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  • Entire mechanism triggered inside the cell. There is no inflammation, no residual or any sign that the cell existed. Infact its close to impossible to pick up under a microscope.
  • Lymphocytes not stimulated by antigens and cytokines Involution of hormone dependent tissues Neurons deprived of nerve growth factors
  • Mito membrane permeability important
    More than 20 proteins
    Prototype is Bcl-2
  • Binding of death ligands (CD95L is used here as an example) to their receptor leads to the formation of the death-inducing signalling complex (DISC). In the DISC, the initiator procaspase-8 is recruited by FADD (FAS-associated death domain protein) and is activated by autocatalytic cleavage. Death-receptor-mediated apoptosis can be inhibited at several levels by anti-apoptotic proteins: CD95L can be prevented from binding to CD95 by soluble 'decoy' receptors, such as soluble CD95 (sCD95) or DcR3 (decoy receptor 3). FLICE-inhibitory proteins (FLIPs) bind to the DISC and prevent the activation of caspase-8; and inhibitors of apoptosis proteins (IAPs) bind to and inhibit caspases. FLIPL and FLIPS refer to long and short forms of FLIP, respectively.
    Viruses produce homologus of FLIP hence keeping cells alive
    This process mainly involved in eliminating self reactive lymphocytes and killing targets by cytoxic T cells.
  • Fundamental event is activation of caspases
  • When p53 is damaged it cannot induce apoptosis so cell survive Subsequent mutations develop Progresses to malignancy
  • Pro-apoptotic BCL2 family proteins are activated by treatment ---- caspase cascade ---- death of cell
  • Apoptosis

    1. 1. Apoptosis
    2. 2. Literal sense
    3. 3.  Programmed cell death  Internally controlled “suicide” program  Cells are removed with minimal disruption of the surrounding tissue
    4. 4. PathologicalPhysiological Potentially harmful cells Outlived their usefulness Maintain a steady number Damaged beyond repair
    6. 6. Growth factor deprivation
    7. 7. Cell loss in proliferating populations • Eliminate self reactive lymphocytes • Cell death by induced cytotoxic T lymphocytes (defense against virus / tumorogenisis)
    8. 8.  Genetically altered beyond repair  DNA damage (Direct or Free radical)  Radiation  Cytotoxic drugs  Extreme tempetature  Hypoxia
    9. 9.  Accumulation of misfolded proteins  Arise because of mutations in the genes coding these proteins or extrensic factors like free radicals  Cell injury in infections  Atropy in parynchyma after duct obstruction
    10. 10. G1 S G2 M p53 Repair Attempted NO YES
    11. 11.  Initiation  Progression  Termination
    12. 12. Cytochrome C antagonists of endogenous cytosolic inhibitors of apoptosis Mitochondria has several proteins capable of inducing apoptosis Bcl 2 and bcl XL antiapoptotic BAX and BAK ( Pro apoptotic)
    13. 13. Dimerize and insert into membrane of mitochondria Permiability transition pore Channel activated (voltage gated channel) Cytochrome C CASPASE ACTIVATION
    14. 14. Surface molecules trigger apoptosis Members of TNF family Ex: Type 1 TNF and Fas (CD95) Normally present in cytoplasmic region On Activation move to surface
    15. 15. FasLigand membrane protein on activated T lymphocytes activate caspase-8 activating caspase cascade by activating member of Bcl-2 family called Bid Bind to Fas
    16. 16. CASPASE Mitochondrial pathway Death receptor pathway Both pathways are not mutually exclusive with p53 causing cross over to death receptor pathway. Cystene protease that cleave proteins after aspartic residues Cell Death by Apoptosis Activation of caspase is the fundamental event of apoptosis
    17. 17. Caspase-1 (ICE) Caspase-2 (ICH-1, Nedd-2) Caspase-3 (CPP32, Apopain, Yama) Caspase-4 (ICH-2, TX, ICEreıı) Caspase-5 (ICErelııı, TY) Caspase-6 (Mch2) Caspase-7 (ICE-LAP3, Mch3, CMH-1) Caspase-8 (FLICE, Mch5, MACH) Caspace-9 (Mch6, ICE-LAP6) Caspase-10 (Mch4)
    18. 18. Caspase activation a. Proteolytic cleavage e.g. pro-caspase 3 b. Induced proximity e.g. pro-caspase 8 c. Oligomerization, e.g. cyt c, Apaf-1 & caspase 9
    19. 19.  Inflammatory Caspases: -1, -4, and -5  Initiator Caspases: -2, -8, -9, and -10  Long N-terminal domain  Interact with effector caspases  Effector Caspases: -3, -6, and -7  Little to no N-terminal domain  Initiate cell death
    20. 20.  But become avid targets to phagocytosis by flip out of phosphotidyl serine on the inner leaflet of plasma membrane  These are recognized by macrophages  Also secrete soluble factors that recruit phagocytes  Express adhesive glycoproteins recognized by phagocytes and macrophages
    21. 21. CASPASE . Cleavage of nuclear LAMINS is involved in chromatin condensation and nuclear shrinkage. Cleavage of the inhibitor of the DNase CAD (caspase-activated deoxyribonuclease), ICAD causes the release of the endonuclease, which fragment DNA. Cleavage of cytoskeletal proteins such as actin, plectin, Rho kinase 1 (ROCK1) and gelsolin leads to cell fragmentation, blebbing and the formation of apoptotic bodies.
    22. 22.  Constant number of cells in an organism.  Cell death = Cell proliferation Cell Death Growth Survival Proliferation
    23. 23. What happens when things become imbalanced? Cell Death Growth Survival Proliferation
    24. 24.  Resistance of tumor cells to apoptosis is an essential feature of cancer development  Loss of apoptosis can promote tumor  Initiation  Progression  Treatment resistance
    25. 25.  p53 induce the expression of proteins involved in  the mitochondrial pathway — BAX, NOXA, PUMA and P53aip1  death receptor pathway — CD95, TRAIL-R1 and TRAIL- R2. G1 S G2 M p53 MalignancyMutation
    26. 26.  overexpression of anti-apoptotic genes  follicular B-cell lymphoma t(14;18), couples BCL2 gene to immunoglobulin heavy chain locus, leading to enhanced BCL2 expression.  EBV and HHV8 encode proteins that are homologues of BCL2.  BHRF1 from EBV and KSbcl-2 (vBcl-2) from HHV8 — have an anti-apoptotic function and enhance survival of the infected cells.
    27. 27.  Adenovirus induce cells to bypass apoptosis and replicate by E1B oncoprotein inducing cells to enter S phase  Cytokines like IL6 can inhibit apoptosis
    28. 28.  soluble receptors that act as decoys for death ligands.  soluble CD95 (sCD95) and decoy receptor 3 (DcR3) shown to competitively inhibit CD95 signaling.  sCD95 is expressed in various malignancies, and elevated levels can be found in the sera of cancer patients.
    29. 29.  associated with poor prognosis in melanoma patients  It is genetically amplified in several lung and colon carcinomas  overexpressed in several adenocarcinomas, glioma cell lines and glioblastomas.
    30. 30.  the pro-apoptotic BCL2 family member BAX is mutated.  Frame shift mutations  loss of expression and function.  Reduced BAX expression is associated with a poor response rate to chemotherapy and shorter survival
    31. 31. Expression of anti-apoptotic proteins  High levels of FLIP which interferes with apoptosis induction at the level of the death receptors  Human melanomas  murine B-cell lymphoma  FLIP:caspase-8 ratio cause resistance to CD95-mediated apoptosis  EBV-positive Burkitt's lymphoma  Viral analogues of FLIP, called viral FLIPs (v-FLIPs)
    32. 32. Expression of anti-apoptotic proteins  Expression of the IAP-family protein survivin is highly tumour specific.  Found in most human tumours but not in normal adult tissues.  In neuroblastoma expression correlates with a more aggressive and unfavourable disease.
    33. 33.  Bcl2 shows discrepancies in effect on tumor management  Studies have shown a correlation between high levels of BCL2 expression and the severity of malignancy of human tumours.  a high level of BCL2 expression is associated with a poor response to chemotherapy and seems to be predictive of shorter, disease- free survival.  While in some cases Bcl-2 has been shown to cause chemo resistance in other settings it is shown to improve prognosis.
    34. 34.  Clinical studies examine single alterations Cannot exclude extragenic mutations in the same pathway Thus almost impossible to determine negative results Ex: murine lymphomas harbouring INK4a/ARF have defective p53 but harbour wild type p53 hence classified as p53 normal  Extracellular survival factors affect cell death Cell density microenvironment
    35. 35. Application in Treatment  Chemotherapy, irradiation and other stimuli can initiate apoptosis through the mitochondrial (intrinsic) pathway.  Pro-apoptotic BCL2 family proteins are activated by treatment
    36. 36.  Disrupted apoptotic cycle can affect sensitivity of cancer drugs  As multiple drugs affect same mechanism it causes multidrug resistance  Sufficient doses of almost all anticancer drugs induce apoptosis by alternate pathway independent of p53 pathway  Contribution of p53 is dependent on  Agent  Dose  mutational background
    37. 37.  The best-defined mechanism of therapy-induced cellular stress induced cancer cell death.  Chemotherapeutic drugs (for example, the nucleotide analogue 5-FU) induce CD95 by a transcriptionally regulated, p53-dependent mechanism.  Leads to upregulation of CD95L  Allows the cells to either commit suicide or kill neighbouring cells.
    38. 38. Targeting in therapy  TNF alpha and Fas are toxic to  Normal cells  Cancer cells But….  TRAIL ---- tumor necrosis factor related ligand preferentially attacks tumor cells
    39. 39.  Most mutations occur upstream  Machinery is retained  Tumor specific alterations in apoptotic programmes  Ex: Adenoviral gene transfer in ovarian cancer  Ad-DF3-Bax eradicated >99% of these tumors in nude mice  Ex2: Taxanes known to phosphorylate and inactivate bcl2  inactivating NF-kb enhances chemo induced cell death  Restoring of pro-apoptotic p53  Activating death ligands hence p53 independent death
    40. 40. A Few Add Ons  Apoptosis limited role in solid tumors  Side effects of cancer drugs are because along with cancer cells they also target normal cells like intestine and bone marrow
    41. 41.  Deregulated proliferation alone is not sufficient for tumour formation  Overexpression of growth-promoting oncogenes — such as c-MYC, E1A or E2F1 — sensitizes cells to apoptosis.
    42. 42. Cell proliferation Causes apoptosis IN MALIGNANCY
    43. 43.  BCL2 family members can be divided into anti-BCL2 family members can be divided into anti- apoptotic (BCL2, BCL-Xapoptotic (BCL2, BCL-XLL, BCL-w, MCL1, A1/BFL1,, BCL-w, MCL1, A1/BFL1, BOO/DIVA, NR-13) and pro-apoptotic proteinsBOO/DIVA, NR-13) and pro-apoptotic proteins (BAX, BAK, BOK/MTD, BCL-X(BAX, BAK, BOK/MTD, BCL-XSS, BID, BAD,, BID, BAD, BIK/NBK, BLK, HRK/DP5, BIM/BOD, NIP3, NIX,BIK/NBK, BLK, HRK/DP5, BIM/BOD, NIP3, NIX, NOXA, PUMA, BMF). Most anti-apoptotic membersNOXA, PUMA, BMF). Most anti-apoptotic members contain the BCL2 homology (BH) domains 1, 2contain the BCL2 homology (BH) domains 1, 2 and 4, whereas the BH3 domain seems to beand 4, whereas the BH3 domain seems to be crucial for apoptosis induction. The pro-apoptoticcrucial for apoptosis induction. The pro-apoptotic members can be subdivided into the BAXmembers can be subdivided into the BAX subfamily (BAX, BAK, BOK) and the BH3-onlysubfamily (BAX, BAK, BOK) and the BH3-only proteins (for example, BID, BAD and BIM).proteins (for example, BID, BAD and BIM).