Insights into normal cell biology Targets for diagnosis and follow-up

1. What can we learn from the identification of specific molecular abnormalities in malignant disease? • Insights into normal cell biology • Targets for diagnosis and follow-up • Targets for rational drug design

2. Conventional cytotoxic drugs mainly act by causing DNA damage and cell death Studying the biology of cancer cells may provide new targets for drug development

3. Signal transduction modules Molecular links between changes in cell environment and cellular responses

4. Signalling pathways control cell functions Replicate Move Live/Die

5. Signal transduction modules Molecular links between changes in cell environment and cellular responses • e.g. • Erythropoietin and prevention of apoptosis in • erythroid progenitors • G-CSF and proliferation in myeloid progenitors

6. The hallmarks of cancer Many of these features may result from abnormalities in signalling components (Hanahan & Weinberg (2000) Cell 100, 57)

7. P P P P Ligand binding dimerizes receptor tyrosine kinases resulting in their activation Monomeric receptor Dimeric receptor No ligand Ligand present

8. P P P P A number of signalling modules link growth factor receptor binding to changes in cell function PI3-kinase Ras MAPK STAT PKB Activation of gene transcription

9. The Ras protein acts as a molecular switch in response to changes in the external environment of the cell Growth factor OFF Ras.GDP Exchange factor e.g. SOS GTPase activating protein e.g. NF-1 Ras.GTP ON Proliferation Survival Movement

10. SH3 SH2 SH3 SH2 SOS SOS P P P P GRB2 GRB2 Recruitment of a Grb2-SOS complex to an activated receptor tyrosine kinase mediates Ras activation RAS GTP GDP

11. Examples of signalling pathway abnormalities in haematological malignancy Aberrant tyrosine kinase Bcr-Abl CML activity Increased Ras activity point mutation AML loss of NF1

14. The constitutive activity of the Bcr-Abl tyrosine kinase bypasses the requirement for growth factors Bcr-Abl Ras PI3-kinase MAPK STAT PKB Activation of gene transcription Increased proliferation/survival

15. Examples of signalling pathway abnormalities in haematological malignancy Aberrant tyrosine kinase Bcr-Abl CML activity Increased Ras activity point mutation AML loss of NF1

16. AML AML Normal

17. Ras proteins are frequently activated by point mutation in human cancers OFF Ras.GDP • Carcinoma • pancreas • colon • thyroid • AML • Myeloma Exchange factor e.g. SOS NF-1 MUTANT Ras.GTP ON Proliferation Survival Invasion

18. Loss of the NF-1 protein results in excessive Ras activation OFF Ras.GDP • Neurofibromatosis • Myeloid leukaemias Ras.GTP ON Proliferation Survival Invasion

19. Molecular targets in leukaemia therapy Signal transduction pathways Dysregulated kinases eg Bcr-Abl Mutant Ras proteins Apoptosis pathways Bcl-2, NF-kappaB, p53 Differentiation pathways Retinoic acid receptor Histone deacetylases

20. Imatinib mesylate inhibits the activity of Bcr-Abl by competing with ATP and is effective in the treatment of CML

21. Addition of a farnesyl (C15) moiety is required for Ras proteins to be active F Plasma membrane Ras -C-OMe Farnesyl transferase active Ras -CAAX Cytoplasm inactive

22. Targeting Ras proteins by inhibiting membrane localisation Plasma membrane FT Inhibitors Farnesyl transferase Ras -CAAX Cytoplasm inactive

23. IkB The transcription factor NF-kB induces transcription of pro-survival genes and is constitutively activated in a variety of tumours NIK Degradation by proteasome NEMO IKK1 IKK2 P IkB NF-kB NF-kB Increased transcription eg Bcl-2

24. IkB Inhibitors of proteasomal activity prevent NF-kB activation by blocking IkB degradation Proteasome inhibitor Eg PS-341 NIK NEMO IKK1 IKK2 P IkB NF-kB IKK inhibitors Reduced transcription

25. What can we learn from the identification of specific molecular abnormalities in malignant disease? • Insights into normal cell biology • Targets for diagnosis and follow-up • Targets for rational drug design