Candidate Microbicides: What we can learn from in vitro work

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Candidate Microbicides What we can learn from
in vitro work

• Guido Vanham, MD PhDgvanham_at_itg.be
• Institute of Tropical Medicine, Antwerp

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Vaginal HIV transmission
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HIV life CyclePotential targets for prevention
Direct DISRUPTION
FUSIONINHIBITORS
HIV
CD-4 CCR-5
TARGET CELL
ITM Y. Van Herrewege
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Possible classes of candidate microbicides

• - Buffers Acidform, Buffergel still in
  trial but only indirect antiviral action
• Virus disrupters Nonoxynol-9, Savvy (C31G)
  obsolete
• Non-specific binding inhibitors
• Cellulose sulphate Carraguard, PRO-2000
  Vivagel
• some failed, some still in trial but even in
  vitro weak anti-HIV activity
• Inhibitors of gp120CD4 (e.g. BMS806, BMS793)
• Inhibitors gp120CCR5 (e.g. TAK-779, Maraviroc)
• Inhibitors of gp120 DC SIGN (e.g. Mannan)
• Fusion inhibitors (e.g. T20, D-peptides)
• Reverse Transcriptase inhibitors (RTI) in trial
  
• Nucleotide RTI PMPA (Tenofovir) FTC (Truvada)
  
• NNRTI TMC120 (Dapivirin), UC781
• Integrase inhibitors e.g. L 870 812 (Raltegravir
  analogue) ?

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Clinical Research Process
Human (clinical)
Animal models
In Vitro

• Safety
• Rabbit vaginal irritation
• Efficacy to prevent infection
• NOD/SCID-PBL mice HIV
• Macaques (SIV or SHIV)

Safety - In low-risk women (Phase I) - In
representative population (Phase
I/II) Effectiveness (Phase III)
Activity against pathogen Cellular toxicity
profile
10 years
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In Vitro models to test HIV Microbicides
- Limited access (HT -) - Risk of damaging
epithelium
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Data on candidate microbicides in in vitro models

• 1) Cell suspension models
• A) Cell line (GHOST) single cycle pseudovirus
  
• B) DC/T4 co-culture monocyte-derived
  dendritic cells
• autologous T4 cells primary replicative
  virus
• 2) Models of female genital tract mucosa
• A) In vitro dual chamber model DC/T4
  epithelial cells on top
• B) Ex vivo cervico-vaginal explant tissue from
  hysterectomy

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Pseudovirus Test Screening
Production of luciferase
Infection
30
48h

Ghost-CD4-CCR5
Microbicide
Ba-L PV
Integrase
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Co-culture model of MO-DC and T4 cells

T4 cells
Cell-associated virus
PBMC-PHA/IL-2 activated cells
Cell-free virus
Compound
MO-DC
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Summary of cell suspension data
EC50 (nM)
Pseudovirus DC/T4 co-culture
Ghost-CCR5 Free HIV Cell-ass. HIV
BMS
5
326
gt 10,000 gt 10,000 4,500
Binding/fusion
TAK 779
42
848
55 140
T20
PMPA
82
92
125
RT inhibitors
TMC-120
2 3 2
7 111 52
UC781
Integrase Inh
L 870 812
9 183
1,250
Concluding All compounds active in PV/GHOST (lt
100 nM) Binding/fusion inhibitors less active
with repicative free HIV inactive with
cell-associated HIV Reverse Transc. Inh. very
active in all conditions Integrase Inhibitors
intermediate profile
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Dual Chamber model
In vivo
In vitro
Nature Rev 2006, Lederman MM
Nature Rev 2006, Lederman MM
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Effect of Binding Inhibitors and NNRTI against
Cell-associated HIV in dual chamber model
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Explant model
Cervical epithelium (Junction zone)
Migratory cells (DC T cells)
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Inhibition of cell-free infection in explant model
Various binding inhibitors
UC781 (NNRTI)
Cervical epithelium
Migratory cells
Conclusion Binding Inhibitors active, but less
against migratory cells NNRTI very active,
especially against migratory cells (From R
Shattocks group J Exp Med 2004 and J Virol
2005)
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Issues in further development of microbicides

• Incomplete knowlegde of transmission process
• - Cell-free or cell-associated virus ?
• - Which are the relevant target cells and
  receptors ?
• - Role of seminal and cervico-vaginal fluid
  factors ?
• - Role of normal vaginal flora/STD and
  vaginal practices ?
• Avoiding unwanted side-effects
• - Enhancing infection by epithelial damage or
  inflammation
• - Limiting therapeutic options by induction of
  resistance

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Which in vitro test is suitable and predictive?

• Impossible to say until first succesful human
  clinical trial,
• In the mean time
• Use several models reflecting aspects of sexual
  transmission
• e.g. DC and T cells ( epithelial cells)
• Explant model
• Inclusion of seminal and vaginal fluid
  factors
• In addition
• - Ensure activity agains cell-free and
  cell-associated HIV
• - Study optimal drug combinations
• - Thorough evaluation of toxicity
• - Study consequences of possible resistance
  development.

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ACKNOWLEDGEMENTS

• Collaborators
• Yven Van Herreweghe Katty Terrazas Youssef
  Gali
• Jo Michiels Laetitia Aerts Leo Heyndrickx
• Funding
• EUROPRISE sponsored this lecture
• EMPRO European Microbicides Program
• ANRS (France)
• IWT and FWO Scientific funds of Flemish
  government
• DGOS Belgian Ministry of Development
• AmfAR American Foundation for AIDS Research
• IPM/TIBOTEC
• CONRAD
• ITM institutional support