Therapeutic strategies: Targeting apoptosis in cancer - PowerPoint Presentation

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Therapeutic strategies:Targeting apoptosis in cancer

Last updated:

September 2020

IntroductionApoptosis, a form of programmed cell death, is a

key aspect of cellular homeostasis1In cancer, the processes and signals that promote apoptosis are inhibited, allowing cancer cells to survive and proliferate in a dysregulated manner2,3A deeper understanding of cell death pathways in cancer is essential for the development of precisely targeted treatments or synergistic treatment combinations4 A number of different strategies are under investigation to optimise outcomes with pro-apoptotic agents31. Fuchs Y, Stellar H. Cell 2011;147(4):74258; 2. Sharma A, et al. Cancers 2019;11:1144; 3. Jan R, Chaudry G. Adv Pharm Bull 2019;9(2):20518; 4. Ricci MS, Zong WX. Oncologist 2006;11(4):34257

.

Cell death and apoptosis

Cell death is a necessary biological function involved in growth, homeostasis and maintenance of healthy cells

Programmed cell death plays a fundamental role in animal development and tissue homeostasis1The cell death mechanism is involved during:1,2 1. Fuchs Y, Steller H. Cell 2011;147(4):74258; 2. Jan R, Chaudhry G. Adv Pharm Bull 2019;9:205–18.Elimination of abnormal cells during pathogenesisArrangement of cells during morphogenesisRegulation of cell number during homeostasis

Cell death typically occurs as a result of either necrosis or a programmed cell death such as apoptosis*

Necrosis occurs when cells die in response to injury or cellular stress, by swelling and rupturing1,2

1. Jacobson MD, et al. Cell 1997;88(3):347



54

; 2. Jan R, Chaudhry G.

Adv

Pharm Bull 2019;9:205-18;

3. Van

Cruchton

S, Van den

Broeck

W

.

Anat

Histol

Embryol

2002;31:214–23.*Apoptosis is one example of programmed cell death; other examples include autophagy and necroptosis.2Figures adapted from Van Cruchton S, Van den Broeck W. 2002.3Apoptosis occurs when cells die during (e.g.) homeostasis, by condensation – without losing membrane integrity - and removal by phagocytosis1,2Injured cell

Cell swellsCell becomes ‘leaky’ (blebbing)Cell breaks downRedundant cellCell and chromatin condense and shrink

Cell ‘buds’ into apoptotic bodies

Apoptotic bodies removed by phagocytosis

1. Fuchs Y, Steller H. Cell 2011;147(4):74258; 2. Elmore S. Toxicol

Pathol 2007;35(4):495–516. chemotherapeutic drugs Normal development Hormone-dependent changes in tissues Cell loss in proliferating populations

Elimination of lymphocytes Elimination of abnormal cells Response to radiation or

OH

HO

H

H

H

Apoptosis

Apoptosis can be triggered by a range of internal

and external circumstances

1,2

Apoptotic triggers act through the intrinsic or extrinsic signalling pathways to cause apoptosis1. Green DR,

Llambi F. Cold Spring Harb Perspect Biol 2015;7:a006080; 2. Zhang M, et al. Sci Rep 2017;7:2635; 3. Bratton SB, Salveson GS. J Cell Sci 2010;123(19):3209–14; 4. Jan R, Chaudhry G. Adv Pharm Bull 2019;9:205–18; 5. Martinez-Ruiz G, et al. J Exp Clin Cancer Res 2008;27:48; 6. Baig S, et al. Cell Death Dis 2016;7:e2058.Apaf, apoptotic protease activating factor; Bak, Bcl-2 antagonist/killer; Bax, Bcl-2-associated X protein; Bcl, B-cell lymphoma; IAP, inhibitor of apoptosis proteins; SMAC, second mitochondrial-derived activator of caspases (also known as DIABLO, direct IAP-binding protein with low pI); XIAP, X-linked inhibitor of apoptosis protein. Figure adapted from Baig et al. 2016.6The

intrinsic pathway

is initiated by

apoptogenic

triggers or cellular stress

1

This leads to the insertion of the Bcl-2 proteins,

Bax

/

Bak

, into the mitochondria, followed by cytochrome C release

2

Cytochrome C combines with Apaf-1 and procaspase-9 to form an

apoptosome

, which activates the

caspase cascade

to terminate the cell

3–6

SMAC, a pro-apoptogenic mitochondrial protein, is also released into the cytosol, where it antagonizes IAPs to allow caspase activation and subsequent apoptosis5BH3protein

Bax/BakSMAC

XIAP

IAP

Mitochondrion

Cytochrome C

Apoptosis

Procaspase-9

Apoptosome

Apaf-1

Caspase 3,6,7

Apoptotic triggers act through the intrinsic or extrinsic signalling pathways to cause apoptosis

1. Green DR, Llambi F. Cold Spring Harb Perspect Biol

2015;7:a006080;

2.

Baig

S, et al. Cell Death Dis 2016;7:e2058.

DISC, death-inducing

signaling

complex; TRAIL,

tumor necrosis factor-related apoptosis-inducing ligand.

Figure adapted from

Baig

et al. 2016.

2

The

extrinsic pathway

is triggered by

external stimuli or ligand molecules such as TRAIL

1Activation of the extrinsic pathway then triggers the caspase cascade to promote apoptosis1Activated death receptor

DISCCaspase 8

Procaspase 3,6,7

Caspase 3,6,7

Apoptosis

Evasion of apoptotic processes in cancer

Cancer cells survive and proliferate via evasion of apoptogenic triggers

Abnormal regulation of programmed cell death is associated with many diseases, including cancer1Defects in DNA repair and chromosome segregation normally trigger cell death to remove these genetically unstable cells2Defects in apoptosis:Allow genetically unstable cells to survive and allow selection of progressively aggressive clones2,3 Allow neoplastic cells to survive beyond their typical lifespan, providing protection from hypoxia and oxidative stress as the tumor mass expands3Allow epithelial cells to survive in a suspended state, detached from the extracellular matrix, which facilitates metastasis3 1. Fuchs Y, Stellar H. Cell 2011;147(4):74258; 2. Hassan M, et al. Biomed Res Int 2014;2014:150845; 3. Reed JC.

Cancer Cell. 2003;3(1):17–22.

chemotherapeutic drugs

Normal development Hormone-dependent changes in tissues Cell loss in proliferating populations

Elimination of lymphocytes Elimination of abnormal cells Response to radiation or

OH

HO

H

H

H

Apoptosis

Cancer cells inhibit the intrinsic pathway by upregulating anti-apoptotic Bcl-2 proteinsBak, Bcl-2 antagonist/killer;

Bax, Bcl-2 associated X protein; Bcl, B-cell lymphoma; BH, Bcl-2 homology; BIM, Bcl-2 interacting mediator of cell death; MOMP, mitochondrial outer membrane permeabilization; PUMA, p53-upregulated modulator of apoptosis; tBID, truncated BH3 interacting domain death agonist.Figure adapted from Baig et al. 2016.41. van Delft MF, Huang DCS. Cell Res 2006;16:203–13; Happo L, et al. J Cell Sci 2012;125(5):1081–7; 3. Sharma A, et al. Cancers 2019;11:1144; 4. Baig S, et al. Cell Death Dis 2016;7:e2058.

BH3

protein

Bax

/

Bak

SMAC

XIAP

IAP

Mitochondrion

Cytochrome C

Apoptosis

Procaspase-9

Apoptosome

Apaf-1

Caspase 3,6,7

Stimulus, e.g. ROS

Release of ‘activators’ (e.g.) PUMA

Activation of apoptotic pathway via MOMP

Bcl-2 proteins – all of whom contain BH1-4 domains – are key regulators of the intrinsic pathway

1,2

The intrinsic pathway is initiated by release of pro-apoptotic

BH3 ‘activators’

(i.e. BIM, PUMA,

tBID

) that activate multi-domain ‘effectors’ (e.g.

Bax

;

Bak

) to promote MOMP

2

Cancer cells often express elevated levels of pro-apoptotic BH3-only proteins

3

Thus, cancer cells will typically upregulate

anti-apoptotic Bcl-2 proteins

in order to sequester unbound pro-apoptotic proteins, preventing MOMP and activation of the apoptotic cascade

3

Cancer cells inhibit the intrinsic pathway by suppressing p53 expressionThe p53 tumor suppressor protein

upregulates pro-apoptotic BH3-only proteins such as PUMA and NOXA1The p53 gene is mutated or deleted in many human cancers, inactivating its suppressor activity2In some cancers with wild-type p53 status, its function is effectively inhibited by amplified expression of MDM2, which is the primary cellular inhibitor of p531,2Thus, by inhibiting or deleting p53 activity, malignant cells remove a key driver for apoptosis and maintain their proliferative stateBH, Bcl-2 homology; MDM2, murine double minute 2; PUMA, p53 upregulated modulator of apoptosis.

1. Aubrey

BJ, et

al. Cell Death Differ 2018;25:104–13; 2,

Shangary

S, Wang S.

Clin

Cancer Res 2008;14(17):5318–24.

3. Nag S, et al. J Biomed Res 2013;27(4);254

–

71; 4. Zhao Y, et al.

Acta

Biochim

Biophys

Sin 2014;46:180

–

9.Tumor cell3,4

Tumor

cellCancer cells inhibit the intrinsic pathway by modifying cell metabolismSpecific Bcl-2 proteins can be regulated by metabolite stresses1Glucose deprivation may induce PUMA via p53 induction1p53 inhibits glycolysis through down-regulation of glucose transporters, glycolytic enzymes, and inhibition of hypoxic-inducible factors1Activation of oncogenes (e.g. RAS, AKT, MYC) as well as loss of genes such as p53

drive

aerobic glycolysis

in cancer cells and promote a glycolytic phenotype,

1,2

thereby

subverting micronutrient-driven apoptotic activation

1.

Sharma A, et al. Cancers 2019;11:1144; 2. Zheng J.

Oncol

Lett

2012

;4:1151–7.

Bcl

, B-cell lymphoma; PUMA, p53-upregulated modulator of apoptosis;

tBID

, truncated BH3 interacting domain death agonist.

p53

Oncogene

Glycolytic environment

Cancer cells inhibit the intrinsic pathway by offsetting oxidative stress mechanisms

Higher ROS can upregulate cell death pathways1 Low ROS may upregulate proliferative pathways and, thus, promote tumorigenesis2 Due to their high metabolic activity, cancer cells often contain increased ROS2,3 Consequently a cancer cell must synthesize antioxidants in order to retain the cell’s redox homeostasis2,3Cancer cells may also increase the uptake of antioxidant nutrients and metabolic enzymes2 to moderate intracellular ROSROS, reactive oxygen species. 1.

Redza-Dutordoir

M, et al.

Biochim

Biophys

Acta

2016;1863:2977–92;

2

.

Sharma A, et al. Cancers 2019;11:1144; 3

.

Liou

GY,

Storz

P. Free

Radic

Res 2010;44(5):479–96.

Increased ROS =

increased cell death activation

Decreased ROS =

increased tumorigenesis

Cancer cells have evolved strategies to evade extrinsic pathway-induced apoptosisThe extrinsic pathway is mediated by the death receptors TNFR1, CD95,

FasR, APO-1, DR4 and DR51Death receptors are activated by regulatory ligands such as TRAIL, which induce DISC formation and initiate the apoptotic pathway1In cancer cells, TRAIL resistance may be caused by various genetic mutations (resulting in altered apoptotic signaling proteins), or overexpression of anti-apoptotic Bcl-2 proteins1APO-1, apoptosis antigen 1; Bcl, B-cell lymphoma; CD95, cluster of differentiation 95; DISC, death-inducing signaling complex; DR, death receptor; FasR, Fas receptor; TNFR1, tumor necrosis factor receptor 1; TRAIL, TNF-related apoptosis-inducing ligand.Figure adapted from Baig et al. 2016.2

1.

Ukrainskaya VM, et al.

Acta

Naturae

2017;9(3):55–63

; 2.

Baig

S, et al. Cell Death Dis 2016;7:e2058.

Activated death

receptor

DISC

Caspase 8

Procaspase-3,6,7

Caspase 3,6,7

Apoptosis

Targeting apoptosis in cancer therapy

Apaf, apoptotic protease activating factor; Bak, Bcl-2 antagonist/killer; Bax, Bcl-2-associated X protein; Bcl

, B-cell lymphoma; DISC, death-inducing signaling complex; DR, death receptor; IAP, inhibitor of apoptosis proteins; MDM2, murine double minute 2; SMAC, second mitochondrial-derived activator of caspases (also known as DIABLO, direct IAP-binding protein with low pI); X-linked inhibitor of apoptosis protein. *These are investigational compounds and have not been approved. Their efficacy and safety have not been established.Figures adapted from Baig et al. 2016.31. Ricci MS, Zong WX. Oncologist 2006;11(4):342–57; 2. Jan R, Chaudry G. Adv

Pharm Bull 2019;9(2):20518; 3.

Baig

S, et al. Cell Death Dis 2016;7:e2058.

DISC

Caspase 8

Procaspase-3,6,7

Caspase 3,6,7

Apoptosis

DR ligands

2

BH3

protein

Bax

/

bak

SMAC

XIAP

IAP

Mitochondrion

Cytochrome C

Apoptosis

Procaspase-9

Apoptosome

Apaf-1

Caspase 3,6,7

p53/MDM2 expression

2

Attenuation of

Bcl-2 proteins

2

SMAC expression

2

Targeted induction

of apoptosis

in cancer cells

A deeper understanding of cell death pathways in cancer is essential for the development of precisely targeted treatments or synergistic treatment

combinations

1*

Bcl, B-cell lymphoma; BH, Bcl-2 homology; Mcl-1. myeloid cell leukemia-1; XIAP, X-linked inhibitor of apoptosis protein.*These are investigational compounds and have not been approved. Their efficacy and safety have not been

established. 1. NCT02427451. Available at: https://clinicaltrials.gov/ct2/show/NCT02427451. Accessed July 2020; 2. NCT04277637. Available at: https://clinicaltrials.gov/ct2/show/NCT04277637. Accessed July 2020; 3. Baig S, et al. Cell Death Dis 2016;7:e2058; 4. Lu AQ, et al. Int J Clin Exp Med 2018;11(7):6767–75; 5. Boffo S, et al. J Exp Clin Cancer Res 2018;37:36; 6. Jan R,

Chaudry

G.

Adv

Pharm Bull 2019;9(2):205–18; 7. Mukherjee N, et al. Cell Death Dis 2018;9:907.

Bcl-2

inhibitors

1,2*

Antisense

oligonucleotides

*

to enhance sensitivity to cytotoxic drugs

3,4

BH3-mimicking

agents

6,7*

Cyclin-dependent kinase

inhibitors

*

to downregulate expression of factors such as Bcl-2, Mcl-1 and XIAP

5

Strategies

used to inhibit

the anti-apoptotic

Bcl-2 family include:

Approaches to target the intrinsic pathway: Bcl-2

attenuation

Spotlight on p53/MDM2The tumor suppressor gene p53 impacts both cell cycle arrest and apoptosis

1p53 activity is inhibited by MDM2 upregulationAntisense oligonucleotides and MDM2 inhibitors* have been developed to reduce MDM2 overexpression and trigger wild-type p53 activity1,2Another approach uses small molecules that reactivate wild-type function* of mutant p531,3MDM2, mouse double-minute 2; wt, wild-type. *These are investigational compounds and have not been approved. Their efficacy and safety have not been established.1. Jan R, Chaudry

G.

Adv

Pharm Bull 2019;9(2):205–18; 2. Rayburn ER, et al. Anticancer Agents Med

Chem

2009;9(8):882–903; 3.

Di Agostino S, et al.

J

Exp

Clin

 Cancer Res 2019;38(1):290

;

4.

Rudolph D, et al. Presented at the Annual Meeting of the American Association for Cancer Research 2018. Abstract 4868 and poster;

5. Nag S, et al. J Biomed Res 2013;27(4);254

–71; 6. Zhao Y, et al. Acta Biochim Biophys Sin 2014;46:180–9.Interaction inhibitor*

Tumor cell4–6

Spotlight on SMACSMAC is released in response to Bcl-2 activity on the mitochondrion, where it binds to and induces IAP degradation to promote apoptosis

1In cancer cells, elevated IAP expression increases cell survival, tumor growth and metastasis1SMAC mimetics* bind to cellular IAPs to restore effector caspase function and are under investigation alone or as combination therapy2,3However, SMAC is overexpressed in some cancers, suggesting a role in non-apoptotic cancer cell survival and a possible role for reducing SMAC expression in some cancers4,51. Bai L, et al. Pharmacol

Ther

2014;144(1):82–95;

2

. Jan R,

Chaudry

G.

Adv

Pharm Bull 2019;9(2):205–18; 3.

Derakshan

A, et al.

Clin

Cancer Res 2017;23(6):1379–87; 4. Zhao XY, et al. Cells 2020;9:1012; 5. Paul A, et al.

Mol

Ther

2018;26(3):680–94

; 6. Baig S, et al. Cell Death Dis 2016;7:e2058. Bcl, B-cell lymphoma; IAP, inhibitor of apoptosis protein; SMAC, second mitochondrial-derived activator of caspases (also known as DIABLO, direct IAP-binding protein with low pI).*These are investigational compounds and have not been approved. Their efficacy and safety have not been established.Figure adapted from Baig et al. 2016.6BH3protein

Bax/bakSMAC

XIAP

IAP

Mitochondrion

Cytochrome C

Apoptosis

Procaspase-9

Apoptosome

Apaf-1

Caspase 3,6,7

Spotlight on TRAIL ligandsDR, death receptor; TNF, tumor necrosis factor; TRAIL, TNF-related apoptosis-inducing ligand

.*These are investigational compounds and have not been approved. Their efficacy and safety have not been established.Figure adapted from Garcia-Martinez et al. 2019.5DR4 and DR5 receptors are candidates for targeted tumor therapy, due to their high expression levels in cancer cells1DR4 and DR5 are activated by TRAIL ligands, which are widely expressed on cells.2 Approaches to targeting TRAIL in cancer therapy include: TRAIL-R novel forms3*TRAIL-R1 and -R2 antibodies1,4*Peptide agonists1*A key feature of targeting TRAIL is its reduced cytotoxicity to normal cells versus, for example,

Fas

ligands or TNF

1

1.

Ukrainskaya VM, et al.

Acta

Naturae

2017;9(3):55–63

; 2.

Ashkenazi A, et al. J

Clin

Invest 1999;104(2):155–62; 3.

Leng

Y, et al. Cancer

Chemother

Pharmacol 2017;79(6):1141–49; 4. NCT04137289. Available at https://clinicaltrials.gov/ct2/show/NCT04137289. Accessed July 2020; 5. Garcia-Martinez JM, et al. American Association for Cancer Research Annual Meeting 2019; Abstract 2051.TRAILR2/CDH17 antibody*Tumor-specific induction of apoptosis

SummaryAbnormal regulation of apoptosis is associated with many diseases, including cancer1Apoptosis is a complex process with

numerous points of regulation that may be targeted to provide therapeutic benefit in cancer2Future cancer treatments may include modulation of strategic points of the intrinsic and extrinsic apoptotic pathways, such as:Inhibition of anti-apoptotic Bcl-2 family3* Upregulation of tumor suppressor gene p534*Regulation of SMAC activity3*Application of TRAIL antibodies5*Bcl, B-cell lymphoma; SMAC, second mitochondrial-derived activator of caspases (also known as DIABLO, direct IAP-binding protein with low pI); TRAIL, TNF-related apoptosis-inducing ligand.*These

are investigational compounds and have not been approved. Their efficacy and safety have not been established.

1.

Fuchs Y, Stellar H. Cell 2011;147(4):742

58; 2.

Fox JL, Macfarlane M. Br J Cancer 2016;115:5–11; 3.

Jan R,

Chaudry

G.

Adv

Pharm Bull 2019;9(2):205–18; 4. Rayburn ER, et al. Anticancer Agents Med

Chem

2009;9(8):882–903; 5.

Ukrainskaya VM, et al.

Acta

Naturae

2017;9(3):55–63.