Understanding Cancer: Biology, Genetics, Types and Colon Cancer Cell Lines

This article provides an in-depth understanding of cancer, including its biology, genetics, types, and colon cancer cell lines. It discusses the differences between normal and cancer cells, genetics of cancer, types of cancer, and colon cancer. It also provides information on HCT116 and HT-29 colon cancer cell lines.

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Cancer
Name of the Student
Name of the University
Author note

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Cancer
Cancer results from a cascade of molecular pathways that alter the characteristics of
normal cells. While understanding the biology of cancer, it is caused when cells inside the body
accumulate due to genetic mutations that interfere with normal cell properties. Cancer cells
prevent the invasion and cell overgrowth where mutated cells divide and grow without any
specific signals (De Martel et al. 2012). As these cells divide, they gain new characteristics
including structural changes, new enzymes production and decrease in cell adhesion. This
heritable change makes the cells and its progenies to grow and divide inhibiting the growth of the
surrounding cells. As a result, they spread and start invading cells in proximity. Cancer cells are
malignant in nature that spread or invade to other parts of the body through blood or lymph
forming new tumours distant from the original site. Unlike these tumours, benign tumours do not
invade or spread to other nearby tissues and large (Meacham and Morrison 2013).
Difference between normal cells and cancer cells
Cancer cells differ from normal cells as they have uncontrollable growth rate and
invasive in nature. Normal cells are distinct and specialized to perform specific functions where
cancers cells are not specialized making them divide unstoppably. Cancer cells grow
continuously forming a cluster called tumour having growth factor proteins that make them grow
continuously. Another difference between cancer and normal cells is that the latter does not
responds to signals to top growing. Moreover, they do not repair or undergo apoptosis or die
when they get old or damaged. Cancer cells lack adhesion molecules that make them float and
travel to distant body arts via bloodstream or lymph gaining the ability to metastasize
(Vogelstein et al. 2013). When they reach new regions like lungs, lymph nodes, bones or liver,

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they start growing and form tumours at the secondary locations. In contrast to normal cells, there
is variability in cancer cell size as they are larger with abnormal shape, nucleus or cell
organelles. The nucleus appears darker and larger containing excess DNA having abnormal
chromosomes arranged in disorganized manner.
Genetics of cancer
Only small proportion of 35,000 human genes is associated with cancer and alterations in
these genes are associated with various cancer forms. This malfunctioning is broadly divided
into three groups: proto-oncogenes, oncogenes and tumour suppressor genes. Proto-oncogenes
in normal cells code for proteins that send signals for cell division to the nucleus initiating signal
transduction pathway. This cascade comprises of membrane receptors for signalling and
intermediary proteins that carry signal to cytoplasm and transcription factors activate cell
division genes in the nucleus. However, altered versions of proto-oncogenes give rise to
oncogenes activating signals stimulating uncontrolled growth. Proto-oncogenes like RAS, MYC,
WNT, TRK and ERK acquire activating mutation that form tumour-inducing molecule called
oncogene through three activation methods involving gain-of-function mutation (Hnisz et al.
2016). Point mutations occur in proto-oncogenes result in constitutively protein product. Gene
amplification or localized reduplication of DNA segment in proto-oncogene also leads to
overexpression of protein. Chromosomal translocation results in growth-regulatory gene that is
controlled by a different promoter resulting in inappropriate gene expression.
Tumour suppressor genes also play an important role in cell growth and division. They
act to inhibit cell proliferation and development of tumour. However, these genes lose control or
gets inactivated that remove negative regulators in cell proliferation and contribute to abnormal

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tumour cell proliferation (Yates and Campbell 2012). Mutations occur in DNA repair genes like
mismatch repair (MMR) genes, nucleotide excision repair (NER) group, DNA crosslink repair
that results in inherited cancer syndromes. DNA repair mechanism is the backbone of survival;
however, decreased efficiency or inactivation occurs by epigenetic gene activation mechanism
that affects DNA repair activity of the genes (Jeggo, Pearl and Carr 2016).
Types of cancer
Carcinoma cancers begin in tissue or skin that covers the surface of glands or internal
organs forming solid tumours. Examples comprises of breast cancer, prostate cancer, colorectal
and lung cancer. Sarcoma begins in the connective tissue that connects and supports the body
developing in nerves, muscles, fat, joints, tendons, cartilage, blood vessels or bone. Leukaemias
are blood cancers that begin when there is change in structure and uncontrollable growth occurs
in blood cells. The four main types of leukaemia are acute and chronic lymphocytic leukaemia,
acute and chronic myeloid leukaemia. Lymphomas are cancer that begins in lymphatic system
causing Hodgkin and non-Hodgkin’s lymphoma. Melanomas are cancers that begin in cells that
form melanocytes that are specialized cells synthesizing melanin, pigment that impart colour to
the skin causing skin cancer (Sandoval and Esteller 2012). Colon cancer is also a type of
adenocarcinoma that is discussed in the subsequent section.
Colon cancer
Colon cancer is the large intestine cancer that makes up the final part of the digestive
tract. Mostly, colon cancer begins as a benign or non-cancerous clump of cells known as
adenomatous polyps. These polyps are small and show few symptoms. They comprise of excess
abnormal and normal appearing cells in the glands that cover colon’s inner wall. These abnormal

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growths later enlarge and degenerate to become adenocarcinomas. This cancer generally occurs
before the age of 40 years and due to lifestyle factors. More than 75-95% colon cancers occurs in
individuals with no or little genetic risk and out of which 10% is linked to sedentary lifestyle and
addictive behaviour (Kandoth et al. 2013). It originates from epithelial cells that lines colon of
the gastrointestinal tract due to mutations in Wnt signalling that enhance signalling activity.
Colon cancer are acquired or inherited that occur in crypt stem cell of the intestine.
Adenomatous polyposis coli (APC) gene is also polyposis 2.5 deleyed protein that is encoded in
humans by APC gene. This protein is a negative regulator of beta-catenin concentrations and E-
cadherin that play an important role in cell adhesion. This protein prevents the beta-catenin
protein accumulation and as a result, it is accumulated to high levels that translocated to nucleus,
DNA binding and activates proto-oncogenes transcription (Burrell et al. 2013). Mostly, APC
gene is important for cell differentiation and renewal, but at inappropriately high levels, it causes
cancer. Apart from Wnt signalling, other mutations also occur that make cells cancerous like p53
gene that monitors cell division. Inherited gene mutations increase the risk of colon cancer that is
present in two most common forms. Hereditary nonpolyposis colorectal cancer (HNPCC) or
Lynch syndrome develop in individuals before the age of 50 years. Familial adenomatous
polyposis (FAP) is a disorder that greatly increases the risk of colon cancer and develops polyps
in the colon and rectum lining (Isik et al. 2014).
Diagnosis of colon cancer is performed through sampling of colon obtained during
colonoscopy that depends on lesion location. The cell line acquires mutation where it transforms
from benign to invasive epithelial cancer cells. To study the phenotype of colon cancer, it is
important to study cell lines that are considered efficient biomedical research tools that
emphasize on genotype characterization and authentication (Kreso et al. 2014). There are 24-

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colon cancer cell-lines that vary in growth characteristics and appearance. They are used to study
the biology of colon cancer and test treatments. The established cell lines maintain and represent
the genetic diversity of the primary colon cancers. DNA copy number and exome mutation
spectra in colon cancer cells closely resemble to hypermutation phenotypes defined by defective
DNA polymerase and DNA mismatch repair, proofreading deficiency and concordant mutations
altered in Wnt, p53 pathways (Cayrefourcq et al. 2015).
Colon cancer cell lines
Among all the cell lines in colon cancer, HCT116 and HT-29 is human colon cancer
cells that are used in drug screening and therapeutic research. HCT-116 has mutation in KRAS
proto-oncogene in codon 3. They are considered suitable transfecting agents or the gene therapy
and research. These cell lines possess epithelial morphology that can be used to study the
tumorigenicity. They have adherent properties having epithelial morphology having posttive
control for PCR mutation assays present in codon 13 (Wang et al. 2013). When these cell lines
are transducted with viral vectors that carry p53 gene, there is arresting of HCT116 in the G1
phase. Colony proliferation of HCT116 was inhibited by P85/5-Fu to study the proliferation of
colon cancer and corresponding inhibitors. This cell line has two important variations: Insp8
gene having large expression and one without it. This gene plays a role in the metabolism of
cell’s energy processing that can in turn also affects the cell phenotype (Sun et al. 2012).
The doubling time of HCT-116 is ~18 hours and is suitable for in vivo and in vitro
experimentation in life studies where it forms tumours and metastasize followed by implantation
of cells (Kim, Lee and Kim 2013). These cell lines are also beneficial in evaluating the effects of
therapies and drugs available with different reporters and facilitate multi-modality imaging. For

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high and constitutive expression of HCT-116 reporter proteins, generation of cell lines are
carried out by lentiviral vector transduction. These vectors are used for carrying out
transductions that are self-inactivating (SIN) vectors where there is deletion of viral promoter
and enhancer. This results in increase in the biosafety of lentiviral vectors through prevention of
mobilization of competent viruses’ replication. These cancer cell lines are genotypic in nature
that results in reduced responsiveness over time that ensures performance and stability of the
cancer cell lines (Sahlberg et al. 2014). This cell line does not express or differentiates CDX1 or
sub-populations of cells having great tumour-forming capacity. This suggests that HCT-116
contains cancer stem cells (CSCs) subpopulations that are characterized by cell surface markers
and morphology of colony having self-renewal ability and differentiation into multiple lineages.
HT-29 was initially derived from Jorden Fogh, 44-year-old Caucasian female in 1964 is
also epithelia in nature having adherent properties. They have a unique model that can be used to
study molecular mechanisms of differentiation of intestinal cells. Under appropriate culture
media conditions, these cell lines can be manipulated to express enterocyte differentiation. This
is the reason HT-29 is considered pluripotent cells that can be used to study molecular and
structural events that are involved in colon cancer cell differentiation. This cell line forms tight
monolayer that exhibit similarity to the enterocytes in the small intestine. These cell lines
increase the production of p53 tumour antigen having a mutation at the 273 position that results
in replacement of arginine to histidine (Bogaert and Prenen 2014).
HT29 cell line is used in preclinical research for the differentiation ability and thus
mimics real colon cells in vitro successful for research of epithelial cells. HT-29 terminates
differentiation into enterocytes replacing glucose by galactose in the cell culture medium.
Moreover, these cells have induced differentiation property that is galactose-mediated that causes

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adherens junction strengthening. These cell lines also proliferate in cell culture media that lack
growth factors with 4 days doubling time, although it can reduced by using fetal bovine serum
(FBS). They have high glucose consumption and remain undifferentiated in standard medium
(Mouradov et al. 2014). They grow in multi-layer comprising of unpolarised differentiated cells
that does not possess characteristics of normal intestinal epithelia cells and low amount of
hydrolases. Biochemical markers physiologically and morphologically characterize the polarised
phenotype of HT-29 colon cancer cells (Martínez-Maqueda, Miralles and Recio 2015). The
differentiation is influenced by growth related factors that starts after confluence forming a
monolayer and tight junctions. The brush border of HT-29 contains proteins that are present in
normal intestinal microvilli like villin. Moreover, these cells also express hydrolases that are as
small intestine; however, enzymatic activity is much lower than normal intestine with no lactase
expression.
HT-29 show relevance to in vivo as there is similar protein expression as human
intestinal cells. The significant differences in metabolic and transporters gene expression from
the human intestinal cells also affect suitability of HT-29 in showing in vivo permeability.
According to Cancer Genome Atlas Network (2012), the expression of 377 genes in this cell line
used as in vitro models in epithelium showed that HT-29 differentiation has similarity with
human colonic tissues and are not different. However, there are significant advantages and
disadvantages of using HT-29 cell line in vitro as summarised by (Wilding and Bodmer 2014).
The differentiated phenotype of this cell line is similar to enterocytes of small intestine in regards
to structure, cell differentiation process and brush-border hydrolases. The amount of villin
expressed in HT-29 differentiated cells shows similarity with freshly prepared colonocytes. On a
contrary, they are malignant cells that have high glucose consumption and glucose metabolism

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impairment. Due to brush-border hydorlases, they do not show any similarity with normal
enterocytes. Altogether, HT-29 colon cancer cell lines are considered valuable model for
studying the biology of colon cancer cells (Calon et al. 2012).
Multi-drug resistance in cancer
Cancers generally develop resistance to the chemotherapies and there is an increased
prevalence of drug resistant cancers. The ability of cancer cells to become resistant is
functionally and structurally unrelated to anti-cancer drugs called multi-drug resistance (MDR).
Chemotherapy is the treatment given to the patients diagnosed with locally, advanced and
metastasized cancer. This poses challenge in administering drug dosage that minimizes treatment
toxicity and maximizes efficacy. MDR is multifactorial and follow cellular pathways that are
involved in drug resistance in cancer. MDR in cancer is a mechanism where they develop
resistance to chemotherapy that results in minimization of cell death and drug-resistant tumours
expansion (Kathawala et al. 2015). This problem affects the treatment of cancers posing major
challenge to the researchers. This resistance occurs against anticancer drugs and as a result, there
is decreased drug uptake, activation of detoxification system, increase in drug efflux, and evasion
of apoptosis (drug-induced) and DNA repair mechanism activation.
MDR either acquired or inherent exist against anticancer drugs developed through
multiple mechanisms. Drug inactivations that occur in vivo undergo complex mechanisms
where drugs interact with different proteins that modify or partially degrade the other molecules
leading to activation. In such cases, anticancer drugs undergo metabolic activation that acquire
efficacy. Due to decrease in drug activation, cancer cells may develop resistance to
chemotherapies. Many anticancer treatments demand metabolic activation and therefore cancer

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cells may develop resistance through change in proteins that are related to apoptosis. For
example, p53 gene is mutated in half of cancers where deletion or mutation of this gene results in
non-functional form that results in MDR. On a contrary, p53 regulators inactivation like
apoptotic protease activating factor 1 (Apaf-1) and caspase-9 and cofactors can result in drug
resistance (Kibria, Hatakeyama and Harashima 2014).
Another mechanism is alteration of drug targets takes place due to modifications in
expression levels or mutations. This can be explained in a manner where efficacy of a drug is
influenced by molecular target and its alterations. One form of such mechanism is that in certain
cases, topoisomerase II is targeted by anticancer drugs that prevents under or super-coiling of
DNA. The complex that is formed between topoisomerase II and DNA is transient, however,
these drugs leads to DNA damage, DNA synthesis inhibition and halting of mitosis. Moreover,
cancer cell lines can also develop resistance to this enzyme through mutations that takes place in
topoisomerase II gene. Another mechanism that can cause MDR in cancer is through signalling
kinases like epidermal growth factor receptor (EGFR) family and downstreaming of signalling
proteins like Raf, Ras, Src and MEK (Wu et al. 2014). These kinases are active in some type of
cancers promoting uncontrolled cell growth. In such circumstances, over-activation of these
kinases is caused by mutations that may also occur due to over-expression of genes. Apart from
alterations that occur in drug targets, the resistance occur through alteration in process of signal
transduction that mediates activation of drugs.
The mechanism that is most commonly studied in MDR in cancer is drug efflux that
involves reduction in drug accumulation through efflux enhancement. ATP-binding cassette
(ABC) transporter family proteins are transmembrane proteins that are classified into two
domains having highly conserved binding domain for nucleotides and variable transmembrane

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domain. When a substrate binds to the transmembrane domain, there is hydrolysis of ATP
occurring at nucleotide binding site that acts as a driving factor for conformation change pushing
the substrate out of the cell. ABC transporters induce efflux considered as normal process;
however, it is also a mechanism for MDR in cancer cells. There are three transporters called
multidrug resistance protein 1 (MDR1), multidrug resistance-associated protein 1 (MRP1) and
breast cancer resistance protein (BCRP) that are responsible for MDR in cancer (Zahreddine and
Borden 2013). These transporters have broad specificity for substrates and as a result, efflux the
xenobiotics from the cells. Therefore, the cancer cells are protected from first line
chemotherapies inducing MDR.
DNA damage repair also performs a role in developing MDR. Chemotherapies indirectly
or directly damage DNA and its response mechanisms that can reverse the damage that is
induced by cancer. For example, few anti-cancer drugs cause DNA crosslink that result in
apoptosis. On a contrary, resistance to these drugs arises due to homologous recombination and
nucleotide excision repair that reverses the effect of this anticancer drug property. Moreover,
impairment or dysregulation of DNA damage response (DDR) due to epigenetic silencing or
mutations can result in increased resistance and failure to chemotherapy (Gillet and Gottesman
2010).
Cell death inhibition is another mechanism where recombinant forms of TNFs related
apoptosis-inducing ligand (TRAIL) induce apoptosis through caspase-8 activation. However,
results showed that prolonged use of these drugs causes resistance and they need to be used in
combination with other cytotoxic drug so that it kills the cancer cells in vulnerable states
(Zahreddine and Borden 2013).

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Epithelial-Mesenchymal Transition and Metastasis (EMT) is a process where solid
tumors metastatic causing angiogenesis. EMT plays an important role in drug resistance
development depending on the metastatic tumor grade defined by EMT degree and level of
differentiation. MDR that occur in cancer cells during differentiation and its signaling process
are essential for the mechanism of EMT. For example, in colon cancer, there is increased integrin
αvβ1 expression that regulates positive transforming growth factor β (TGFβ) expression, EMT
expression and acts as survival signal for the cancer cells against anticancer drugs (Housman et
al. 2014).
Cancer cell heterogeneity is a mechanism involved in MDR where fraction of cells
presents in heterogenous populations possess stem cell properties and drug resistant. Moreover,
small fraction of adult stem cells (ASC) also has MDR capabilities. Cancer treatment kills drug
sensitive cells and result in drug resistance where cancer cells survive, grow, expand and
metastasize. Drugs that may go into circulation and form secondary tumours in the distant organs
in the body kill some of these resistant cells. As a result, heterogeneity is witnessed in cancer in
both solid tumours and circulation (Zahreddine and Borden 2013).
Drug repurposing
Drug repurposing is commonly referred to as therapeutic switching or re-tasking
encompasses the application of drugs and compounds that are known, with the aim of treating
particular diseases. It has been identified as a promising pharmaceutical strategy that effectively
reduces the resources required for development of therapies for certain diseases. The process also
amplified the probability of the drug entering the market from phase I, for new indications
(Strittmatter 2014). This approach of drug repositioning is found to primarily capitalize on and

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utilise the fact that a range of abandoned, approved drugs, and other compounds have been
already tested in humans. In addition, the detailed information widely available on the
formulation, pharmacology, potential toxicity and dose of the drugs help in redevelopment of the
drug for use in a different disease or illness. The process is underpinned by the fact that a
plethora of molecular pathways has been recognised to play an important role in the onset of
several diseases.
The major advantages of repositioning over conventional drug discovery procedures are
associated with the significant reduction in the cost and time of development, owing to the fact
that safety demonstration of most of these drugs in humans often negates the requirement for
conducting phase I clinical trials (Andrews, Fisher and Skinner-Adams 2014). In addition, there
exist huge numbers of potential drugs that never reach stages of clinical testing. Moreover, less
than 15% of the pharmaceutical compounds that enter the clinical development are able to obtain
an approval, majority of them being considered safe. Formulating new drug indications are
considered beneficial for patients, both for failed and approved drugs, the safety of which has
already been established (Xu and Wang 2013). Drug repurposing approaches span a plethora of
disease areas, namely sildenafil, the phosphodiesterase inhibitor, cures coronary artery disease
treat cyclooxygenase inhibitor aspirin, erectile dysfunction and gastric motility is cured by
erythromycin. In addition, drugs that are said to create adverse effects also merit reconsideration,
as is evidenced by an effective reprofiling of thalidomide, the antiemetic for treating multiple
myeloma.
While there are, several benefits of drug repositioning, success till date are mostly
unanticipated. Large scale, systemic drug re-profiling have not be quite possible, owing to the
lack of drug collection by physical means, low drug annotation quality and lack of sufficient

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readouts related to drug activity, from which a range of drug indications can be easily predicted.
Recent studies have provided evidence for gene expression profiling techniques that have
enabled drug repurposing discoveries, which includes sirolimus, used for acute lymphocytic
leukemia that is glucocorticoid-resistant, topiramate for treating inflammatory bowel disease, and
imipramine for the treatment of small cell lung cancer (Bertolini, Sukhatme and Bouche 2015).
Quite recently, an assay has also been developed with the use of barcoded cell lines, commonly
referred to as PRISM, for cancer therapeutics that enables the quick testing of several drugs
against huge cancer cell lines. Approximately 10 drug candidates are required to enter human
investigation, with the purpose of producing a fresh molecular entity product. This process
generally follows screening of thousands of members of the molecular library that are
structurally optimised and tested to determine the animal toxicology effects (Stenvang et al.
2013).
Thus, risks of R&D are greatly reduced upon starting with products that have already
been passed through the stages of development. In other words, upon comparison of drug
repositioning with use of conventional approaches, the former has been found to have a high
success rate (25% versus 10%). Thus, one substantial advantage of the procedure is related to its
ability of obtaining ‘method-of-use’ patents designed in a way that promotes the discovery of
secondary benefits of the compounds.
Although cancer is, a major health issue on a global basis, limited success obtained by
implementation of current therapies have resulted in greater investments for drug development.
A decline has also been observed in the average FDA approvals for anti-cancer drugs. The
failure to meet the needs of appropriate cancer treatment drugs have increased the interest in
drug reposition where already approved drugs, commonly used for other indications, will be used

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to treat cancer (Gupta et al. 2013). In the context of treating cancer, terminal or rare oncological
manifestations are found to afford less restrictions on the safety of the drugs and compounds,
owing to the ever increasing demand of novel treatment therapies. In addition, cancer is also
manifested in the form of a multistage disease, with interventions targeting different phases of
carcinoma initiation, rapid growth, metastasis and recurrence.
These features provide the indication that drug repurposing that is focused on cancer
treatment would have mutual benefits for both the pharmaceutical companies and the patients
alike. Studying the ability of a drug to bring about changes in the expression profile of cancer
cells have also allowed drawing inferences about their mechanism-of-action (Xu et al. 2014).
The approach has eventually resulted in the discovery of major antitumor properties of
trifluoperazine, which was previously approved in the form of an antidepressant for treating
schizophrenia (Koch et al. 2014).
Some of the conventional use of the repurposed drug anisomycin is associated with
inhibition of protein and DNA synthesis. It has also gained potential as a psychiatric drug and
has been identified beneficial for selective removal of memories, brought about by injection of
the drug into the hippocampus. Anisomycin also seems useful during transition of normal to
fibrotic lungs, both during stages 1 and 2 of idiopathic pulmonary fibrosis, due to its role in
inhibiting ER stress induction (Karatzas et al. 2017). Owing to the fact that repurposing is a
major alternative to conventional use of drugs, anisomycin has also been subjected to re-profiling
in several studies (Monaghan et al. 2014).
The antibiotic anisomycin has been recognised as one of the most effective compounds,
during secondary screening of compounds. Anisomycin has been found highly effective against

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specific subsets of TNBC cell lines, in addition to some in vitro cell lines of prostate cancer. The
drug has also shown its benefits in inducing ribotoxic stress in MDA-MB-468 TNBC cell line,
evidenced by JNK activation (Cruickshank 2016). This activation generally occurs by inducing
caspase-3 processing and caspase-3 like activity. This antibiotic anisomycin has been found to
induce death of the tumor cells, along with display of metastatic activity, when coupled with
apoptosis (Boyer et al. 2018). Thus, even low doses of anisomycin have the ability of inhibiting
protein synthesis in cases of melanoma.
Cell death
Cell death refers to the event that encompasses death of a biological cell that
subsequently ceases to conduct all its functions (McIlwain et al. 2013). Cell death might occur
due to natural process of death of old and damaged cells that are replaced by new ones, or might
also result due to localised injuries, diseases or death of the organism.
Apoptosis
This refers to the form of programmed cell death occurring in multicellular organisms
where a set of biochemical events bring about changes in the cell morphology and its subsequent
death. Some of the most characteristic changes that occur in apoptosis include blebbing, nuclear
fragmentation, cell shrinkage, chromosomal DNA fragmentation and chromatin condensation.
Upon comparison with traumatic cell death or necrosis, apoptosis has been identified to be
highly controlled and regulated that confers several advantages during the lifecycle of an
organism. The intrinsic pathway of apoptosis involves an event where the cell kills itself due to
sense of stress. On the other hand, the extrinsic pathway is followed when the cells kills itself
due to reception of signals from adjacent cells. Apoptotic pathways are induced by activation of

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caspases, a class of protease that are responsible for degradation of proteins (Mukhopadhyay et
al. 2014). Excess apoptosis most often leads to atrophy or reabsorption and breakdown of the
cells and tissues.
However, less apoptosis has been found responsible for uncontrolled cell proliferation,
commonly referred to as cancer. Disruption in the apoptotic pathway results in increase in the
progeny of cells that have faulty machinery, which in turn increases the likelihood of the cells
becoming cancerous. Damage of DNA due to biochemical factors leads to the accumulation of
tumor suppressor protein p53 (Motawi et al. 2014). Induced transcription of p53 results in its
increase and cancer cell-apoptosis enhancement. Disruption in p53 regulation is responsible for
apoptosis in impairment and possible tumor formation.
Figure 1: Induction of autophagy and apoptosis
Source- (Mukhopadhyay et al. 2014)

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Necroptosis
This is a programmed pattern of inflammatory cell death or necrosis and is associated
with cell death that occurs due to damage of the cells or pathogen infiltration. This is generally
defined as a form of defence mechanism, initiated by viruses. This facilitates undergoing of
cellular suicide in a manner that is independent of the caspase enzymes. Necroptosis has also
been identified as an integral component of cvarious inflammatory diseases such as, pancreatitis,
myocardial infarction and Chron’s disease (Kaczmarek, Vandenabeele and Krysko 2013). TNFα,
the cell signalling protein is produced during viral infection that results in stimulation of the
TNFR1 receptor. RIPK1 is signalled by TRADD, the TNFR-associated death protein, which
sends signal to the RIPK3, to form the necrosome. Necroptosis is generally characterised by
necrotic cell death morphology that is controlled by RIP3, RIP1, and kinase domain-like proteins
of mixed lineage.
In addition, the physiological role of necroptosis is associated with serving as a "fail-
safe" cell death form for specific cells failing to undergo apoptosis, at the time of disease defence
and embryonic development. Cells that undergo necroptosis often rupture and leak the contents
in intercellular space (Pasparakis and Vandenabeele 2015). Necroptosis has also been associated
with initiation of cancer and its progression. Elevation in necroptosis often increases the risks of
metastasis and proliferation of the cells by generating reactive oxygen species and suppressing
immune response. Recent studies have established necroptosis based cell therapies as essential
antitumor treatment strategies.

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Figure 2: Pathogen-Mediated Necroptosis
Source: (Kaczmarek, Vandenabeele and Krysko 2013)
Pyroptosis
This refers to an inflamed pattern of programmed cell death that is found to occur more
upon acquiring infections caused by intracellular pathogens. Pyroptosis is most likely to become
a part of antimicrobial response in the body. The process involves recognition of danger signals
that trigger the release of cytokines, followed by swelling, bursting and death of the cells. These
cytokines are found to play a crucial role in attracting other immune cells that help in fighting
infections and contribute to tissue inflammation. Pyroptosis also promotes clearance of different
viral and bacterial infections that is facilitated by the removal of intracellular replication niches,

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thereby enhancing the defensive responses of the host (Doitsh et al. 2014). However, in case of
pathogenic chronic diseases, these inflammatory response fail to eliminate or eradicate primary
stimuli, as is normally found in most cases associated with injury or infection. This ensues a
chronic and severe form of inflammation that is found to potentially result in tissue damage.
Pyroptosis initiation in infected macrophage is brought about by flagellin component
recognition of Shigella and Salmonella species with the help of NOD-like receptors that
recognise antigen present within cells. Caspase-1 function is also crucial in the process of
pyroptosis (Case et al. 2013). The enzyme gets activated in the process with the help of major
supramolecular complexes called pyroptosome, composed of ASC protein dimers. In other
words, the process is a pathway that is linked to cell death, mediated by caspase-1 activation.
Failure of the cells to respond to particular stimulus and die subsequently results in organ
dysfunction and abortive embryogenesis, all of which contribute to cancer initiation.

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Figure 3: Caspase-induced pyroptosis
Source: (Shi, Gao and Shao 2017)
Ferroptosis
This kind of programmed cell death is primarily characterised by lipid peroxidase
accumulation and is dependent on iron. The process is biochemically and genetically distinct
from other cell death forms. Failure of the antioxidant defences that are glutathione dependent
initiate the process of ferroptosis and remove the check on lipid peroxidation, thereby resulting
in cell death. Iron chelators and lipophilic antioxidants (Jiang et al. 2015) can generally prevent
Ferroptotic cell death. The process is also initiated by direct loss of GPX4 activity that is
mediated by inhibition of Xc- system. This kind of cell death is found to occur during events that

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involve uptake of electrons by the free radical molecules from some lipid molecule. This
facilitates the degradation or degeneration of the lipid molecule. Oxidation of the lipid molecules
result in loss of electrons to free radical molecules, thereby acting as a trigger for the
degradation.
Activation of ferroptosis has been identified to play an essential role that regulates the
growth of tumor and carcinoma cells in the human body. Thus, ferroptosis can significantly
contribute to the field of research and medicine in devising cancer treatments that involve
induction of this kind of cell death in the human body (Angeli et al. 2014). Ferroptosis induced
by small molecules such as, erastin, has been found to act as strong inhibitors of tumor growth
and subsequently enhance sensitivity of certain chemotherapeutic drugs namely cisplatin,
temozolomide and adriamycin, mostly in conditions that are associated with drug resistance.

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Angeli, J.P.F., Schneider, M., Proneth, B., Tyurina, Y.Y., Tyurin, V.A., Hammond, V.J.,
Herbach, N., Aichler, M., Walch, A., Eggenhofer, E. and Basavarajappa, D., 2014. Inactivation
of the ferroptosis regulator Gpx4 triggers acute renal failure in mice. Nature cell biology, 16(12),
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