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Increased Herpes Simplex Virus-2 Shedding in HIV-1 Infected Persons is due to Poor Immunologic Control in both Ganglia and Genital Mucosa

Joshua T. Schiffer, MD, MSc

Study questions

• HIV infected persons with low CD4+ T-cell count shed HSV-2 at higher rates and are more likely to develop persistent genital ulcer disease

1. In which anatomic site is anti-HSV-2 immunity decreased in patients with HIV (ganglia and / or mucosa)?

2. In mucosa, is immune deficiency due to impaired clearance of infected cells, or impaired clearance of free virus?

HSV-2 genital shedding is frequent, episodic and heterogeneous

Mark KE, JID, 2008.

Schiffer JT, Sci Trans Med, 2009.

HSV-2 ulcers are associated with an intense, highly-localized & persistent CD4+ / CD8+ response

Zhu J, Nat Med, 2009.

Zhu J, JEM, 2007.

Peng T, JV. 2012.

Zhu J, Nature, 2013.

CD8αα(+) T cells display a memory effector phenotype weeks after lesion healing

Ma JZ, Russell TA, Spelman T, Carbone FR, Tscharke DC (2014) Lytic Gene Expression Is Frequent in HSV-1 Latent Infection and Correlates with the Engagement of a Cell -Intrinsic Transcriptional Response. PLoS Pathog 10(7): e1004237. doi:10.1371/journal.ppat.1004237http://127.0.0.1:8081/plospathogens/article?id=info:doi/10.1371/journal.ppat.1004237

Ganglionic reactivation occurs frequently

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There are high numbers of activated CD8+ effector memory cells directed against HSV-1

in trigeminal ganglia

Verjans G M G M et al. PNAS 2007.

HSV-1 LAT CD3+

CD3+ CD8+ CD4+ CD69+

CD8+ GzB+

• 4 cohorts of HSV-2 seropositive men:

– HIV negative (n=98)

– HIV positive, CD4>500 (n=34)

– HIV positive, CD4=200-499 (n=43)

– HIV positive, CD4<200 (n=21)

• Daily swabs x 60 days for HSV qPCR

Which site of immunity is decreased

in HIV infected patients?

HIV negative CD4>500 CD4 200-499 CD4<200

Total Swabs 6448 1960 2495 1556

Shedding rate 15% 19%*** 21%*** 31%***

Episode rate / year (95% CI)1 13 (11-16) 13 (10-18) 17 (13-22)* 20 (14-26)**

Median episode duration (days) 3.7 4.6 4.1 5.5**

* p<0.1, ** p<0.05, *** p<0.0011. Episode rate is underestimated with daily sampling.

Shedding rate, episode rate &

episode duration increase with

decreasing CD4 count

Schiffer JT et al. JID. 2012

Schiffer JT et al JAC, 2012.

Kinetics features of episodes are

surprisingly equivalent across all

CD4 strata

0

10

20

30

40

50

60

2-3 3-4 4-5 5-6 6-7 7-8 8-9 >9

Perc

en

t ep

iso

des

Peak log10 HSV DNA copies

Orange: HIV negativeMaroon: CD4>500Green: CD4 200-499Blue: CD4<200

Cell-associated HSV2

Neuronal HSV2

Cell-free HSV

CD8+ T-cell

Viral plaque

Plaque forming region

be

S I

E

f

c

a

d

l

F

q

c

bi

p

Cell-associated

HSV (Vi)

Cell-free

HSV (Ve)

Neuronal

HSV

(Vneu)

a

Schiffer JT, Elife, 2013.

Mathematical model of HSV-2 pathogenesis

The model’s most important limitation is oversimplification of immunity

Schiffer and Corey. Nat Med. 2013 Mar; 19(3): 280–290.

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The model predicts that mucosal T-cell density determines episode heterogeneity

0

1

2

3

4

5

6

7

8

9

0 2000 4000 6000 8000 10000 12000

CD8+ lymphocytes / mm3

HS

V D

NA

lo

g1

0 c

op

ies

/mL

tr

an

sp

ort

me

diu

m

High-copy episodes

Medium-copy episodes

Low-copy episodes

Schiffer JT, PNAS, 2010.

0

1

2

3

4

5

6

7

8

9

0 1 2 3 4 5 6 7 8 9 10

Days

log

10

HS

V D

NA

co

pie

s

The model predicts extremely rapid HSV-2

containment within single ulcers

Schiffer JT, Elife, 2013.

Model fit to HIV (-) & (+) data

S I

E

f

ca

d

l

F

q

bi

p

Cell-associated

HSV (Vi)

Cell-free

HSV (Ve)

Neuronal

HSV

(Vneu)

Varied parametersQ = mucosal CD8+ expansion rater = Number of HSV infected cells prior to CD8+ expansiond = mucosal CD8+

decay rate c = Free viral decay rateF = Neuronal HSV drip rate

Model fit to HIV (-) & (+) data

0123456789

10

2-3 3-4 4-5 5-6 6-7 7-8 8-9 >9

Perc

en

t sw

ab

s

log10 HSV DNA copies

0123456789

10

2-3 3-4 4-5 5-6 6-7 7-8 8-9 >9

Perc

en

t sw

ab

s

log10 HSV DNA copies

0123456789

10

2-3 3-4 4-5 5-6 6-7 7-8 8-9 >9

Perc

en

t sw

ab

s

log10 HSV DNA copies

0123456789

10

2-3 3-4 4-5 5-6 6-7 7-8 8-9 >9

Perc

en

t sw

ab

s

log10 HSV DNA copies

HIV negative

HIV positiveCD4 200-499

HIV positiveCD4 ≥500

HIV positiveCD4 <200

SimulatedHIV negativeshedding

SimulatedHIV positiveCD4<200shedding

The model also reproduces shedding episode rate and kinetic characteristics Neuronal release rate of HSV increases as CD4 decreases

S I

c

F

Neuronal

HSV

(Vneu)

Results represent top 5% of fits for 500 tested parameter sets

Negative CD4>500 CD4=200-499 CD4<2000

100

200

300

HS

V D

NA

co

pie

s / d

ay

p=0.01

p=0.002

p=0.64

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T-cell mucosal decay rate increases when CD4<200

I

E

f

dq

Results represent top 5% of fits for 500 tested parameter sets

Negative CD4>500 CD4=200-499 CD4<200-4.5

-4.0

-3.5

-3.0

-2.5

-2.0

log

10

T-c

ell d

ec

ay

/ d

ay

p=0.01

p=0.33

p=0.009

Viral clearance rate decreases when CD4<200

I

a

p

Cell-

free

HSV

(Ve)

Results represent top 5% of fits for 500 tested parameter sets

Negative CD4>500 CD4=200-499 CD4<2000

5

10

15

HS

V D

NA

cle

ara

nc

e / d

ay

p=0.01

p=0.34

p<0.001

p=0.02

c

Each parameter is only weakly predictive of increased shedding rate:

immune dysfunction is therefore likely to be multifactorial

0

20

40

60

80

100

0 50 100 150 200 250

Sh

ed

din

g ra

te (

%)

HSV DNA copies / day

0102030405060708090

100

-4 -3 -2

Sh

ed

din

g ra

te (

%)

log10 CD8+ T-cell / day

0

20

40

60

80

100

0 5 10 15

Sh

ed

din

g ra

te (

%)

HSV DNA decay / day

Neuronal release rate Mucosal T-cell decay rate Mucosal free viral decay rate

Implications

• Three broad areas of immunodeficiency in HIV patients:1. Decreased ganglionic immune protection against

reactivation

2. Decreased lifespan of mucosal T-cells

3. Decreased clearance of cell-free HSV-2 in genital mucosa

• Enhancement of these parameters may be a key feature of a therapeutic vaccine

Thank you!

• VRC clinicians

• Study Participants

• NIH / NIAID