10. Apoptose celular

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Apoptose
Profa. Dra. Adrianne C Palanch
UNIMEP
Apoptose é um processo importante
Apoptose (morte celular programada) tem um importante papel no desenvolvimento e na homeostasia
Apoptose é ativada por duas vias de sinalização diferentes : intrínseca e extrínseca
Células normais entram em apoptose em resposta a eventos induzidos por extresse celular que levam ao dano do DNA
Erros no processo de apoptose podem definir o desenvolvimento do câncer e a sobrevivência das células tumorais 
Resumo das duas principais vias de Apoptose
Apoptose
Caspases
Receptor Pró-apoptótico 
Ligante Pró-apoptótico 
Via extrínseca
Via intrínseca
BAX
BCL2
PUMA
Estresse
Mitocôndria
Adaptado de Ashkenazi A. Nat Rev Cancer 2002;2:420–430. BAX, proteína associada a BCL2; BCL2, linfoma 2/leucemia linfótica crônica de células B; PUMA, modulador de apoptose inibidor de p53
Apoptose desencadeada por Caspases 
(Cysteine Aspartate Specific ProteASEs)
Iniciador de caspases
Estímulo pró-apoptótico
Efetor de caspases
Cascata de Caspases
Adaptado de Thornberry NA, Lazebnik Y. Science 1998;281:1312–1316. Caspase, cysteine aspartase.
Via intrínseca da Apoptose
Adaptado do Ashkenazi A. Nat Rev Cancer 2002;2:420–430. APAF1, fator de ativação da protease apoptótica-1; BAK, antagonista homólogo ao BCL2/killer; BAX, proteína associada ao BCL2; BCL2, B-cell chronic lymphocytic leukemia/lymphoma 2; BCLXL, BCL2-like 1; IAP, inhibitor of apoptosis protein; MCL1, myeloid cell leukemia sequence 1 (BCL2-related); PUMA, p53-upregulated modulator of apoptosis; SMAC/DIABLO: second mitochondria-derived activator of caspase/direct IAP binding protein with low pI.
Quimioterapia
Radioterapia
Caspase 3, 6, 7
Via intrínseca
BCL2, BCLXL, MCL1
Mitocôndria
SMAC/DIABLO
p53
Caspase 9
Dano do DNA 
PUMA, NOXA
APAF1
Citocromo c
Dano do DNA 
IAP
Via intrínseca da Apoptose
A via intrínseca é iniciada pelo p53 tumor-suppressor em resposta ao dano do DNA e outros tipos de estresse celular severo
Terapias anticancer convencionais, como quimioterapia e radioterapia, ativam esta via dependente de p53
p53 ativa a via intrínseca através da inibição transcricional dos membros pró-apoptóticos da família BCL2, proteínas como PUMA e BAX
BAX causa a liberação do citocromo c da mitocôndria, o qual junto com o adaptador APAF1, ativando o iniciador caspase 9
Caspase 9 ativa os efetores caspases 3, 6 e 7, os quais são responsáveis pela destruição dos componentes críticos da célula e induz apoptose
p53 é inativado por mutações em mais das metades dos cânceres humanos
A via extrínseca da Apoptose
Adaptado de Ashkenazi A. Nat Rev Cancer 2002;2:420–430. FADD, Fas-associated death domain; FLIP, FLICE (FADD-like interleukin 1β-converting enzyme) inhibitory protein.
Ligante Pró-apoptótico
Caspase 3, 6, 7
Apoptose
FADD
FLIP
DR5
DR4
Via extrínseca
Pró-caspase 8, 10
Caspase 8, 10
Via extrínseca da Apoptose
A via extrínseca ativa a apoptose em resposta a ativação de receptores pró-apoptótico, como DR4 e DR5, por ligantes específicos pró-apoptóticos, como Apo2L/TRAIL
Esta via estimula apoptose independente do p53
Ativação induzida pelo ligante ao DR4 e DR5 leva a um arranjo rápido do complexo de morte induzida (death-inducing signaling complex-DISC) e o recrutamento do iniciador da caspases 8 e 10 por meio do adaptador do domínio de morte associado do Fas (Fas-associated death domain-FADD)
Caspases 8 e 10 ativa o efetor de caspases 3, 6 e 7 levando a apoptose.
As duas vias principais de Apoptose
Adaptado do Ashkenazi A. Nat Rev Can 2002;2:420–430. APAF1, apoptotic protease activating factor-1; BAK, BCL2 homologous antagonist/killer; BAX, BCL2-associated protein; BCL2, B-cell chronic lymphocytic leukemia/lymphoma 2; BCLXL, BCL2-like 1; BID, BH3-interacting domain death agonist; DR, death receptor; FADD, Fas-associated death domain; FLIP, FLICE (FADD-like interleukin 1β-converting enzyme) inhibitory protein; IAP, inhibitor of apoptosis protein; MCL1, myeloid cell leukemia sequence 1 (BCL2-related); PUMA, p53-upregulated modulator of apoptosis; SMAC/DIABLO: second mitochondria-derived activator of caspase/direct IAP binding protein with low pI.
Caspase 3, 6, 7
Apoptose
Ligante Pró-apoptótico
FADD
DR5
DR4
Via extrínseca
Pró-caspase 8, 10
Caspase 9
Caspase 8, 10
p53
BAX, BAK
Mitocôndria
SMAC/DIABLO
Quimioterapia
Radioterapia
Dano no DNA 
PUMA, NOXA
APAF1
Citocrome c
Dano do DNA
BID
IAP
Via intrínseca
BCL2, BCLXL, MCL1
Apoptose via Receptores Pró-apoptóticos (DR4 e DR5)
 Apo2L/TRAIL Endógeno
DR4
DR5
Apo2L/TRAIL, apoptosis-inducing ligand 2/tumor necrosis factor-related apoptosis-inducing ligand; DR, death receptor.
Apo2L/TRAIL Endógeno ativa receptores pró-apoptóticos (DR4 e DR5)
DR4
DR5
DR, death receptor.
Receptores Pró-apoptóticos (DR4 e/ou DR5) recruta FADD
DR4
DR5
DR, death receptor; FADD, Fas-associated death domain.
FADD recruta o iniciador de Caspase 8 e/ou 10 
para o DISC
DR4
DR5
Pró-caspase 8, 10
Pró-caspase 8, 10
DISC
DR, death receptor; DISC, death-inducing signaling complex.
As Caspases 8 e 10 ativadas são liberadas dentro do citoplasma
Caspase 8,10
Caspase 8,10
DR4
DR5
DR, death receptor.
Caspase 3, 6 e 7 são ativadas
DR4
DR5
Caspase 3, 6, 7
Caspase 3, 6, 7
Caspase 8, 10
Caspase 8, 10
DR, death receptor.
Caspases 3, 6 e 7 quando ativadas estimulam Apoptose em Células tumorais
Adapted from Ashkenazi A. Nat Rev Cancer 2002;2:420–430. APAF1, apoptotic protease activating factor-1; BAK, BCL2 homologous antagonist/killer; BAX, BCL2-associated protein; BCL2, B-cell chronic lumphocytic leukemia/lymphoma 2; BCLXL, BCL2-like 1; BID, BH3-interacting domain death agonist; DR, death receptor; FADD, Fas-associated death domain; FLIP, FLICE (FADD-like interleukin 1β-converting enzyme) inhibitory protein; IAP, inhibitor of apoptosis protein; MCL1, myeloid cell leukemia sequence 1 (BCL2-related); PUMA, p53-upregulated modulator of apoptosis; SMAC/DIABLO: second mitochondria-derived activator of caspase/direct IAP binding protein with low pI.
Caspase 3, 6, 7
Apoptose
Ligante Pró-apoptótico
FADD
DR5
DR4
Via extrínseca
Pró-caspase 8, 10
p53
p53
Caspase 9
Caspase 8, 10
p53
BAX, BAK
Mitocôndria
SMAC/DIABLO
Quimioterapia
Radioterapia
Dano no DNA 
PUMA, NOXA
APAF1
Citocrome c
Dano no DNA
BID
IAP
Via intrínseca
BCL2, BCLL, 
MCL1
Vias da Apoptose
filme apoptose
Células em Apoptose
Referências
Ashkenazi A. Targeting death and decoy receptors of the tumor-necrosis factor superfamily. Nat Rev Cancer 2002;2:420–430.
Thornberry NA, Lazebnik Y. Caspases: Enemies within. Science 1998;281:1312–1316.
Ghobrial IM, Witzig TE, Adjei AA. Targeting apoptosis pathways in cancer therapy. CA Cancer J Clin 2005;55:178–194.
Cells have a built-in cell death program called apoptosis, which is essential for normal development and homeostasis. Apoptosis is activated through two major signaling pathways.1–2 The intrinsic pathway is triggered from within the cell by developmental cues or severe cell stress, such as DNA damage. The extrinsic pathway is activated when a pro-apoptotic ligand, such as endogenous Apo2L/TRAIL (apoptosis-inducing ligand 2 or tumor necrosis factor-related apoptosis-inducing ligand), binds to key functional pro-apoptotic receptors, such as death receptor (DR) 4 and DR5. The apoptotic cell death program is carried out by intracellular protease enzymes called caspases that, on activation through the intrinsic and/or extrinsic pathways, destroy a large set of cellular proteins important for cell viability.
Normal cells switch on apoptosis in response to stress-inducing events such as irreparable DNA damage. Dysregulation of apoptosis is critical for cancer development and tumor cell survival.3
Abbreviations:
BAX, BCL2-associated protein;
BCL2, B-cell chronic lymphocytic leukemia/lymphoma 2; PUMA, p53-upregulated modulator of apoptosis.
There are two tiers of caspase activation during apoptosis. First, initiator caspases are activated through the apoptosis signaling pathways. In turn, the initiator caspases activate, in cascade fashion, the effector caspases, which carry out the apoptosis program.4
Abbreviations:
Caspase, cysteine aspartase.
The intrinsic apoptosis pathway is triggered in response to DNA damage and other types of severe cell stress and involves the release of pro-apoptotic factors from the mitochondria.1 p53-mediated activation of the pro-apoptotic BCL2 family proteins, including PUMA, NOXA, BAX and BAK, leads to the release of cytochrome c and SMAC/DIABLO from the mitochondria. Cytochrome c binds APAF1, forming an “apoptosome” that activates the apoptotic protease caspase 9. Caspase 9, in turn, activates downstream caspases, including caspase 3, 6, and 7, leading to apoptosis. SMAC/DIABLO, a pro-apoptotic protein, directly interacts with inhibitor of apoptosis (IAP) proteins, preventing them from attenuating apoptosis.
Anti-apoptotic members of the BCL2 family, BCL2, BCLXL, and MCL1, regulate the mitochondria-initiated caspase activation pathway by preventing the release of cytochrome c in the presence of apoptotic stimuli.
The tumor suppressor gene p53 is inactivated by mutations in more than half of human cancers.5 Failure of cancer cells to enter apoptosis, often through p53 mutations, can result in resistance to conventional anticancer therapies (chemotherapy and radiotherapy).6–8
Abbreviations:
APAF1, apoptotic protease activating factor-1; BAK, BCL2 homologous antagonist/killer; BAX, BCL2-associated protein; BCL2, B-cell chronic lymphocytic leukemia/lymphoma 2; BCLXL, BCL2-like 1; IAP, inhibitor of apoptosis protein; MCL1, myeloid cell leukemia sequence 1 (BCL2-related); PUMA, p53-upregulated modulator of apoptosis; SMAC/DIABLO: second mitochondria-derived activator of caspase/direct IAP binding protein with low pI.
The extrinsic apoptosis pathway triggers apoptosis independently of p53 in response to pro-apoptotic ligands, such as Apo2L/TRAIL. These ligands activate specific pro-apoptotic receptors, such as DR4 and DR5. Ligand-induced activation of DR4 and DR5 leads to the recruitment of the adaptor Fas-associated death domain (FADD) and initiator procaspases 8 and 10 to rapidly form the death-inducing signaling complex (DISC). Procaspase 8 and 10 are self-cleaved, and activated caspase 8 and 10 activate the effector caspases 3, 6, and 7, so triggering apoptosis.
Abbreviations:
FADD, Fas-associated death domain; FLIP, FLICE (FADD-like interleukin 1β-converting enzyme) inhibitory protein.
The two major apoptosis signaling pathways converge at the level of the effector caspases. Moreover, there can be cross-talk between the extrinsic and intrinsic pathways. Caspase 8 cleaves the pro-apoptotic BCL2 family member BID, which in turn engages the mitochondria through BAX and BAK, leading to activation of caspase 9 and further activation of caspases 3, 6, and 7. The effector caspases also can activate caspase 8 and 10, forming a positive amplification loop.
Abbreviations:
APAF1, apoptotic protease activating factor-1; BAK, BCL2 homologous antagonist/killer; BAX, BCL2-associated protein; BCL2, B-cell chronic lymphocytic leukemia/lymphoma 2; BCLXL, BCL2-like 1; BID, BH3-interacting domain death agonist; DR, death receptor; FADD, Fas-associated death domain; FLIP, FLICE (FADD-like interleukin 1β-converting enzyme) inhibitory protein; IAP, inhibitor of apoptosis protein; MCL1, myeloid cell leukemia sequence 1 (BCL2-related); PUMA, p53-upregulated modulator of apoptosis; SMAC/DIABLO: second mitochondria-derived activator of caspase/direct IAP binding protein with low pI.
The key functional pro-apoptotic receptors DR49 and DR510 are cell-surface transmembrane receptors that are expressed in a broad range of malignancies. Endogenous Apo2L/TRAIL is a protein capable of triggering apoptosis in a variety of tumor cells,11–12 regardless of p53 status. Apo2L/TRAIL does not induce detectable apoptosis in normal tissue cell types including epithelial, endothelial, fibroblastic, smooth muscle, and astrocytic cells and hematopoietic stem cells.12–15
Abbreviations:
Apo2L/TRAIL, apoptosis-inducing ligand 2/tumor necrosis factor-related apoptosis-inducing ligand; DR, death receptor.
Endogenous Apo2L/TRAIL triggers apoptosis by binding to the extracellular component of DR4 and/or DR5 and activating the extrinsic signaling cascade. The intracellular domains of DR4 and/or DR5 assemble the DISC.16 The DISC consists of endogenous Apo2L/TRAIL, DR4 and/or DR5, FADD, and caspase 8 and/or caspase 10.
Abbreviations:
DR, death receptor.
The first stage in assembly of the DISC is the recruitment of the endogenous adaptor protein FADD.16–17
Abbreviations:
DR, death receptor; FADD, Fas-associated death domain.
FADD then recruits the endogenous apoptosis-initiating enzymes, procaspase 8 and procaspase 10, to the DISC.16,18
Abbreviations:
DR, death receptor; DISC, death-inducing signaling complex.
Procaspase 8 and 10 are self-cleaved to form activated caspase 8 and 10, and are released from the DISC into the cytoplasm.18
Abbreviations:
DR, death receptor.
Cleaved caspase 8 and caspase 10 activate effector enzymes such as caspase 3, caspase 6, and caspase 7.19–20
Abbreviations:
DR, death receptor.
The two major apoptosis signaling pathways converge at the level of the effector caspases. Moreover, there can be cross-talk between the extrinsic and intrinsic pathways. Caspase 8 cleaves the pro-apoptotic BCL2 family member BID, which in turn engages the mitochondria through BAX and BAK, leading to activation of caspase 9 and further activation of caspases 3, 6, and 7. The effector caspases also can activate caspase 8 and 10, forming a positive amplification loop.
Abbreviations:
APAF1, apoptotic protease activating factor-1; BAK, BCL2 homologous antagonist/killer; BAX, BCL2-associated protein; BCL2, B-cell chronic lymphocytic leukemia/lymphoma 2; BCLXL, BCL2-like 1; BID, BH3-interacting domain death agonist; DR, death receptor; FADD, Fas-associated death domain; FLIP, FLICE (FADD-like interleukin 1β-converting enzyme) inhibitory protein; IAP, inhibitor of apoptosis protein; MCL1, myeloid cell leukemia sequence 1 (BCL2-related); PUMA, p53-upregulated modulator of apoptosis; SMAC/DIABLO: second mitochondria-derived activator of caspase/direct IAP binding protein with low pI.