Programmed cell death

(Apoptosis)

Nidhika
M.Pharm.
(Pharmacology)
IInd sem.
Introduction
• Apoptosis, or programmed cell death, is a highly
regulated process that allows a cell to self-degrade in
order for the body to eliminate unwanted or
dysfunctional cells. During apoptosis, the genome of
the cell will fracture, the cell will shrink and part of
the cell will disintegrate into smaller apoptotic
bodies.
• Between 50 and 70 billion cells die each day due to
apoptosis in the average human adult. For an average
child between the ages of 8 and 14, approximately 20
billion to 30 billion cells die a day.
History

• German scientist Carl Vogt was first to describe the

principle of apoptosis in 1842. In 1845, anatomist
Walther Flemming delivered a more precise description
of the process of programmed cell death.

• Kerr received the Paul Ehrlich and Ludwig

Darmstaedter Prize on March 14, 2000, for his
description of apoptosis. He shared the prize with Boston
biologist Robert Horvitz.
• The 2002 Nobel Prize in Medicine was awarded to Sydney
Brenner, Horvitz and John E. Sulston for their work
identifying genes that control apoptosis. The genes were
identified by studies in the nematode C. elegans and these
same genes function in humans for apoptosis.
• Apoptosis has since been recognized and accepted as
a distinctive and important mode of “programmed”
cell death, which involves the genetically determined
elimination of cells.
• Apoptosis is essential to embryonic development and
the maintenance of homeostasis in multicellular
organisms. In humans, for example, the rate of cell
growth and cell death is balanced to maintain the
weight of the body. During fetal development, cell
death helps sculpt body shape and making the right
neuronal connections.
• Tissue homeostasis mainly depends on the balance
between cell proliferation and cell death. Programmed
cell death or apoptosis is an intrinsic death program that
occurs in various physiological and pathological
situations .
• Apoptosis or self destruction is necessary for normal
development and homeostasis of multicellular organisms.
Apoptosis plays a major role in many diseases like
cancer,
AIDS and
neurodegenerative disorders.
 Cell death
• Cell die by one of two mechanisms-

- Necrosis - Death By Injury

- Apoptosis - Death By Suicide

• Apoptosis and necrosis have different

characteristics
NECROSIS Vs APOPTOSIS
APOPTOSIS ( Physiological cell death) NECROSIS ( Pathological cell death)

Functional form of cell death Accidental form of cell death

Occurs under physiological conditions Seen under pathological conditions

Energy (ATP)- dependent No energy requirement

Cell shrinks and pulls away from its Cell swelling in a defining features
neighbours
Nucleus ruptures Entire cell balloons and ruptures

Induced by physiological stimuli( lack of Induced by non-physiological disturbances

growth factor, changes in harmonal lytic viruses, hypothermia,hypoxia,
environment ischaemia,metabolic poisons

No inflammation follows apoptosis Necrosis is followed by inflammation

 Need Of Apoptosis
• Apoptosis is needed for proper development
Examples:
The resorption of the tadpole tail
The formation of the fingers and toes of the fetus
The formation of the proper connections between neurons in the
brain
• Apoptosis is needed to destroy cells
Examples:
Cells infected with viruses
Cells with DNA damage
Cancer cells
Incomplete differentiation due to lack
of Apoptosis
 Reasons of apoptosis
• Withdrawal of positive signals
examples :
– growth factors for neurons
– Interleukin-2 (IL-2)

• Receipt of negative signals

examples :
– increased levels of oxidants within the cell
– damage to DNA by oxidants
– death activators :
• Tumor necrosis factor alpha (TNF-α)
• Lymphotoxin (TNF-β)
• Fas ligand (FasL)
 Inducers of Apoptosis
• TNF family

• Growth factor withdrawl

• Calcium

• Oncogenes

• Nutrient deprivation

• Toxins

• UV radiation

• Gamma radiation
 Apoptosis in physiological
conditions

In human body about 100,000 cells are produced

every second by mitosis and a similar no. die by
apoptosis
• Important in normal physiology
– Development: Immune systems maturation,
Morphogenesis, Neural development
– Adult: Immune privilege, DNA Damage and wound
repair.

• Excess apoptosis
– Neurodegenerative diseases

• Deficient apoptosis
– Cancer
– Autoimmunity
 Caspases

• Caspases stands for cysteine aspartate-specific protease.

• Caspases have the characteristics of high specificity for

substrates containing Asp, and use a Cys for catalyzing peptide

bond cleavage.

• Synthesized in the cell as precursors named procaspase.

• Caspases are the major executioners in apoptosis.

Caspase Structure
• NH2-terminal
domain
• Large subunit
(~20kD)
• Small subunit
(~10kD)
Caspase Activation
 Caspase Role in Apoptosis
• Cut off contact with surrounding cells
• Reorganize cytoskeleton
• Shut don DNA replication and repair
• Interrupt splicing
• Destroy DNA
• Disrupt nuclear structure
• Induce cell to display signals marking it for
phagocytosis
• Disintegrate cells into apoptotic bodies
 Two types:
Types
- those involved in apoptosis:
Initiators – activate downstream effector caspases to initate
activation cascades:
Caspases2
Caspases9
Caspases8
Caspases10
Effectors - cleave target proteins resulting in
morphological and biochemical markers of
apoptosis:
Caspases3
Caspases6
Caspases7
Caspases14
 Apoptosis: Pathways
“Extrinsic Pathway”

Death Death Initiator

Ligands Receptors Caspase 8

Effector
“Intrinsic Pathway”
Caspase 3 PCD

DNA Initiator
Mitochondria/
damage Cytochrome C Caspase 9
& p53
 Intrinsic Pathway
• Initiated from within the cell.
• Activated in response to signals such as DNA
damage, loss of cell survival factors ,cell stress.
• Hinges on balance brtween pro and
antiapoptotic signals of Bcl-2 family.
• Apaf-1,cytochrome-c,ATP(apoptosome) activate
procaspase-9 complex.
• pro apoptotic proteins released which activate
caspase proteases
 Extrinsic Pathway
• Begins outside the cells.

• Activation of death receptors (Fas-R,TNF-R ,DR-

3,DRY/DR5) by death ligands (Fas-L,TNF-
alpha,Apo3L,Apo2L)play major role.

• Death induced signalling compex (DISC) activated.

• On DISC activation same effctor pathway as intrinsic

pathway is adopted
Disorders where apoptosis is
inhibited (decreased
apoptosis,increase in cell survival)
1.Cancer
Colorectal cancer
Glioma
Liver
Lymphoid malignancies
Neuroblastoma
Prostate cancer
Follicular lymphomas
Carcinomas with p-53 mutations
2. Autoimmune disorders
Mysthenia gravis
Systemic lupus erythematous
3 Inflammatory disease

Bronchial asthma
Inflammattory bowel disease
Pumonary inflammation

4 Viral infections

Herpesviruses
Poxviruses
Adenoviruses
Baculovirus
Cowpox
Disorders where apoptosis is
excessive
increase in cell death (Means
hyperactive apoptosis).
1 AIDS
CD4+ cells
T- lymphocytes

2. Neurodegenerative disorders
Alzheimer’s disease
Epilepsy
Parkinson’s disease
Amyotrophic lateral sclerosis
Retinitis pigmentosa
Cerebellar degeneration
Role of Caspases in Alzheimer's Disease
Role of Caspases in HUNTINGTON DISEASE

3
 AIDS

Autoimmune deficiency syndrome (AIDS) is an

example of an autoimmune disease that results from
infection with the human immunodeficiency virus
(HIV) . This virus infects CD4+ T cells by binding to
the CD4 receptor. The virus is subsequently
internalized into the T cell where the HIV Tat protein is
thought to increase the expression of the Fas receptor,
resulting in excessive apoptosis of T cells .
So what kills so many uninfected CD4+ cells?
Answer is: Apoptosis.

There are several possibilities. One of them:

 All T cells, both infected and uninfected,
express Fas.
 Expression of a HIV gene (called Nef) in a HIV-
infected cell causes
 The cell to express high levels of FasL at its
surface
 While preventing an interaction with its own
Fas from causing it to self-destruct.
 However, when the infected T cell encounters
an uninfected one (e.g. in a lymph node), the
interaction of FasL with Fas on the uninfected cell
kills it by apoptosis.
 Recent advances
• Caspase inhibitors such as N-benzyloxycarbonyl-Val-Ala-
Asp fluoromethyl-ketone (Z-VAD) were being investigated
for the treatment of a number of neurodegenerative
disorders including:-
 Alzheimer disease
 amyotrophic lateral sclerosis (ALS)
 Huntington’s disease
 Parkinson’s disease (PD)
 and finally acute neurologic diseases including ischemia or
traumatic injury .
Unfortunately, clinical development of Z-VAD was
discontinued following the recognition that metabolism of Z-
VAD produces liver damage following the production of the
toxic compound, fluoroacetate .
Following the disappointment of Z-VAD, a number of
other caspase inhibitors have been developed with the
goal in mind of being safer and more selective-

• Quinolyl-valyl-Omethylaspartyl-[- 2, 6-
difluorophenoxy]-methyl ketone (QVD-OPh)-
-Q-VD-OPh appears to be able to cross the blood-
brain barrier, which is always a central issue when
developing a drug for treatment of a CNS disorder.
-including greater potency, selectivity,stability and
cell permeability
-After acute treatment of mice with Q-VD-OPh, all
organs were normal suggesting a lack of toxicity.
• Another promising compound that is currently
under evaluation is Minocycline, a second-
generation tetracycline. In comparison to Q-VD-
OPh, minocycline is much further along as an
investigational drug for the treatment of
neurodegenerative disorders and in some cases
human clinical trials have begun. For example,
- Phase II for PD
- Phase I/II for spinal cord injury
- Phase III trail for ALS
• Caspase-6:A Potential Therapeutic Target for
Neurodegenerative Disease:
Many studies, as recently reviewed byGraham et. al. in
Trends in Neurosciences, have revealed Caspase-6 as a
possible contributor to neurodegenerative pathology associated
with Huntington’s Disease (HD), Alzheimer’s Disease (AD),
aging, and stroke.

zVADfmk ,zVEIDfmk & zDHRDfmk (under research) are

peptide agents to inhibit CASP-6.

5. Inflammatory caspase inhibitors, such as VX-740

(Pralnacasan) and VX-765(caspase-1 inhibitor VX-765 is
undergoing phase 2 clinical trials by Vertex) in clinical studies
for rheumatoid arthritis and osteoarthritis.
• The present findings indicate that nonlipid LPA₂-
specific agonists represent an excellent starting
point for development of lead compounds with
potential therapeutic utility for preventing the
programmed cell death involved in many types of
degenerative and inflammatory diseases.
Lysophosphatidic acid (LPA) is a highly potent
endogenous lipid mediator that protects and rescues
cells from programmed cell death. Earlier work
identified the LPA₂ G protein-coupled receptor
subtype as an important molecular target of LPA
mediating anti apoptotic signaling
• GRI977143, which was effective in reducing
activation of caspases 3, 7, 8, and 9 .

• On the basis of the present results, it is concluded that

vitamin D displays anti-apoptotic effects in human
osteoblast-like osteosarcoma cells in vitro. This
observation suggests that besides regulating growth and
differentiation, vitamin D exerts its anabolic effects on
bone by protecting osteoblastic cells from undergoing
apoptosis.
Thank You
So Much