Conclusions

Abstract

Materials and MethodsIPA 7.5 software platform was used to perform all data uploads and in silico analyses.

Where applicable, publicly available data sets from the referenced articles were

uploaded into IPA, and analyzed. Fold change of 1.5 of treated-to-control experiments

and p<0.05 cutoffs were used where applicable to analyze the data. Ingenuity

Knowledge base was used as a reference set. Only direct interactions were included

where downstream molecules were analyzed.

References and Acknowledgements1. “Potential Ebola Therapies from Phenotypic screening,”

,

Feb 2008, ASM poster

2. “Genome-wide identification of estrogen receptor alpha-binding sites in mouse

liver,” Gao H, Falt S, Sandelin A, Gustafsson JA, Fahlman-Wright K, Mol

Endocrinol. 2008 Jan;22(1):10-22

3. “

”

4. “

,”

The authors would like to thank Dr. Chan Whiting, Dr. Dan Richards and Dr. Megan

Laurance for their valuable contributions, and to acknowledge that the datasets used in

this poster are publically available and are obtainable from the reference cited here in

this poster.

Results

Antoaneta Vladimirova1, Gene Olinger2

1Ingenuity Systems, Redwood City, CA and 2USAMRIID, Ft. Detrick, MD

Figure 2. Top Impacted Pathways and Functions by Genes with ER-alpha binding Sites in

Mouse Liver. A.) Top Clomiphene-impacted pathways in Pathogen-influenced signaling, Humoral

and Cellular Immune Response and Cytokine Signaling categories B.) Top significantly impacted

functions and genes involved. C.) Detailed view of specific functions, genes and the corresponding

p-values involved in impacted T lymphocyte response. Blue bars indicate significance. Orange line

indicates ratio.

In Silico

Figure 3. Clomiphene Gene Expression Data in Rat Liver Reveals Key Modes of Drug-induced

Protective Immune Responses. A) Top Impacted Signaling Pathways in Pathogen-influenced

signaling, Humoral and Cellular Immune Response and Cytokine Signaling categories B.) The

expression levels of the genes that were measured in this dataset are indicated near their respective

nodes. B) Il-10 Signaling pathway with Clomiphene 5 day expression data overlay. The expression

bar charts next to each gene show expression in 1d and 3d experiments as well. Up-regulated genes

are depicted in red and down-regulated – in green color. C) Selected significantly impacted functional

categories, functional annotations, p-values and respective genes that are impacted by Clomiphene

5d. D.) Processes consistently increased by gene expression by Clomiphene 5d and their

corresponding p-values and genes. E.) Top Induced molecules by Clomiphene along with bar chart

showing expression for all three doses. F.) Key impacted pathways by top induced molecules by

Clomiphene 5d and the directly-interacting molecules downstream of them.

A.

Figure 4. Gene expression data from Ebola-infected non-human primate PBMCs were

compared to Clomiphene-expressed genes in rat liver to identify key impacted pathways

significantly modulated by both treatments. A.) Top impacted pathways by both Ebola and

Clomiphene. Blue bars indicate significance. Orange line indicates ratio. B.) IL-10 Signaling with

expression data overlaid from Ebola-infected NHP PBMCs. Up-regulated genes are depicted in red

and down-regulated – in green color.

Figure 1. Biological processes and Pathways regulated by ER1 and ER2. A.) Genes

Downstream of ER1 and ER2 surface receptors (360 genes). Highlighted are genes expressed in

immune cells, kidney and liver. B.) B. Top impacted IPA Signaling Pathways by genes signaling

downstream of ER1 and ER2. in Pathogen-influenced signaling, Humoral and Cellular Immune

Response and Cytokine Signaling categories. Blue bars indicate significance. Orange line indicates

ratio.

A. B.

B.

B.

A.

C.

Function Annotation Molecules # Molecules P-value

increases developmental process of antigen presenting cells (9/11) BCR, DLL1, EGR1, FOS, IL6, IL1B, RACGAP1, RB1, TYROBP 9 4.12E-03

increases differentiation of B lymphocytes (7/8) CD14, DLL1, EGR1, FOS, IL6, INPP5D, TCF12 7 3.89E-03

increases transactivation of AP1 consensus site (4/4) FOS, JUN, JUNB, RUNX1 4 1.01E-03

increases activation of IL-6 response element (3/3) IL6, JUN, STAT3 3 1.44E-05

increases sensitization of macrophages (2/2) IL6, IL1B 2 3.44E-03

Figure 5. Depiction of T Helper Cell Differentiation pathway with gene expression data

overlay. A.) Expression data from Ebola-infected NHP PBMCs at day 2. B.) Clomiphene 5 day

gene expression data . Up-regulated genes are depicted in red and down-regulated – in green

color.

C.

D.

E. F.

A. B.

A. B.

B.

•

•

and may confer the therapeutic effect of Clomiphene

against

•

•

• Analysis of

Function Category Function Annotation P-value MoleculesAntigen Presentation Sensitization of macrophages 3.44E-03 IL6, IL1B

Cell Signaling

Protein kinase cascade 9.18E-04 ADORA2B, ADRA1B, EGFR, GJA1, IL6, IL1B, IL1RL1, JAK3, MAP3K7IP1,

OTUD7B, PDGFRB, PEBP1, PRKAG2, PSEN1, REN, SHANK3, STAT3,

TGM2, ZFP36

Development of Th17 cells 1.52E-03 IL6, IL1B, STAT3

Differentiation of Th2 cells 1.16E-03 DLL1, GATA1, IL6, IL1RL1, JUNB, RUNX1

Infiltration of T lymphocytes 3.57E-04 CCR5, CXCL10, IL6, IL1B, REN, STAT3, TNFRSF9

Trafficking of T lymphocytes 1.08E-03 CCR5, CXCL10, TNFRSF9

Activation of lymphocytes 1.13E-03 BCR, C1QA, CD1D, DLL1, EGR2, HSPA4, IL6, IL1B, IL1RL1, INPP5D,

JAK3, PLXNB1, PTPRJ, REN, TNC, TNFRSF9, TNFSF13, TYROBP

Activation of mononuclear leukocytes 3.28E-04 BCR, C1QA, CD14, CD1D, DLL1, EGR2, HSPA4, IL6, IL1B, IL1RL1,

INPP5D, JAK3, LBP, PLXNB1, PTPRJ, REN, TNC, TNFRSF9, TNFSF13,

TYROBP

Developmental process of antigen

presenting cells

4.12E-03 BCR, DLL1, EGR1, FOS, IL6, IL1B, LIFR, RACGAP1, RB1, STAT3,

TYROBP

Developmental process of phagocytes 4.37E-04 BCR, DLL1, EGR1, FOS, IL6, INPP5D, LIFR, RACGAP1, RB1, RUNX1

Differentiation of leukocytes 2.80E-04 BCR, CD14, CD1D, DLL1, EGR1, FOS, GATA1, IL6, IL1B, IL1RL1,

INPP5D, JAK3, JUN, JUNB, LAMP1, LIFR, RACGAP1, RB1, RUNX1,

STAT3, TCF12, TNFSF13, ZFP36

Infiltration of leukocytes 9.71E-04 CCR5, CNP, CXCL10, IL6, IL1B, IL1RL1, INPP5D, PLEC1, REN,

SERPING1, STAT3, TG, TGM2, TNFRSF9, UCP2

Infiltration of lymphocytes 3.41E-04 CCR5, CXCL10, IL6, IL1B, REN, STAT3, TG, TNFRSF9, UCP2

Invasion of macrophages 1.75E-03 CXCL10, IL1B

Activation of AP1 consensus site 7.61E-05 ATF3, FOS, IL1B, ILK, JUN, JUNB, RB1, RUNX1

Activation of ATF binding site 2.25E-05 ATF3, FOS, JUN, JUNB

Activation of Ets element 1.01E-03 ALAS2, FOS, GATA1, JUN

Activation of IL-6 response element 1.44E-05 IL6, JUN, STAT3

Activation of NF-E2 binding site 1.38E-04 ALAS2, GATA1, NFE2

Transactivation of AP1 consensus site 1.01E-03 FOS, JUN, JUNB, RUNX1

Differentiation of B lymphocytes 3.89E-03 CD14, DLL1, EGR1, FOS, IL6, INPP5D, TCF12, TNFSF13

Differentiation of macrophages 1.97E-03 BCR, DLL1, EGR1, IL6, LIFR, RACGAP1, RB1

Differentiation of phagocytes 2.41E-04 BCR, DLL1, EGR1, IL6, INPP5D, LIFR, RACGAP1, RB1, RUNX1

Quantity of hematopoietic progenitor cells 3.89E-04 BCR, GATA1, GJA1, IL6, INPP5D, JAK3, STAT3, TNFRSF9

Infection Mechanism Expression of virus 1.49E-03 FOS, GRN, GTF2B, RB1, TCERG1

Inflammatory Response Sensitization of macrophages 3.44E-03 IL6, IL1B

Cellular Movement

Cellular Development

Gene Expression

Hematological System

Development and Function

Cell-mediated Immune

Response

Cell-To-Cell Signaling and

Interaction