Transmission of HIV-1Where Epidemiology Meets Biology

Myron S. Cohen, MD

J. Herbert Bate Professor

Medicine, Microbiology and Immunology and Epidemiology

The University of North Carolina at Chapel Hill

Epidemic Spread of HIV

Ro = bDC

When Ro >1 epidemic is sustained

b = Efficiency of transmission (…a biological event)

D = Duration of infectiousness

C = Number of people (partners) exposed

Transmission of HIV:Biological Requirements

Infectious Susceptibility

Inoculum (concentration) Hereditary resistance

Phenotypic factors Innate resistance

Acquired (immune)

resistance…from Cohen and Galvin

Nature Microbiology Reviews 2:33-42, 2004

UNC HIV Transmission Group

• The Swanstrom Lab (and CFAR)

• The Fiscus Lab

• The Eron Clinical Research Group (ACTG)

• The Kashuba Pharmacology Group

• The Hobb’s STD Research Group (CRC)

• The Hoffman Malawi Research Team

…and most recently the Margolis Lab

semensemenSI HIV (T-tropic)SI HIV (T-tropic)

NSI HIV (M-tropic)NSI HIV (M-tropic)

lamina proprialamina propriadendritic celldendritic cellCD4+CD4+CCR5+CCR5+DC-SIGN+DC-SIGN+

exposureexposureat mucosalat mucosalepitheliumepithelium

T cellT cell

CD4CD4

CCR5CCR5DC-SIGNDC-SIGN

migrationmigrationto lymphoid organsto lymphoid organs

V3 isn't variable for subtype C.

Routes of Exposure and H.I.V. INFECTION ROUTE RISK OF INFECTION

Sexual Transmissiona. Female-to-male transmission…………1 in 700 to 1 in 3,000b. Male-to-female transmission……...…..1 in 200 to 1 in 2,000c. Male-to-male transmission………...….1 in 10 to 1 in 1,600d. Fellatio??…………………………….. .0 (CDC) or 6% (SF)

Parenteral transmissiona. Transfusion of infected blood………….95 in 100b. Needle sharing………………………….1 in 150c. Needle stick…………………………..…1 in 200d. Needle stick /AZT PEP…………………1 in 10,000

Transmission from mother to infanta. Without AZT treatment………...…….1 in 4b. With AZT treatment………………….Less than 1 in 10

Royce, Sena, Cates and Cohen, NEJM 336:1072-1078, 1997

Coital Frequency per Month by Age

10.02

8.98 9.11

7.44

15-24 25-29 30-34 35-59

Age

4

5

6

7

8

9

10

11

Co

ital

fre

qu

ency

per

Mo

nth

2.6%

9.4%

2.0%

6.0%

2.1%

4.0%3.6%

4.1%

2.3%

11.0%12.2%

7.9%5.8%

13.8%

4.1%

14.4%

31.2%

25.0%

28.9%

26.3%

0%

5%

10%

15%

20%

25%

30%

35%

40%

45%

50%

55%

Age 15

Males

Females

95% Confidence Interval

Age 16 Age 17 Age 18 Age 19 Age 20 Age 21 Age 22 Age 23 Age 24

HIV Prevalence by Age and Gender among South African youth age 15-24 years

Pettifor A, et al. AIDS 2005, 19: 1525,

Pettifore et al. AIDS 2007 21:2007 862.

0

5

10

15

20

25

<=14 15 16 17 18 19 20+

Age of coital debut

Pe

rce

nta

ge

of

yo

un

g w

om

en

ou

t o

f to

tal p

op

ula

tio

n SA girls

US girls

Comparison of age of coital debut among young women aged 18-24 years in South Africa (SA) in 2003 and the United States (US) in 2002 (Pettifore et al., submitted)

Hypothesis

1) Estimated transmission rates from earlier studies are too low to explain the epidemic

2) HIV transmission is intermittently

AMPLIFIED by increased genital tract shedding

3) AMPLIFIED transmission is critical to the spread of HIV

Factors that Amplify HIV Transmission INFECTIOUSNESS• Stage of Disease … ACUTE INFECTION?• Systemic co-infections

-Malaria, Tuberculosis, Helminthic infections (?)• Genital Tract Changes -Bacterial vaginosis, STDs!!!• Circumcision • Hormonal contraception• Genetic factors (HLA B and C)SUSCEPTIBILITY• STDs!!!• Bacterial vaginosis• Inate immunity• Circumcision• Hormonal contraception• Genetic factors (CCR5)

A. Fauci, 2006

Why is Acute HIV Important? Pilcher and Cohen, J Clin Invest, 2005

• Vaccine Development (CHAVI)

• Prevention Opportunities (HPTN)

• Treatment

-for secondary prevention (HPTN)

-for viral load reduction (ACTG)

- to attack “persistance” (ACTG)

The acute retroviral syndrome

• 49-89% of patients “symptomatic” within 3 mos. Schacker Kinloch-de Loes

Fever 93% 87%Fatigue 93 26Pharyngitis 70 48Weight loss 70 13Myalgias 60 42Headache 55 39

….BUT LESS THAN 1,000 subjects with acute/early HIV have been reported out of 60,000,000 infected!!

EclipsePhase I II III IV V VI

Western + (p31+)

Western blot +/-

(Fiebig, AIDS 2003)v RNA+

Western blot + (p31-)*

* Env SGA sequence analysis of plasma virus populationTransmitted virus

Fiebig Classification of HIV-1 Infection

A

Individual testing on 10 specimens

10 pools of 10 screened

20 Screening Pools Tested

N=2000

Pooling Serum Specimensto Detect HIV RNA

(Pilcher et al JAMA 288:216-221, 2002)

NAAT+

-

F/U Testing(Ab+NAAT) - +

Acute HIVHIV Negative

-

EIA/WB +

LS-EIA at UNC

Likely recent “Detuned”

+

Unknown duration

-

Long Term HIV Positive

HIV Testing in North Carolina (n=109,250)

(Pilcher, Cohen et al NEJM 352: 1873, 2005

106 477 23

0123456789

10

Established HIV+ (n=66)

Acutely HIV + (n=21)

Lo

g H

IV R

NA

cp

/ml

median 209,18329,347

Viral Loads at Initial Detection

ADDED BENEFIT OF ROUTINE AHI SCREEINGLOCATION TESTING

POPULATIONN ANTIBODY + HIV

PREVALENCE, %INCREASED YIELD

WITH AHI, %

NORTH CAROLINA1 ALL PUBLIC TESTING

109,250 0.5 3.6

SAN FRANCISCO2 STD CLINIC 3075 3.4 10.5

LOS ANGELES3 STD CLINIC 1712 0.8 7.1

SEATTLE4 MSM ONLY 6395 2.4 13.5

ATLANTA5 VCT AND

STD CLINIC

2202 3 6.1

JOHANNESBERG, SOUTH AFRICA6

VCT AND

STD CLINIC

1906 35.2 1.8

LILONGWE, MALAWI7

STD CLILIC

MALE

929 46.8 5

LILONGWE, MALAWI8

STD CLINIC

ALL

1450 40.5 3.6

PORTO ALEGRE, BRAZIL9

VCT CLINIC 933 19.1 2.8

1PILCHER, 2KLAUSNER, 3PATEL, 4STEKLER, 5PRIDDY, 6STEVENTS, 7PILCHER, 8FISCUS, 9 deSOUZA

Infection by testing site: NC(Pilcher et al. NEJM, 2006)

0

50

100

150

200

250

STD HIV Testing

“Other” Prison,Jail

N= 44656 11688 7575 3053

Not shown: Prenatal/OB FP Drug Treatment General Medical TB Field visits

Acute

Recent

Unknown Duration

16/23 Acute Infections from STD Clinics

STDs Amplify HIV-1 Transmission• Reducing physical/mechanical barriers

• Increasing HIV in genital lesions, semen or both

• Evoking a more infectious HIV variant

• Increasing the number of receptor cells or the density of receptors per cell

…and co-transmission of HIV and STDs leading to clusters of subjects with acute HIV infection

Malawi Overview

• Population 10 million

• 90% rural

• Per Capita income $190

AIDS impact• 900,000 people living with HIV

• 15% adult prevalence

• STD Clinic: 47% prevalence

The Tidziwe Center Lilongwe Central HospitalLilongwe, Malawi

Dean Roper Visits Malawi

Acute HIV Infection and STDsPilcher et al. AIDS 18:1-8, 2004

• 1,361 men screened in STD and Dermatology Clinics in Lilongwe Malawi

• 47% antibody + (chronic HIV Infection)• 2.1% (28) with acute HIV (antibody -, RNA +)

Inguinal nodes: 11.4% acute HIV

Genital ulcer (HSV): 7.8% acute HIV

CSW exposure: 9.1% acute HIV

Acute HIV was detected ONLY in symptomatic STD patients (!!) implying co-transmission or “staged” transmission of an STI followed by HIV, or vice-versa.

0

2

4

6

8

10

Lo

g H

I V R

NA

cp

/ ml

HIV Ab+ HIV Ab- (acute)

Viral Loads in Malawi: Chronic and Acute HIV Infection

1

3

5

7

9

0 4 8 12 16 20 24 28

log

10 H

IV-1

RN

A c

op

ies

per

mL

Weeks Since HIV infection

HIV-1 viremia (grey) and Semen (black) Pilcher et al. AIDS August 2007

Grey=Blood and Black=semen (95% CI)

HIV-1 Viremia and SheddingPilcher et al. AIDS August 2007

wk4 wk8 wk16

Established HIV Infection

CD4<350

1

3

5

7

9

CD4>350

Acute HIV Infection

log

10H

IV-1

RN

A c

opie

s p

er m

L

HIV

RN

A in

Sem

enH

IV R

NA

in S

emen

(Log

(Log

1010

copi

es/m

l) c

opie

s/m

l)

Acute Infection

Acute Infection

3 wks3 wks Asymptomatic

Asymptomatic

InfectionInfection

HIV Progression

HIV ProgressionAIDSAIDS

00

33

55

77

1/1000 - 1/1000 - 1/10,0001/10,000

1/500 - 1/500 - 1/20001/2000

1/100-1/100-1/10001/1000

Risk of TransmissionRisk of TransmissionReflects Reflects Genital Genital Viral Burden Viral Burden

1/30-1/30-1/2001/200

Sexual Transmission of HIV (Cohen and Pilcher, JID May 2005)

HIV

RN

A in

Sem

enH

IV R

NA

in S

emen

(Log

(Log

1010

copi

es/m

l) c

opie

s/m

l)

Acute Infection

Acute Infection

3 wks3 wks STD Episode

STD Episode

STD Episode

STD EpisodeAIDSAIDS

00

33

55

77

Acute HIV and STD episodes(Cohen and Pilcher, JID, 2005

Prevention of HIV1. STD control, behavior change, condoms2. Vaccines (Trials Ongoing) 3. Treatment of Bacterial Vaginosis (Planning)4. Topical microbicides (Trials Ongoing)5. The diaphragm (Trial Completed)6. Male circumcision (Trials Ongoing)7. HSV-2 treatment/prevention (Trials Ongoing) 8. Antiretroviral therapy (Trials Ongoing)9. Societal (structural) interventions: needle

exchange, poverty reversal, etc…ALL WORK

Impact of MC on HIV : Evidence from observational studies and RCTs

85 80 70 60 .50 1

Reduction of risk

(95% CI)

South Africa (RCT) 60 ( 76, 33)

Kenya (RCT) 59 ( 76, 30)

Uganda (RCT) 51 ( 82, 14)

Overall 58 ( 66, 48)15

17

1

1

Reduction of risk (0%)

Bailey et al. Lancet 2007; 369: 643–56

Weiss et al. AIDS 2000, 14:2361-70

Auvert et al. PLoS Med 2005(11): e298.2006

Gray et al. Lancet, 2007, 657–66

Antiviral TherapyCohen et al. Annals Internal Med. 2007

Effect on Sexual Transmission of HIV

?

ART to Prevent Sexual Transmission of HIV

• Treatment of the infected person?

• Post-exposure prophylaxis (PEP)?

• Pre-Exposure Prophylaxis (PREP)?

NFV (5%)

LPV (5%)

SQV (3%)

EFV (3%)

RTV (3%)

d4T (2%)

ENF (ND)

Male Genital Tract Exposure percent of blood plasma

Kashuba et al. and CROI 13 Abstract 569 (Vourvahis), 13th CROIAbstracts 396 (Stekler), Abstract 618 (Katzenstein)

IDV (100%)

NVP (70%)APV (20%)

0 50% 100% 150% 200% 500% 600%

TDF (500%) 3TC (600%)ZDV (200%)ABC (150%)

NNRTIPI FINRTI

NVP (80%)

APV (50%)

ABC (40%)

ABC (150%)

Female Genital Tract Exposure(percent of blood plasma)

Dumond et al. Abstract 129, 13th CROI

ZDV (200%)

IDV (200%)

0 200% 400% 600%

TDF (400%)

3TC (400%)

NNRTIPINRTI

LPV (30%)

ATV (30%)

RTV (20%)

DLV (20%)

FTC (600%)ddI (100%)

SQV(ND)

EFV (0.6%)

d4T (4%)

HIV-RNAHIV-RNA HIV-DNAHIV-DNA

00

2020

4040

6060

8080

100100

Pa

tient

s (%

) w

ithP

atie

nts

(%)

with

det

ect

ab

le H

IV in

se

me

nd

ete

cta

ble

HIV

in s

em

en

n=55n=55

n=114n=114

Controls (drug naive) Controls (drug naive)

Potent ARTPotent ART

p<0.0001p<0.0001

p=0.025p=0.025

ART Suppresses HIV in Semen:ART Suppresses HIV in Semen:Biological PlausabilityBiological Plausability

Vernazza, Cohen Vernazza, Cohen et al.,et al., AIDS, 2000 AIDS, 2000

0 2 4 6 8 10 12 140

25

50

75

100

Number of rectal exposures

% U

nin

fect

ed a

nim

als

Controls (n = 18)

Injectable FTC (n = 6); p = 0.005 [HR = 3.9]

Injectable FTC/Tenofovir (n = 6)

Oral truvada (n = 6); p = 0.0004 [HR = 7.8]

Oral TDF (n = 4); p = 0.095

Potential end-points of HIV-vaccine efficacy trials

UNAIDS–97100 1 August 1998

“normal” infectionwith variable levels

of viral load

no protection

no infection

protection against HIVsterilizing immunity

protection against disease (modification of the course of HIV infection in vaccine recipients)

initial infection“controlled”

establishment of chronic infection with low viral load

This CHAVI web site is at http://www.chavi.org

Interdisciplinary CHAVI 001 Studies

CHAVI 001Acute HIV-1 Infection

T Cell DiscoveryGood vs. Bad T cells

B Cell DiscoveryAntibody Studies

Genetic Data

Sexual BehavioralSexual NetworksSexual Dynamics

Transmitted SequencesViral Biology

Structural Biology

Apoptosis

Computational BiologyRo Analyses

Mucosal ImmunityInnate Immunity

Cytokines

HIV- Acute HIV

• Massive reduction in mucosal CD4+ T cells even in acute infection

The Consequences of Acute Infection

Douek and Schacker 2004J Exp Med

Onset of Innate Immune Responses During Acute HIV-1

Infection

(Kessler/Borrow/Norris)

Summary of changes in plasma cytokine levels in AHI

-10 -5 0 5 10 15 20

Time relative to T100

Vira

l loa

d or

cyt

okin

e le

vel

IFN

Plasma viraemia

IL-15

IL-18 TNF

IFN

IL-10

SAA elevated

Apoptosis of CCR5+, CD4+ T Cells is a Hallmark of

Pathogenic Retrovirus Infections

10-5

10-4

10-3

10-2

10-1

0

101

102

103

104

105

106

107

108

Transmission

Vir

us

Conce

ntr

ati

on in E

xtr

ace

llula

r Fl

uid

or

Pla

sma (

Copie

s/m

l)

Tumor Necrosis Factor Related Apoptosis Inducing Ligand (TRAIL) And Apoptosis In

Acute HIV-1 Infection

Time Post Exposure (days)

0 5 10 15 20 30 3525 40 45 50 55 60 65 70

Pla

sma M

arke

rs

TRAIL

Plasma Apoptotic Microparticles

Window of Opportunity

Viral Load

Smith, Crossman, Haynes, 2007

Onset of Antibody Responses in Acute HIV-1 Infection

Summary Of Antibody Responses Immediately Following Acute HIV-1

Infection

0

2

4

6

8

10

-20 -10 0 10 20 30 40 50 60 70 80

Viral Load

gp41 gp140 Env V3 CD4BS

2F5, 4E10,2G12

Transmission

Non-Neutralizing MPER

Cluster II

Vir

al L

oa

d O

r A

ntib

ody

Re

spo

nse

s

Days Of Observation

Autologous NAb

10-5

10-4

10-3

10-2

10-1

0

101

102

103

104

105

106

107

108

Transmission

Vir

us

Conce

ntr

ati

on in E

xtr

ace

llula

r Fl

uid

or

Pla

sma (

Copie

s/m

l)CD8 T Cell Responses In Acute HIV-1

Infection: Too Little Too Late

Time Post Exposure (days)

0 5 10 15 20 30 3525 40 45 50 55 60 65 70

Anti-HIV-1CTL Activity

CTL A

ctivity

Viral Load

Window of Opportunity

Limit of detection of

assay for plasma virus

R. Koup, D. Watkins, A. McMichael et al.

Acute HIV and the Virus

• A unique signature sequence using single gene amplification?

• Glycosylation sites?

• Antibody neutralization resistance?

• Envelope length?

• Transmitted drug resistance?

The Transmitted Virus HIV Env Characterization by SGA

The Transmitted Virus HIV Env Characterization by SGA

cDNAsynthesiscDNA

synthesis

1:21:2

1:41:41:81:8

1:161:161:321:321:641:64

1:1281:1281:2561:256

100%100%

100%100%100%100%92%92%67%67%33%33%17%17%0%0%

sequencesequencegenotypegenotype

phenotypephenotypeclone/

pseudotypeclone/

pseudotype

1:641:64

serial dilutionsserial

dilutions

The Transmitted/Early Virus-The First 15 Days of Infection

• In 77 subjects (80%), one monophyletic env lineage, suggesting productive infection by a single viral infectious unit.

• In 19 subjects (20%), more than one monophyletic env lineage was identified, indicating productive infection by multiple viral infectious units.

• In 13 (14%) multiply infected subjects, recombinants were identified.

The Transmitted/Early Virus-The First 15 Days of Infection

• Sensitive to MPER antibodies 2F5, 4E10

• Sensitive to HIV-1 entry drugs, Fuzeon (T20) and T1249

• Variably sensitive to sCD4, HIV Ig, Mab 1b12

• Insensitive to V3 antibodies

• R5 co-receptor usage

Phylogenetic analysis of SGA-derived env from donor blood/semen vs. recipient plasma

X4-tropic

R5-tropic

1. 36 amplicons from donor blood—4.7% nucleotide diversity 11-X4 tropic, 25 R-tropic2. 15 amplicons from donor semen (S)—1.5% nucleotide diversity all X4-tropic3. 29 amplicons from recipient blood—0.2% nucleotide diversity all X4-tropic

Results:

Conclusions:1. Single X4-tropic variant transmitted2. Recipient env populations were extremely homogenous3. Donor env populations are heterogenoeous4. Semen compartment has significantly less diversity and localized enrichment of variants vs. blood plasma5. Transmission of X4-tropic variant that was detected in donor blood and not semen, suggests cell-associated vs. transmission of minor variant

Treatment of Acute HIV??Walker et al., 2004, Kinloch-de Loes, JID 2006

• Many examples of lack of benefit measured by REBOUND VIREMIA (Streck et al. JID, 2006)

• AIDRP and CASCADE: Some modest benefit?• SPARTAC ongoing (Fidler, Weber et al.)

…But remember limited by

i) small n

ii) subject heterogeneity

iii) rebound as the definition of success/failure

Effects of ART on “The Latent Pool” Margolis: AHI (CROI 2007, Triangles) Chun: Early HIV (JID 2007, Squares)

UNC HIV Transmission Group

• The Swanstrom Lab (and CFAR)

• The Fiscus Lab

• The Eron Clinical Research Group (ACTG)

• The Kashuba Pharmacology Group

• The Hobb’s STD Research Group (CRC)

• The Hoffman Malawi Research Team