: 2 15 - 17 2553

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: 2 15 - 17 2553

: ., PhD.

: MSc.

: MSc. ., PhD. MSc. MSc. MSc.

ISBN : 978-616-11-0706-2 : 500 : : 4 2

. . . . 11000 0 2590 3196-9 0 2965 9152http://www.nvco.go.th

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.. . Scientific Affairs Manager Bionet-Asia Co., Ltd.

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2 15-17 2553 7

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Dr.Stephen J. ThomasDepartment of Virology,US Army Medical Corps Commander-ArmedForces Research Institute of Medical Sciences(USAMC-AFRIMS)

: 8

3 4 5 61. Key note lecture : Vaccine development in Developing world, 11

past, present and future: SEAR perspective.

2. History of vaccine development 16. .

3. Vaccine Development and Protective Immunity 30..

4. New Paradigm in Immunology: A Personal Perspective 48. ..

5. New Vaccine Design 71...

6. From Pre-clinical Research to Vaccine Development: Examples of 81Go-no-go Decisions - a case study of rotavirus vaccine...

7. (Essential Practical Aspects of 94Conducting Clinical Trials)Dr. Stephen J. Thomas

8. Pandemic threat : from policy to implementation 102. .

2 15-17 2553 9

9. Round table discussion: H1N1 development, lesson learnt in Thailand 115Moderator: . Discussants: .

. .. 10. Master donor strain 125

... 11. Registration preparedness for new vaccine licensing 139

. 12. Round table debate: Regulatory and Intellectual property barriers 156

e.g. new adjuvants, gene transfer productModerator: .. Discussants: . ..

... 13. HIV vaccine trial: Why Thailand?, Is it a true success story? 170

. 14. Light up the candle in the dark: HIV Vaccine Hope 177

Moderator: . : HIV Vaccine: New Design and the Challenges 178 .. : Community view on HIV vaccine development 191 : Clinical trial and public health benefit 194 .

15. Role of Cell culture technology in new vaccine development 201..

: 10

16. The challenges of developing new tuberculosis vaccines 216..

17. G(M)...P : Standard VS barriers 249Moderator: . Discussants: . . .

1: . 259

2: . 260 3: 2 262

2 15-17 2553 11

Vaccine development in developing world, past,present and future: SEAR perspective

2

.

cost effectiveness 1980-1990 (Fully immunization) 15 30 26

: 12

1.7 1 disease burden 7 1980 81 1.4 90 25 1990 (UN) 1

(UNICEF)

tradi-tional vaccine traditional vac-cine (Combination vac-cine) Hib pen-tavalent (DTP) base DTP traditional vaccine traditional vaccine

DPaT(Acellular pertussis)

2 15-17 2553 13

DPT (Whole-cell pertussis)

HPV 3-5 HPV

traditional vaccine 2000-2010 10 14 traditional vaccine UNICEF

2010 15 21 82 12 88 18

: 14

secondaryprevention primary prevention

GMP GMP

Developing Country Vaccine Manufac-turers Network: DCVMN 28 1.7 promising country DTP 80 DCVMN traditional vaccine 2 3 1 2000 International Finance Facility forImmunization (IFF), International VaccineInstitute (IVI), Malaria Vaccine Initiative

2 15-17 2553 15

oral chlorella, rota virus, anthrax,pneumococcal, meningococcal, measlesaerosol H1N1 pre-clinical trial H1N1 H1N1, Dengue, H5N1 phase IIIclinical trial HIV vaccine

technologytransfer patent intellectual property right

NationalRegulatory Authority

(PublicPrivate Partnership: PPP)

: 16

History of vaccine development

. .

2519

( 1)

1

3 Vaccines

4

2550

2

2525 2548 ( 2) 2550 Stanley Plotkin, Walter Orenstein PaulOffit Vaccines ( 3)

2 15-17 2553 17

2550 ( 4)

(Lymphocyte) 2 Bone marrow-derived lymphocyte (B-cell) (Humoral immune re-sponse) Thymus-derived lymphocytes (T-cell)

26 2522

(Smallpox) 2 2520

(Cell-mediated immune response)

Anthrax, Bacterial meningitis, Chickenpox,Cholera, Diphther ia, Haemophi lusInfluenzae type b, Hepatitis A, HepatitisB, Human Papilloma, Influenza, JapaneseEncephalitis, Measles, Mumps, Pertussis,Pneumococcal pneumonia, Polio, Rabies,Rotavirus Diarrhea, Rubella, Tetanus,Typhoid Fever Yellow fever Bordettlapertussis 2449

: 18

2491 42 Polio virus 2451 2498 47 100 Sallmonella typhi

1

2427 2532 105 Mycobacterium tuberculosis 126 1

2520 2535

(Herd immunity)

2 15-17 2553 19

(Basic research) 1, 2 3

1 ()

2 ()

3

Pneumocystis 5 5 2524 HIV

3 2546 24 2552

Times Magazine 8 The 50 Best Inventions ofthe Year 5

100

(King of Pontus)

(Variolation) 2264 Lady Mary Wortley Montagu

Anton van Leeuwenhoek ( 2175-

2266) ( 5)

: 20

Lazzaro Spallanzani ( 2272-2342)( 6) biogenesis

Edward Jenner ( 7) 2339 (Cowpox-milkers node) 6 vacci-nation vacca vaccine

Louis Pasteur ( 2365-2438) ( 8) (Vaccinology) (Attenuation)

Robert Koch ( 2386-2453) ( 9) (Anthrax) Bacillus anthracis

Elie Metchnikoff ( 2388-2459) (10) phagocytes

Walter Reed ( 2394-2445) ( 11) (Yellow fever)

Emile Roux ( 2396-2476) ( 12)

Emil von Behring ( 2397-2460) ( 13)

Maurice R. Hilleman, Ph.D. ( 14) .. 2545 36 (Japanese Encephalitis B), (Mumps), (Measles), (Rubella), (MMR) (Jeryl Lynn) 8 1 . 2

2 15-17 2553 21

5Anton van Leeuwenhoek

6Lazzaro Spallanzani

7Edward Jenner

8Louis Pasteur

9Robert Koch

10Elie Metchnikoff

11Walter Reed

12Emile Roux

13Emil von Behring

14Maurice R. Hilleman, Ph.D.

: 22

18

1798 Smallpox 19

1885 Rabies1897 Plague

201917 Cholera Typhoid vaccine (parenteral)1923 Diphtheria1926 Pertussis1927 Tuberculosis (BCG) Tetanus1935 Yellow Fever1940 DTP1945 influenza vaccines (flu) 1955 Inactivated polio vaccine (IPV) Tetanus

diphtheria toxoids1959 World Health Assembly 1961 Monovalent oral polio vaccine 1963 Trivalent oral polio vaccine (OPV) Measles vaccine 1964 Advisory Committee on Immunization Practices (ACIP)

CDC 1964-1965 Congenital Rubella

Syndrome 20,000 1966 1967 Mumps vaccine

2

2 3

2 15-17 2553 23

1969 Rubella vaccine

57,600 1970 Anthrax vaccine The Michigan Department of Public Health1971 Measles, Mumps, Rubella vaccine

(MMR) 1976 Guillain-Barre syndrome Swine flu1977 1978 Fluzone Aventis pasteur 1979 wild virus 1980 1981 Meningococcal polysaccharide vaccine, groups A,

C, Y, W135 (Menomune)1982 Hepatitis B vaccine1983 Pneumococcal vaccine, 23valent1986 The National Childhood Vaccine Injury Act

1988 WHO, UNICEF,Rotary International, CDC

1989-1991 55,000 1983 1,497 MMR 2

1990 Typhoid vaccine (oral) Haemophilus influenzae type B (Hib)polysaccharide conjugate vaccine

1991 Hepatitis B vaccine Acellularpertussis vaccine (DTaP) 15 6

1993 Japanese encephalitis vaccine1994 Vaccines

for Children (VFC) program 1995 Advisory Committee on Immunization Practices (ACIP) First

harmonized childhood immunization schedule the American Academy of Family Physicians and the American Academyof Pediatrics

: 24

1995 Hepatitis A vaccine Varicella vaccine

Varicella vaccine 4 1996 Acellular pertussis vaccine (DTaP) 1998 First rotavirus vaccine 1999 Rotavirus vaccine Lyme

disease vaccine (FDA) FDA thimerosal

212000 800,000

endemic area Pneumococcal conjugate vaccine (Prevnar)

2001 11

2002 Lyme disease vaccine

2003 endemic area First live attenuated influenza vaccine (FluMist) 5-49

2004 Inactivated influenza vaccine 6-23 DTaP, IPV, Hep B vaccines Pediarix

2005 endemic area Tdap vaccines Boostrix and Adacel Meningococcal vaccine Menactra 11-55

2006 Rotavirus vaccine RotaTeq MerckMMR vaccine Varivax vaccines (measles, mumps, rubella chicken pox) 1 HPV vaccine Gardasil

2007 Chickenpox vaccine Varivax booster dose Flu vaccine, Flumist 2

2 15-17 2553 25

Bacillus anthracis Anthrax InactivatedCoxiella burnetii Q fever InactivatedHepatitis B Liver disease, cancer InactivatedPoliovirus Poliomyelitis, paralysis Inactivated/attenuatedSalmonella typhi Typhoid fever InactivatedYersinia pestis Plague InactivatedBordetella pertussis Whooping cough InactivatedHepatitis A Liver disease InactivatedInfluenzavirus Respiratory disease InactivatedJapanese encephalitis virus Brain infection InactivatedRabies virus Rabies InactivatedVibrio cholerae Cholera InactivatedAdenovirus Respiratory disease Live attenuatedInfluenzavirus A Respiratory disease Live attenuatedVibrio cholerae Cholera Live attenuatedMycobacterium tuberculosis Tuberculosis Live attenuatedMeasles virus Respiratory tract infection, SSPE Live attenuatedMumps Virus Mumps, meningitis, orchitis Live attenuatedPoliovirus Poliomyelitis, paralysis Live attenuatedRubella virus German measles, fetal malformations Live attenuatedSalmonella typhi Typhoid fever Live attenuatedVaccinia virus Smallpox Live attenuated

3

2008

Rotavirus vaccine, Rotarix 2 DTaP, IPV, Hib Pentacel DTaP IPV Kinrix 4-6

: 26

Varicella-zoster virus Chickenpox Live attenuatedYellow fever virus Jaundice, kidney and liver failure Live attenuatedHaemophilus Meningitis, epiglottitis, Polysaccharide (conjugated)

pneumonia type bNeisseria meningitidis Meningitis Polysaccharide (conjugated)Streptococcus pneumoniae Pneumonia, otitis media, Polysaccharide (conjugated)

meningitisSalmonella typhi Typhoid fever PolysaccharideStreptococcus pneumoniae Pneumonia, otitis media, Polysaccharide

meningitisBorrelia burgdorferi Lyme disease SubunitBordetella pertussis Whooping cough SubunitHepatitis B Liver disease, cancer SubunitInfluenzavirus A Respiratory disease SubunitClostridium tetani Tetanus ToxoidCorynebacterium diphtheriae Diphtheria Toxoid

2476 Smith, Andrewes, Laidlaw (Nasal washings) (Ferrets) Influenza A

2478 Wilson Smith (Embryo-nated eggs) WSWstrain

2477-2478 ThomasFrancis Jr. Frank

2 15-17 2553 27

Macfarlane Burnet 2483Thomas Francis Jr. Influenza B A B

2483-2485 Frank MacfarlaneBurnet (Allantoic sac) (Amniotic sac) Burnet

George Hirst Rockefeller Institutes, New York Hemagglutination Hemagglutinins

Hemagglutinating unit

2482-2488 Hemagglutinins (Formalin Betapropiolactone) 80 whole cellinactivated influenza vaccine splitvirus vaccine subunit vaccine

2486 2490 A 2491 A1 A prime strains

Taylor RM influenza C

: 28

A, B C A B C

2500 flu pandemic variance A A2 Asian virus A H2N2 A A1

2503 Himmelweit Medical Research Council Influenza Vaccine Committee

2512 Glezen Zonal centrifugation

30 2 0.2 . 6 0.5 . 8

2510 (Sharples centrifugation; zonalcentrifugation) (Chemical disruption - split virus vaccine) 2513 Davenport subunit process subunit vaccine 2525

A strain H1N1,H3N2 B TrivalentInactivated Vaccine TIV killed vaccine

2 live attenuatedvaccine killed vaccine

Live attenuated vaccine

Killed vaccine 3

- Whole virus vaccine

2 15-17 2553 29

- Split virus vaccine

-Subunit vaccine haemagglutinin (HA) neuraminidase (NA)

adjuvant subunit vaccine immunogenicity adjuvant synergic effect

Salk Alacel A mineraloil water in oil adjuvant

adjuvant Aluminium hyroxide gel, Aluminiummonostearate, Mannite mono-oleate,Arlacel - vegetable oil (A-65), BayolF - vegetable oil, Peanut oil, Drakreol,MF 59, AS03, AS04

: 30

Vaccine Development and Protective Immunity

..

(Preventivevaccine) (Therapeuticvaccine) (Autoimmune disease) degenerative diseases

(Sterilization)

1) (Eradication)

2) carrier latent infection

3) delay disease progression 2) 3)

chronic infectious disease HIV vaccine slow disease progression

Immunology antigen specific immune

response 1. Humoral immune response

B cell B cell differentiate plasma cell neutralization

2 15-17 2553 31

complement-killing, antibody-dependent cell-mediated cytotoxicity(ADCC) -killing epitope receptor NKcell NK cell antibodydependent cell-mediated viral inhibition(ADCVI) ADCC neutralize (NK cell) viralreplication

2. Cell-mediated immune response T cell CD4+ CD8+ CD4+ helper cell B cell CD8+

macroph-age monocyte granulomatous in-flammation CD8+ CTL-kill-ing cytotoxic T lymphocyte CD8+

cytok ine/chemok ine viral replication Non-lyticinhibition

Neutralizing antibodyProf. Vincent Racaniello

neutralization mechanism 1

1. aggregation receptor

2. attachment fusion

3. endocytosis uncoding

immunogenicity in vitro neutralizing assay in vivo good correlation vaccine protection

1. Cell line in vitro in vivo

2. Biological functions compli-ment ADCC Fc por-tion in vitro

: 32

Specific Cell-Mediated Immune Re-sponse

cellmediated immune response antigen presentation dendritic cell antigen presenting cell T cell CD4+ MHC Class II CD8+ MHC Class I interaction antigen presenting cell T cell T helper cell

Nature of Disease and VaccineDevelopment

1 () ()

identify protective immuneresponse

- acute infection small-pox, polio, heppatitis A influenza

- (Latent) / (Chronic) hepatitis B, TB, malaria

- HIV

specific immunity response (Acquiredimmunity) humoral

2 15-17 2553 33

cellmediated immune response regulatory T (Treg) cells hu-moral immune response neutralizingantibody live vaccine cell-mediated humoralimmune response kill sub-unit vaccines humoral immuneresponse

protective immunity (Assess clinical safetyand efficacy) (Smallpox vaccine) (BCG)

identify protective immunity target antigen

3 1. Basic research

basic science Virology, Immu-nology, Pathology Epidemiology

2. Targeted Research in vitro in vivo pre-clinical trial immunogenicity (Clinical trial) 1, 2 3

3. Product Development

10 clinical trial

: 34

Vaccine Years to developTyphoid 110H. influenzae 97Pertussis 94Polio 52Measles 47HBV 21HPV 30

1 *

* Markel H. NEJM 2005; 353:753-57

2

2

1

live attenuated / recombinantantigen

2 15-17 2553 35

(Human vac-cine)

1. Live attenuated vaccine Measles, Mumps, Rubella, Polio (Sabin),Rotavirus, Tuberculosis (BCG), Varicella Yellow fever

2. Killed vaccine Cholera, Influ-enza, HAV, Plague, Polio (Salk) Ra-bies

3. Toxoid Diphtheria Teta-nus

4. Subunit HBV, Pertussis Streptococcal pneumonia

5. Conjugate vaccine Hemophillus influenzae type b (Hib) Streptococcal pneumonia

6. DNA7. Recombinant vector DNA recom-

binant vector

(US FDA) 22 13 9 2 3

2 US FDA

: 36

2 3 cocktail combine vaccine

, , -- (DPT), (OPV), - (dT),-- (MMR)

3 US FDA

Identify Protective Immunity

identify protective immunity protective immunity antigen specific immune re-sponse humoral-mediated cell-mediated immunity (Progressive disease) strongcorrelate of protection animal

2 15-17 2553 37

model active immunization passive immunization

neutralizing an-tibody protectiveimmunity cell mediated killing cyto-toxic T lymphocyte, antibody-dependentcell-mediated cytotoxicity (ADCC)

antibody dependent cell mediated viralinhibition (ADCVI) protective immunity cell-associated ()

neutralizingmediated protection cut-off level protectiveantibody seroprotective level 4

4 neutralizing mediated protection seroprotective level

1) Dagan et al. Pediatr Infect Dis J. 2007 Sep; 26(9): 787-93 2) McCullers et al. CID 2010; 50: 1487-1492, Rose et al. Clin. Diagn. Lab. Immunol 2005, Pichichero et al. J Pediatr 2007; 151: 43-9, Barton et al.

Vaccine Cut-off levelHAV 20 mIU/mLDiphtheria 0.1 IU/mLTetanus 0.1 IU/mLPertussis FHA 2 FDA unitsH. influenzae type b 0.15 g/mLPoliomyelitis titer 1: 4 to 1: 8Measles 300 mIU/mLMumps 1: 230Rubella 10 IU/mLInfluenza HAI antibody titer of 1: 40Pneumococcus Ab 1.0 g/ml (other study 0.05) and opsonophagocytic titer of >1: 64Varicella 5 gpEUSmallpox HAI titer >1: 10 (suggested)

: 38

1

3 .. 2339 Dr. Edward Jenner vacca Dr. Jenner (Cowpox) royalsociety 2 Dr. Jenner

Protective immunity neutralizing antibody 95 develop neu-tralizing antibody hemagglutination (HA)inhibition antibody titer 1: 10 ( cut-off antibody

level) 10 pre- post-exposure post-exposure 7 protection rate protective immunity major reaction (Global eradication) .. 2520 ..2523

3 types 1, 2 3

90-92 (Asymptomatic) 4-8 0.1-2 acute flaccidparalysis acute flaccid paralysis

Poliovirus protective immunity neutralizing antibody seroprotectivetiter 1: 4 - 1: 8 .. 2508 poliovirus neutralizing

2 15-17 2553 39

antibody assay control neutralizing antibody , viral stocks, cell line monkey kidney cell strain LLCMK2,micro-culture system, immune serum identify cytopathogeniceffect (CPE)

antibody response 3 serotypes (IPV) 2 3 antibody response 90 100 Bulletin of the WHO .. 2539

3 3 1) OPV 4 2) OPV 4 IPV 3 3) IPV 3 3 IPV response 1 2 60 OPV seropositive 100 pharmacogenomic, immunogenetic / 3 multi-national study

3 Seroprevalence 1, 2 3 24 (10 )

: 40

globalization

Human Papillomavirus

HPV ..2549 30 .. 2519 HPV (CA cervix) papillomavirus

papillomavirus 200 (HPV-linked genitalcancer)

1) highest risk 16, 18,31 45

2) Other high-risk 33,

35, 39, 51, 52, 56, 58 59 3) Probably high-risk

26, 53, 66, 68, 73 82Gardasil

HPV Merck (US-FDA) 8 .. 2549 Virology, Immunology,Pathology Epidemiology clinical study design

identify neutral-izing epitope structural pro-tein L1 pentamer 4 5 neutralizing antibody (B ind ing) v i ra l par t ic le Papillomavirus Merck viral like particle (VLP) bioengineer L1 5 pentamer VLP

immunoge-nicity protection neutralizing antibody passive transfer IgG

2 15-17 2553 41

4 Papillomavirus neutralizing epitope L1 pentamer

5 neutralizing antibody () Papillomavirus particle

: 42

/ (Mucosal protection) serum IgG (Transudation) (Inflamma-tion) (Exudate) (Local protection) mucosal immunity response

HPV 16 18 genital cancer 70 analcancer genital cancer 70 6 11 90 genital warts 6, 11,16 18 tetrava-lent VLP (End point)

end point endpoint initial HPV infection70 - 90 continuous infection 1 - 3 earlycancer change (Sign) in situ neoplasia classiccervical cancer 20 clinical trial primary endpoint cervical intraepithelial neoplasia 2 3 (CIN2/3) Adenocarsinoma in situ (AIS) CIN 2/3 AIS genitalwarts clinical marker HPV in-fection 16 - 26 bridging 9 -15 clinical trial 16 - 26 9 - 15

2 15-17 2553 43

pathologist diagnosis war t s cerv ica lintraepithelial neoplasia (CIN) adenocarsinoma in situ (AIS) tissue section multiplexPCR HPV vaccine positive specifictype early cancer positive case multinationalstudy 33 5 27,004

4 protocols Protocols 005, 007, 013 015 HPV 9,342 placebo 9,400 end point HPV 16/18 early cancerplacebo 52 positive case 100 3 GSK

US-FDA .. 2552 recombinant protein ASO4 adjuvant

Unmet-needed Vaccines /

, ,,

BCG .. 2464

90 Tuberculosis (TB) vaccine bovinevaccine antigen TB eff icacy effectiveness pulmonary TB BCG severe TB form TBmeningitis, milliary TB (Cross protection) (Leprosy) 2 BCG 100 TB

: 44

protective immunity Mycobacterium tuberculosis Th1 response TB 10 phase IIB Oxford MVA85A modified vaccinia ankara vector (MVA)base vaccine phase III TB prevent infection,prevent disease, prevent reactivation, shortcourse improve response chemo-therapy (Adherence) (Drug resistance)

Journal of Infec-tious Diseases .. 2551 Oxford groupvaccine (Immu-nogenicity) TB 85A MVA (Modifiedvaccinia ankara) live vector TB neutralizing base

vaccine immunogenic-ity Gamma interferon (IFN-)ELISPOT assay ( T cell base assay) T cell IFN- 7 immunogenicity 6

new TB vaccine rBCG30 recombinant BCGvaccine TB BCG 85b 30 kD (Over expressing) protective immunity guinea pig BCG rBCG 30 BCG guinea pig (Uninfected) (Control) guinea pig rBCG 30 guinea pig BCG 7 rBCG30 CD4 CD8 BCG ( 85b specificgamma inteferon response BCG)

2 15-17 2553 45

6 MVA85A IFN- ELISPOT assay

7 guinea pig rBCG 30 BCG uninfected guinea pig

: 46

DNA vaccine

DNA vaccine 1 3

protective immunity humoralbased vaccine (Neutralizing antibodybased vaccine) (Targetgene) (Target protein) 4 prM, E,NS1 NS3 prM E structural protein DNA vaccine plasmid/vector express prM/E 4serotype codonoptimization express

1 - 4 neutralizing antibody recombinant DNA express prM/E ( se-rotype 2) dose dependent needle free injection 100mg titer 1,280 10 vectorpCMV titer 640 vector NationalCancer Institute: NCI National Insti-tutes of Health: NIH

DNA vaccine (Delivery system) in vivo electroporation, needlefree, (Intradermal) (Intramuscular)

chimeric vaccine multi-national study IIB 5

2 15-17 2553 47

22 13 9 BCG protective immunity neutralizing antibody neutralizing antibody

T cel l base vaccine

5

BCG T-cell base vaccine TB

new vaccine de-signs new adjuvant

: 48

New Paradigm in Immunology: A Personal Perspective

. ..

Immunol-

ogy Vaccinology Immunology

HistoryAdaptive immunity

1960

: Proc Jpn Acad, Ser B 85: 143, 2009

adaptive immunity innate immu-nity 1996 receptor adaptive immunity innate immunity innateimmunity adaptive immunity innate immu-nity adaptive immunity 1

1 innate adaptive immunity

2 15-17 2553 49

Innate and adaptive immunityInnate adaptive immunity

innateimmuni ty adaptive immunity pathogen innate immune system neutrophil, NK cell,macrophage antigen presenting cells( dendritic cell) (Interact) pathogen lymph node naive B cell naive T cell adaptive immune system naive B cell naive T cell differentiate antibody T cell

innate immune system innate immune

system 2 pathogen associated molecular patterns pattern recognition receptors

- Pathogen associated molecularpatterns (PAMPs) lipopolysaccharide

(LPS) doublestranded RNA

- Pattern recognition receptors(PRRs) receptors innate cell dendritic cell macrophages

pattern contact antigenpresenting cell (APC) naive T cell adaptive immune active Th1 Th2 1 innate immune innate immune patternrecognition receptors innate cell adaptive immune Th17, Th19, Th22, TFH regulatory T cell

innate immunity innate-like cell sub-population B cell T cell B1, MZB, NKT dgT adaptive immune

: 50

innate immunity immuneregulation

1959 innate adap-tive Science lymphocyte receptor self non-self ( B cell) recep-tor B cell signal 1 (Sig 1) B cell (Proliferation) 2a 1969 receptor Sig 1 B cell helpersignal (Sig 2) T cell 2b

: Nat Rev Immunol 1:135, 2001

1 PAMP PRR

1975 Tcell B cell co-stimulation signal (Sig 2) antigen presenting cell Sign 1 B cell Sig 1 T cell 2c

1989 immune regulation Charles A. Janeway antigenpresenting cell T cell potentialreceptor surface antigenpresenting cell microbe infectious non-self antigen presenting cell self non-self 2d antigen presenting

2 15-17 2553 51

cell phagocyte self non-self immune regulation PollyMatzinger ligand antigenpresenting cell

damage 2e

Stranger model of immune activation antigen presenting cell

interact microbe pathogen-asso-

2 Immune regulation antigen presenting cell T cell B cell

: Science 296: 301, 2002

: 52

ciated molecular patterns (PAMPs) lipopolysaccharide, peptidoglycan, singlestranded RNA double stranded RNA PAMPs pattern recogni-tion receptor antigen present-ing cell antigen presenting cell regional lymphnode T cell B cell 3

Pattern recognition receptors (PRRs) antigen presenting cell PAMPs mi-

3 Stranger model of immune activation

crobe lipopolysac-charide , double-stranded RNA receptors toll-like receptors (TLRs) 4

Toll-like receptors Toll receptor mammalian TLRs TLRs interact PAMPs microbe signal (Turn

: Nat.Rev. Immunol. 8:279, 2008.

2 15-17 2553 53

4 Types and location of microbial sensors (Pattern-RecognitionReceptors: PRRs)

: Cell 124: 823, 2006

5 General components of innate immune receptor

: 54

on) anti-microbial peptide proinflammatorycytokine

antigenpresenting cell/macrophage non-toll-like receptors (non-TLRs) C-type lectin (CLEC) endocyticreceptor endocytosis microbe microbe T cell

5 diagram Toll-like receptors extracellular unit (Recognition unit) leucine-rich repeat receptor signal signal kinase (kinase network) transcrip-tion factor

transcription factor 5 NF-KB (NF-kappaB) activation antimicrobial peptides,enzyme, inflammatory cytokines (IL-8,

TNF-), apoptosis adhesion mol-ecules

Toll-like receptors adapter 1 activation pathway pathway effector components

Pathway activation path-way 2 pathways 6

1. MyD88 (Myeloid differentiationprimary response gene 88) dependentpathway proinflammatory cytokines IL-1,TNF-

2. MyD88 independent pathway interferon-

Toll-like receptors (TLRs) MyD88-dependent pathway inflammatory cytokine TLR3 doublestranded RNA TLR4 adapters MyD88-independent path-way interferon-b production interferontype I antiviral (Immunomoduratingactivity)

2 15-17 2553 55

Danger model of immune activation(New paradigm in Immunology)

1989 antigenpresenting cell self non-self 1994( 2e) Danger model antigen presentingcell microbe PAMPs antigenpresenting cell receptor alarm signal

: Nature Reviews Immunology 4: 499, 2004

6 TLR signaling MyD88-dependent () MyD88-independent ()

stress ( physical stress / biologi-cal stress) infection host cell components host cell components antigen presentingcell 7

(Paradigm) antigen presenting cell host cell

: 56

components damaged tissue non-infectious self antigen presenting cell receptor infectiousagent host cell components infectious agent exogenouscomponents host cell components endogenous components

innate immune system PAMPs PRRs Damage associated mo-

7 Danger model of immune activation

lecular pattern molecules (DAMPs) endogenous components dangersignal host cell

- TNF-a,- IL-1- ATP- HSP (Heat shock proteins)

cytoplasm- Proteases- Necrosis/Apoptosis- Uric acid crystal

: Nat.Rev. Immunol. 8: 279, 2008.

2 15-17 2553 57

inflammatory cytokines (Gout)

- Cholesterol crystal inflammation (Atherosclerosis)

- HMGB1 (High mobiloty group box1) nuclear protein stress

- Oxidized LDL (Low densitylipoprotein) infectious agent oxidized LDL oxidized LDL receptor macrophage

- Advanced glycation end prod-ucts () ligand receptor antigenpresenting cell

(Apoptosis) 8 necrosis DAMPs antigen presenting cell/macroph-age proinflammatory cytokines necrosis death DAMPs 9

1) internal cylosolic compo-nent

2) internal component complement system complement receptor sur-face macrophage

3) necrotic cell protease extracellular matrix extracellularmatrix recep-tor receptor sig-nal

macrophage receptor DAMPs

8 living cell,apoptosis necrosis receptor innate immune cell

: at Rev Immunol 8:279, 2008

: 58

10 receptor RAGE (Receptor for advanced glycationend product) HMGB1 receptor receptor oxidized LDL

9 inflammation necrotic cell

: Nat Rev Immunol 8: 279, 2008

turn on inflammatory cytokine inflammation receptors TLR4,TLR9 TRL7 receptors microbe DAMPs heat shock protein, hostcell DNA, host cell RNA

2 15-17 2553 59

Host component TLRs TLRs microbe endog-enous ligands host HSP60,HSP70 Gp96 (96 kDa glycoprotein ofthe endoplasmic recticulum) TLR2 TLR2 peptidoglycan TLR signal

10 receptors innate immune cell DAMPs PAMPs

necrosis oxidized LDL, amyloid-b, oxidized phos-pholipid b-defensin-2

binding & signaling ofself ligands to TLR

tissue damage cell host DNA (Self DNA), HMG-B1 LL37 cathelicidins white blood cell

: Cell 124: 823, 2006

: 60

self DNA, HMG-B1 LL37 complex TLR9 endosome endocytosis complex RAGE

HMG-B1 complex anti-DNA antibody immunecomplex Fc receptor 11

11 TLR self ligands

: Cell Host Microbe 3: 352, 2008

Endogenous components exog-enous components

TLRs en-dogenous components exogenouscomponents components TLRs co-receptor

modulate signal exogenous components microbe lipopolysaccharide TLRs receptor signal inflammatory cytokinegene inflammation tissue repair

2 15-17 2553 61

: Science 327: 291, 2010.

adaptive immune recep-tor endogenous ligands signal signal signal exogenous components

12 TLR recognize interact exogenous (Microbe) endogenous (self) ligands

TLR endogenouscomponents inflamma-tory response tissue repair adaptive immune 12

Co-receptor

- PAMPs microbe TLR inflammation pathogenelimination, collateral tissue damage adaptive immune system

- DAMPs TLR limitation of tissue damage tissue

reconstruction DAMPs co-receptor co-receptor immunosuppressive signal signal signal DAMPs co-receptor 2 CD24 siglec G/10 transcriptionfactor SHP-1 SHP-1 inflammatory cytokine 13

: 62

innate immune adaptive immune

14 antigenpresenting cell signal TLR c type lectin receptor non-TLRs naiveT cell signal 1 PAMPs TLR co-stimulation molecule signal 2 T cell signal 2 signal T cell

13 inflammatory cytokines PAMP DAMP

signal signal 3 differentiation T cell T cell Th1 Th2 cytokine microenvironment IL-12 antigen presenting cell activated T cell Th1 IL-4 () activated T cell Th2 Th signal 3 14

: Trends Immunol. 30: 557, 2009

2 15-17 2553 63

14 activated T cell signal 3 innate immune

cytokines interleukin IL-23, IL-6, IL-1 TGF-b IL-12 (Stimuli), microbe, microenvironment antigen presenting cell interleukin activated T cell Th17 IL-17 chemokine polymorphnuclear leucocyte (PMN) (Chemotaxis) 15 IL-17 fibroblast fibroblast ligand

: Nat. Rev. Immunol. 1: 135, 2001

(Osteoclast) (Rheumatoid arthrit is) (Bone loss)

pathogenesis regulation newstrategy vaccine development antigen presenting cell IL-23 TGF- Regulatory T (Treg) cells Th CTLs infected cell

: 64

: Nat. Rev. Immunol 10: 159, 2010

15 Th 17 signal IL-23, IL-6, IL-1 TGF-b innate immune

16 Differentiation CD4+ T cell

: Nat. Rev. Immunol. 1: 135, 2001

2 15-17 2553 65

naive T cell differentiate Th cytokine 16 model model cytokine subpopulation Th cell Th9, Th22 TFH

cytokine modulation antigen presenting cell activation

1) subpopulations dendritic cell

2) maturation stage dendriticcell dendritic cell mature

3) recognition signal receptor

4) microenvironment

microen-vironment mucosal immunity epithil ial cell maturation stage dendritic cell microenvironment epithilial cell aller-gen microbe Thymic stromal lymphopoietin (TSLP) dendritic cell

dendritic cell Tcell Th2 17

signaling PAMP TLR cytokine 18 () IL-1 DAMP/infected cell viral infected cell apoptotic death type I interferon (type 1 IFN) type 1IFN dendritic cell TLR PAMP cytokine signal 18 () TLR innateimmune cell inf lammatoryresponse infected dead cell type 1 IFN ( 18 ) innate immunity

innate immunity surface microbial detector receptors innate immune cell TLR, non-TLR receptors cytoplasmic receptor cytoplasm microbial detector intracellular

: 66

: Science 327: 29, 2010

17 Central role of TSLP in driving DC maturation for Th2-cell response

: Nat. Rev. Immunol.8: 193, 2008

18 Cell-extrinsic recognition PAMP () infected dead cell ()

2 15-17 2553 67

organisms microbe

19 endosomal pathways cytoso-lic pathways endosomal pathway microbe, apoptotic cell intrac-ellular parasite endocytosis endosomal TLR (TLR3, TLR7, TLR8 TLR9 membrane-associated) proinflammatory cytokines type I IFN

19 endosomal cytosolic pathway

: Nat Immunol 11: 367, 2010

cytosolic pathways receptors 2 subgroup

- RLR (RIG-I-like receptors) viral nucleic acid IFN-b, IFN-a (Type I IFN) proinflammatory cytokines

- NLR (NOD-like receptors) 20 Nod1 Nod2

Nod1 Nod2 pepti-doglycan

: 68

cytoplasm Nod1 Nod2proIL-1 proIL-18 receptors receptor inflammasomes caspase I proIL-1 mature IL-1 inflammation (inflammatory response) intracel lularbacterial infection viral infection

receptor AIM2 (absent in melanoma 2) intracytoplasmic DNA receptor CpG TLR9 AIM2 main receptor TLR9 CpG cytoplasm

Receptor NALP3inflammasome receptor microbe toxin signal host

ATP, uric acid crystals uricacid crystals NALP3 NALP3 caspase I pro-IL 1 pro-IL18 IL 1 IL18 cholesterolcrystal NALP3 ( 20) immuneresponse microbe host compo-nents NALP3 autoim-mune disease

AIM2 detector/receptor self non-self nucleic acid cytoplasm AIM2 susceptible infection Francisella infection vaccinia infection AIM2 detector DAI(DNA dependent activator of interferonregulatory factor) nucleic aciddetector cytoplasm adjuvant

2 15-17 2553 69

: Nat. Immunol 7: 1250, 2006.

20 inflammasome NALP3

: 70

DAI nucleic acid inflammasome IFN- IFN- IFN- autoactivate AIM2

inflammation hostresponse IFN- CpG 21

modulate host response TLR ligand action

21 self non-self nucleic acid cytosolic receptors

: Nat. Immunol. 11: 997, 2010.

agonist immuneresponse antagonist sepsis host response

2 15-17 2553 71

New Vaccine Design

...

(Vaccine design)

(Immune response)

1.

2. (Live vaccine) (Vector) (Liveattenuated vaccine)

3.

downstream (Purification)

Flumist Flumist (Intellec-

: 72

tual property) (Killed vaccine) Deep sequencing

(Cell culture) (Purification) endotoxin (Egg-basedtechnology) (Cell-basedtechnology)

4. o (Route)

(Live-attenuatedvaccine) (Mucosal) (Herdimmunity) (Host cell) 1 (Primary vaccination) (Booster) 2

o (Schedule)

o (Combi-nation) (EPI) vector (Large vector)

2 15-17 2553 73

5. (Shelf-life) (Cold chain)

2

o Live or Killed vaccine

o Intrinsic or Extrinsic antigen

o Protective antigen/epitope HIV protective epitope epitope

epitope

epitope surfaceglycoprotein glycopro-tein epitope epitope protective( neutralize) enhance neu-tralize enhance

o Variability protective immune response

Neutralizing antibody protective antibody HIV neutralizing antibody neutralize (Invitro) (Assay) neutralization neutralize enhancement

: 74

(Artificial)

HIV

conserved antigen protective antigen protectiveantigen haemagglutinin antigenic drift

branch

protective antigen conserved (Measles) protective conserved

(Innate immune response) (Specific immune response) innate immune response Toll-Like-Receptor (TLR) (Inflamma-

2 15-17 2553 75

tion) Adaptive immune response Innate Adaptive im-mune response Toll-Like-Receptor (TLR) adaptive innate im-mune response

Majorhistocompatability complex (MHC) class I II

1 MHC class I II

T lymphocytes cytotoxic T - cell (CD8) helper T cell (CD4) T cell receptor (T cellreceptor: TCR) (Cytoplasmic antigen) MHC Class I MHC ClassII 1

MHC Class I Class II MHC class I cytotoxic lymphocyte

DNA

: 76

DNA toxin pore forming toxin toxin 2subunits subunit subunit (Lipo-some) (Fuse)

2 innate adaptive immunity

MHC Class I

correlateimmune correlate immune protective immune

2 15-17 2553 77

innate

adaptive immune response innate antiviral response T cell Th 1 Th 2 B cell T cell adaptiveimmune response 2 cytotoxic T cell innate response innate response adaptive immuneresponse

Nature medicine 2009 Lackof antibody affinity maturation due topoor Toll-like receptor stimulation leads toenhanced respiratory syncytial virusdisease Delgado, M. F.

Respiratory Syncytial Virus (RSV) (Pneumonia)

RSV RSV (Formaldehyde) (Enhanced respiratory disease: ERD) epitope protective TLR

: 78

3 (d) viral load( FIRSV: Formaldehyde inacti-vated, UVRSV: UV inactivated, RSV) RSV wild type RSV challenge wild type RSV parenteral 3 (b) RSV (Placebo group) RSV viral load

3 RSV (Nonreplicating vaccine) prime enhanced respiratory disease: ERD

FIRSV (Formaldehyde inactivated) UVRSV (UV inactivated) viral load eosinophils titration

(Correlate) 50%Plaque reduction: PRNT50 ( 4) RSV wild type neutralization antibody FIRSV neutralize titer neutralizingantibody

2 15-17 2553 79

4 RSV-specific neutralization 50% Plaque reduction: PRNT50

UV inactivated RSV(UVRSV) formalin inactivated RSV(FIRSV) EL ISA (Enzyme- l inkedImmunosorbent Assay) affinities maturation(Lymph node) affinities matu-ration killed vi-rus affinities maturation

(Signal) TLR ligand

TLR ligand 5 (c) RSV UVRSV ( viral load ) LPS (Lipopolysac-charide) (Live virus) viral load RSV RSV (UVRSV+TLR)5 (d)

: 80

immune response

5 UVRSV TLR ERD

immune response correlate of protection

2 15-17 2553 81

From Pre-clinical Research to Vaccine Development:Examples of Go-no-go Decisions - a case study of rotavirusvaccine

...

(Rotavirus vac-cine) 20

5

: 82

3-5 90

/

(Rotavirus)

Dr. Ruth Bishop

. 1973

(Seasonal pattern)

(segmented genome) 2 VP7 VP4 (VP viral protein) neutralizing antibody ant igenicity neutralization assay serotype G

2 15-17 2553 83

serotype VP7 P serotype VP4 sero-type neutralization group subgroup VP6

(Influenza virus) hemag-glutinin (HA) neuraminidase (NA) hemagglutinin neutralization VP7 (G serotype) VP4(P serotype) neutralization VP7 VP4 G sero-type serotype 1,2, 3 4 G serotype 8 9 Pserotype 7 1A, 1B, 2 3

G serotype 1, 2, 3 4 G 1, 2, 3 4 G serotype 8 9 G9 8.6 G8 12.8

serotype serotype 1 sero-type serotype serotype

: 84

(Rotavirusvaccine development)

20 (host cell) 3

Jennerian approach

Modified Jennerian approach (animal/humanrotavirus reassortant)

Human live, attenuated vaccineapproach

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cross protec-tive immunity vaccinia

Jennerian approach (rhesus monkey)

Jennerian ap-proach VP7 VP4

Modified Jennerian approach

2 15-17 2553 85

Modified Jennerian approach (reassortant vi-rus) VP7 VP4 VP7 VP4 (seasonal influenza)

(reassortant) hemagglutinin neuraminidase

Human live, attenuated vaccineapproach

(Rotavirus VaccineCandidate)

3 1

: 86

1 (rotavirus vaccine candidate) approach

.. 1983 RIT 4237 1 bovine RV strain (NCDV), G6 P(6)

1986 WC3 1 bovine RV strain, G6 P(7)

1986 MMU18006, RRV 1 rhesus RV strain, G3 P(5B)

1991 RRV-TV, RotaShield 1 RRV, G3 + 3 rhesus/human reassortants, G1, G2, G4

2000 RV5, PRV, RotaTeq 5 WC3/human reassortants, G1, G2, G3, G4, P(8)

1999 RIX4414, Rotarix 1 attenuated human RV strain (HRV, RV1), G1 P(8)

2009 RV3 1 attenuated human neonatal RV strain, G3 P(6)

2010+ 116E 1 natural bovine/human RV reassortant, G9 P(11)

2010+ BRV(UK)-TV/HV 4 - 6 bovine (UK)/human RV reassortants,G1, G2, G3, G4, P(4), P(8) + G8, G9

1

2

3

( .. 1983- 1986)

Jennerianapproach (bovine rotavirus) RIT4237 WC3 (rhesus rotavirus) MMU18006

IgA cross protective immunity G serotype P serotype

2 15-17 2553 87

0 - 58 VP7 VP4 crossprotective antibody prime booster effect

Jenner ianapproach

( .. 1990- 2000)

Modified Jennerian approach Human

live, attenuated vaccine approach 1. reassortment

G P serotype RotaShield(RRV-TV) G genotype 4 RotaTeq (RV5, PRV) 5 genotype

2. Rotarix serotype G1 P(8) cross protective antibody serotype

(licensure) RotaShield RotaTeq Rotarix RotaShield (intussusception)

: 88

RotaTeq Rotarix

RotaShield (RRV-TV) Dr. Albert Z. Kapikian

Laboratory of Infectious Diseases NIAID,NIH rhesus G serotype G serotype 1, 2 4 G serotype 3 serotype VP7 G serotype1, 2 4 3 tet-ravalent rotavirus vaccine

Tetravalent reassortant virus 80 license RotaShield

(intussusception) 2 RotaShield 20 1-2 1 10,000

500,000 .. 1999

RotaShield

2 15-17 2553 89

rhesus RotaShield 3 3 3 3

RotaTeq (RV5 PRV) Dr. H. Fred Clark

Dr. Paul A. Offit Childrens Hospi-tal of Philadelphia Merck RotaTeq WC3 G serotype G1, 2,3 4 P serotype P(8) 98 63 .. 2006

RotaTeq (placebo) 3 RotaTeq

Rotarix (RV1 HRV) Dr. Richard L.

Ward David I. Bernstein CincinnatiChildrens Hospital Medical Center, Uni-versity of Cincinnati primary AfricanGreen Monkey kidney cell 26 GlaxoSmithKline Vero cell RIX4414 85

: 90

42

liveattenuated vaccine coldchain RotaTeq Rotarix

(EPI program) ..2009 3 Rotarix 300 2 600

1 4 10 Can it lead to cost-saving? 3 ( )

2 15-17 2553 91

(.. 2009 - )

serotype G8 G9 serotype 10 (cost effectiveness)

RV3

(attenuated) serotype G3 P(6)

116E

(Bharat Biotech International Limited) (natural bovine/human RVreassortant) serotype G9 P (11) G9

BRV (UK)-TV/HV BRV(UK)-TV/HV

Dr. Albert Z. Kapikian RotaShield bovine rotavirus UK RotaTeq bovine (UK)/human RVreassortant serotype G1,G2, G3, G4, G8 G9

: 92

(academia) (industry) (production facility) (clinical facility)

(criteria)

Bernstein DI. Live attenuated human

rotavirus vaccine, Rotarix. Semin.Pediatr. Infect. Dis. 2006; 17: 188-94.

Bishop RF. Discovery of rotavirus: implica-t ions fo r ch i ld heal th . J .Gastroenterol. Hepatol. 2009; 24Suppl. 3: S81-5.

Bishop RF, Davidson GP, Holmes IH, RuckBJ. Virus particles in epithelial cellsof duodenal mucosa from chil-

dren with acute non-bacterialgastroenteritis. Lancet 1973; 2:1281-3.

Centenaria C, Gurgel RQ, Bohland AK, OliveiraDMP, Faragher B, Cuevas LE. Rotavirusvaccination in northeast Brazil: a laud-able intervention, but can it lead tocost-savings? Vaccine 2010; 28: 4162-8.

Clark HF. Portrait of a leading vaccinologist; abrief account of experiences lead-ing to RotaTeq vaccine. Hum. Vac-cines 2008; 4: 256-9.

2 15-17 2553 93

Kapikian AZ, Simonsen L, Vesikari T, Hoshino Y,Morens DM, Chanock RM, LaMontagne JR, Murphy BR. Ahexavalent human rotavirus-bovinerotavirus (UK) reassortant vaccine de-signed for use in developing coun-tries and delivered in a schedulewith the potential to eliminate therisk of intussusception. J. Infect. Dis.2005; 192: S22-9.

Midthun K, Kapikian AZ. Rotavirus vaccines:an overview. Clin. Microbiol. Rev.1996; 9: 423-34.

Parashar U, Hummelman E, Bresee J, Miller M,Glass R. Global illness and deathscaused by rotavirus disease inchildren. Emerg. Infect. Dis. 2003; 9:565-72.

: 94

(Essential Practical Aspects of Conducting Clinical Trials)

Department of Virology, US Army Medical Component-AFRIMSStephen J. Thomas, MD

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2 15-17 2553 95

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2 15-17 2553 99

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2 15-17 2553 101

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Pandemic threat: from policy to implementation

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2 15-17 2553 103

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2 15-17 2553 105

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2 15-17 2553 107

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2 15-17 2553 109

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2 2552 2553

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2 15-17 2553 111

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2 15-17 2553 113

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2 15-17 2553 115

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2 15-17 2553 117

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2 15-17 2553 119

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2 15-17 2553 121

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2 15-17 2553 123

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2 15-17 2553 125

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: 126

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2 15-17 2553 127

(Measles) antigenic site antigenicdrift

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: 128

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2 15-17 2553 129

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: 130

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2 15-17 2553 131

4 reverse genetics

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2 15-17 2553 133

(Split subunitvaccine)

cold-adaptation, temperature-sensitive at-tenuation phenotype 3 marker masterdonor strain reassortant

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: 134

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2 15-17 2553 135

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2. preclinical

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: 136

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2 15-17 2553 137

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: 138

3. master donor strain typeA master donor strain mas-ter donor strain splitvaccine PR8 master donor

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2 15-17 2553 139

Registration preparedness for new vaccine licensing

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2 15-17 2553 141

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2 15-17 2553 145

1 2552 ASEANHarmonization Product on Pharmaceuti-cal Registration

ASEAN Harmonization(ACTD: The ASEAN Common TechnicalDossier) 4

1 (Administrative data andproduct information)

2 (Quality Document)

3 : (Safety: Non-clinical Document)

4 : (Efficacy: Clinical Document)

www.fda.moph.go.th 1 (Administrative data andproduct information) 3 (Section) A, B C

A (Introduction)

ASEANHarmonization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ............................

B (Table of contents) C

(Documents required for registration)

1. ( ..1 .1) ..1 ASEAN Harmonization .1 ..1 .1 ..1 .1

2. (Good Manufacturing Practice:GMP) GMP (Certificate of Pharmaceuti-

: 146

cal Product: CPP) (Certificate of Free Sale: CFS)

3.

4.

5. .. 8 .. 8

6.

7. - //

8.

9.

ASEAN Harmo-nization

ACTD 3

1. (Unit carton)2. (Inner label)3. (Blis-

ter/Strips) 2

1. (Unit carton)

1.1 (Productname)

1.2 1.3 1.4 1.5 1.6 1.7

// /

1.8 / (Intramuscular: IM) (Subcutaneous: SC)

1.9 1.10 1.11

/

2 15-17 2553 147

1.12

1.13 SMP

1.14 ()

1.15 2. (Inner label)

vial am-poule

2.1 (Productname)

2.2 2.3 2.4 2.5 vial /

ampoule 3 Exp.

2.6 2.7 2.8

vial ampoule

ASEANHarmonization 3

1. Summary of Product Character-istics: SPC

2. Package Insert: PI3. Patient Information Leaflet : PIL 1. NCE

SPC PI PI

2. NCE 1 SPC PI

3. PIL SPC PI PIL

SPC

1. (Name of Medici-nal Product)

: 148

1.1 (Product Name)1.2 (Strength)1.3 (Pharmaceuti-

cal Dosage Form)2.

(Quality and Quantitative Composition)2.1 (Qualita-

tive Declaration)2.2 (Quantita-

tive Declaration)3.

(Pharmaceutical Form)

lyophilized

4. (Cl in icalParticulars)

4.1 (Therepeutic indication)

4.2 (Poso-logy and method of administration)

4.3 (Contraindication)4.4

(Special warning and precau-tions for use)

4.5 (Interaction with othermedicinal products and other forms ofinteractions)

4.6 (Pregnancy and lac-tation)

4.7 (Effects onability to drive and use machine)

4.8 (Undesir-able effects)

4.9 (Over-dose)

5. (Pharma-cological properties)

5.1 (Pharmacodynamic properties)

5.2 (Pharmacokinetic properties)

5.3 (Preclinical safety data)

6. (Pharma-ceutical particulars)

6.1 (List ofexcipients)

6.2 (Incompatabilities)

6.3 (Shelf life)

6.4

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(Special precautions for storage)6.5

(Nature and contents of con-tainer)

7. (Marketingauthorization holder)

8. (Marketingauthorization number)

9. (Date of authorization)

10. (Date of revision of the text)

PI

1. (Product Name)2.

( Name and strength of active ingredient(s))

3. (Productdescription)

4. / (Pharmacodynamic / Pharmacokinetics)

5. (Indication)6. (Recommended

dose)7. (Mode of adminis-

tration)8. (Contraindication)

9. ( Warningand precaution)

10. (Interactionswith other medicaments)

11. (Pregnancy and lactation)

12. (Undesirableeffects)

13. (Overdose and treatment)

14. (Storage con-dition)

15. (Dosage forms and packagingavailable)

16. (Name and address of manufacturing /marketing authorization holder)

17. (Date of revision of package insert) PI SPC SPC PI PI

: 150

2 (Quality document)

4 (Section) A, B, C D

A (Table of contents) B

(Quality overall summary)

- (Drug sub-stance)

- (Drug product)

C (Body of data) (Drug substance) (Drug product)

D (Keyliterature reference)

(Body of data) (Drug substance) (Drug product)

(Drug substance)

-

-

- -

(Specification) (Batch analysis)

- -

(Drug product)

- -

-

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1 formulate ASEAN Harmonization( .1) GMP

1 (Batch formula) activeingredient excipient

flowchart

purification

(Process validation)

-

adjuvant, preservative excipients (specification)

- 3

- (Reference stan-dards) reference stan-dards

- (Containerclosure system) ampoule/ vial

- (stability data)

3 : (Safety: non-clinicaldocument)

new

: 152

adjuvant

5 (Section) A, B, C, D E

A (Table of contents) B

(Non-clinical overview) model

C (Non-clinical summary: Written andtabulated)

D ()(Non-clinical study report: as requested)

E (List of key literature references)

(Safety:non-clinical document) (Pharmacology) (Pharmacokinetics) (Toxicology)

4 : (Efficacy: clinical document)

6 (Section)

A, B, C, D, E F A (Table of contents) B

(Clinical overview) C

(Clinical summary) D

(Tabular listing of all clini-cal studies)

E (Clinical study reports) () (Pharmacovigilanceplan)

F (List of key literature references)

- -

-

-

2 15-17 2553 153

ICH GCP

(check-list)

- . B1 (Administrativedata and product information)

- . 2 (Quality document)- . B3 (Safety: non-clini-

cal document)- . B4 (Efficacy: clinical

document)

- 1 ()- 2 ()- 4 (

)

5

: 154

checklist 3 5

USFDA EMEA (European MedicinesAgency) / 2

( SMP) ( SMP)

2 pre-marketing phase post marketingphase ( 6) pre-marketing phase (Good ManufacturingPractice: GMP) post

2 15-17 2553 155

6

marketing phase (EPI)

(Nationalregulatory system)

: 156

Regulatory, Intellectual Property and Trade Barriers forVaccine Development

..

5 1) Knowledge value chain 2) Regulatoryrequirement 3) Commercialization 4) Man-agement 5) IP deal making

IP ..

.

.. 2548 Intellectual Property (IP)

new comer IP (Technol-ogy transferring)

Regulatory and IP: implications for vaccine R&D

(.)

2 15-17 2553 157

2

Stem cell stem cell

adjuvant new component ( antigen ) adjuvant adjuvant

Gene transfer DNA fragment

knowledge valuechain value chain value chain (Basicscience)

(Busi-ness management)

regulatory requirement knowledge value chain regulatory agency Food and Drug Administration (USFDA) regulatoryrequirement require-ment (Over requirement) new comer IP

: 158

1

knowledge value chain 1 2 basic research (Efficacy data) .

2 15-17 2553 159

3

2

3

: 160

2009

dihydroartemisinin (DHA) (. ; Biotec) . (.)

4 dihydroartemisinin (DHA) 1999

4 1999 (New drug application) . (Formulationstudy) (Clinicalstudy) 1 4 5

2 15-17 2553 161

(ClinicalTrial:CT) 2 6 Thai-GMP cGMP (c = current)

5 dihydroartemisinin (DHA)

. CT GMP . CT (Toxicity study) (Pre clinical study)

CT

: 162

. CT impurity X X derivativedegradative product active ingredi-ent DHA po-tency impurity X

X

regula-tory agency

regulatory requirement

2 15-17 2553 163

..

regulatory requirement GLP (Good Laboratory Practice), GCP(Good Clinical Practice) GMP (GoodManufacturing Practice) 1 1 official report

- -

- regulatory requirement

(Tacit knowledge)

Regulatory, Intellectual Property and Trade Barriers forVaccine Development

regulatoryrequirement plant-based vaccine regulator USFDA USFDA

- - -

: 164

- . 6 1 CT

-

(IP) regulatory requirement 2 /

-

IP - Plasmid seed virus

vaccine plasmid - Host cell system (rapid growth,

high yield, low pyrogens, etc.) IP

- Seed vaccine system - Production system- Purification system- Adjuvant

IP (Inventor) IP system

2 15-17 2553 165

IP system (Invention) IP system (Scientificpublication) (Sign MOA)

IP (Abuse) IP IP

1. (Preemp-tive) 2 Preemptive fil-ing

2.

3.

4.

IP IP

IP patent patent Oseltamivir Roche

: 166

()

(Trade Barriers)

- 1 (Sustainability)

- 2 captive market

- 3

- 4

2 15-17 2553 167

. . .

2

60 10 (.. 2544-2553) 50,000-100,000 ( 100-200 )

.. 2515 Walter Reed Army

Ins t i tu te of Research Fetal Rhesus Lung cells

10-15

.. 2523 Dog Kidney cells( Fetal Rhesus Langcells Walter Reed ) . 12

: 168

.. 2536 4 / 250 2 1 1 ()

1.

International PeerReview Committee 12 2-3

audit Dog Kidney cells cell

copy process Dog Kidney cell Technology know-how copy know-how

2.

2 15-17 2553 169

2 30 2-5 1,000 100,000

2 2 (backbone) 2 (

) Collaborative Research Agreement 10 2 patent ownership

2-3

: 170

HIV vaccine trial: Why Thailand?,Is it a true success story?

.

3 - 2546 2552

HIV vaccine trial: Why Thailand 2533

136,000 5 2543 30,000

375 2533 81 2543

HIV

2 15-17 2553 171

HIV

HIV

E (CRF01_AE) 5 2536,2541 2544-2549 CRF01_AE

HIV

2536 1 4 (AFRIMS) (RIHES) 1 2 - 1 2 3

: 172

Stage 1: Laboratory development

1-2

Stage 2: Pre-clinical study

2 Stage 3: Clinical trial

6-10 3 Phase I:

1. 2.

3. HIV 5% 4.

() 1-2

Phase II: () 2-3

Phase III: 3-5

Stage 4: Delivery

2 15-17 2553 173

3

prime boost HIV viral load CD4

Community-based, Ran-

domized, Double-blind, Placebo controlled1: 1 Intention to treat 18-30 16,400 heterosexual transmission ( )

(Dataand Safety Monitoring Board) 8 interimefficacy analysis 36

incident rate 0.34 in-

fection/100 person-years 4 40 screening sites 8 clinical sites 1 2

0, 1, 3

6 0 1 prime vac-cine ALVAC 3 6 prime boost vaccine ALVAC AIDSVAX immuniza-tion reaction 72 6 3 follow up rate 95% 6

- (Prime-boost concept) (E&B)

Prime vaccine ALVAC-HIVvCP1521 (Canary pox virus) (Vec-tor) HIV (Cell MediatedImmunity; CMI)

Boost vaccine AIDSVAX B/E

: 174

(VaxGen) (gp120) E B (HumoralMediated Immunity)

Placebo ALVAC carrier AIDSVAX adjuvant

26,676 40 16,402 7 HIV 16,395 modified ITT (mITT) 8,197 - 8,198 48 28 24

per protocol(PP) protocol 4 6 mITT 6,176 - 6,366

mITT

ALVAC-HIV AIDSVAX B/E 31.2 (p = 0.04, 95% CI(OBF) 1.1, 52.1) HIV 0.28 0.34 PP mITT

HIV vaccine trial: Is it a true successstory?

2

1.

2.

3.

4.

2 15-17 2553 175

(Dataand Safety Monitoring Board) 80 65 end point HIV infection 130 interim analysis 30 10

5. protocol 20 10 1 protocol

6.

7.

3

8. product

9. end point accredit American College of Pathologists

10.

: 176

modified ITT (mITT) 31.2

1. 2552 The New England Journal of Medicine

2. Times Magazine 8 The 50 Best

Inventions of the Year 2 2552

3. Bangkok break through

4. Dr.Anthony S. Fauci, Profes-sors Francoise Barre-Sinoussi, Dr.NormanLatvin

3

2 15-17 2553 177

Light up the Candle in the Dark: HIV Vaccine Hope

.

. ..

(HIV Vaccine: New Design and the Challenges) (Community view on HIV vaccine development). (Clinical trial and public health benefit)

2544

Shots in the Dark: The Way-ward search for an AIDS Vaccine Dr. Jon R. Cohen Thai Trial 24 2552 prime-boost 31.2 Candle in the Dark: The WayForward of Searching for an AIDS Vaccine

3 .. 2536 . 3 licensed vaccine

: 178

new design 29 21 (Identification)

3

1. parenteral

2.

3.

HIV Vaccine: New Design and the Challenges

..

3

3

antibody-basedHIV vaccine

- AIDSVAX (B/B)

- AIDSVAX (B/E)

T cell-based HIVvaccine

- STEP study (Merck V520-023/HVTN 502)

2 15-17 2553 179

- PHAMBILI study (HVTN 503) STEP PHAMBILI

recombinant Adeno 5 (Ad 5)Vector gag, col naf clade B Ad 5 im-munogenicity T cell T cell-based vaccine

3

Thai trial prime-boostvaccine (Prime) ALVAC (Boost) AIDSVAX recombinant protein 1 3 protective immunity cohort

3

1. Critical Timing: Early Transmis-sion and Infection Events (window ofopportunity to prevent HIV infection)

1.1 target cell dendritic cell T cell immunize prime boost protective level protective immunity

1.2 het-erosexual vaccine design submu-cosal gut mucosa CD4 ractal chal-lenge false protection false protection ractal chal-lenge vagina vaginal mucosa sub-mucosa CD4 recruit CD4

: 180

CD4 dendritic cell Dr.Gary Nabel VaccineResearch Center NIH

2. AIDS Vaccine: Pendulumapproach

neutralizing antibody-based HIV vaccine monomeric GP120 160 neutralizing antibody cross protection subtype strian T cell vaccine T cell vaccine neutralizing antibody T cell vaccine HIV

3. neutralizing antibody basedvaccine

neutralizingantibody-based vaccine

- structure HIV envelop

glycoprotein- neutraliz-

ing antibody HIV - monoclonal

antibody broad neu-tralizing antibody

James A. Hoxie Toward an Antibody-Based HIV-1 vaccine Annual Review of Medi-cine 2010 structure HIV envelop glycoprotein HIV neutraliz-ing antibody HIV

neutralizing anti-body vaccine HIV envelopprotein binding receptor co-receptor fusion binding immunesystem

envelop glycoprotein HIV GP120 trimeric ( 3 ) GP41 1 epitope heavily glycosylated anti-body

2 15-17 2553 181

2 interaction binding trimerenvelop HIV CD4 co-recep-tor GP120 CD4 co-receptor GP120 GP41 fusion protein GP41 fuse membrane membrane membrane RNA binding fusion antibody

1 HIV envelop glycoprotein

neutralizing antibody (High ratemutation) antibody

Andrew McMichael Nature Review in Immunology Vol-ume 10 2010 immune response 5 100 3 Y copies/ml X () HIV-1transmission innate immunity 1-7 accute phase protein cytokine

: 182

antibody 2-3 antibody non-neutralizing an-tibody, anti-GP120 anti-GP41 30 cytotoxic Tcell (CD4) infected cell replication 5 (Vi-ral escape) immune individual 80 neutralizingantibody ( type specific) recog-

2 HIV

nize infect 7 high titer antibody T cell broad specific type spe-cific

John Mascola (Chronic infec-tion) broad neutralizingantibody (3 neutralizing antibody

2 15-17 2553 183

type specific) broad specific cross neutralize 13-20 ( 1/10 - 1/4 chronic infection broadneutralizing antibody)

B cell broadneutralizing antibody monoclonal high throught put screening monoclonal mapping 4 broad neutralizing

3 immune response HIV 100

antibody GP41 GP120 ( GP41 trimer 3 GP120) 4E10, 2F5, Z13e1, b12 2G12 monoclonal antibody 5 broad spectrum protect 4E10, 2F5 Z13e1 antibody GP41 b12 2G12 antibody GP120 antibody potency broad monoclonal

: 184

2 PG9 PG16 2G12 potency newstructure binding designimmunogen 5 structure PG9 PG16 new

4 broad neutralizing antibody GP41 GP120

5 broad neutralizing antibody PG9 PG16

monoclonal antibody broad neu-tralizing antibody Proc NatlAcad Sci U S A. 2010

neutralizing antibody animal model

2 15-17 2553 185

11 protective immunity serum passively immunize protect protect protective immunity broadspectrum neutralizing 2009 AJHessell, Dr Burton 2 neutralizing antibody broad spectrum 2G12 low titer (Protect) broad spectrum neutralizingantibody protection

design neutralizing-basedvaccine ( review James A. Hoxie)

1. particles-based design Virallike particles (VLPs) HPV HPV pentamer VLPs solid-phaseproteoliposomes (PLs)

2. soluble envelop trimers3. mutation technique4. modify carbohydrate envelop5. biological modified envelop6. structure-based design

antibody function neutralize antibody-dependent cell-mediated cyto-toxicity (ADCC) antibody-dependentcell-mediated viral inhibition (ADCVI) viral inhibition antibody-comple-ment mediated lysis activity

4. T cell-based HIV-1 vaccine T

cell-based vaccine promising data T cell-basedvaccine neutralizing antibody-based vaccine vaccineresearch center Dr.Nabel DNA vaccine strong T cell response 50(efficacy) 88 44 neutralizing antibody T cell (Com-bine)

replicating vector CMV-based vaccine

: 186

challenge rhesus 4 12 33

T-cell1. Replicating vector VS non-repli-

cating vector (Chronic latent in-fection) (Disease progression) livevector non-replicating vector immunogenicity replication antigen replicat-ing vector vaccinia replicate immunogenicity 1

2. Non-human viral based vector adinovirus human viral based vector adinovirus 70-80 antibody ad-enovirus memory T cell adinovirus

activate HIV vector non-human vector

3. In vivo electroporation DNA vac-cine delivery system ( HIV-1 DNA Vaccine )

4. long-lived T cell anti-body TLR-ligands

5. HIV-1 DNA Vaccine

CHULA Initiated HIV-1 DNA Vaccine Vaccine & CellularImmunology Research (VCI)

HIVDNA vaccine NIH recognize HIV antigen gag, nef, pol en-velopment recognition 70-80 6 pro-tein expression humanizedHIV-1 gag DNA vaccine

2 15-17 2553 187

humanleukocyte antigen (HLA)

6 Higher protein expression humanized HIV-1 gag DNA vaccine

(BIOTEC) (.)

DNA vaccine NIH vaccine research center DNA vaccine immunogenic 1 2 1 dosedependent dose antibodyresponse 70-80 T cell response 60-80

CHULAInitiated HIV-1 DNA Vaccine needle-free injection 7 antigen minuteinjury priming cytokinechemokine dendritic cell uptake antigen

: 188

DNA vaccine AE B South-East Asia Globalmosaic

7 Needle-free injection () Needle injection

sequence 100 base recombinants cocktail 3 cocktails Mosaic cock-tail 8

8 Global Mosaic

2 15-17 2553 189

gagepitopes recognization CD4 CD8response mosaic consensus recognize gagepitopes 5 epitopes mosaic vaccine global mosaic

consensus vaccine 5 7 mosaic vaccine CD4 re-sponse consensus vaccine re-sponse 2 7 CD8mosaic vaccine 3 7 consensus vaccine response 7 9

9 recognize epitopes mosaic consensus

South-East Asia mosaic AE B Los Alamos mosaic

immune response needle-freeinjection 10 prime-boostmosaic DNA - vaccinia ELISPOT 1800 spot AE cross B 900 spot

: 190

sign agreement electroporation

electroporation DNA vaccine eletroporation

10 Prime-boosted Mosaic HIV-1 AE/B gag + Vaccinia

immune response 10 1,000 spot eletroporation 10,000 spot

(Protection) 1-3 correlate immune protection immu-nogenicity

Humoral immune responses- Nab assays

- ADCC- ADCVI

Cell-mediated immune responses- Lymphoproliferative assay- CD4 ICS by flow cytometry- CD8 : ICS by flow cytometry,

ELISPOT

2 15-17 2553 191

( 31.2)

200 TIME

Community View on HIV Vaccine Development

10 / /

: 192

24 2552

31.2

3

2 15-17 2553 193

HIV

7-8 ()

: 194

3 2 (Japanese en-cephalitis: JE Nakayama)

Clinical trial and public health benefit

(UNFPA).

15 15 3 15 7-8

placebo 3 1 2

2 15-17 2553 195

public health risk .. 2000 Dr. Cohen Shot inthe Dark 3 public health ben-efit HIVvaccine

Public health risk 1. False expectation 2

2. Guinea pig phenomenon 20 /

(MOU) Guinea pig phenomenon

: 196

Guinea pig phenom-enon public health risk

3. HPV vac-cine 4,000 3 Hepatitis B HIV vaccine

Public health benefit

1. (Reduce HIV trans-

mission) HIV HIV

HIV

2. (Delay disease

progression)3.

4. benefit risk

reduceinfectiousness benefit

Efficacy vs Effectiveness clinical trial efficacy

effectiveness efficacy effectiveness intervention

Economic determinants

2 15-17 2553 197

proposal 8 () RVI44 efficacy 31.2 10 1000 : 1 efficacy

transmissibi l i ty

32 4 100 32 4

WHO-UNAIDS Consultation ..2000 .. 2001 3 10 2 (Health benefit) (Societal benefit)

Health benefit healthbenefit benefit estimated saving percase . 1000 . 10 . 1000 . 10 model

: 198

Societal benefit

benefit medicalexpenditures . 1000 . 10 benefit

benefit 1benefit