Fundamental research on breast cancer in Belgium Rosita Winkler

1. Fundamental research on breast cancer in BelgiumRosita Winkler

2. Medline search for « breast cancer » and Belgium limits: english, posted in the last 5 years. • Result: 484 papers - fundamental / clinical - according to the abstracts ???? • Identified the following topics • A. The breast cancer cell : • The estrogen receptor (ER) • ERBB2 gene • Expression of other breast cancer associated genes • Breast cancer genomics B. Invasion, metastasis • Epithelial to mesenchymal transition (EMT) • Proteases • Metastasis

3. 1. Estrogen Receptor (G. Leclercq, Bordet Inst.) ER17p P295 T311 • The P295-T311 sequence: • involved in ER transcriptional activity and stability • Calmodulin (CaM) binding site -> transcriptional activity, stability

4. ER17p • ER17p: estrogenic response in ERexpressing breast cancer cells • proliferation, transcription • ER expression • CaM binding • Binds ER • Dissociates the ER /HSP70 complex-> activation of ER • hypothesis: P295-T311 sequence in the full length ER has a repressive function • Hypothesis: ER17p could be produced endogenously by ER degradation by the proteasome and exert estrogenic effects in breast cancer cells • ; production could be enhanced by treatment with hormone and hormone agonist -> sustained estrogenic effect.

5. 2. Mechanisms of ERBB2/HER2 overexpression in breast cancers • In ± 20% breast cancers erbB2/HER2 protein level is very high. • Main mechanisms: • gene amplification • Increased transcription rates

6. HER2/ERBB2 gene amplification(Vanden Bempt I., De Wolf-Peeters C, KUL) Characterization of the ERBB2 amplicon - Identification of 4 chromosomal regions (≠ ERBB2 gene) over-expressed in ERBB2 gene amplified breast cancers; ex : MMP9 Other co-amplified chromosomal regions but no pattern common to all breast cancers with ERBB2 gene amplification. High level over-expression associated with gene amplification not with chromosome 17 polyploidy

7. ERBB2 ERBB2/HER2 gene expression regulation (ULg) • Identification of : • Several enhancer regions, bound by • Activator proteins - AP-2 family transcription factors - overexpressed in ERBB2 positive breast cancer cells and in subpopulation of ERBB2 positive primary breast cancers

8. 3. pRB/E2F: regulate the expression of Brip1, CHK1 genes in breast cancers: (Verstuyf A., KUL) S-phase - proliferation Brip1/BRCA1 • Brip1 - (BRCA1 interacting protein) / CHK1mutations -> increased breast cancer risk. • Brip1 and CHK1 expression: • associated with cell proliferation • controlled by E2F transcription factor • Brip1 Increased in grade 3 ER-/PR-/HER2+ cancers • CHK1 increased in grade 3 ER-/PR-/HER2- cancers

9. A.4 Identification of new breast cancer susceptibility genes: rat model(Szpirer, ULB) • 10-15% of human breast cancers are associated with susceptibility genes - only 30% of these have been identified. How identify low penetrance genes? • Rat mammary tumours interesting model : • similar developments and histological features as human breast cancers • strongly hormone dependent • no virus involvement • susceptible and resistant strains • Comparison of the genome of resistant / susceptible strains: • Different genes involved in chemically/hormone induced cancers • Different loci control tumour multiplicity, latency, and aggressiveness.

10. B. Breast cancer cells invade and metastasize They acquire the capacity to move (EMT) They produce or stimulate the stromal cells to produce proteases necessary for invasion, angiogenesis, metastasis They form distant metastasis

11. Epithelial to Mesenchymal Transition - EMT (G. Berx Ghent, C. Gilles, Liège) Epithelial cells are tightly bound by “adherens junction” and their basal surface lies on a basal membrane.

12. EMT: epithelial to mesenchymal transition: transient process during which epithelial cells loose their epithelial characteristics and acquire mesenchymal properties: loss of adherens junctions, increased motility; described during embryogenesis - gastrulation. Involved in the metastatic process. Probably followed by mesenchymal to epithelial transition (MET).

13. For EMT cells are reprogrammed by re-expressing “Master regulators”: Twist, Snail , Slug transcription factors. • Twist, Snail , Slug: • control embryonic EMT • upregulated in metastatic cancers. • induce the replacement of the epithelial specific proteins by mesenchymal proteins.

14. Sip1 inhibits E-cadherin expression and stimulates vimentin expression in breast cancer cells (Berx G. RUG, Gilles C. Ulg) Nanos (Van Roy, Ghent) Sip1 E-cadherin (Berx G., Ghent) MCP-1 (Gilles C, Lg) Sip1 Vimentin (Gilles C., Liège)

15. MMP produced by tumoral and stromal cells during tumour progression

16. Proteases and breast cancer progression uPAR/ uPA/PAI/plasmin system controls angiogenesis MT1-MMP expressed by myofibroblasts in contact with the cancerous cells MMP2 (activated by MT1-MMP) expressed by several cell types MT4-MMP protein is over-expressed in primary breast cancers and node metastasis but mRNA levels are not increased MT4-MMP over-expression accelerates the growth of mammary cancer cells in immunodeficient mice. (Noel A., Foidart JM, Liege) SNP in MMP8 gene promoter associated with the expression level of the gene (Paridaens and Ye, KUL).

17. Bone metastasis (Castronovo, Lg) Osteotropic breast cancer cells express bone extracellular matrix proteins Regulatory factors and promoter regions are being identified