1. STAT3 and the Immune System Maureen Sherry Lynes February 29, 2012

2. Overview  STAT family; STAT3 signaling and structure  Role of STAT3 revealed by tissue specific knockouts  STAT3 and the anti-tumor immune response  Therapeutic approaches for targeting STAT3  Paper discussion

3. The STAT Family of Transcription Factors  7 mammalian STAT proteins  latent cytoplasmic transcription factors  Important in a wide range of physiological processes Adapted from Darnell and Levy, 2002

4. How is the STAT3 pathway activated? Cytokines, growth factors Secreted by diverse cell types in response to diverse processes Receptor specific Minegishi et al 2011

5. The STAT3 Signaling Pathway Negative Regulation of STAT3 signaling 1.Cytoplasmic phosphatases 2.SOCs proteins 3.Pias Proteins 4.STAT3ß

6. Domain Structure of STAT3

7. STAT3 Target Genes STAT3 Immortalization (hTERT) Apoptosis (Survivin, bcl-xl) Growth (C-myc, cyclin D) Angiogenesis (VEGF) Differentiation

8. Tissue Specific STAT3 Knockouts Brain: obesity, inability to control temperature, diabetes Skin: impaired wound healing, resistance to skin cancer Mouse Embryonic Stem Cells: loss of pluripotency Lung Macrophages and Neutrophils Liver Th17 development Gao et al, 2004.

9. STAT3 and the Acute Phase Response Liver: site of integration of signals from tissue, toxins and bacterial products in blood, etc Macrophages in the periphery: cytokine release Hepatocytes respond to cytokines (IL-6, IL-1 for example) and upregulate appropriate response proteins (clotting, innate responses to bacterial infection, toxins, etc) Hepatocyte deletion of STAT3- polymicrobial sepsis Similar bacterial load; decreased acute phase proteins, increased inflammatory cytokines such as IL-6 and TNFa LPS injection or cecal ligation and puncture: Sakamori et al 2007

10. STAT3 in macrophages and neutrophils Takeda et al, 1999. STAT3 deleted in cells expressing lysozyme M Mice were susceptible to LPS-induced endotoxic shock Macrophages constitutively activated

11. STAT3 in macrophages and neutrophils STAT3 -/- macrophages are resistant to IL-10 IL-10 normally dampens macrophage activation and response LysmCre STAT3 mice develop chronic enterocholitis

12. STAT3 in the lung epithelium House dustmite extract: induces allergic inflammation in the lung, as well as STAT3 activation Lung specific STAT3 KO: appears normal STAT3 lung KO: decreased airway hyperresponsiveness, immune infiltration upon HDM challenge Simeone et al, 2007.

13. STAT3 in Th17 Differentiation CD4+, defense against extracellular bacteria, fungi Pathogenic autoimmune responses Celiac, IBD, etc Fischer 2008

14. STAT3 in hyper IgE syndrome activation DNA binding

15. STAT3 in hyper IgE syndrome Recurrent skin and lung infections (Th17, lung epithelial defects?)

16. Activation, Duration, and extent of STAT3 signaling is tightly regulated Brain: obsesity, inability to control temperature, diabetes Skin: impaired wound healing, resistance to skin cancer Mouse Embryonic Stem Cells: loss of pluripotency Lung- promotes allergic inflammation Macrophages and Neutrophils- prevents constitutve activation, Liver: actue phase response; integration of signals from tissues Th17 differentiation Gao et al, 2004.

17.  STAT3 is constitutively activated in many human cancers  Inappropriately activated or overexpressed  Prevents apoptosis, stimulates migration, stimulates proliferation, increases angiogenesis  Upregulates cytokines that perpetuate STAT3 activation in the tumor and surrounding tissue STAT3 and tumorigenesis Wang, Rich et al. Stem Cells 2009

18. DN STAT3 induces melanoma regression in a mouse model  Transfection of B16 tumors with dominant negative STAT3 induces tumor regression  Bystander effect: regression disproportionate to transfection Niu et al Cancer Research 1999

19. STAT3 Suppresses Anti-Tumor Immunity  Cultured B16 cells transfected with DN STAT3: increased production of pro- inflammatory cytokines and chemokines (TNFa, RANTES, IP-10)  In vivo: enhances macrophage and neutrophil infiltration Wang et al Nature Med 2004

20. STAT3 Suppresses Anti-Tumor Immunity Yu et al Nature Reviews 2007

21. STAT3 as a therapeutic target

22.  Activation  EGFR antibodies; small molecules  Jak inhibitors  Multi-receptor kinase inhibitors  Protein-protein interactions  Small molecules to prevent dimerization  phosphoTyr peptides  DNA binding  Double stranded oligonucleotides  Nuclear translocation, natural inhibitors

23. Summary  STAT3 is a cytokine and growth factor activated transcription factor  STAT3 has a wide range of functions, both anti and pro inflammatory depending on the tissue and physiological context  STAT3 is also involved in disease states, such as asthma, colitis, and cancer  STAT3 can suppress anti-tumor immunity  STAT3 directed therapeutics is an active area of research