Special approaches of tumor biology and chemotaxis

Orsolya Láng

2012.

TUMOR CELLS AND MIGRATION

METASTASISPRIMARY TUMOR

AngiogenezisAdhesion

CELL and CELL CYCLE

Growth factors

Adhesion Adhesion moleculesmolecules

ChemokineChemokiness

Regulatory Regulatory proteinsproteins

ApoptosisApoptosis

TUMOR CELL

CELL KINETICS

Doubling time of the tumor volume (Td)

Time of the Cell cycle (Tc):Tc= Ts / Li

Ts: S phaseLi: labeling index

(proportoin of cells in S phase)

Growth fraction(GF):GF=P / (P+Q’)

P: number of the mitotic cellsQ: number of the cells in interphase

Rate of the cell loss ():= 1-Tpd / Td Tpd= *Ts/Li

Tpd: Potential tumor volume doubling timeTd: tumor volume doubling time

Lymphoma 48 h

Lung cancer 108 h Usually 15-125 h

Lymphoma 4 weeksColon adenoma 90 weeksUsually 18-200 days

Volume of the tumor tissue

~10 division =*1000 cell number increase (210 =1024)~20 division= 106 cells = 1 mg= 1 mm3

~30 division= 109 cells = 1 g= 1 cm3

~40 division= 1012 cells = 1 kg

1 tumor cell ~30-33,25 division=1-10 cm3

Time of the clinical symptomes / diagnosis

~27 division= 0,1cm3

Earliest time of diagnosis

~40 division= 1012cellFatal

BIOLOGY OF THE TUMORPROGRESSION

Exogen and endogen factorsGenom instability

Activation of the oncogeneInactivation of tumorsuppressors

Epithelialcell

Hiperplasticadenoma

DisplaticCarcinoma in situ

Invasive and metastasis carcinoma

Local and systemic factorsinhibition acceleration

Growth rate

Ectopic survival capacity

Invazivity

De-Differentation

Tumorigenesis

Important steps of tumor progression

Transformation of the microenvironment: stromal cells,ECM components,proteolytic degradation

Induction of the angiogenesis Escaping from immune-mediated rejection Formation of metastasis

MICROENVIRONMENT – STROMAL CELLS

Cell types:fibroblasts, myofibroblasts, endothelial cells, lymphocytes, macrophages

Function: host defence

! MALT - B cell helps to maintain lymphomas

! Growth factors are released by the stromal cells (VEGF-angiogenesis)

ANGIOGENESIS

Hypoxia formation of new vessels, proliferation of the endothelial cellsTypes: vessels

arteriovenous shunts„dead end”

/lack of smooth muscle , weak vessel wall, irregular shape(insuficient endothelial cell and basement membrane layers)/

sinuses /wall is formed by tumor cells/Venous circulation

VEGF induces angiogensisincreases permeability

Lack of lymphatic vessels

OEDEMA, decresed blood flow

Strategies that tumors use to escape from immune-mediated rejection are:

To decrease the antigen expressionTo inhibit the immune-reactive cells:

degrade the chemoattractansdecrease their cell adhesioninhibite their phagocytotic activity

Angiogenesis

Local invasion

ECM

AdhesionProteolysisMigration

Intravasation

Extravasation

circulation

Metastasis

Tumor cell

Primary tumor

AdhesionProteolysisMigration

Angiogenesis

VEGFAngiogeni

nFGF

spreading

METASTATIC CASCADE

INVASION

In situ carcinoma

DECREASED CELL ADHESION, INCREASED MOTILITY

ECM proteolysis

Angiogenesis

Local invasion

ECM

AdhesionProteolysisMigration

Intravasation

Extravasation

circulation

Metastasis

Tumor cell

Primary tumor

AdhesionProteolysisMigration

Angiogenesis

VEGFAngiogeni

nFGF

spreading

METASTATIC CASCADE

CELL ADHESION

Significant change in cell-cell and cell-ECM interactionsMolecules:

selectinsintegrinsimmunoglobulin superfamily cadherins catenins

SELECTINS

Cell-cel junctionsTypes:

E- endothelial cellsP- trombocytesL- leukocytes

Extracellular C-lectin domain Ca2+ dependent anchorage It binds Sialyl-Lex carbohydrates

„ROLLING”

! Tumor cells express increased amount of sialil-Lex or -Lea

INTEGRINS

Transmembrane receptorsForm cell-ECM interaction8 , 14 subunites ~20 heterodimerCa2+, Mg2+ dependent anchorage„RGD” sequence is the specific substrateSignalling: outside-in – signalling

inside-out – adhesionIncreased expression of integrins promotes angiogenesis and helps to bind MMPs at the cell surfaceEXTRAVASATION, ATTACHMENT

D

G

R

Integrin or celladhesion regulated signalling pathways

cellproliferation

PTEN

RAC PI(3)K

CDC42

integrin

ECM

ILK

-cateninCiklin D1BAD

PKB/AKT

FAK

RASRAFMEKMAPK

GSK3

motilitygene expression

cellcycleapoptosis

SHCGRB2/SOS

Integrin or celladhesion regulated signalling pathways

integrin

ECM

ILK

-kateninCiklin D1BAD

PKB/AKT

FAK

cellproliferation

RASRAFMEKMAPK

GSK3

motilitygene expression

cellcycleapoptsis

SHCGRB2/SOS

PTEN

RAC PI(3)K

CDC42

Molecular partners of the integrins

Cytoskeletal components:actinin, talin,F- actin, filamin

Adaptors:rack 1, ICAP-1

Calcium binding proteins:CIB, calreticulin

Protein kinases:pp125FAK, p59 ILK

Membrane proteins:CD9, CD16,CD47…caveolin, urokinase-plazminogen-activator receptor

Ligands in ECM:collagen, laminin, fibronectin, fibrinogen, von

Willebrand factor, osteopontin, elastin

IMMUNGLOBULIN SUPERFAMILY

has 5 Ig-like domains at the extracellular regionforms cell-cell junction interacts with integrins

VCAM - 41, PECAM - v3

takes essential part in extravasation

! ! Over expression of ICAM-1, MUC18 increased inavsion

! ! Down-regulation of VCAM-1 increased metastatic potential (faster detachment)

CADHERIN

Is a transmembrane glycoproteinForms homophyl cell-cell junctionsCa2+ dependent anchorageClassical types: E- epithelial

P- placentaN- neural,

Intracellular part interacts with catenins to connect aktin filaments

! Increased expression invasion

CATENINCATENIN

Is an intracellular moleculeFixes cadherins to F-actin

! Catenin expression is often decreased in carcinomas

! -catenin binds to the az APC gén termékéhez

Increases

N-Cadherin

B-Catenin

SrC

Ras

Ihibits

E-Cadherin

aE-Catenin

cMET

FGFR

PTEN

Adhesionmolecules

Signal pathways

Factors influencing the metastatic potential of the melanoma cells

Angiogenesis

Local invasion

ECM

AdhesionProteolysisMigration

Intravasation

Extravasation

Circulation

Metastasis

Tumor cell

Primary tumor

AdhesionProteolysisMigration

Angiogenesis

VEGFAngiogeni

nFGF

spreading

METASTATIC CASCADE

IntegrinscadherinsSelectins

CAM

PROTEOLYSISComponents of the basement membrane(BM) and ECM: IV collagen, laminin, proteoglycanesTumorcells (stromal cells) secrete proteasesCathepsinMatrix metalloproteinase (MMP)Plazmin, tPA ,Urokinase (plasminogen activator inhibitor 1&2)TIMP

INVASION

Tissue inhibitor of metalloproteinases

MATRIX METTALLOPROTEINASESMATRIX METTALLOPROTEINASES (MMP)(MMP)

MOLECULAR STRUCTURE OF THE MATRIX MOLECULAR STRUCTURE OF THE MATRIX METTALLOPROTEINASESMETTALLOPROTEINASES

SUBSTRATE OF TIMP

MMP/TIMP EXPRESSION IN BREAST MMP/TIMP EXPRESSION IN BREAST CANCERCANCER

MMP – TUMORPROGRESSION?!?

Angiogenesis

Local invasion

ECM

AdhesionProteolysisMigration

Intravasation

Extravasation

Circulation

Metastasis

Tumor cell

Primary tumor

AdhesionProteolysisMigration

Angiogenesis

VEGFAngiogeni

nFGF

spreading

METASTATIC CASCADE

IntegrinscadherinsSelectins

CAM

MMP/TIMPCathepsin

Plasminogen

MIGRATORY MECHANISMS IN TUMOR

Small-cell lung cancer

FORMS OF MIGRATORY ADAPTATION

2D –3D MIGRATIONS

STEPS OF 3D MIGRATION

1. Pseudopod protrusion2. Formation of focal contact3. Focal ECM proteolysis4. Actomyosin contraction5. Detachment

Cell-cell interactions visualized in tumorigenesis

Angiogenesis

Local invasion

ECM

AdhesionProteolysisMigration

Intravasation

Extravasation

Circulation

Metastasis

Tumor cell

Primary tumor

AdhesionProteolysisMigration

Angiogenesis

VEGFAngiogeni

nFGF

spreading

METASTATIC CASCADE

IntegrinscadherinsSelectins

CAM

MMP/TIMPCathepsin

Plasminogen

AMF/gp78AutotaxinHGF/c-MET

!! Tumor markers e.g. cytokeratin, mucin

HEMATOGENIC DISSEMINATION

EXTRAVASATION

?

Attachment

Migration

LOCALISATION OF THE METASTASIS

CHEMOKINES – TISSUE SPECIFIC LOCALISATION

adhesion

moti

lity

?