P53

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P53

  1. 1. p53 andchemotherapy Blossom Sabi
  2. 2. p53 and chemotherapy 1.Introduction 1.Introduction 2. Relationship between p53 and chemotherapy 2. Relationship between p53 and chemotherapy 3. Summary 3. Summary
  3. 3. p53 p53 Introduction Introduction Regulation RegulationHistoryHistory Introduction Activation Activation Function Function
  4. 4. p53 introduction • A 53 kD nuclear phosphoprotein • Encoded by TP53 gene • Short arm of chromosome 17 • Regulates cell growth and proliferation • Prevents unrestrained cell division after chromosomal damage • The absence of p53 increases the risk of developing cancers
  5. 5. History
  6. 6. Function • Apoptosis • Cell circle arrest • Senescence • DNA repair • Metabolic homeostasis • Antioxidant defence
  7. 7. Stabilization Activation Antirepression Promoter- specific activation Next NextKruse JP, et al. Cell 2009; 137; 609-22
  8. 8. More More More More Back Back
  9. 9. Regulation • Phosphorylation of N-terminal domain • Protein kinases targeting p53 More More MAPK family (JNK1-3, ERK1-2, p38 MAPK) ATR, ATM, CHK1 and CHK2, DNA-PK, CAK Oncogenes p14ARF • Mdm2 More More
  10. 10. Back Back
  11. 11. Back Back
  12. 12. p53 and chemotherapy 1.Introduction 1.Introduction 2. Relationship between p53 and chemotherapy 2. Relationship between p53 and chemotherapy 3. Summary 3. Summary
  13. 13. p53 and chemotherapy Chemotherapy Prognosis P53
  14. 14. Controversy Relationship between p53 and chemotherapy Pros • p53 is favourable to chemotherapy More More Cons • p53 is unfavourable to chemotherapy More More Neutral • p53 is irrelevant with chemotherapy More More
  15. 15. p53 is favourable to chemotherapy Yamasaki M, et al. Ann Surg Oncol 2010, 17, 634-42
  16. 16. p53 is favourable to chemotherapyPatients with mutations inp53 showed significantlypoorer prognosis thanthose without mutant p53.p53 IHC staining did notcorrelate with prognosis. Back Back
  17. 17. p53 is unfavourable to chemotherapy TP53 status was strongly associated with responseBertheau P, et al. PLoS Med 2007, 4, 585-92
  18. 18. p53 is unfavourable to chemotherapy ·TP53-mutant patients have a more favorable event-free survival · Patients with wild-type TP53 tumors had favorable overall survival, despite the lack of complete response. · Patients with TP53-mutant tumors who did not reach complete response had a significantly shorter overall survival Back Back
  19. 19. p53 is irrelevant with chemotherapy· p53 proteinoverexpression is asignificant marker of p53 IHC results p53 IHC results,prognosis (B) observation· Patients with p53positive tumors deri-ved significantbenefit from chemo-therapy(C).· Patients with p53negative tumors hadno survival benefit p53 IHC positive p53 IHC negativefrom chemotherapy(D). Tsao MS, et al. J Clin Oncol 2007, 25,5240-7
  20. 20. p53 is irrelevant with chemotherapy· p53 mutation wasnot prognostic forsurvival in the ob-servation arm(B)· Patients with p53 p53 mutation results, observationmutations did notderive significantsurvival benefitfrom chemotherapy(C).· Chemotherapysignificantlyprolonged survivalin patients with wild mutant p53 wild-type p53type p53compared withobservation (D), Back BackTsao MS, et al. J Clin Oncol 2007, 25,5240-7
  21. 21. Clinical data Basic research
  22. 22. p53 and chemotherapy Induce cell death in response Induce early cell to drug-induced cycle arrest, DNA damages protecting tumor cells from further damages More More More More
  23. 23. Cyclophosphamide Inhibits TumorProliferation and Stimulates Apoptosis InVivo · In response to CPM, Ki- 67 labeling decreased dramatically even by 24 hours (C). · Control tumors had negligible staining for the apoptosis marker cleaved caspase-3 (D), · High levels of cleaved caspase-3 were observed 3 and 6 hours after treatment with CPM (E F ) · A wave of apoptosis throughout the tumor was demonstrated at low power (H–J ), with maximal staining visible at 6 hours after treatmentChesler L ,et al. Neoplasia 2008, 10, 1268-74
  24. 24. Apoptosis in Tumors Treated with CytotoxicChemotherapy Is Driven by p53 · Activation of the p53 pathway and caspase-3 cleavage were minimal in vehicle-treated tumors, consistent with the low-level caspase-3 staining · Rapid induction of p53 was observed at 3 hours after treatment with CPM, with a peak at 6 hours after treatment · Cleaved caspase-3 and -9 were maximal at 6 hours after treatment and were sustained during a 12-hour period.Chesler L ,et al. Neoplasia 2008, 10, 1268-74
  25. 25. CPM Treatment of Murine NeuroblastomaActivates Apoptosis · Bax, a downstream target of PUMA and a critical effector of myc- induced mitochondrial apoptosis, was strongly expressed and PARP occurred concurrently, indicating high levels of apoptosis. · The proapoptotic effect of CPM proceeds through the intrinsic p53 regulatory pathway of BH3 proteins, implicating PUMA, Bax, and Bim as key effectors of p53 function in primary murine neuroblastoma tumors.Chesler L ,et al. Neoplasia 2008, 10, 1268-74
  26. 26. Conclusion • The p53 pathway plays a significant role in opposing MYCN-driven oncogenesis in a mouse model of neuroblastoma and that basal inactivation of the pathway is achieved in progressing tumors. • Chemotoxic agents induce p53-dependent apoptosis in such tumors. It is consistent with clinical observations that therapy-associated mutations in p53 are a likely contributor to the biology of tumors at relapse and secondarily mediate resistance to therapy. Back BackChesler L ,et al. Neoplasia 2008, 10, 1268-74
  27. 27. Treatment-induced senescence-like phenotype isdependent on TP53 status · In the TP53 wild-type tumor Ki67 immunostainings were not statistically different before and after treatment . · SA-b-gal staining was negative before treatment in all tumors. After treatment, 5-10% tumor cells in TP53wt showed SA-b-gal cytoplasmic staining, as soon as Day 1 (Figs. d–2). · p21 immunostained cells increased starting at Day 3 with a mean number of stained cells over 50% (Figs. g–i).
  28. 28. Treatment-induced mitotic catastrophe andapoptosis is dependent on TP53 status · The TP53 mutant tumors remained negative for SA-b- gal at all time points after treatment (Figs. d–f). · No significant change for p21 immunostaining was observed in the TP53 mutant tumors (Figs. g–i). · The number of abnormal mitoses on semi-thin sections significantly increased between Day 3 and Day 5 (Figs. p–r).
  29. 29. Treatment-induced mitotic catastrophe andapoptosis is dependent on TP53 status The results were simi- lary to TP53mut1 tumor
  30. 30. Treatment-induced mitotic catastrophe andapoptosis is dependent on TP53 status · No significant change in Bax mRNA was found in TP53mut1 and TP53mut2 (Figs. e and h). · Similarly PUMA showed a strong mRNA overexpression at D3and D5 in TP53wt, but no change in mutant tumors (Figs. c, f and i)
  31. 31. Conclusion • Treatment-induced senescence-like phenotype is dependent on TP53 status. • Treatment-induced mitotic catastrophe and apoptosis is dependent on TP53 status • The lack of response in TP53 wild-type tumors may be due to the induction of cell cycle arrest, allowing tumor cells to reinitiate proliferation at the end of chemotherapy Varna M, et al. Int J Cancer 2009, 124, 991-7
  32. 32. Other agents
  33. 33. P53- and p21-deficient cells undergo apoptosisafter treatment with DNA-damaging agents Cells with wild type p53 were quite sensitive to 5-FU, and a large proportion underwent apoptosis. Cells with p21 deletions were as sensitive to 5-FU as p53 wild-type cells. Bunz F, el at. J Clin Invest 1999, 104,263-9
  34. 34. Cells with targeted p53 deletion are resistantto apoptosis induced by 5-FU · Apoptosis was not observed in p53-deficient cells with 5-FU treatment. · Tight control of 5-FU sensitivity by p53. · p21 does not play a role in the ability of p53 to modulate the response to 5- FU.Bunz F, el at. J Clin Invest 1999, 104,263-9
  35. 35. Cell cycle distribution of drug-treated cellswith wild-type p53 and disrupted p53 genes · Adriamycin: p53-deficient cells accumulated in a single peak with 4N DNA content. · 5-FU: regardless of p53 genotype, cellsaccumulated in a single peak that spanned the G1/S phase boundary. · Tomudex: both p53-proficient and p53- deficient cells exhibited an S-phase block. · Methotrexate: induced identical responses in all cells, with an increase in the S-phase fraction Bunz F, el at. J Clin Invest 1999, 104,263-9
  36. 36. Characteristics of 5-FU–induced apoptosis · Both adriamycin and 5-FU caused increases in p53 protein levels over a similar time course. · The stabilization of p53 was associated with increased levels of p21 · 5-FU treatment led to the dramatic induction of death in cells with intact p53, a relatively small proportion of cells apparently survived and gave rise to colonies upon replating.
  37. 37. Growth of xenograft tumors Tumors with intact p53 genes regressed during the treatment, whereas the tumors with deleted p53 genes continued to grow.Bunz F, el at. J Clin Invest 1999, 104,263-9
  38. 38. Conclusion • p53 had profound effects on drug responses, and these effects varied dramatically depending on the drug. • The p53-deficient cells were sensitized to the effects of DNA-damaging agents as a result of the failure to induce expression of the cyclin-dependent kinase inhibitor p21. • p53 disruption rendered cells strikingly resistant to the effects of the 5-FU.Bunz F, el at. J Clin Invest 1999, 104,263-9
  39. 39. PFTA potentiates the antitumor effect ofcyclophosphamide PFTa greatly potentiated the antitumor effect of cyclophosphamide, due to the effect of the p53 inhibitor on stromal cells. Burdelya LG ,et al. Cancer Res 2006, 66, 9356-61
  40. 40. Experimental design and model
  41. 41. · The levels of GSE56expression were similarbetween virus-producing andnonproducing populations(B)· No EYFP expressiondetected in the endotheliumof tumors formed by the cellstransduced with retroviralvector lacking the packagingsignal as judged byimmunofluorescent stainingof tumor sections byantibodies against mouseendothelium marker CD31 (C,bottom).· Both cells producing andnot producing GSE56 viruswere equally sensitive totreatment withseveralchemotherapeutic drugs in
  42. 42. Dependence of tumor chemosensitivity andradiosensitivity on the p53 status of stroma· No differences in the growth rate of tumors were formed (A).· Tumors formed by the cells that released the p53-inhibit- ing GSE56 virus showed a much stronger response to cyclophosphamide treatment (B).· For radiation, GSE56 virus- producing tumors showed significant size reduction, whereas Δ ψ-GSE56 tumors responded only with a slight decline in growth rate (C.D)· PFTa strongly sensitized Δ ψ-GSE56 tumors to radiation.
  43. 43. Conclusion • Tumors with p53-deficient stroma were significantly more sensitive to experimental chemotherapy and radiotherapy. • Potentiation of the anticancer effect of chemotherapy and radiotherapy by p53 suppression in the tumor stroma is likely to be due to the increased sensitivity of p53-deficient endothelium to genotoxic stress as shown both in cell culture and in experimental tumors.Burdelya LG ,et al. Cancer Res 2006, 66, 9356-61
  44. 44. p53 and chemotherapy 1.Introduction 1.Introduction 2. Relationship between p53 and chemotherapy 2. Relationship between p53 and chemotherapy 3. Summary 3. Summary
  45. 45. Summary • p53 pathway plays a significant role in chemotherapy. • p53 Induce cell death and cell circle arrest. Their balance contribute to the response of chemotherapy. • Effects of p53 in chemotherapy varied dramatically depending on the drug. • Tumors with p53-deficient stroma were significantly more sensitive to chemotherapy.

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