Collaborating Institutions:

University of Nairobi

Seattle/Kenya Collaboration- MTCT

Julie Overbaugh

Fred Hutchinson Cancer Research Center

University of Washington

Target cell (e.g. T Cell)

Antibody

Designing an effective vaccine: -> Identifying immune correlates

• A major focus of HIV vaccine efforts is trying to elicit HIV-

specific NAbs.

• Most successful vaccines are thought to work through

antibody-mediated protection.

• Do NAbs protect against HIV?

(Mascola et al. Nat. Med. 2000)

NAbs can block HIV (SHIV) infection in macaques

Controls Given HIV NAb (HIVIG/2F5/2G12)V

irus leve

ls

• High levels of antibodies were used, much higher than in HIV infection.• Challenge dose was high• Antibody was matched to the virus (SHIV89.6PD), but would not have ‘matched’ most viruses in the ‘real world’

There is surprisingly little evidence that NAbs contribute

to protection from HIV in humans

HIV is both genetically and antigenically diverse

Neutr

aliz

atio

n s

ensitiv

ity

Easy to neutralize

Hard to neutralize

•At any given time, most people harbor a mixture of neutralization sensitive and neutralization resistant viruses due to the dynamic process of immune escape.

Studies of HIV-exposed cohorts: MTCT

� MTCT occurs in the presence of a HIV-specific NAb

response in the mother.� HIV-specific NAbs are present in the infant due to

passive transfer, especially near the time of delivery and

during breastfeeding

MTCT

Nairobi breastfeeding clinical trial 1992-1998:

Ruth Nduati

University of Nairobi

• Randomized clinical trial comparing infant HIV-1 infection between breast-fed and formula-fed groups (N=425) to determine the frequency and timing of breast milk HIV-1 transmission from a mother to her infant.

• In 1992: Risk was unclear and ARVs were not available.

Joan

KreissUniversity of Washington

Dorothy Mbori-Ngacha

Grace John-Stewart

Barb Richardson

Child age0%

10%

20%

30%

40%

6 wks

14 wks

6 months12 months

24 months

HIV Infection rate-Breastfeeding

HIV Infection rate-Formula feeding

Nduati, et al. Effect of Breastfeeding and Formula Feeding on Transmission of HIV-1.

JAMA 2000:283:1167-1174

Nairobi Breastfeeding Clinical Trial (1992-1998)

Breast feeding doubles the risk of infant infectionThe majority of breastfeeding transmission occurs in the first 6 weeks postpartum

% infe

cte

d

~ Sixty % of infants do not get infected despite

exposure in utero, intrapartum and through

breastfeeding

100%

80%

60%

40%

20%

0%

Do NAb contribute to protection?

uninfected

Do NAb play a role in protection of infants?

100%

80%

60%

40%

20%

0%

N=32

N=68

Select 100 infants in the

breastfeeding arm who

were HIV negative at birth

Examine the breadth of the NAb response

near delivery: mom and baby

Hypothesis: NAb contribute to protection

If a broad and potent NAb response

provides protection, then these infants are

less likely to be infected

To measure breadth and potency of the NAb

response, we use a panel of circulating HIV variants

Neutr

aliz

atio

n s

ensitiv

ity

This heterologous panel gives a measure or breadth and potency of the NAb

response

Easy to neutralize

Hard to neutralize

• A range of NAb sensitivities

Pla

sm

a R

NA

• Variants from early in infection representing circulating viruses

IC50s are defined as the reciprocal dilution of plasma or MAb required to inhibit infection by 50% (based on the dose response curve, log function).

Higher IC50 means more potent neutralization

Virus 1 IC50 = 620Virus 2 IC50

20 55

148

403

1096

IC50

Lynch et al. J. Virol. 2011

100 infa

nt pla

sm

a a

t birth

There was no difference in the breadth or potency of HIV NAb in infants who became infected versus those who remained

uninfected

Lynch et al., JV 2011

Similar results were obtained when we examined the breadth of the

maternal Nab response in relation to infant infection. Majiwa, submitted. J. Lynch

M. Majiwa

100%

80%

60%

40%

20%

0%

Do NAbs protect against the most neutralization sensitive viruses?

} Infected

Baby virus (week 6)

Mom virus(week 0)

Isolate maternal and infant envelope variants and

test neutralization sensitivity (N= 12 pairs, 96 envs)

NAbS

NAbR

NAbR?

Maternal antibodies (wk 0)

10

100

1000

Neutr

aliz

atio

n s

ensitiv

ity

IC50

va

lue

s v

ers

us a

uto

logou

s A

b

ne

ar

the

tim

e o

f in

fection

Transmitting

mothers

Infected

infants

Infant viruses variants are poorly neutralized by maternal NAbs.

Viruses

neutralized by

maternal

plasma

No detectable

neutralization

�p=0.02

Wu, JV, 2006X. Wu

10

100

1000

Neutr

aliz

atio

n s

ensitiv

ity

IC50

va

lue

s v

ers

us a

uto

logou

s A

b

ne

ar

the

tim

e o

f in

fection

Transmitting

mothers

Infected

infants

Infant viruses variants are poorly neutralized by maternal NAbs.

Sensitive variants are

not transmittedWu, JV 2006

Easy to neutraliz

e

Hard to neutraliz

e

Nab elicited by HIV infection may have adequate potency to block the most neutralization sensitive variants

Ab n

eed

ed t

o n

eutr

aliz

e

10

100

1000

Transmitting

mothers

Infected

infants

A broad and potent Nab response per se does not correlate with transmission because the mothers harbor a mix of neutralization sensitive and resistant viruses and the Nab are only able to get the ‘low hanging fruit”

Ab n

eed

ed t

o n

eutr

aliz

e

Mom’s virus

How does this fit with the first study, showing a lack of association

between breadth and potency and risk of infant infection?

(Mascola et al. Nat. Med. 2000)

NAbs can block HIV (SHIV) infection in macaques

Controls Given HIV NAb (HIVIG/2F5/2G12)V

irus leve

ls

• Antibody was matched to the virus (SHIV89.6PD), which was a very neutralization sensitive virus.

Easy to neutraliz

e

Hard to neutraliz

e

These data suggest together with animal model studies, suggest that Nab perhaps can protect against the more neutralization sensitive viruses

> The Nab breadth/potency needed for protection remains unclearA

b n

eed

ed t

o n

eutr

aliz

e

10

100

1000

Transmitting

mothers

Infected

infants

Current

SHIVs

Perhaps the protective antibodies are acting

locally, e.g. in breast milk to reduce

infectiousness?

Courtesy, Grace John-Stewart

Breastmilk NAbs are low

• Low level neutralizing activity is detected in BMS,

but much of this is non-specific.

• NAbs are only rarely detected in purified antibody

fractions (IgG, IgA).

Mabuka, J et.al PloS Path 2012J. Mabuka

1.11

1.67

Rousseau, JID 2004

Transmitting mothers have higher levels of cell-associated

breast milk HIV than non transmitting mothers.

Non transmitting Transmitting

Cell-

associa

ted B

M levels

(Log

10

BM

cell

DN

A/1

06 c

ells

)

Adjusted

p =0.002

C. Rousseau

Cell-free virus

RNA form

Cell-associated virus

DNA form

Infected

cell

Fc Receptor

ADCC : Antibody-dependent cell cytotoxicityLeads to elimination of infected cells

Antibody neutralizationPrevents new rounds of infection

Target cell (e.g. T Cell)

Effector cell (e.g. NK cell)

granzymes, perforin

Infected cell

ADCC assay measures killing of HIV coated cells in the presence of Ab.

HIV Envelope protein

(subtype A)

Target cell (CEM.NKr cells)

CFSE

PKH-26Effector cell

(PBMCs

Envelope Coated

Target cells

Uncoated

Target cells

CFSE

PK

H-2

6

BMS 1:100

*ADCC % Killing= %PKH+ cells, CFSE- cells - 2x background

42.8 57.24.0 96

+ +

BM Abs

Breast Milk ADCC is correlated with

risk of infant infection

J. Mabuka

Non transmitting Transmitting

p =0.039

Mabuka, J et.al PloS Path 2012

• NAb can protect against highly sensitive variants in MTCT, but the amount needed and the breadth to achieve a

protective Nab responses is unclear.

• Suggest the breadth and potency needed to protect against diverse HIV variants circulating globally is likely higher than elicited by chronic Hiv infection.

• Antibodies that act through ADCC may contribute to protection, especially in settings where cell-associated virus is contributing to transmission.

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