Epithelial Mesenchymal Transitions (EMT)In Cancer Metastasis

Greg Longmore, February 19, 2008

Post-Transcriptional Regulation of Snail/EMT

1. A. Cano et al., Nat. Cell Biol. 2:76-83, 2000 2. B.P. Zhou et al., Nat. Cell Biol. 6:931-40, 20043. Z. Yang et al., Cancer Res. 65:3179-84, 20054. H. Peinado et al., EMBO J. 24:3446-58, 20055. J.I. Yok et al., Nat. Cell Biol. 8:1389-406, 2006 6. E. Langer et al., Dev. Cell March 11, 2008

Reviews

1. J.P. Thiery and J.P. Sleeman Nature Rev. Mol. Cell Biol. 7:131-142, 2006 2. H. Peinado et al., Nature Rev. Cancer 7:415-428, 2007 3. A. Barrallo-Gimeno and M.A. Nieto. Development 132:3150-61, 20054. J.P. Thiery. Nature Rev. Cancer 2:442-54, 2002

Breast Cancer

1. S.E. Moody et al., Cancer Cell 8:197-209, 20052. N. Fujita et al., Cell 113:207-19, 20033. C. Xue et al., Cancer Res. 63:3386-94, 20034. A. Dhasarathy et al., Mol. Endocrinology 21:2907-18, 2007

1. Cancer Metastasis

2. EMT - MET - definitions

3. In Normal Development 4. In Adult Pathology

5. Signals that Induce EMT

6. Snail Family - Transcriptional regulators of EMT

7. Clinical - Breast Cancer

OUTLINE

1. Primary tumors (10%) rarely kill, metastases do (90%)

2. Primary tumor size often predicts for metastasis

3. Some tumors don’t metastasize (skin SCC, brain glioblastoma) while other do frequently (melanoma)

4. Some tumors have a propensity for specific tissue metastasis(breast, prostate - bone), while others are excluding from tissues

- when considering blood flow as a single variable

5. “micrometastases” at diagnosis - breast, colon - worse outcomes

6. Organ fibrosis is a significant risk factor for the development ofaggressive cancers (hepatic cirrhosis, lung fibrosis)

7. The metastatic process (Fig.)

Cancer Metastasis

INVASIONEMT

MET

EMT in DevelopmentGastrulation Neural Crest Delamination

Epithelial Mesenchymal

EMT in the Adult

- epithelia wound healing (skin)- tissue fibrosis in response to injury (lung, kidney, liver)- epithelial cancer metastasis

Skin wound healingSlug expression

Epithelial Mesenchymal Transition (EMT)

Altered Cell Morphology

Breakdown of Intercellular Junctions

Increased Cell Motility / Invasiveness

Mesenchymal Epithelial Transition (MET)

Lost or decreased1. Epithelial adhesion receptors - E-cadherin, Occludin, Claudins2. -catenin, -catenin frequently translocates to nucleus (Wnt) 3. Circumferential F-actin fibers4. Epithelial cytokeratins5. Apico-basal polarity

Acquired1. Intermediate filament protein - Vimentin2. Matrix metalloproteinases secreted, produced 3. Fibronectin secretion4. N-cadherin5. -smooth muscle actin (myofibroblasts)6. v6 integrin7. Motility, Invasiveness

Cellular changes during EMT

Tight junctionAdherens junction

Gap junctionDesmosome

Apical Surface

focal adhesions

Basolateral Surface

Epithelial Cells

Epithelial cell-cell adhesive complexes: general organization

Adherens Junctions: Cadherins - catenins - Actin

transmembrane receptor

cytoplasmic plaque proteins“scaffolding / adapter proteins”

Cytoskeletal elements

outside

inside

signal

transduction

polarityproliferation

cell fate

E-cadherin and Cancer pathogenesis“A metastasis tumor suppressor gene?”

1. Mouse models - TAG-insulinomas2. Germline mutations in CDH1 strongly predispose individuals to

gastric cancer and breast cancer3. Somatic inactivating mutations in CDH1 in gastric cancers and

infiltrative lobular breast cancers

4. But in the majority of cancers where CDH1 expression is lostmutations are rare or absent

(? Epigenetics or trans-acting factors)

E-cadherin (brown)

Colon Cancer

transformedhuman mammary cells

implanted in amouse

Does EMT occur in vivo?

Lung Fibrosis model: - -gal transgenic mice + TGFgenerate-gal + myofibroblasts

PyV-mT, FSP1.TK mice - less invasion and Metastasis following treatment with GCV

Other Data

Extrinsic Signals that Induce EMT:

- Tumor-derived (autocrine), Stromal Cell-derived (paracrine) - FGF, TGF-, EGF, HGF (scatter factor), Wnt, TNF- - E-cadherin cleavage (MMPs)- E-cadherin endocytosis

Intracellular Pathways:

- PI3K - Ras - MAPK, - GSK3, NF-B, p38, Smads, STAT3- Rac1b - ROS (MMP-3)

Transcriptional regulation:

- E2a/E47, FOXC2, SIP1, Snail, Slug, Twist

SIGNALING

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+ Snail

The Snail family of transcriptional repressors

Snail 264aa

Slug 269aa

SNAG Domain Zinc Fingers

Scratch 348aa

Smuc 292aa

Snail or Slug

FGF

Neural crestGastrulationLimb dev’pt

Tumor metastasis

Wnt

Neural crestHeart dev’pt

Tumor metastasis

TGFSkin

Palate fusionTissue fibrosisHeart dev’pt

Tumor metastasis

BMP

Neural crestL/R asymmetry

Mammary dev’ptgastrulation

Tumor metastasis

EGF

GSK3-mediatedphosphorylation

MTA3

Estrogens

BUT, There is only a modest inverse relationship between

Snail and E-cadherin expression (IHC, mRNA) in many metastatic cancers

With possibly one exception - breast cancer (see later)

SNAG Domain

Zinc Fingers

93 - SDEDSGKGSQPPSPPSPAPSSFSSTSVSSLE- 122

GSK3 nuclear export

GSK3cytoplasmicdestruction

S246

Pak1

K98 K137

LOX 2/3

Ajuba LIM proteins:- adapters that assemble

repressor complex (co-repressors)

Snail Modification/Function

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Inhibit GSK3 - increase Snail - decrease E-cadherin - metastasis

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Wnt - Axin2(mRNA) - GSK3 nuclear-cytoplasm - Snail nuclear - EMT/Invasion

How a Wnt signal cooperates with Snail to influence metastasis

Remember Wnt also inhibits GSK3 - stabilizes Snail, and - results in nuclear translocation of -catenin

EpithelialMarkers

E-cadherinClaudinsOccludins

DesmoplakinCytokeratins

Mesenchymalmarkers

FibronectinVitronectinVimentin

Cell shape changesCell movements, invasion

RhoBMMPs

Proliferation

Cyclin DCDK4

Rb phosphp21

Survival

PI3K activityERK activityCaspases

P53BID

Snail or Slug functions