MOLECULAR BASIS of CANCER

1. MOLECULAR BASISof CANCER • NON-lethal genetic damage • A tumor is formed by the clonal expansion of a single precursor cell (monoclonal) • Four classes of normal regulatory genes • PROTO-oncogenes • Oncogenes Oncoproteins • DNA repair genes • Apoptosis genes • Carcinogenesis is a multistep process

2. TRANSFORMATION &PROGRESSION • Self-sufficiency in growth signals • Insensitivity to growth-inhibiting signals • Evasion of apoptosis • Defects in DNA repair: “Spell checker” • Limitless replicative potential: Telomerase • Angiogenesis • Invasive ability • Metastatic ability

3. Oncogenes Cell cycle Apoptosis Tumor Suppressor Inv. and Mets Angiogenesis Hanahan and Weinberg, Cell 100: 57, 2000

4. ONCOGENES • Oncogenes are mutated forms of cellular proto-oncogenes. • Proto-oncogenes code for cellular proteins which regulate normal cell growth and differentiation.

5. Five types of proteins encoded by proto-oncogenes participate in control of cell growth: Class I: Growth Factors Class II: Receptors for Growth Factors and Hormones Class III: Intracellular Signal Transducers Class IV: Nuclear Transcription Factors Class V: Cell-Cycle Control Proteins

6. Functions of Cellular Proto-Oncogenes 1. Secreted Growth Factors 2. Growth Factor Receptors 4. Nuclear Proteins: Transcription Factors 3. Cytoplasmic Signal Transduction Proteins 5. Cell Growth Genes

7. ONCOGENES • Are MUTATIONS of NORMAL genes (PROTO-oncogenes) • Growth Factors • Growth Factor Receptors • Signal Transduction Proteins (RAS) • Nuclear Regulatory Proteins • Cell Cycle Regulators • Oncogenes code for Oncoproteins

8. Mutations that confer these properties fall into two categories • Oncogene • : a cancer-causing gene that has been mutated to cause an increase in • activity, or the activity becomes constitutive, or a new activity is acquired. • -a mutation in a single allele is sufficient to transform cells (dominant). • -originally identified as viral proteins that resembled normal human proteins. • -the term "proto-oncogene" refers to the normal protein that has not been mutated

10. tumor Suppressor gene • : cancer-causing gene that has been mutated to cause a loss of activity. • -mutations are required in both alleles to transform cells (recessive)

11. 4 types of genetic mutations that contribute to cancer 1 2 4 3

12. Categories of oncogenes • A. Growth factors • -generally not directly involved transformation, but increased expression seen as part of • an autocrine loop due to changes in other steps in the same pathway

13. growth factor receptors • -They are transmembrane proteins with an external ligand binding domain and an • internal tyrsosinekinase domain. • -oncogenic mutations can result in dimerization and activation in the absence of • ligand • -more commonly, increased activity is a result of overexpression of receptors

14. Growth factor receptors • They are transmembrane proteins with an external ligand binding domain and an • internal tyrsosinekinase domain. • -Oncogenicmutations can result in dimerization and activation in the absence of • ligand • -More commonly, increased activity is a result of overexpression of receptors.

15. signal transducers • -Activated directly or indirectly by growth factor receptors • -Activation of signal transducers triggers a phosporylation cascade that ultimately • results in changes in gene expression at the transcriptional level. • -mutations in RAS • , a GTPase, are the most common oncogenic abnormality in tumors • -failure to hydrolyze GTP locks RAS in its active form.

16. Transcription factors • -Transcription factors contain DNA binding domains. • Sequences • Regulate expression of genes essential for passage through the cell cycle, or • regulation of apoptosis. • -

17. Normal CELL CYCLE Phases INHIBITORS: Cip/Kip, INK4/ARF Tumor (really growth) suppressor genes: p53

18. cyclins and cyclin-dependent kinases • -cyclins are only expressed at specific stages of the cell cycle • -cyclin-dependent kinases are expressed constitutively, but must bind cyclins for • activation; phosphorylation of target proteins essential for progression through • cell cycle

19. Regulation of G1/S cell cycle transition Cell cycle arrest at G1/S (in response to DNA damage or other stressors) is medicated through which gene? p53 (levels of p53 under negative regulation by MDM2 and p14 ARF)

20. a second level of control is achieved by CDK inhibitors • -p21 family (broad specificity) and the INK4 (p16) family (CDK4/6 • specific) • -overexpression of cyclin D and CDK4 common. • -phosphorylate and inactivate • Rb

25. 2) Activation Growth-Promoting Oncogenes Which signal transduction pathway is continuously activated by mutant RAS? MAP kinase pathway Point mutations of ras are seen in what % of all human malignancies? 15-20%

26. Tumor supressor gene • . Tumor suppressor were originally identified as inherited mutations that confer a • predisposition to cancer (familial form). • -inheritance is dominant, meaning a single defective allele is sufficient to confer • the predisposition

27. Inactivation of tumor suppressors can occur • Sporadically • -sequential inactivation of both alleles in somatic cells • You may hear the term • haploinsufficiency • , which refers to inactivation of a single • allele contributing to malignancy. • -usually not the initiating event, but exacerbating. • Viral inactivation • -HPV expresses proteins that inhibit Rb and p53 function.

28. P53 and RAS p53 RAS H, N, K, etc., varieties Single most common abnormality of dominant oncogenes in human tumors Present in about 1/3 of all human cancers • Activates DNA repair proteins • Sentinel of G1/S transition • Initiates apoptosis • Mutated in more than 50% of all human cancers

29. RB gene • a.Lossof RB function confers a predisposition to retinoblastoma. • occurs in both the familial form (early onset) and sporadic fromthe basis for tissue specificity of some tumor suppressors is unknown, but • presumably is due to the transcriptional profile of the tissue, determined by tissue • function

30. P53 • p53 is the most commonly mutated gene in tumors • -over 50% of all tumors lack functional p53 • - • Li-Fraumeni syndrome • : inheritance of a single defective copy of p53 results in a • predisposition to a wide spectrum of cancers. • -p53 is a transcription factor.

31. 1: Failure of DNA Repair (acquired) Normal function of p53 is to upregulate activity of which 2 genes to allow repair of DNA? p21 GADD45

32. Unlike Rb, p53 inhibits G1 progression only in response to DNA damage • -normally p53 is very unstable, due to proteolytic degradation triggered by • mdm2 • . • -p53 is phosphorylated in response to DNA damage; mdm2 no longer binds p53 • -p53 upregulates expression of p21, which in turn inhibits G1/S CDKs. • c. In response to excessive DNA damage, p53 can trigger apoptosis

33. Some other tumor suppressors found to be inactivated in tumors inhibit proliferation by • various mechanisms: • -APC: degradation of • b • -catenin, a transcriptional activator anchored to E-cadherins • -NF-1: activates GTPase activity of ras • -TGF- • b • receptor: a tyrosine kinase that upregulates expression of CDK inhibitors • -

34. -PTEN: dephosphorylatesinositol phospholipids, which act as dockingsites for • intracellular signaling proteins • VHL: transcriptional elongation • -WT-1: transcriptional regulator

35. MYC • Encodes for transcription factors • Also involved with apoptosis

36. Tumor (really “GROWTH”) suppressor genes • TGF-β COLON • E-cadherin STOMACH • NF-1,2 NEURAL TUMORS • APC/β-cadherin GI, MELANOMA • SMADs GI • RB RETINOBLASTOMA • P53 EVERYTHING!! • WT-1 WILMS TUMOR • p16 (INK4a)  GI, BREAST • BRCA-1,2  BREAST • KLF6  PROSTATE

37. Evasion of APOPTOSIS • BCL-2 • p53 • MYC

38. DNA REPAIR GENE DEFECTS • DNA repair is like a spell checker • HNPCC (Hereditary Non-Polyposis Colon Cancer [Lynch]): TGF-β, β-catenin, BAX • XerodermaPigmentosum: UV fixing gene • Ataxia Telangiectasia: ATM gene • Bloom Syndrome: defective helicase • Fanconi anemia

39. LIMITLESS REPLICATIVE POTENTIAL • TELOMERES determine the limited number of duplications a cell will have, like a cat with nine lives. • TELOMERASE, present in >90% of human cancers, changes telomeres so they will have UNLIMITED replicative potential

40. TUMOR ANGIOGENESIS • Q: How close to a blood vessel must a cell be? • A: 1-2 mm • Activation of VEGF and FGF-b • Tumor size is regulated (allowed) by angiogenesis/anti-angiogenesis balance

41. TRANSFORMATIONGROWTHBM INVASIONANGIOGENESISINTRAVASATIONEMBOLIZATIONADHESIONEXTRAVASATIONMETASTATIC GROWTHetc.

42. Invasion Factors • Detachment ("loosening up") of the tumor cells from each other • Attachment to matrix components • Degradation of ECM, e.g., collagenase, etc. • Migration of tumor cells

44. METASTATIC GENES? • NM23 • KAI-1 • KiSS

45. CHROMOSOME CHANGESin CANCER • TRANSLOCATIONS and INVERSIONS • Occur in MOST Lymphomas/Leukemias • Occur in MANY (and growing numbers) of NON-hematologic malignancies also

47. Carcinogenesis is “MULTISTEP” • NO single oncogene causes cancer • BOTH several oncogenes AND several tumor suppressor genes must be involved • Gatekeeper/Caretaker concept • Gatekeepers: ONCOGENES and TUMOR SUPPRESSOR GENES • Caretakers: DNA REPAIR GENES • Tumor “PROGRESSION” • ANGIOGENESIS • HETEROGENEITY from original single cell

48. Carcinogenesis: The USUAL (3) Suspects • Initiation/Promotion concept: • BOTH initiators AND promotors are needed • NEITHER can cause cancer by itself • INITIATORS (carcinogens) cause MUTATIONS • PROMOTORS are NOT carcinogenic by themselves, and MUST take effect AFTER initiation, NOT before • PROMOTORS enhance the proliferation of initiated cells

50. Q: WHO are the usual suspects? • Inflammation? • Teratogenesis? • Immune Suppression? • Neoplasia? • Mutations?