Non-Infectious Papilloma virus–like Non-Infectious Papilloma virus–like Particles (VLPs) inhibit HIV-1 Particles (VLPs) inhibit HIV-1 replication: Implications for immune replication: Implications for immune control of HIV-1 replication by IL-27.control of HIV-1 replication by IL-27.

Laboratory of Human Retrovirology CSP / NIAID-Frederick

J.Mohamad FakruddinJ.Mohamad Fakruddin

BACKGROUNDBACKGROUNDImmune system uses a range of soluble molecules to suppress viral infections.

Action of non-cytolytic antiviral factors to control HIV-1 replication.

Some viral infections elicit the secretion of soluble HIV-1 suppressor factors.

HPV-like particles(VLPs) bind to dendritic cells and induce the expression of anti-viral cytokines such as IFN-, IL-10 and IFN-.

VLPs have been used as a vaccine to prevent HPV infection.

BACKGROUNDBACKGROUND

VLP StructureVLP Structure

To study the potential ofVLP induced soluble HIV-1 suppressive factors.

AIMAIM

Cells+PHA 3-days

+ HIV-1 X4(NL4.3) or R5(AD8) 2hr/370C/CO2

Cultured with VLP

7 or 10 days

HIV-1 replication evaluated by p24 ELISA

METHODSMETHODS

VLP inhibits HIV-1 replication in PBMCsVLP inhibits HIV-1 replication in PBMCs

X4 virus R5 virus

VLP suppresses HIV-1 replication regardless of the virus tropism

HIV-1 inhibition is Papilloma virus specificHIV-1 inhibition is Papilloma virus specific

SV40 VLP has no effect on replication of HIV

Pretreatment of PBMC with VLP inhibits HIV-1Pretreatment of PBMC with VLP inhibits HIV-1

24 hr pretreatment is sufficient for inhibition

VLP does not alter the expression of HIV receptors

Effect of VLP on HIV-1 receptorsEffect of VLP on HIV-1 receptors

VLP does not inhibits HIV in CD4VLP does not inhibits HIV in CD4++ T cells and CD8 T cells and CD8++ cells not required cells not required

Effect of VLP on subset of PBMCsEffect of VLP on subset of PBMCs

Media

VLP

VLP binds strongly to Monocytes

VLP exerted its inhibitory activity on PBMCs, but not on CD4+ cells.

VLP inhibitory effect is due to anti-HIV soluble factors or by cell to cell interaction.

To study the mechanism- The effect of conditioned medium(CM) derived from VLP treated PBMCs on HIV infected CD4+ T-cells was investigated.

Is there a role for soluble factors?

Effect of VLP treated CM on CD4Effect of VLP treated CM on CD4++ T cells T cells

VLP induces soluble anti-HIV factor(s) from both PBMCs and MDMs

Global gene response by microarrayGlobal gene response by microarray

One point study to examine transcriptional changes.

MDMs and PBMCs at 24h or 36h after exposure to VLP.

Fold Increase

Gene ID Gene name PBMC MDM

24678 IL-27 14.0 20.0

6355 MCP-2 31.0 30.0

10673 TNSF13B 2.6 2.5

3586 IL-10 NS 2.3

8743 TNSF-10 6.4 111.0

3600 IL-15 NS 4.7

6373 CXCL11 85.0 54.0

3627 CXCL10 10.0 58.0

6347 MCP-1 15.0 NS

3469 INF-

143.0 NS

3456 IFN-

135.0 NS

Expression of cytokine genesExpression of cytokine genes

Detection of of anti-HIV cytokines and IL-27 in CMDetection of of anti-HIV cytokines and IL-27 in CM

Significant increase of IL-27 in both PBMCs and MDMs

Blocking the activity of anti-HIV factorsBlocking the activity of anti-HIV factors

VLP stimulated anti-HIV activity is partially blocked by -IL-27

IL-27 expression is specific for HPV-VLPIL-27 expression is specific for HPV-VLP

SV40 VLP has no effect on IL-27 expression

IL-27 is key factor of VLP mediated HIV inhibition

- many fold increase of IL-27 by VLP was observed at both RNA and protein levels.

- in neutralization experiments only IL-27 blocked VLP inhibition.

IL-27 is a member of the IL-6/IL-12 family of cytokines.

IL-27 produced by dendritic cells and monocytes.

Expression of IL-27R can be detected in NK, B cells and T cells.

Nature of the IL-27/IL-27R interaction is pleiotropic.

IL-27 is an immunomodulator as it promotes pro- and anti-inflammatory response.

Biology of IL-27

IL-27 suppresses X4 replication in IL-27 suppresses X4 replication in CD4CD4++ T cells T cells

IL-27 suppresses X4 replication in IL-27 suppresses X4 replication in PBMCsPBMCs

IL-27 suppresses R5 replication in IL-27 suppresses R5 replication in MDMsMDMs

Conclusions Conclusions VLP displayed anti-HIV activity in PBMCs and MDMs.

This inhibitory effect was lost in the enriched CD4+

T-cells.

Soluble factors play a major role in VLP mediated anti-HIV effect.

Neutralization experiments implicated IL-27 to be one of the mediator anti-HIV VLP effect.

Further studies identified IL-27 as a novel anti-HIV cytokine.

Elucidation of the molecular mechanism of action of IL-27 may lead to HIV therapeutics.

ConclusionsConclusions

Tomozumi Imamichi Cliff Lane

Ligia Pinto Alfonso Pineres

Richard Lempicki Joseph Adelsberger

Jun Yang Catherine Watkins

Cathi Yeager

AcknowledgementsAcknowledgements