oncogenic viruses ppt.

basic understanding regarding oncoviruses

1. ONCOGENIC VIRUSES By- Dr. Ravi Bhushan

2. INTRODUCTION HISTORY EPIDEMIOLOGY MECHANISM OF ONCOGENECITY BY VIRUSES VIRUSES ASSOCIATED WITH HUMAN TUMORS DIAGNOSIS PREVENTION

3. A  virus  is a small infectious agent that can replicate only inside the living cells of an organism. Viruses can infect all types of organisms, from animals and plants to  bacteria .   Viruses are found in almost every ecosystem on Earth and are the most abundant type of biological entity .   Virus particles (known as  virions ) consist of two or three parts: i ) the genetic material made from either DNA or RNA, ii) a protein coat that protects these genes; and in some cases iii) an envelope of lipids that surrounds the protein coat when they are outside a cell. 

4. What is Cancer? Definition: Any disorder of cell growth that results in invasion and destruction of surrounding healthy tissue by abnormal cells. Cancer cells arise from normal cells whose nature is permanently changed. They multiply more rapidly than healthy body cells and do not seem subject to normal control by nerves and hormones. They may spread via the bloodstream or lymphatic system to other parts of the body, where they produce further tissue damage (metastasis).

5. Regulation of Cell Division In Normal Cells

6. 6 Changes in cell that are at the roots of cancer Genetic and epigenetic alterations: Mutations Deletions Recombinations Transpositions Epigenetic alterations (DNA methylation, imprinting) Acquisition of viral genetic material Various combinations of these lead to the development of cancers – ( some viruses contribute single hits while others contribute multiple hits .)

7. 7 Inherited Somatic Random Transposition Exposure to deleterious environmental agents Radiation carcinogenic chemicals Viruses Other persistent infections Source of genetic alterations

8. Multistep Carcinogenesis Multistep genetic changes must occur to convert a normal cell into a malignant one. Tumors usually develop slowly over a long period of time. 3 – 8 mutations are thought to underlie this process; resulting in activation of multiple cellular oncogenes and inactivation of tumor suppressor genes . Tumor virus acts as a cofactor in the carcinogenesis process. Viruses are necessary but not sufficient for development of tumors with a viral etiology .

9. ONCOGENIC VIRUSES Viruses that produce tumours in their natural host / experimental animals or which induce malignant transformation of cells on culture. Features of viral oncogenesis - cause cancer in humans & animals - long latency between viral infection and tumorigenesis - modulate growth control pathways in cells - viral markers are present in tumor cells

10. 10 Integrations that cause activation or inactivation of oncogenes or tumor suppressors (e.g. RNA viruses) Expression of genes that alter key signal transduction pathways. Chronic activation of inflammatory responses How do Viruses contribute to cancer?

11. Genes and Cancer Mutations that result in cancer typically occur in 3 types of genes. – Proto-oncogenes: (genes whose products stimulate cell multiplication ) – Tumor-suppressor genes: (genes whose products inhibit cell multiplication ) – Mutator genes: (genes whose products ensure accurate DNA replication and DNA repair)

12. Oncogenes A gene that is capable of transforming a normal cell into a cancerous cell. Oncogenes result from the mutation of normal genes (proto-oncogenes). Oncogenes are also seen in oncogenic viruses. (Viral oncogenes are derived from normal host genes that have become incorporated into the viral genome and subsequently undergo mutation .) 2 types: C - onc V- onc

13. Tumor suppressor genes These are negative regulators of cell growth. They form complexes with oncoproteins of certain DNA tumor viruses- causes inactivation or functional loss -leads to tumor formation. The prototype of these genes are retinoblastoma ( Rb ) gene and P53 gene. Function: - p53 acts as transcription factor and blocks cell cycle progression . -p53 causes cells with DNA damage to undergo apoptosis. p53 gene is mutated in over half of all human cancers.

14. Research History In 1908, Ellerman & Bang first discovered virus, producing leukemia in chicken. In 1911 Peyton Rous 1 st shows the presence of filterable sarcoma material that induce the CANCER.

15. YEAR SIG NIFICANCE 1908 Ellerman & Bang reported that cell free filtrates from chicken with leukemia could transmit disease to healthy birds 1911 Rous - first described association of viruses with malignancy - Fowl sarcoma caused by virus - Nobel Prize in 1966 1932 Shope – demonstrated viruses causing tumors in animals - isolated Rabbit Fibroma virus (1932) & Pappiloma virus (1933) 1957 Stewart & Eddy – discovered Polyoma virus 1962 Trentin – demonstrated sarcoma in newborn mice by human Adenovirus Discovery of tumorigenic potential of Simian virus 40 1965 Burkitt – identified Epstein Barr virus as causative for Burkitt’s lymphoma. First human tumor virus 1975 Blumberg et al – linked chronic hepatitis B infection to hepatocellular carcinoma 1980 Second-generation recombinant HBV surface antigen subunit vaccine (against HBV & HCC) 1974 Harald zur Hausen – proposed HPV as etiologic agent of cervical cancer - Nobel Prize in 2008 1981 Gallo et al – proposed causal role of HTLV 1 in adult T cell Leukemia 1989 Houghton et al – proposed association between chronic HCV infn and HCC 1994 Chang et al – isolated kaposi’s sarcoma virus (HHV8)

16. EPIDEMIOLOGY Viruses are responsible for 20% of malignant conditions in humans, including some of the most common cancers worldwide, and are especially common in immunosuppressed patients.

17. Classification

18. ONCOGENIC VIRUSES TAXONOMIC GROUPING EXAMPLES PRIMARY TUMOR TYPES RNA VIRUSES 1.Flaviviridae Hepatitis C virus Hepatocellular carcinoma 2.Retroviridae Alpha-retroviruses Rous sarcoma virus(RSV) Sarcoma Rous associated virus(RAV) B-cell lymphoma, erythroleukemia Avian myeloblastosis virus (AMV) Myeloid/ erythroid leukemia Avian erythroblastosis virus (AEV) Erythroid leukemia Myelocytoma virus (MC29) Myeloid leukemia Beta-retroviruses Mouse mammary tumor virus(MMTV) Mammary carcinoma Jaagsiekte sheep retrovirus Lung carcinoma

19. Gamma-retroviruses Murine leukemia virus( MuLV ) Leukemia , lymphoma Murine sarcoma virus( MuSV ) Sarcoma Feline leukemia virus Leukemia,lymphosarcoma Feline sarcoma virus Sarcoma Simian sarcoma virus Sarcoma Koala retrovirus T cell leukemia Delta-retroviruses Human T lymphotropic virus(HTLV) Adult T cell leukemia Bovine Leukemia virus B cell leukemia Epsilon-retroviruses Walleye dermal sarcoma virus Sarcoma

20. DNA VIRUSES 1.Adenoviridae Types 2,5,12 Various solid tumors 2. Hepadnavirus Hepatitis B virus (HBV) Hepatocellular carcinoma 3.Herpes viridae Epstein-Barr virus(EBV) (HHV4) Burkitt’s lymphoma, nasopharyngeal carcinoma Kaposi sarcoma Herpes virus(KSHV) (HHV8) Kaposi sarcoma 4.Polyoma viridae SV40,polyoma virus Various solid tumors 5.Papillomaviridae HPV 6,11,16,18 Bovine papilloma virus Papilloma,carcinoma 6.Poxviridae Shope fibromavirus Myxoma,fibroma

21. CELL CYCLE OF RETRO VIRUSES

22. Tumor Viruses Genome all viral proteins Replication Lysis Progeny virions Lytic Life Cycle For most viruses: www.freelivedoctor.com

23. Tumor Viruses Virus Cell Integration (often) Transformation Latent Life Cycle Some virus-specific proteins expressed (early functions) - No mature virus Viral structural proteins are not expressed Changes in the properties of host cell - TRANSFORMATION Sometimes latency may terminate – cell must be infected by complete virus

24. MECHANISM OF ONCOGENECITY Introduction of new Alteration of expression of ‘Transforming gene’ pre-existing cellular gene into the cell Loss of normal growth regulation processes Affection of DNA repair mechanisms Genetic instability Mutagenic phenotype DIRECT ACTING INDIRECT ACTING

25. Growth parameters and behavior of transformed cells Immortal (can grow indefinitely) Reduced requirement for serum growth factors Loss of capacity for growth arrest upon nutrient deprivation Loss of contact inhibition (can grow over other cells) Increased ability to grow in suspension Anchorage independence (can grow in soft agar) Altered morphology (appear rounded and refractile ) Tumorigenicity Induction of DNA synthesis Chromosomal changes

26. VIRAL TRANSFORMATION The changes in the biological functions of a cell that result from REGULATION of the cell’s metabolism by viral genes and that confer on the infected cell certain properties characteristic of NEOPLASIA These changes often result from the integration of the viral genome into the host cell DNA.

27. VIRUSES CAUSING HUMAN CANCERS VIRUS FAMILY VIRUS HUMAN CANCER Papillomaviridae Human Papilloma virus Genital tumors Squamous cell carcinoma Oropharyngeal carcinoma Herpesviridae Epstein –Barr virus Nasopharyngeal carcinoma Burkitt’s lymphoma Hodgkin disease B cell lymphoma Human Herpes virus 8 Kaposi sarcoma Hepadnaviridae Hepatitis B virus Hepatocellular carcinoma Flaviviridae Hepatitis C virus Hepatocellular carcinoma Retroviridae Human T cell lymphoma virus Adult T cell lymphoma Human immunodeficiency virus AIDS related malignancies

28. RNA VIRUSES

29. RETROVIRUS Properties Virion – spherical, helical nucleoprotein within icosahedral capsid Genome – 2 copies of single stranded RNA Possess reverse transcriptase(RNA dependent DNA polymerase) Enveloped virus Not cytolytic (except lentivirus )

30. Retrovirus (alpha, gamma) – no viral oncogene gag – encodes core proteins(group specific) pro – encodes protease enzyme pol – encodes reverse transcriptase env – encodes envelope glycoproteins Influence proto-oncogene by insertional mutagenesis Deltaretrovirus , Lentivirus tax/tat – transactivating regulatory gene .

31. HUMAN T- CELL LEUKEMIA VIRUS Epidemiology - First retrovirus implicated in human disease. -20 million infected worldwide. -High endemic areas – Latin America , Caribbean , Africa ,Japan - IV drug users - U.S & Europe Adult T-cell leukemia /lymphoma Oncoproteins Tax - Enhanced proliferative potential of T – lymphocytes - interference of cell regulation pathways & DNA repair mechanisms.

32. Accessory protein p12 - alters MHC 1 & T-cell receptor (TCR) cascade activation Accessory protein p13II - targets mitochondria - affects cell proliferation , apoptosis , ROS production

33. HIV Accessory protein Tat - interferes with DNA repair mechanisms - interfere with Rb gene mediated growth regulation pathway HCV Flaviviridae . Single stranded RNA. Over 170 million chronic carriers Hepatocellular carcinoma Non Hodgkins B cell lymphoma Mechanism- inflammation cirrhosis

34. DNA VIRUSES Hepatitis B virus HBV Human Papillomavirus

35. DNA virus oncogenesis Inhibition of tumor suppressor genes - Rb gene mutation - inactivates p53 mediated growth regulation Oncoproteins interact with specific targets on the host cell

36. DNA virus Oncoproteins and their major targets VIRUS TARGETS Adenovirus E1A Rb family members Adenovirus E1B19K Bak , Bax Adenovirus E1B55K p53 Adenovirus E4 orf 6 p53 Epstein Barr virus EBNA2 Glycogen synthetase kinase,RBP -J kappa/CBF 1 Epstein Barr virus Lmp 1 PI3K ,TNF signalling components Epstein Barr virus LMP 2 Src family members Hepatitis B virus X protein p53 Human Papilloma virus E5 EGF receptor Human Papilloma virus E6 P53, PDZ proteins, Human Papilloma virus E7 Rb family members, p21,p27,p600 KSHV ORF 50 ,KSHV K- bZIP p53 KSHV vCyclin Cyclin dependent kinase 6

37. HPV Small Non-enveloped Virion – Icosahedral Genome – double stranded ,circular DNA (8000bp ) Closely related to Polyomaviruses 16 genera (5 – human infections) Nearly 140 types are identified Classified using molecular criteria Epidemiology - HPV induced cervical cancer is 2 nd most common cancer worldwide - 16% of all female cancers are linked to HPV - Papilloma virus is found in 90% of women with cervical cancers

38. HUMAN PAPILLOMA VIRUS TYPE CLINICAL LESION SUSPECTED ONCOGENIC POTENTIAL 1 Plantar warts Benign 2,4,27,57 Common skin warts Benign 3,10,28,49,60,76,78 Cutaneous lesions Low 5,8,9,12,17,20,36,47 Epidermodysplasia verruciformis Benign -  malignancy 6,11, 40,42,43,44,54,61,70,72,81 Anogenital condylomas , Laryngeal papillomas , Mucaosal dysplasia & intraepithelial neoplasia Low 7 Hand warts in butchers Low 16,18 ,30,31,33,35,39,45,51-53,56,58,59,66,68,73,82 High grade dysplasia,genital carcinoma ,laryngeal & esophageal carcinomas High ( espl with cervical cancer )

39. HPV REPLICATION High tropism for epithelial cells of skin & mucous membranes V iral replication are dependent on sequential differentiation states of epithelial cells . DNA synthesis is supported only in basal cells of the squamous epithelium. Hence, difficult to propagate in vitro Opening Reading Frames(ORF) – encode viral proteins . Located on only one of the 2 viral DNA strands No viral DNA polymerase--Dependent on host cell replication machinery for viral genome replication.

40. ORF FUNCTION L1 L1 protein-major capsid protein ( VLP vaccine ) L2 L2 protein- minor capsid protein E1 Initiation of viral DNA replication,helicase,ATPase E2 Trancriptional regulatory protein, genomic maintenance E4 Late protein. Disrupts cytokeratins E5 Membrane transforming protein, interacts with specific growth factor receptors E6 Transformation. Degradation of p53, telomerase activation E7 Transformation. Inactivation of pRb .

41. HPV pathogenesis Sexually transmitted Peak incidence – adolescents & young adults Episomal HPV DNA – in skin carcinoma, premalignant lesions Integrated HPV DNA – in cervical cancer cells Oncogeneticity – transforming oncoproteins interacting with tumor suppressors(p53, Rb )

42. HERPES VIRUS EPSTEIN BARR VIRUS (HHV 4) KAPOSI SARCOMA HERPES VIRUS (HHV 8 ) Properties Large viruses Genome – linear double stranded DNA Icosahedral capsid with lipid containing envelope Acute infection followed by latency Recurrence from latent infection Latency genes

43. EBV Infectious mononucleosis Burkitt’s lymphoma Nasopharyngeal carcinoma Non Hodgkin’s lymphoma Remain latent in lymphoblast cell lines. Epidemiology - Ubiquitous - Burkitt’s lymphoma – children in Central Africa - Nasopharyngeal carcinoma – Cantonese China , Alaskan Eskimos Malaria - cofactor Tumors contain integrated & episomal forms of viral DNA

44. Oncoproteins LMP1 (Latent Membrane Protein 1) - membrane protein with 6 domains - receptor - TNF alpha TRAF TRADD (TNF Recceptor ass. factor) (TNF receptor ass. death RIP domain) (Receptor interacting protein) Stimulation of transcription factors for Anti-Apoptotic proteins LMP 1

45. EPSTEIN BARR NUCLEAR ANTIGEN (EBNA) EBNA 1 EBNA 2 EBNA LP Conversion of EBV infected peripheral EBNA 3A blood cells to lympho- blastoid cell lines EBNA 3C P53 mutation

46. KAPOSI SARCOMA HERPES VIRUS HHV 8 KAPOSI’S SARCOMA- affects multiple organs with prominent vascular endothelial component PRIMARY EFFUSION LYMPHOMA MULTICENTRIC CASTLEMAN DISEASE Epidemiology - Mediterranean & African countries - Elderly men at high preponderance

47. ONCOPROTEINS - Seven latent genes - Latency Associated Nuclear Antigen(LANA) Impairs p53 & Rb up-regulates Beta – catenin pathway V - FLIP( Flice inhibitory protein) inhibits caspase activity

48. HBV Hepadnaviridae Genome – circular, ds DNA Epidemiology - endemic in Africa ,China, South east Asia - over 250 million persistently infected Primary infections in neonates & children- 90% chronic Hepatocellular carcinoma Hepatitis B vaccination has lowered incidence of infection & HCC

49. Oncoprotein 1. X gene - encoded by ORF X - affects cellular gene expression - interferes with p53 function 2. Mutated proto-oncogene N-myc2 . Aflatoxin - co-factor - Africa & China . ROS - generated by inflammatory cells of Chronic active hepatitis - DNA damage & mutagenesis

50. POLYOMA VIRUS MERKEL CELL VIRUS Merkel cell carcinoma - highly lethal skin cancer - immuno-compromised Oncoproteins - Large T antigen (LT) - Small T antigen (ST) - inactivate Rb tumor suppressor pathway

51. SIMIAN VIRUS 40 Osteosarcoma, lung carcinoma, brain tumors EPIDEMIOLOGY - primary infection in Asian macaques -H/O accidental exposure through contaminated poliovirus vaccines b/w 1955 and 1963

52. DETECTION DETECTION OF TRANSFORMATION – studied in established cell lines-- immortal GENETIC MAPPING - to identify specific viral genes with transforming activity GENE SEQUENCING PCR - HTLV 1 - HBV , HCV - EBV (LMP 1,2) SOUTHERN BLOTTING - HBV - EBV--EBNA1

53. ANIMAL INOCULATION MICROARRAY IN SITU HYBRIDISATION - HPV (E6,E7) - KSHV

54. VACCINES HPV Non-infectious, recombinant vaccines - Virus Like Particles( VLP ) composed of L1 proteins(major capsid protein) - VLPs generate high titre type specific neutralizing antisera GARDASIL - FDA approved (Merck) - Quadrivalent vaccines (particles derived from HPV 6,11,16,18) - also used in men to prevent genital warts (approved in 2009)

55. CERVARIX - FDA approved (GlaxoSmithKline) - Bivalent vaccines ( HPV 16,18) - immunity for 5 years - contraindicated in pregnancy. Immune response is Type specific No cross protection against other HPV types Expensive Heat labile.

56. HBV Subunit, recombinant vaccine HBsAg HCV Genome is highly variable No effective vaccine HTLV No vaccine Interferons

57. EBV Chemotherapy Vaccine – in pipeline KSHV Gancyclovir – inhibits viral DNA polymerase Incidence lowered in AIDS patients treated with HAART No vaccine

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