Cancer Genetics

1. Cancer Genetics SDK December 17, 2013

2. Objectives • Upon completion of this session the students should be able to • Define necrosis and apoptosis • Explain the process of apoptosis & necrosis • Define& understand and what is cancer • Explain the genetic nature of cancer • Elaborate the causes of cancer such as genetic mutation • Know the genes that play a role in cancer • To know and understand the role of following in cancer • Oncogenes • Tumor Suppressor Gene • DNA Repair Gene • Viruses and Cancer

3. Cancer & its Molecular Basis • Cancer is a collection of diseases characterized by abnormal and uncontrolled growth • Cancer arises from a loss of normal growth control • In normal tissues, the rates of new cell growth and old cell death are kept in balance • In cancer, this balance is disrupted • This disruption can result from 1) Uncontrolled cell growth or 2) Loss of a cell's ability to undergo apoptosis

4. Types of Cell Death • Apoptosis (Programmed Cell Death) • During apoptosis cells die in a controlled, regulated fashion. • Apoptosis, is a process in which cells play an active role in their own death (which is why apoptosis is often referred to as cell suicide) • Necrosis • A form of cell death that is caused by a range of toxic chemicals, biological agents, or physical damage. • It is associated with breaking of cellular membranes, swelling of the cells and destruction of the cellular structures

5. Types of Cell Death • Cell death by injury(Necrosis) • Mechanical damage • Exposure to toxic chemicals • Cell death by suicide(Apoptosis) • Internal signals • External signals

6. Apoptosis or Suicidal cell death • Apoptosis or programmed cell death, which is carefully coordinated collapse of cell, protein degradation , DNA fragmentation followed by rapid engulfment by neighboring macrophage cells. • Essential part of life for every multicellular organism. • Apoptosis plays a major role from embryonic development to old age. • It is single cell death or death of selected cell.

7. Why Should a Cell Commit Suicide • Apoptosis is needed for proper development • The formation of the fingers and toes of the fetus by removing inter digital membrane • Formation of barrel of esophagus • The sloughing off of the inner lining of the uterus during menstruation • Apoptosis of lactating mammary gland after cessation of lactation. • The formation of the proper connections between neurons in the brain • Bone marrow production of cells. • GIT mucosa, process of apoptosis is continue all the time Apoptosis is needed to destroy cells • Cells with DNA damage such as by Radiation, thermal injury or Hypoxia • Cells infected with viruses • Cells of the immune system • Cancer Cellsand activation of P53 genes

8. What Makes A Cell Decide To Commit Suicide The balance between: • The withdrawal of positive signals; that is, signals needed for continued survival, and • The receipt of negative signals When this balance is disturbed cell decide to commit suicide

9. What Makes A Cell Decide To Commit Suicide • Withdrawal of positive signals • Growth factors for neurons • Interleukin-2 (IL-2) an essential factor for the mitosis of lymphocytes • Receipt of negative signals • increased levels of oxidants within the cell • damage to DNA by oxidants • Death Activators : • Tumor necrosis factor alpha (TNF-). that binds to the TNF receptor • Lymphotoxin (TNF-β) . that also binds to the TNF receptor. • Fasligand (FasL). a molecule that binds to a cell-surface receptor named Fas (also called CD95).

10. Necrosis vs Apoptosis Necrosis Apoptosis • Cellular condensation • Membranes remain intact • Requires ATP • Cell is phagocytosed, no tissue reaction • DNA fragmentation • Individual cells or selected cells are affected • Cellular swelling • Membranes are broken • ATP is depleted • Cell lyses, eliciting an inflammatory reaction • DNA fragmentation is random, or smeared • Whole areas of the tissue are affected

11. How Does Apoptosis Occur? • The cell receives specific signals with instruction to undergo apoptosis • A number of changes occur in the cell • A family of proteins (enzymes) known as Caspasesare produced • These proteins breakdown cellular components such as those in the cytoskeleton and nuclear proteins such as DNA repair enzymes • Caspases also activate other enzymes such as nuclease, which begin to breakdown the DNA in the nucleus

12. Major Players In Apoptosis • Caspases • Adaptor proteins • TNF & TNFR family • Anti apoptotic genes • Bcl-2 family i.e. Bcl2 and BclX • Proapoptotic genes • Bad • Bax • Back

13. Bcl-2 FAMILY Anti-apoptotic Pro-apoptotic • Bax • -Bcl-XS • Bak • Bad • Bcl-2 • Bcl-XL • Mcl-1

14. Bad Bad Bcl-XL Bcl-2 CELL SURVIVAL Bcl-XL Bcl-2 Bax Bax Bax Bax CELL DEATH

15. What is observed during apoptosis • The cell begins to shrink following the breakdown of the cytoskeleton. Cells lose contact with surrounding cells • The breakdown of chromatin in the nucleus • Cells continue to shrink • Parts of the membrane break off to make membrane blebs • Finally the cell is fragmented into small membrane-enclosed structures, called 'apoptotic bodies' which contain cytosol, the pieces of chromatin, and organelles

16. Apoptosis Events • Cell shrinkage • Organelle reduction • Mitochondrial leakage • Chromatin condensation • Nuclear fragmentation • Membrane blebbing

17. Blebbing & Apoptotic bodies The control retained over the cell membrane & cytoskeleton allows intact pieces of the cell to separate for recognition & phagocytosis by Macrophages. Bleb Apoptotic body Macrophage

18. Death Receptors Initiator Caspase 8 EffectorCaspase 3 PCD Initiator Caspase 9 Mitochondria/Cytochrome C Apoptosis: Pathways “Extrinsic Pathway” Death Ligands “Intrinsic Pathway” DNA damage & P53

20. Ligand-induced cell death Ligand Receptor FasLFas (CD95) TNF TNF-R TRAIL DR4 (Trail-R)

21. Death Domains Death Effectors Induced proximity of Caspase 8 Activation of Caspase 8 Ligand-induced cell death “The death receptors” Ligand-induced trimerization FasL Trail TNF

22. Why do we need apoptosis • Apoptosis has the functions of: • Keeping the number of cells relatively constant through cell death and division • Replacing cells when they malfunction or become infected • Stopping the spread of viral infection • The advantage of apoptosis is that it packages the dead cell into small blisters, which can be easily digested by immune cells

23. Why do we need apoptosis • Important in normal physiology / development • Development: Immune systems maturation, Morphogenesis, Neural development • Adult: Immune privilege, DNA Damage and wound repair. • Excess apoptosis • Neurodegenerative diseases • Deficient apoptosis • Cancer • Autoimmunity

24. p53 and Apoptosis • p53 also known  tumor suppressor p53 is a protein that in humans is encoded by the TP53 gene. • The p53 protein is crucial in multicellular organisms, where it regulates the cell cycle and, thus, functions as a tumor suppressor, preventing cancer protein. • As such, p53 has been described as "the guardian of the genome" because of its role in conserving stability by preventing genome mutation • n humans, the TP53 gene is located on the short arm of chromosome 17 (17p13.1).

25. p53 and Apoptosis • Activate DNA repair proteins when DNA has sustained damage. • It can arrest growth by holding the cell cycle at the G21/S regulation point on DNA damage recognition (if it holds the cell here for long enough, the DNA repair proteins will have time to fix the damage and the cell will be allowed to continue the cell cycle). • It can initiate apoptosis, the programmed cell death, if DNA damage proves to be irreparable.

26. Mutation of p53 Gene • The tumor suppressor gene p53 is one of the most frequently mutated gene in human cancers. • "Mutations in p53 or the pathway that directly regulates it have been found in over 80% of human tumors..." • A mutant p53 will no longer bind DNA in an effective way, and, as a consequence, it will not be able to act as the "stop signal" for cell division and lead to over production of cell.

27. Cancer • Caner is defined as the continuous uncontrolled growth of cells

28. Cancer Cells • Cancer cells do not respond normally to body’s control mechanisms: • Ignore density dependent inhibition, cells continue to multiply until nutrients are exhausted • Divide excessively and invade other tissues

29. How Cancer Develops? • Cancer arises from a loss of normal growth control. In normal tissues, the rates of new cell growth and old cell death are kept in balance. In cancer, this balance is disrupted • This disruption can result from uncontrolled cell growth or loss of a cell’s ability to undergo cell suicide by a process called “apoptosis”

30. Loss of Normal Growth Control Normal cell division Cell Suicide or Apoptosis Cell damage that couldn’t be repaired Cancer cell division First mutation Second mutation Third mutation Fourth orlater mutation Uncontrolled growth

31. Properties of cancer cells They keep growing And growing And growing And growing

32. Cell Cycle

33. G1 - Growth phase 1 S - DNA synthesis G2 - Gurther growth M - Cell division • Mitosis: • prophase, prometaphase, metaphase, anaphase • and telophase • Cytokinesis Cell Cycle Interphase Mitotic phase

34. Cell Cycle Regulation • For all living eukaryotic organisms it is essential that the different phases of the cell cycle are precisely coordinated • Errors in this coordination may lead to chromosomal alterations. Chromosomes or parts of chromosomes may be lost, rearranged or distributed unequally between the two daughter cells or may lead to uncontrolled growth.

35. Cell Cycle Regulation Cell-cell Contact Growth Factors Cell Size Regulatory Proteins & Protein Kinases Nutrients

36. Is cell big enough? Is environment favourable? G2 Checkpoint Is all DNA replicated? M G2 Metaphase Checkpoint G1 S G0 G1 Checkpoint Is cell big enough? Is environment favourable? Are all chromosomes aligned on spindle?

38. Characteristic's of Cancer Cells • Undifferentiated (Dysplasia) • Less adherent • Loss of cell cycle control • Tissue invasion and metastasis • Escaping apoptosis • Insensitivity to antigrowth factor • Increased mutation rate • Induce local blood vessel formation angiogenesis.

39. Tumors &Cancer Tumor is mass of abnormal growth of tissue resulting from uncontrolled, progressive multiplication of cells i.e Hyperplasia. Some tumors are beningn while somw are malignant. • Benign, e.g., moles • Slow growth • Expands in the same tissue; does not spread • Cells look nearly normal • Malignant • Rapid growth • Invades surrounding tissue and metastasizes • Cell differentiation usually poor

40. A Cancer Cell’s Structure Is Abnormal • Cancer is a result of a series of mutations in the cell’s genes • Larger cell nucleus and less cytoplasm • Loss of structural specialization • Cytoskeleton shrinks • Plasma membrane proteins could be lost or altered • New plasma membrane proteins may appear • Changes passed on to cell’s descendants

41. Cancer Cells Also Do Not Divide Normally • Cancer cells don’t necessarily divide faster than normal cells; more cancer cells are dividing than dying • Cancer cells do not respond to crowding; loss of contact inhibition • Leads to a disorganized mass; cells may have extensions • May produce HCG and angiogenin • Metastasis: makes a cancer malignant

42. What causes Cancer.Cancer & Genetics • Cancer is a genetic disease resulting from a disruption of one or more genes • As genes switch on and off, they determine when and how fast the cell will grow and divide, when it will stop dividing, and even when it will die. • 5-10% caused by inherited genetic mutations • Majority is caused by cumulative effect of somatic mutations caused by • Carcinogenic chemicals • Radiation • Some viruses

44. Carcinogenesis • Transformation of a normal cell into a cancerous cell • Step-by-step transformation

45. Carcinogenesis • Genetic changes can occur at many levels, from gain or loss of entire chromosomes to a mutation affecting a single DNA nucleotide. • Epigenetics such as DNA methylation and microRNAs.

46. Genetic Mutation • Germlinemutations • Are responsible for 5% to 10% of cancer cases. • This is also called familial cancer (occurring in families). • Inherited and thus present in all cells from birth. • Acquired mutations (Sporadic cancer or somatic mutation). • Most of these cancers are caused by a series of mutations that develop during a person's lifetime. • Acquired mutations are not in every cell of the body and are not passed from parent to child. • Acquired mutations are caused by; Tobacco, Over-exposure to UV radiation, and Toxins and chemicals

47. Genes that play a role in cancer • 3 Normal regulatory genes; • Growth promoting(protooncogene) • Growth inhibiting(tumor suppressor gene) • Gene regulate apoptosis • 4th category is DNA repair gene

49. Genes and Cancer

50. Association with human cancers • Growth Factor Receptor Increased numbers in 20 percent of breast cancers • Ras Protein Activated by mutations in 20 to 30 percent of cancers • AblKinase Activated by abnormal chromosomes in chronic myelogenous leukemia • SrcKinase Activated by mutations in 2 to 5 percent of cancers • p53 Protein Mutated or deleted in 50 percent of cancers