Introduction to Cancer Biology part II – Oncogenes and tumour viruses

1. Introduction to Cancer Biology part II – Oncogenes and tumour viruses Stjepan Uldrijan Department of Biology Faculty of Medicine Masaryk University Brno uldrijan@med.muni.cz

2. Genes commonly altered in cancer • Proto-oncogenes / Oncogenes – they get activated in cancer cells. • Code for proteins that are usually involved in signal transduction and regulate cell growth, proliferation, or differentiation. • Mutations or increased expression of these proteins lead to the increase in their activity in cell – a proto-oncogene becomes an oncogene – a tumour-inducing agent. • Examples: Myc, Ras, Src, HER2, etc. • Tumour suppressors – their function is commonly lost in cancer cells • Prevent growth and survival of aberrantly growing or damaged cells. • Some tumour suppressors take part in DNA repair and prevent the accumulation of potentially oncogenic mutations. • Examples: p53, p16, Rb, APC, MLH1, MSH2, etc.

3. Part 1: Cellular oncogenes

4. Examples of cellular oncogenes IV. VI. I. II. III. VI. V. IV. VI. • Autocrine growth factors • Receptor tyrosine kinases • Ras oncogene • Non-receptor tyrosine kinases • Raf kinase • Transcription factors Hanahan D, Weinberg RA. The hallmarks of cancer. Cell. 2000;100:57-70, Adapted

5. Autocrine growth factors • In general, normal cells do not produce a growth factor ligand whose receptor they also display on the surface • Cancer cells sometimes activate the expression of genes coding for such growth factor. This creates an auto-stimulatory loop (autocrine signalling). Example: HGF (hepatocyte growth factor) and HGF-receptor (c-Met) overexpression in invasive breast cancer Weinberg RA. The biology of cancer, Garland Science 2007

6. Examples of cellular oncogenes IV. VI. I. II. III. VI. V. IV. VI. • Autocrine growth factors • Receptor tyrosine kinases • Ras oncogene • Non-receptor tyrosine kinases • Raf kinase • Transcription factors Hanahan D, Weinberg RA. The hallmarks of cancer. Cell. 2000;100:57-70, Adapted

7. Growth factor receptors – receptor tyrosine kinases • Normally functioning growth factor receptor emits cytoplasmic signals in response to binding ligand • Cancer cells sometimes achieve increased mitogenic stimulation by overexpression of a growth factor receptor • Mutations in the genes coding for receptors can cause the receptor to become always active, i.e in the absence of ligand binding Weinberg RA. The biology of cancer, Garland Science 2007

8. Weinberg RA. The biology of cancer, Garland Science 2007

9. Dimerization induced by ligand binding is required for the activation of many growth factor receptors Weinberg RA. The biology of cancer, Garland Science 2007

10. Gene fusion can cause constitutively dimerized and active receptors Chromosomal translocations and fusions are common in most cancer cells In a number of malignant tumours, receptor dimerization occurs when the genes encoding growth factor receptor become fused to unrelated genes that code for proteins that normally dimerize. The receptor portions of these hybrid fusion proteins are dragged together by the dimerizing proteins to which they have become joined. Weinberg RA. The biology of cancer, Garland Science 2007

11. Examples of cellular oncogenes IV. VI. I. II. III. VI. V. IV. VI. • Autocrine growth factors • Receptor tyrosine kinases • Ras oncogene • Non-receptor tyrosine kinases • Raf kinase • Transcription factors Hanahan D, Weinberg RA. The hallmarks of cancer. Cell. 2000;100:57-70, Adapted

12. Receptor tyrosine kinase auto-phosphorylation enables the formation of specific protein complex that activates protein Ras Lynda Chin Nature Reviews Cancer 3, 559-570 (August 2003)

13. Activation of Ras requires the release of GDP and binding of GTP Ras binds GDP in its inactive state and GTP in its active, signal-emitting state. Inactive Ras protein is stimulated by a GEF (guanine nucleotide exchange factor) to release GDP and acquire GTP instead, placing Ras into its active, signalling configuration. This active state is transient and Ras soon returns to its inactive state after GTP is hydrolyzed. Oncogenic Ras mutations block the hydrolysis of GTP and lead to constitutively active Ras. Weinberg RA. The biology of cancer, Garland Science 2007

14. Weinberg RA. The biology of cancer, Garland Science 2007

15. Examples of cellular oncogenes IV. VI. I. II. III. VI. V. IV. VI. • Autocrine growth factors • Receptor tyrosine kinases • Ras oncogene • Non-receptor tyrosine kinases • Raf kinase • Transcription factors Hanahan D, Weinberg RA. The hallmarks of cancer. Cell. 2000;100:57-70, Adapted

16. Non-receptor tyrosine kinases - Abl kinase www.antigenics.com/diseases/cml.html Weinberg RA. The biology of cancer, Garland Science 2007 The Philadelphia chromosome is detected in > 95% of cases of chronic myelogenous leukemia. Reciprocal chromosomal translocations between chromosomes 9 and 22 result in the formation of hybrid Bcr-Abl proteins, which retain the tyrosine kinase activity but their activity is not properly regulated.

17. Examples of cellular oncogenes IV. VI. I. II. III. VI. V. IV. VI. • Autocrine growth factors • Receptor tyrosine kinases • Ras oncogene • Non-receptor tyrosine kinases • Raf kinase • Transcription factors Hanahan D, Weinberg RA. The hallmarks of cancer. Cell. 2000;100:57-70, Adapted

18. B-Raf • a serine/threonine kinase involved in transduction of mitogenic signals • mutated in various types of cancer, including malignant melanoma, ovarian carcinoma, thyroid cancer, and colorectal cancer • B-Raf is mutated in about 70 % malignant melanoma • B-Raf mutation is an early event in melanoma development – it is present in 80% of nevi, which are the first lesions associated with this tumour Lynda Chin Nature Reviews Cancer 3, 559-570 (August 2003)

19. Examples of cellular oncogenes IV. VI. I. II. III. V. VI. VI. IV. • Growth factors • Receptor tyrosine kinases • Ras • Non-receptor tyrosine kinases • Raf • Transcription factors Hanahan D, Weinberg RA. The hallmarks of cancer. Cell. 2000;100:57-70, Adapted

20. Transcription factors • Their expression in cancer cells is often increased by gene amplification • within a chromosome (homogeneously staining region) • in autonomously replicating fragments of chromosome (double minutes) Multiple copies of myc oncogene (yellow) in a human tumour cell line (red - chromosomes). One homogeneously staining region is indicated by the arrow. Weinberg RA. The biology of cancer, Garland Science 2007

21. RTK = receptor tyrosine kinase, TF = transcription factor Weinberg RA. The biology of cancer, Garland Science 2007

22. N-myc amplification and childhood neuroblastoma prognosis A B A: The N-myc gene is amplified in about 40% of pediatric neuroblastomas. This amplification, which is associated with the formation of either double minutes or homogeneously staining regions (this example – yellow), represents a bad prognosis for the patient. B: Event free survival (ESF) of children suffering from neuroblastoma, i.e. no clinically significant cancer-related observations or occurrence in the indicated years following initial diagnosis and treatment. Those who have minimal or no N-myc amplification have a very good prognosis, while those who have extensive N-myc amplification have a dramatically poorer prognosis and therefore short survival times after diagnosis. Weinberg RA. The biology of cancer, Garland Science 2007

23. c-myc overexpression in Burkitt’s lymphoma • Malignant cancer of the lymphatic system (B lymphocytes) • The most common malignancy of children in equatorial Africa. • The etiologic agents of this disease include chronic infections both by Epstein-Barr virus (EBV = human herpes virus 4, causes infectious mononucleosis) and by malarial parasites. In the genomes of Burkitt’s lymphoma cells, the expression of the c-myc gene is placed under control of the transcription-controlling enhancer of an immunoglobulin gene as a direct consequence of reciprocal chromosomal translocations. The immunoglobulin enhancer directs high constitutive myc expression. Weinberg RA. The biology of cancer, Garland Science 2007

24. Part 2: Tumour viruses and viral oncogenes

25. Oncogenic retroviruses (RNA viruses) The genetic information of a retrovirus is stored in RNA. Inside infected cells, the RNA sequence is transcribed into DNA by viral reverse transcriptase and integrated into the genome of the cell (= provirus). Weinberg RA. The biology of cancer, Garland Science 2007

26. Insertional mutagenesis Retroviruses that lack acquired oncogenes can induce cancer by theintegration of their proviral DNA adjacent to a cellular proto-oncogene. In this case the expression of the proto-oncogene can be very high because it is driven by a strong, constitutively active viral promoter. Example: Avian leukosis virus (ALV) induces B-cell lymphomas in chicken. Analysis of numerous ALV-induced lymphomas revealed that the ALV provirus becomes often integrated into the c-myc proto-oncogene, mostly between the first non-coding exon and the second exon of the c-myc gene Weinberg RA. The biology of cancer, Garland Science 2007

27. Weinberg RA. The biology of cancer, Garland Science 2007

28. Rous sarcoma virus (RSV) and oncogene v-src A A: The genome of RSV is closely related to the genome of ALV. However, in addition to the common retroviral genes gag, pol, and env, it carries src gene (v-src) that specifies Src protein, which causes cell transformation B B: The viral src gene (v-src) was originally a cellular gene (c-src) that was captured by the virus probably when ALV provirus became integrated (by chance) next to a c-src proto-oncogene in an infected chicken cell. c-src gene codes for a non-receptor tyrosine kinase. Weinberg RA. The biology of cancer, Garland Science 2007

29. Rous sarcoma virus-induced transformation of chicken fibroblasts A B A: Normal fibroblasts grow in a layer one cell thick – monolayer. This is because when these cells touch one another, they cease proliferating = contact inhibition. However, when they get infected by RSV, they acquire a rounded morphology and lose contact inhibition, i.e. they continue to proliferate in spite of touching one another. B: RSV-induced trasformation (normal fibroblasts left panels, RSV-transformed fibroblasts right panels) Weinberg RA. The biology of cancer, Garland Science 2007

30. Viral oncogene erbB Oncogene erbB was discovered in the genome of avian erythroblastosis virus (AEV), a transforming retrovirus that rapidly induces a leukemia of red blood cell precursors (erythroleukemia). The gene codes for a truncated version of chicken EGF receptor. Such a truncated receptor (v-ErbB) can emit mitogenic signals constitutively, i.e. without stimulation by EGF ligand. Weinberg RA. The biology of cancer, Garland Science 2007, Adapted

31. Weinberg RA. The biology of cancer, Garland Science 2007

32. Some oncogenic DNA viruses code for proteins that inhibit tumour suppressors