Seattle/Kenya Collaboration- MTCT Julie Overbaugh Collaborating Institutions : University of Nairobi Fred Hutchinson Cancer Research Center University of Washington
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? Antibody Target cell (e.g. T Cell)
NAbs can block HIV (SHIV) infection in macaques Controls Given HIV NAb (HIVIG/2F5/2G12) Virus levels ( Mascola et al. Nat. Med. 2000) • 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 •At any given time, most Easy to people harbor a mixture of neutraliz neutralization sensitive and e Neutralization sensitivity neutralization resistant viruses due to the dynamic process of immune escape. Hard to neutraliz e
Studies of HIV-exposed cohorts: MTCT 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
Nairobi breastfeeding clinical trial 1992-1998: Dorothy Mbori-Ngacha Grace John-Stewart Ruth Joan Barb Richardson Nduati Kreiss University of University of Nairobi Washington • 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.
Nairobi Breastfeeding Clinical Trial (1992-1998) Breast feeding doubles the risk of infant infection The majority of breastfeeding transmission occurs in the first 6 weeks postpartum 24 months 12 months 6 months 40% 14 wks 6 wks 30% % infected 20% 10% 0% Child age 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
~ Sixty % of infants do not get infected despite exposure in utero, intrapartum and through breastfeeding Do NAb contribute to protection? 100% 80% uninfected 60% 40% 20% 0%
Do NAb play a role in protection of infants? Select 100 infants in the breastfeeding arm who 100% were HIV negative at birth 80% N=68 60% 40% 20% N=32 0% 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 • A range of NAb sensitivities Easy to neutraliz e Neutralization sensitivity • Variants from early in infection representing circulating viruses Plasma RNA Hard to neutraliz e This heterologous panel gives a measure or breadth and potency of the NAb response
Neutralization is quantified by determining IC50 values IC50s were determined for each virus/plasma combination Virus 1 IC50 = 620 Virus 2 IC50 <100 1:100 1:200 1:400 1:800 1:1600 1:3200 Plasma dilution 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
Lynch et al. J. Virol. 2011 100 infant plasma at birth 20 55 IC50 403 148 1096
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 J. Lynch maternal Nab response in relation to infant infection. Majiwa, submitted. M. Majiwa
Do NAbs protect against the most neutralization sensitive viruses? 100% 80% 60% } 40% Infected 20% 0%
Isolate maternal and infant envelope variants and test neutralization sensitivity (N= 12 pairs, 96 envs) Mom virus (week 0) Baby virus (week 6 ) NAb R ? NAb S NAb R Maternal antibodies (wk 0)
Infant viruses variants are poorly neutralized by maternal NAbs. � p=0.02 IC50 values versus autologous Ab Neutralization sensitivity Viruses 1000 near the time of infection neutralized by maternal plasma 100 No detectable 10 neutralization Transmitting Infected mothers infants Wu, JV, 2006 X. Wu
Infant viruses variants are poorly neutralized by maternal NAbs. Sensitive variants are IC50 values versus autologous Ab Neutralization sensitivity not 1000 near the time of infection transmitted Wu, JV 2006 100 10 Transmitting Infected mothers infants
Nab elicited by HIV infection may have adequate potency to block the most neutralization sensitive variants Easy to neutraliz e 1000 Ab needed to neutralize 100 10 Transmitting Infected mothers infants Hard to neutraliz e
How does this fit with the first study, showing a lack of association between breadth and potency and risk of infant infection? A broad and potent Nab response per se does not Ab needed to neutralize 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” Mom’s virus
NAbs can block HIV (SHIV) infection in macaques Controls Given HIV NAb (HIVIG/2F5/2G12) Virus levels ( Mascola et al. Nat. Med. 2000) • Antibody was matched to the virus (SHIV89.6PD), which was a very neutralization sensitive virus.
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 unclear Easy to neutraliz e Current SHIVs 1000 Ab needed to neutralize 100 10 Transmitting Infected mothers infants Hard to neutraliz e
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 2012 J. Mabuka
Transmitting mothers have higher levels of cell-associated breast milk HIV than non transmitting mothers. Cell-free virus RNA form Non transmitting Transmitting Cell-associated BM levels 1.67 (Log 10 BM cell DNA/10 6 cells) Adjusted 1.11 p =0.002 Infected cell Cell-associated virus DNA form C. Rousseau Rousseau, JID 2004
Antibody neutralization ADCC : Antibody-dependent cell cytotoxicity Prevents new rounds of infection Leads to elimination of infected cells Effector cell Fc Receptor (e.g. NK cell) granzymes, perforin Target cell (e.g. T Cell) Infected cell
ADCC assay measures killing of HIV coated cells in the presence of Ab. CFSE Effector cell PKH-26 (PBMCs + + HIV Envelope protein (subtype A) BM Abs Targ et cell (CEM.NKr cells) BMS 1:100 4.0 96 42.8 57.2 PKH-26 Uncoated Envelope Coated Target cells Target cells CFSE *ADCC % Killing= %PKH + cells, CFSE - cells - 2x background
Breast Milk ADCC is correlated with risk of infant infection p =0.039 Non transmitting Transmitting Mabuka, J et.al PloS Path 2012 J. Mabuka
Take home • 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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