Genetic differences between VZV wild-

type and Oka strains

Judith BreuerProfessor of Virology

St Barts and The London Hospital Medical School

UK

1Rome, Italy - March 18, 2005

•masque

. . . .

Oka vaccine Live attentuated virus

A

B

CBang +

Bang BAN7

Reproduced from Barrett-Muir W, et al. J Virol 2002;76:1971-9with permission from the American Society for Microbiology

J95

Bang +

Bang BAN7

Loparev VN, et al. J Virol 2004;78: 8349-58.

E

M

J95

Bang +

Bang BAN7

Grose, J clin Virol 2005; in press

E

A

95

pOka vs Dumas

1/700

MSP vs Dumas

1/2000

BC vs Dumas

1/2000

BC vs

MSP

1/2700

Glycoproteins

5,14, 31, 37, 50, 60, 67 68

6 (8) 7 4 5(1)

transcriptional

4,10 21 61,62,63,64

6 0 1 1 (42)

replication

6, 8, 16, 18, 19, 28,29, 48,51,52,55,59

14 (18) 1 2 (2) 1

Kinases proteases

33,36, 47, 66

3 (2) 1 0 0

Tegument

11, 12,, 22, 54

9 (19) 3 4 4(1)

pOka vs Dumas

1/700

MSP vs Dumas

1/2000

BC vs Dumas

1/2000

BC vs

MSP

1/2700

pOka vs vOka-

1 in 2-3000

Glycoproteins

5,14, 31, 37, 50, 60, 67 68

6 (8) 9 6 5(1) 3(2)

transcriptional

4,10 21 61,62,63,64

6 0 1 1 (42) 10 (8)

replication

6, 8, 16, 18, 19, 28,29, 48,51,52,55,59

14 (18) 1 1 (2) 1 6 (1)

Kinases proteases

33,36, 47, 66

3 (2) 0 0 0 0 (1)

Tegument

11, 12,, 22, 54

9 (19) 3 4 4(1) 2 (1)

Which are the genetic correlates of vaccine attenuation?

Reduced ability of vOka to spread from T-cells to melanoma cell lines (Soong et al, 2000)

Similar to ORF 47 mutant

Evidence that ORF 47 p may be a virulence factor

Soong W, et al. J Virol. 2000;74:1864-70.

Vaccine preparations in comparison to pOka

6 9A 21 31 39 50 51 52 54 55 59 6 2

Nuc

leo

tide

po

siti

on

5745

1090

0

1277

9

3173

2

5859

5

7125

2

8730

6

8973

4

9053

5

9416

7

9774

8

1010

89

1053

10

1055

44

1057

05

1062

62

1067

10

1071

36

1072

52

1081

11

1088

38

O ka Parent A T C C A T T A A T G A A A T T A T T T A

O ka Biken G Y Y Y R Y Y G R C R R R G C C R C C C R

Nuc

leot

ide

posi

tion

5745

1090

0

1277

9

3173

2

5859

5

7125

2

8730

6

8973

4

9053

5

9416

7

9774

8

1010

89

1053

10

1055

44

1057

05

1062

62

1067

10

1071

36

1072

52

1081

11

1088

38

O ka Parent A T C C A T T A A T G A A A T T A T T T AO ka Biken G Y Y Y R Y Y G R C R R R G C C R C C C RO ka GSK MQO ka GSK MQO ka GSK AS

Vaccine preparations in comparison to pOka

Vaccine preparations in comparison to pOka

6 9A 21 31 39 50 51 52 54 55 59 6 2

Nuc

leot

ide

posi

tion

5745

1090

0

1277

9

3173

2

5859

5

7125

2

8730

6

8973

4

9053

5

9416

7

9774

8

1010

89

1053

10

1055

44

1057

05

1062

62

1067

10

1071

36

1072

52

1081

11

1088

38

O ka Parent A T C C A T T A A T G A A A T T A T T T AO ka Biken G Y Y Y R Y Y G R C R R R G C C R C C C RO ka GSK MQO ka GSK MQO ka GSK ASO ka GSK AS

Vaccine preparations in comparison to pOka

6 9A 21 31 39 50 51 52 54 55 59 6 2

Nuc

leot

ide

posi

tion

5745

1090

0

1277

9

3173

2

5859

5

7125

2

8730

6

8973

4

9053

5

9416

7

9774

8

1010

89

1053

10

1055

44

1057

05

1062

62

1067

10

1071

36

1072

52

1081

11

1088

38

O ka Parent A T C C A T T A A T G A A A T T A T T T AO ka Biken G Y Y Y R Y Y G R C R R R G C C R C C C RO ka GSK MQO ka GSK MQO ka GSK ASO ka GSK ASO ka Merk concensus

Fig. 1. Gel electrophoresis of polymerase chain reaction (PCR) products. A: Amplification across BssHII site in ORF 62 and restriction enzyme digest. Lane 1: Parental Oka virus; lane 2, UK wild type J virus; lane 3, Oka Merck vaccine; lanes 4–6, vaccine zoster viruses;lanes 7–18, vaccine rash viruses; lane 19, molecular weight marker (band sizes 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 kb).B: Amplification across Alu1 site in ORF 6 and restriction digest. Lane 1: 100 bp molecular weight marker; lane 2, parental Oka virus; lane 3, UK wild type J virus; lane 4, Oka Merck vaccine; lanes 5–7, vaccine zoster viruses; lanes 8–19, vaccine rash viruses; lane 20, 100 bp molecular weight marker.

Which mutations are important for attenuation?

Vaccine preparations in comparison to pOka

6 9A 21 31 39 50 51 52 54 55 59 6 2

Nuc

leot

ide

posi

tion

5745

1090

0

1277

9

3173

2

5859

5

7125

2

8730

6

8973

4

9053

5

9416

7

9774

8

1010

89

1053

10

1055

44

1057

05

1062

62

1067

10

1071

36

1072

52

1081

11

1088

38

O ka Parent A T C C A T T A A T G A A A T T A T T T AO ka Biken G Y Y Y R Y Y G R C R R R G C C R C C C RO ka GSK MQO ka GSK MQO ka GSK ASO ka GSK ASO ka Merk concensus

Is IE62 necessary for attenuation ?

Does the vaccine protect against wild type viruses?

IE 62 . Overlapping peptide pools

CD8 + cells assayed by Ifn staining

Peptide pool 5 recognised by A2 restricted PBLs

Conclusion

1. wtVZV varies geographically, and between related strains

2. vOka vaccine is attenuated biologically and clinically

3. The vOka vaccine is made up of a mixture of strains

4. Attenuation is likely to be multigenic and involve ORFs 30-54

5. vOka immune protection may be enhanced by the mixture of wt and vaccine mutations in the vaccine preparation.