- Carcinogenesis is the process by which normal cells are transformed into cancer cells through genetic and epigenetic changes. It involves multiple steps including initiation, promotion and progression. - Key events include transformation of cells, uncontrolled proliferation, invasion of surrounding tissues, and metastasis. - DNA damage and defects in DNA repair can lead to mutations and epigenetic alterations that accumulate and drive carcinogenesis. - Oncogenes and mutations in proto-oncogenes disrupt normal cell growth control and promote uncontrolled cell division.

1. CARCINOGENESIS AND
CARCINOGENIC FACTORS
DR. ASHUTOSH SINGH
ASSISTANT PROFESSOR

2. Natural history of carcinomas
Can be divided in several steps :
- Transformation of a normal
cell into a tumour cell
- Clonal proliferation of this
tumour cell
- Growth of the tumour until
clinically becoming detectable and
local invasion
- Spread of the tumour away
from the primary site : metastasis

5. • The natural history of carcinoma is a lenghty and sequential process
which can be divided in several steps.
- Transformation of a normal cell into a tumor cell after occurrence
of genetic abnormalities
- Clonal proliferation of this tumor cell during the pre-invasion step
- Growth of the tumor until becoming detectable and local invasion.
Invasion refers to the direct migration of tumor cells into the
surrounding tissue through the basal membrane
- The spread of the tumour away from the primary site to distant site
via blood vessels or lymphatic vessels : the metastatic step (lymph
node, lung, bone, liver….)
• Metastasis refers to the ability of cancer cells to penetrate into
lymphatic and blood vessels, circulate through the bloodstream and
the invade normal tissue elsewhere in the body

6. DYSPLASIA
• Correspond to the steps before invasion of
surrounding tissue. No disruption of basement
membrane
• Corresponds to the intra-epithelial step of
carcinogenesis
• The first microscopically detectable
anatomic change in the neoplastic process

7. • Dysplasia : Histologicaly unequivocal neoplastic
epithelium without evidence of tissue invasion
> Microscopic term
> Used only in epithelia (digestive tract,
breast, lung, uterine cervix, urinary tract,
pancreas)
> Corresponds to an excessive and
uncontroled proliferation of cells
> Results from genetic abnormalities that
alter cell proliferation and differentiation.
> Occasionally they gradually become
malignant (invading surrounding tissue) by
aquisition of other genetic abnormalities

8. PRENEOPLAS
IA
NEOPLASIA

9. Dysplasia can be observed in
• Inflammatory conditions
- Chronic gastritis caused by Helicobacter pylori
- Inflammatory bowel diseases (Crohn’s disease,
ulcerative colitis)
• Virus infections
- Human papilloma virus (HPV) infection in
uterine cervix
• Some benign tumours
- Colonic adenoma

10. Microscopic criteria of dysplasia
Architectural Cytological
Increased number of cells Increased number of
mitosis (dividing cells)
Loss of cell differentiation Increased
nuclear/cytoplasmic ratio
Loss of normal epithelium
organization
Anisocytosis (cell of
irregular size)
Loss of cell polarity Anisokaryosis (nuclei of
irregular size)
Maturation
Dysplastic squamous epithelium
Normal stratified squamous
epithelium

11. DYSPLASIA – GRADING SEVERITY
• Refers to the intensity/ severity of architectural and cytological
abnormalities .
• Two or 3 grades depending on the organ :
> low/high (colon)
> cervical intra-epithelial neoplasia (CIN) I, II, III (cervix)
• The more high grade dysplasia, the more high risk of evolution to
cancer : Grade is a prognostic factor
• High grade dysplasia is synonymous with in situ carcinoma in
most organs

12. • Carcinogenesis, also called oncogenesis or tumorigenesis, is the
formation of a cancer, whereby
normal cells are transformed into cancer cells
• The process is characterized by changes at the cellular, genetic,
and epigenetic levels and abnormal cell division
• Cell division is a physiological process that occurs in almost
all tissues and under a variety of circumstances
• Normally, the balance between proliferation and programmed cell
death, in the form of apoptosis, is maintained to ensure the integrity
of tissues and organs
• According to the prevailing accepted theory of carcinogenesis, the
somatic mutation theory, mutations in DNA and epimutations that
lead to cancer disrupt these orderly processes by interfering with the
programming regulating the processes, upsetting the normal balance
between proliferation and cell death

13. • This results in uncontrolled cell division and
the evolution of those cells by natural
selection in the body
• Only certain mutations lead to cancer whereas
the majority of mutations do not

14. CARCINOGENIC FACTORS
• Genetic and epigenetic
• There is a diverse classification scheme for the various genomic
changes that may contribute to the generation of cancer cells
• Many of these changes are mutations, or changes in
the nucleotide sequence of genomic DNA
• There are also many epigenetic changes that alter whether genes are
expressed or not expressed
• Aneuploidy, the presence of an abnormal number of chromosomes,
is one genomic change that is not a mutation, and may involve either
gain or loss of one or more chromosomes through errors in mitosis
• Large-scale mutations involve either the deletion or duplication of a
portion of a chromosome
• Genomic amplification occurs when a cell gains many copies (often
20 or more) of a small chromosomal region, usually containing one
or more oncogenes and adjacent genetic material

15. • Translocation occurs when two separate chromosomal regions
become abnormally fused, often at a characteristic location
• A well-known example of this is the Philadelphia chromosome, or
translocation of chromosomes 9 and 22, which occurs in chronic
myelogenous leukemia, and results in production of the BCR-
abl fusion protein, an oncogenic tyrosine kinase
• Small-scale mutations include point mutations, deletions,
and insertions, which may occur in the promoter of a gene and affect
its expression, or may occur in the gene's coding sequence and alter
the function or stability of its protein product
• Disruption of a single gene may also result from integration of
genomic material from a DNA virus or retrovirus, and such an event
may also result in the expression of viral oncogenes in the affected
cell and its descendants.

16. DNA damage
• DNA damage is considered to be the primary
cause of cancer
• More than 60,000 new naturally-occurring
instances of DNA damage arise, on average,
per human cell, per day, due to endogenous
cellular processes

17. • Additional DNA damage can arise from exposure
to exogenous agents
• As one example of an exogenous carcinogenic agent,
tobacco smoke causes increased DNA damage, and this
DNA damage likely cause the increase of lung cancer due to
smoking
• In other examples, UV light from solar radiation causes
DNA damage that is important in melanoma, Helicobacter
pylori infection produces high levels of reactive oxygen
species that damage DNA and contribute to gastric
cancer, and the Aspergillus flavus metabolite aflatoxin is a
DNA damaging agent that is causative in liver cancer

18. • DNA damage can also be caused by substances
produced in the body. Macrophages and
neutrophils in an inflamed colonic epithelium
are the source of reactive oxygen species
causing the DNA damage that initiates
colonic tumorigenesis,and bile acids, at high
levels in the colons of humans eating a high-fat
diet, also cause DNA damage and contribute to
colon cancer

19. • A deficiency in DNA repair would cause more DNA
damage to accumulate, and increase the risk for cancer
• Such germline mutations (which cause
highly penetrant cancer syndromes) are the cause of
only about one percent of cancers
• The majority of cancers are called non-hereditary or
"sporadic cancers“
• About 30% of sporadic cancers do have some
hereditary component that is currently undefined, while
the majority, or 70% of sporadic cancers, have no
hereditary component

20. • In sporadic cancers, a deficiency in DNA
repair is occasionally due to a mutation in a
DNA repair gene; much more frequently,
reduced or absent expression of DNA repair
genes is due to epigenetic alterations that
reduce or silence gene expression

21. • When expression of DNA repair genes is reduced, this causes a
DNA repair deficiency
• With a DNA repair deficiency, DNA damage persists in cells at a
higher than typical level ( this excess damage causes an increased
frequency of mutation and/or epimutation
• Experimentally, mutation rates increase substantially in cells
defective in DNA mismatch repair or in Homologous
recombinational repair (HRR)
• Chromosomal rearrangements and aneuploidy also increase in HRR-
defective cells
• During repair of DNA double-strand breaks, or repair of other DNA
damage, incompletely-cleared repair sites can cause epigenetic gene
silencing

22. • The somatic mutations and epigenetic alterations
caused by DNA damage and deficiencies in DNA
repair accumulate in field defects
• Field defects are normal-appearing tissues with
multiple alterations and are common precursors to
development of the disordered and over-
proliferating clone of tissue in a cancer
• Such field defects may have numerous mutations
and epigenetic alterations

23. • It is impossible to determine the initial cause for most specific cancers. In a few
cases, only one cause exists: for example, the virus HHV-8 causes all Kaposi's
sarcomas
• However, with the help of cancer epidemiology techniques and information, it is
possible to produce an estimate of a likely cause in many more situations
• For example, lung cancer has several causes, including tobacco use and radon gas
Men who currently smoke tobacco develop lung cancer at a rate 14 times that of
men who have never smoked tobacco: the chance of lung cancer in a current
smoker being caused by smoking is about 93%; there is a 7% chance that the
smoker's lung cancer was caused by radon gas or some other, non-tobacco cause
• These statistical correlations have made it possible for researchers to infer that
certain substances or behaviors are carcinogenic
• Tobacco smoke causes increased exogenous DNA damage, and this DNA damage
is the likely cause of lung cancer due to smoking
• Among the more than 5,000 compounds in tobacco smoke, the genotoxic DNA-
damaging agents that occur both at the highest concentrations, and which have the
strongest mutagenic effects are acrolein, formaldehyde, acrylonitrile, 1,3-
butadiene, acetaldehyde, ethylene oxide and isoprene

24. • Using molecular biological techniques, it is possible to characterize
the mutations, epimutations or chromosomal aberrations within a
tumor, and rapid progress is being made in the field of predicting
certain cancer patients' prognosis based on the spectrum of
mutations
• For example, up to half of all tumors have a defective p53 gene
• This mutation is associated with poor prognosis, since those tumor
cells are less likely to go into apoptosis or programmed cell
death when damaged by therapy
• Telomerase mutations remove additional barriers, extending the
number of times a cell can divide
• Other mutations enable the tumor to grow new blood vessels to
provide more nutrients, or to metastasize, spreading to other parts of
the body

25. Genome instability
• Cancers are known to exhibit genome instability or a
"mutator phenotype“
• The protein-coding DNA within the nucleus is about 1.5%
of the total genomic DNA
• Within this protein-coding DNA (called the exome), an
average cancer of the breast or colon can have about 60 to
70 protein altering mutations, of which about 3 or 4 may be
"driver" mutations, and the remaining ones may be
"passenger" mutations
• However, the average number of DNA sequence mutations
in the entire genome (including non-protein-coding regions)
within a breast cancer tissue sample is about 20,000

26. Non-mainstream theories
• Cancer has also been considered as a metabolic disease, in
which the cellular metabolism of oxygen is diverted from
the pathway that generates energy (oxidative
phosphorylation) to the pathway that generates reactive
oxygen species
• This causes an energy switch from oxidative
phosphorylation to aerobic glycolysis (Warburg's
hypothesis), and the accumulation of reactive oxygen
species leading to oxidative stress ("oxidative stress theory
of cancer")
• Another concept of cancer development is based on
exposure to weak magnetic and electromagnetic fields and
their effects on oxidative stress, known as
magnetocarcinogenesis

27. Clonal evolution
• Just as a population of animals undergoes evolution, an unchecked
population of cells also can undergo "evolution". This undesirable process
is called somatic evolution, and is how cancer arises and becomes more
malignant over time
• Most changes in cellular metabolism that allow cells to grow in a disorderly
fashion lead to cell death
• However, once cancer begins, cancer cells undergo a process of natural
selection: the few cells with new genetic changes that enhance their
survival or reproduction multiply faster, and soon come to dominate the
growing tumor as cells with less favorable genetic change are out-competed
• This is the same mechanism by which pathogenic species such
as MRSA can become antibiotic-resistant and by which HIV can
become drug-resistant), and by which plant diseases and insects can
become pesticide-resistant
• This evolution explains why a cancer relapse often involves cells that have
acquired cancer-drug resistance or resistance to radiotherapy)

28. Biological properties of cancer cells
• Acquisition of self-sufficiency in growth signals, leading to
unchecked growth
• Loss of sensitivity to anti-growth signals, also leading to unchecked
growth
• Loss of capacity for apoptosis, allowing growth despite genetic
errors and external anti-growth signals
• Loss of capacity for senescence, leading to limitless replicative
potential (immortality)
• Acquisition of sustained angiogenesis, allowing the tumor to grow
beyond the limitations of passive nutrient diffusion
• Acquisition of ability to invade neighbouring tissues, the defining
property of invasive carcinoma
• Acquisition of ability to seed metastases at distant sites, a late-
appearing property of some malignant tumors (carcinomas or others)

29. Oncogenes
• Oncogenes promote cell growth through a variety of ways. Many
can produce hormones, a "chemical messenger" between cells that
encourage mitosis, the effect of which depends on the signal
transduction of the receiving tissue or cells
• In other words, when a hormone receptor on a recipient cell is
stimulated, the signal is conducted from the surface of the cell to
the cell nucleus to affect some change in gene transcription
regulation at the nuclear level
• Some oncogenes are part of the signal transduction system itself, or
the signal receptors in cells and tissues themselves, thus controlling
the sensitivity to such hormones
• Oncogenes often produce mitogens, or are involved
in transcription of DNA in protein synthesis, which creates
the proteins and enzymes responsible for producing the products
and biochemicals cells use and interact with

30. • Mutations in proto-oncogenes, which are the normally quiescent
counterparts of oncogenes, can modify their expression and
function, increasing the amount or activity of the product protein
• When this happens, the proto-oncogenes become oncogenes, and
this transition upsets the normal balance of cell cycle regulation in
the cell, making uncontrolled growth possible
• The chance of cancer cannot be reduced by removing proto-
oncogenes from the genome, even if this were possible, as they are
critical for growth, repair and homeostasis of the organism
• It is only when they become mutated that the signals for growth
become excessive

31. • One of the first oncogenes to be defined in cancer
research is the ras oncogene
• Mutations in the Ras family of proto-
oncogenes (comprising H-Ras, N-Ras and K-Ras) are
very common, being found in 20% to 30% of all human
tumours
• Ras was originally identified in the Harvey sarcoma
virus genome, and researchers were surprised that not
only is this gene present in the human genome but also,
when ligated to a stimulating control element, it could
induce cancers in cell line cultures

32. Proto-oncogenes
• Proto-oncogenes promote cell growth in a variety of ways.
Many can produce hormones, "chemical messengers"
between cells that encourage mitosis, the effect of which
depends on the signal transduction of the receiving tissue or
cells
• Some are responsible for the signal transduction system
and signal receptors in cells and tissues themselves, thus
controlling the sensitivity to such hormones
• They often produce mitogens, or are involved
in transcription of DNA in protein synthesis, which create
the proteins and enzymes responsible for producing the
products and biochemicals cells use and interact with

33. • Mutations in proto-oncogenes can modify their expression and function, increasing
the amount or activity of the product protein
• When this happens, they become oncogenes, and, thus, cells have a higher chance
of dividing excessively and uncontrollably
• The chance of cancer cannot be reduced by removing proto-oncogenes from
the genome, as they are critical for growth, repair and homeostasis of the body
• It is only when they become mutated that the signals for growth become excessive
• It is important to note that a gene possessing a growth-promoting role may increase
the carcinogenic potential of a cell, under the condition that all necessary cellular
mechanisms that permit growth are activated
• This condition also includes the inactivation of specific tumor suppressor genes
• If the condition is not fulfilled, the cell may cease to grow and can proceed to die
• This makes identification of the stage and type of cancer cell that grows under the
control of a given oncogene crucial for the development of treatment strategies

34. Tumor suppressor genes
• Tumor suppressor genes code for anti-proliferation signals and
proteins that suppress mitosis and cell growth
• Generally, tumor suppressors are transcription factors that are
activated by cellular stress or DNA damage
• Often DNA damage will cause the presence of free-floating genetic
material as well as other signs, and will trigger enzymes and
pathways that lead to the activation of tumor suppressor genes
• The functions of such genes is to arrest the progression of the cell
cycle in order to carry out DNA repair, preventing mutations from
being passed on to daughter cells
• The p53 protein, one of the most important studied tumor suppressor
genes, is a transcription factor activated by many cellular stressors
including hypoxia and ultraviolet radiation damage

35. • p53 clearly has two functions: one a nuclear
role as a transcription factor, and the other a
cytoplasmic role in regulating the cell cycle,
cell division, and apoptosis
• The Warburg hypothesis is the preferential use
of glycolysis for energy to sustain cancer
growth. p53 has been shown to regulate the
shift from the respiratory to the glycolytic
pathway

36. • However, a mutation can damage the tumor
suppressor gene itself, or the signal pathway
that activates it, "switching it off“
• The invariable consequence of this is that
DNA repair is hindered or inhibited: DNA
damage accumulates without repair, inevitably
leading to cancer

37. • Mutations of tumor suppressor genes that occur in germline cells are passed along
to offspring, and increase the likelihood for cancer diagnoses in subsequent
generations
• Members of these families have increased incidence and decreased latency of
multiple tumors
• The tumor types are typical for each type of tumor suppressor gene mutation, with
some mutations causing particular cancers, and other mutations causing others
• The mode of inheritance of mutant tumor suppressors is that an affected member
inherits a defective copy from one parent, and a normal copy from the other
• For instance, individuals who inherit one mutant p53 allele (and are
therefore heterozygous for mutated p53) can develop melanomas and pancreatic
cancer, known as Li-Fraumeni syndrome
• Other inherited tumor suppressor gene syndromes include Rb mutations, linked
to retinoblastoma, and APC gene mutations, linked to adenopolyposis colon cancer
• Adenopolyposis colon cancer is associated with thousands of polyps in colon while
young, leading to colon cancer at a relatively early age. Finally, inherited mutations
in BRCA1 and BRCA2 lead to early onset of breast cancer

38. Knudson two-hit hypothesis
• An inherited, germ-line mutation in a tumor
suppressor gene would cause cancer only if
another mutation event occurred later in the
organism's life, inactivating the other allele of
that tumor suppressor gene

39. • Usually, oncogenes are dominant, as they contain gain-of-
function mutations, while mutated tumor suppressors
are recessive, as they contain loss-of-function mutations
Each cell has two copies of the same gene, one from each
parent, and under most cases gain of function mutations in
just one copy of a particular proto-oncogene is enough to
make that gene a true oncogene
• On the other hand, loss of function mutations need to
happen in both copies of a tumor suppressor gene to render
that gene completely non-functional
• However, cases exist in which one mutated copy of a tumor
suppressor gene can render the other, wild-type copy non-
functional. This phenomenon is called the dominant
negative effect and is observed in many p53 mutations

40. Multiple mutations
• In general, mutations in both types of genes are required for cancer
to occur. For example, a mutation limited to one oncogene would be
suppressed by normal mitosis control and tumor suppressor genes,
first hypothesised by the Knudson hypothesis
• A mutation to only one tumor suppressor gene would not cause
cancer either, due to the presence of many "backup" genes that
duplicate its functions
• It is only when enough proto-oncogenes have mutated into
oncogenes, and enough tumor suppressor genes deactivated or
damaged, that the signals for cell growth overwhelm the signals to
regulate it, that cell growth quickly spirals out of control
• Often, because these genes regulate the processes that prevent most
damage to genes themselves, the rate of mutations increases as one
gets older, because DNA damage forms a feedback loop

41. • Mutation of tumor suppressor genes that are passed on to the next generation of not
merely cells, but their offspring, can cause increased likelihoods for cancers to be
inherited
• Members within these families have increased incidence and decreased latency of
multiple tumors
• The mode of inheritance of mutant tumor suppressors is that affected member
inherits a defective copy from one parent, and a normal copy from another
• Because mutations in tumor suppressors act in a recessive manner (note, however,
there are exceptions), the loss of the normal copy creates the cancer phenotype
• For instance, individuals that are heterozygous for p53 mutations are often victims
of Li-Fraumeni syndrome, and that are heterozygous for Rb mutations
develop retinoblastoma
• In similar fashion, mutations in the adenomatous polyposis coli gene are linked
to adenopolyposis colon cancer, with thousands of polyps in the colon while young,
whereas mutations in BRCA1 and BRCA2 lead to early onset of breast cancer

42. Non-mutagenic carcinogens
• Many mutagens are also carcinogens, but some
carcinogens are not mutagens. Examples of carcinogens
that are not mutagens include alcohol and estrogen
• These are thought to promote cancers through their
stimulating effect on the rate of cell mitosis
• Faster rates of mitosis increasingly leave fewer
opportunities for repair enzymes to repair damaged
DNA during DNA replication, increasing the likelihood
of a genetic mistake
• A mistake made during mitosis can lead to the daughter
cells' receiving the wrong number of chromosomes,
which leads to aneuploidy and may lead to cancer

43. Role of infections
BACTERIAL
• Helicobacter pylori can cause gastric cancer
• Overall about 1% to 3% of people infected
with Helicobacter pylori develop gastric
cancer in their lifetime compared to 0.13% of
individuals who have had no H.
pylori infection

44. • Infection by H. pylori causes no symptoms in
about 80% of those infected
• About 75% of individuals infected with H.
pylori develop gastritis
• Thus, the usual consequence of H.
pylori infection is chronic asymptomatic gastritis
• Because of the usual lack of symptoms, when
gastric cancer is finally diagnosed it is often fairly
advanced. More than half of gastric cancer
patients have lymph node metastasis when they
are initially diagnosed

45. Viral
• Many cancers originate from a viral infection; this is especially true in animals such
as birds, but less so in humans
• 12% of human cancers can be attributed to a viral infection
• The mode of virally induced tumors can be divided into two, acutely
transforming or slowly transforming
• In acutely transforming viruses, the viral particles carry a gene that encodes for an
overactive oncogene called viral-oncogene (v-onc), and the infected cell is
transformed as soon as v-onc is expressed
• In contrast, in slowly transforming viruses, the virus genome is inserted, especially
as viral genome insertion is obligatory part of retroviruses, near a proto-oncogene in
the host genome
• The viral promoter or other transcription regulation elements, in turn, cause over-
expression of that proto-oncogene, which, in turn, induces uncontrolled cellular
proliferation
• Because viral genome insertion is not specific to proto-oncogenes and the chance of
insertion near that proto-oncogene is low, slowly transforming viruses have very
long tumor latency compared to acutely transforming virus, which already carries
the viral-oncogene

46. • Viruses that are known to cause cancer such
as HPV (cervical cancer), Hepatitis B (liver cancer),
and EBV (a type of lymphoma), are all DNA viruses
• When the virus infects a cell, it inserts a part of its own
DNA near the cell growth genes, causing cell division
The group of changed cells that are formed from the
first cell dividing all have the same viral DNA near the
cell growth genes
• The group of changed cells are now special because
one of the normal controls on growth has been lost

47. Helminthiasis
• Certain parasitic worms are known to be
carcinogenic
• These include:
• Clonorchis sinensis (the organism
causing Clonorchiasis) and Opisthorchis
viverrini (causing Opisthorchiasis) are associated
with cholangiocarcinoma
• Schistosoma species (the organisms
causing Schistosomiasis) is associated
with bladder cancer.

48. Epigenetics
• Epigenetics is the study of the regulation of gene expression through
chemical, non-mutational changes in DNA structure
• The theory of epigenetics in cancer pathogenesis is that non-
mutational changes to DNA can lead to alterations in gene
expression
• Normally, oncogenes are silent, for example, because of DNA
methylation
• Loss of that methylation can induce the aberrant expression
of oncogenes, leading to cancer pathogenesis
• Known mechanisms of epigenetic change include DNA methylation,
and methylation or acetylation of histone proteins bound to
chromosomal DNA at specific locations
• Classes of medications, known as HDAC inhibitors and DNA
methyltransferase inhibitors, can re-regulate the epigenetic signaling
in the cancer cell

49. THANK YOU