
Migration to lymph node delayed by hydrophobic
adjuvant
Inflammatory cues wane
Antigen-driven (TH2) response
Microspheres degrade, releasing antigen
DC Migration to lymph node not
delayed
Antigen-driven (TH2) response
Unencapsulated immunogen
DCsMigration to
lymph node not delayed
Inflammatory cytokine context
results in balanced responseIL-12
Summary• Low polymer doses provide adjuvant effect• TT-Loaded microspheres provide immunity
– Preserved antigenicity– Sustained exposure to antigen provides
secondary immune response– Antibody titers and avidity show efficacy in
single-dose formulation
• Immune response mechanism can be modulated by altering formulation