CANCER

A disease in which a group of cells divide uncontrollably and invade other tissues, resulting from an
accumulation of mutations in their DNA.

Genetic alterations often occur in a preferred sequence but the total accumulation of
changes rather than order is responsible for determining tumour’s biologic properties

- CELL CYCLE
G1: Cell has received certain growth signals, and is replicating organelles, synthesising proteins and
aerobically respiring to prepare for division.
*checkpoint* 1/ sufficient nutrients 2/ cell size 3/ growth factors 4/ DNA damage

S: Cell will replicate is DNA (see below)

G2: Cell now synthesises a more extensive microtubule network, which will form the mitotic spindle
which allows for cell division.
*checkpoint* 1/ DNA damage 2/ DNA replication completeness

Mitosis + Cytokinesis:
Prophase = Chromosomes condense, mitotic spindle begins to form, nucleolus disappears, nuclear
envelope begins to break down
Metaphase = Chromosomes line up attached by spindle fibres on metaphase plate at cell’s equator,
chromosomes also attached to spindle poles at opposite ends of the cell
*checkpoint* chromosomes correctly attached to spindle fibres i.e. not floating in cytoplasm
Anaphase = Motor proteins drive pulling apart of sister chromosomes, by contraction of
kinetochores to their respective poles
Telophase = Mitotic spindle begins to break down, chromosomes decondense, nuclei begin to form
around chromosomes

Cytokinesis = Cleavage furrow forms in middle of cell, actin filaments pinching in and contracting till
two cells made ANIMALS
Cell plate forms in between two cells, made of cellulose, which extends until two cells are separated
PLANTS

- DNA REPLICATION
A semi-conservative process, which begins with DNA gyrase and helicase unwinding the DNA double-
helix and breaking the hydrogen bonds between complementary base pairs.
Then, free DNA nucleotides in the nucleoplasm will hydrogen bond to their complementary base pair
on the template strand.
DNA polymerase will then form phosphodiester bonds between adjacent nucleotides, synthesising
new strands.
However, DNA polymerase only works in the 5’ to 3’ direction, only attaching nucleotides to the 3’
end. As the DNA double-helix is anti-parallel, one strand can be continuously replicated (leading) but
the other must be replicated in fragments (lagging), called Okasaki fragments. For the lagging strand,
DNA polymerase requires RNA primers to polymerise the nucleotides.
These are replaced by DNA to form a complete strand, except for when the end of the chromosome
is reached. At this point, the template strand cannot be covered by the fragment, and the last primer
cannot be replaced by DNA because DNA polymerase requires hydroxyl groups from neighbouring
Okasaki fragments for this (of which there are none because end reached).
Therefore, to prevent chromosomes from being shortened, they are capped with repeating
sequences of the base sequence TTAGGG called telomeres. These are shortened instead which each
cell division and DNA replication.
Telomerase is the enzyme that synthesises more telomeres.

- APOPTOSIS
Lysosomes within cell lyse organelles using digestive enzymes and break down cellular contents,
meanwhile cell releases extracellular signals to extracellular matrix. Cell shrinks and eventually
macrophage will engulf it.
Enzymes key to this process are tyrosine kinases and caspases, notably Cas8 and Cas10. There is
an extrinsic and intrinsic pathway to apoptosis.
The extrinsic pathway involves turning on ‘death’ cascade, which involves the Fas receptor.
There is also a survival pathway, in which the Bax and Bak proteins are key. Bax triggers
apoptosis, and is bound to by Bcl-2, which inhibits it. In B-cell lymphoma, Bcl-2 is overexpressed,
resulting in apoptosis evasion. However, Bcl-2 itself can be inhibited by the protein BH3. By
synthesising mimetics of BH3, such as the drug venetoclax, the surplus Bcl-2 present in this
cancer can be inhibited, thus allowing apoptosis to occur, and causing the cancer cells to die.
Cellular membranes disrupted, cytoplasmic and nuclear skeletons broken down, cytosol
extruded, chromosomes degrade, nucleus fragments, macrophage engulfs cell.

Hallmarks [Hanahan et al. Hallmarks of Cancer, 2000 & 2011]

SELF-SUFFICIENT GROWTH SIGNALS
Mitogenic growth signals (GS) allow proliferation for normal cells, and in their absence cells will not
divide. They are transmitted by transmembrane receptors, and oncogenes often synthesise growth
signal mimetics.

(1) Alteration of extracellular growth signals: causes positive feedback loop; autocrine
stimulation e.g. PDGF (platelet derived growth factor) TGF (tumour growth factor) in
glioblastomas and sarcomas [cell migration, proliferation and angiogenesis, skip G1
checkpoints in order to divide]
(2) Alteration of transcellular transducers of those signals; e.g. elevating receptor proteins,
integrins extracellular links affect apoptosis and cell mobility, ligand-activated, tyrosine
kinase
(3) Alteration of intracellular circuits which translate signals into action, Ras
(4) Increased cell growth during G1 then stimulation of tumour associated stroma to produce
growth factors

INSENSITIVITY TO GROWTH INHIBITORY SIGNALS

Soluble growth inhibitors and immobilised inhibitors in extracellular matrix also received by
transmembrane cell surface receptors and intracellular signalling circuits may result in cells moving
into a) G0 b) post-mitotic state: differentiation associated traits
Cell cycle clock in cancers deregulated e.g. pRB [tumoursupressor], hypophosphorylation will block
proliferation, TGF to prevent phosphorylation that inactivates pRB, Smad, E7 encoding transcription
factors [E7 inactivates p53], TP53

ABILITY TO REPLICATE INFINITELY

Upregulation of telomerase which adds hexanucleotide repeats onto ends of telomeric DNA. ~90%
of cases. Telomerase’s cell proliferation protein subunit is TERT and allows end to end fusion.
Otherwise, ALT: recombination-based inter-chromosomal exchanges.
 EVASION OF APOPTOSIS
Sensors and effectors.
Cell surface receptors bind survival e.g. IGF1&IGF2 /death factors e.g. FAS (cascade of proteolysis)
Detect e.g. DNA damage, signalling imbalance, survival factor insufficiency, hypoxia. Many signals
that elicit apoptosis converge on the mitochondria, which respond to proapoptotic signals by
releasing potent apoptosis catalyst cytochrome C. Apoptosis inhibitors include Bcl-2 and p53.
Cancers down regulate Bax, disrupts mitochondrial membrane, Noxa, Puma, BH3, Bak Bim, Puma,
loss of tp53 tumour suppressor.

ANGIOGENESIS
Soluble factors, receptors, integrins, adhesion molecules, VEGF vascular endothelial growth factor
(bind to tyrosine kinase receptors), anti VEGF antibodies potential treatment? p53. Angiogenic
activators and inhibitors.

ABILITY TO INVADE AND METASTASISE

Pioneer cells which invade. 90% of cancer deaths. Changes in physical coupling of cells to their
microenvironment and activation of extracellular proteases.
CAMs and integrins. E-cadherin: homotypic cell to cell interaction molecule. Immunoglobulin.
Antigrowth and other signals. Lost in epithelial cancers. Proteases. NF2 neurofibromatosis.

DIFFERENT ENERGY METABOLISM PATHWAY

Adjustments of energy metabolism in order to fuel cell growth and division.
Limit energy metabolism largely to glycolysis, which may seem counter-intuitive because lower ATP
yield. To balance this out they upregulate glucose transporters e.g. GLUT1 and glycolytic pathway
enzymes.
This effect may be accentuated under hypoxic conditions in which many tumours operate.
Why? Increased glycolysis allows diversion of intermediates into various biosynthetic pathways e.g.
generating nucleosides which thus facilitate biosynthesis of amino acids, macromolecules and
organelles for new cells.

Some tumours have subpopulations with 2 energy pathways

 Glucose dependent which secrete lactate
 Lactate dependent which import lactate from neighbouring cells
This change in energy metabolism is largely orchestrated by proteins which program core hallmarks
of cancer, and the extent to which this hallmark is independent from others is uncertain.

ALTERATION OF IMMUNE RESPONSE

Deficiencies in CD8+ cytotoxic T lymphocytes and CD 4+ Tn1 helper T cells led to demonstrable tumour
increase

[HIGH MUTABILITY]

[CREATION OF TUMOUR MICROENVIRONMENT]

Heterotypic interactions from stromal cells. Macrophage at. Tumour periphery can foster local
invasion by providing matrix-degrading enzymes. Mesenchymal epithelial transition MET and ET.

N.B.// necrotic cells bloat and explode: releasing pro-inflammatory signals, can be actively tumour
promoting bioactive regulator factors e.g. ~40% melanomas B-Raf, Myc, Ras, Raf to MAP, kinase
pathway mitogen-activated protein

Causes
 CARCINOGENS
Tobacco: polycyclic aromatic hydrocarbons [interaction with microsomal enzymes yields reactive
epoxides which react with DNA by e.g. producing covalent bonding]
nitrosamines
Asbestos: microfibres endocytosed by cell accumulate in perinuclear region and affect
chromosome segregation during mitosis, causing anaphase abnormalities.
crocidolite is a fibrous mineral deeply associated to lung and mesothelial cancer, known
as blue asbestos.
Alcohol  acetaldehyde: results in oxidative stress

Oxidative stress: free radicals are big mutagen

OBESITY
Abnormal levels of metabolic proteins including insulin-like growth factors and sex hormones
which promote abnormal cell proliferation.
Adipose tissue also contributes to an inflammatory microenvironment characteristic of the
tumour microenvironment.

INHERITED PREDISPOSITIONS
Genetic mutation in tumour suppressor genes, oncogenes, blood vessels genes, DNA repair
genes
e.g. BRCA1, BRCA2 : breast and ovarian cancers. BRCA1 tumour suppressor, involved with DNA
damage repair and ubiquitination

INFECTION
Usually via causing DNA damage /genomic instability.
e.g. oncoviruses: human papillomavirus HPV cervical cancer [upregulates and down regulates
genes, down regulation of cell growth regulation, E6&E7 high affinity for p53 and pRB]
e.g. bacteria: Helicobacter pylori gastric carcinoma by chronic inflammation

Chronic Inflammation may also promote DNA damage [cytokines release free radicals/ reactive
oxygen species [mutagens] and stimulate angiogenesis], and a weakened immune system may
enable abnormal cell proliferation.

RADIATION
Non-ionising: e.g. UV causes dimerization
Ionising: e.g. medical imaging, gamma/ / radiation

Strong mutagens: May break chromosomes, cause chromosomal translocations, or smaller base
mutations, single strand breaks etc.

OTHER
Disrupting body clock
Organ transplant
Physical trauma
Maternal-foetal transmission

Treatments
SURGERY
Cryotherapy, laser therapy, hypertherapy, photodynamic. Trying to bring to a minimum because
induces inflammatory response which promotes cancerous proliferation by increasing blood
supply.

RADIOTHERAPY
High doses of radiation kill cells. Either external beam or internal, and can be intraoperative
radiation. Use pre, during, and post operationally as is often adjuvant.
Damages DNA of cancerous cells by forming free radicals. As DNA can only be damaged ruing S
phase, the majority of cells are less affected than cancerous cells. Photon radiation creates free
radicals which break DNA bonds.
Radiotherapy has been found to act as a type of immunotherapy by the off-target effect. In this
when a tumour is irradiated other tumours not exposed to radiation undergo remission. This is
because the radiation shears off small nuclear fragments from the tumour cells, which very
potently stimulate the immune response. STING.

CHEMOTHERAPY
Cytotoxic, kills cells that divide quickly therefore side-effect of hair loss and mouth sores.
Mechanisms of action mainly by causing DNA synthesis to go awry/ inhibiting microtubule
function.
e.g. may covalently bond to DNA or form intra and inter-strand DNA crosslinks. Or interfere with
DNA synthesis by breaking DNA strands by intercalation in the DNA. May also inhibit
topoisomerase. Tubulin inhibition which prevents assembly of mitotic spindle, or finally
enhanced microtubule production which also affects mitotic spindle assembly.

STEM CELL TRANSPLANT

Autologous/allogeneic/syngeneic. Sometimes graft vs. tumour preference of cells as treatment
for multiple myeloma and leukaemia but mainly used for symptom alleviation.

HORMONE THERAPY
Blocking hormones required for cell growth, associated to mutations.
E.g. tamoxifen. Binds to oestrogen receptors and in some cells has anti-oestrogenic properties,
while in others pro. It is strongly anti-oestrogenic on mammary epithelial cells, thus preventing
the proliferative actions of oestrogen on the mammary epithelium.
Breast cancer.

IMMUNOTHERAPY
Checkpoint inhibitors. Adoptive cell transfer. Monoclonal antibodies. Treatment vaccines.
Interferons and interleukins. BCG: bacterium.
In a group of melanoma nodules directly injected with BCG, 90% were observed to regress.

Cornell researchers have developed a nanoparticle made of lipids similarly to cells, with the
substance e-selectin to make it ‘sticky’ and a chemical named trail which is cancer specific. These
nanoparticles injected into the bloodstream immediately bind to white blood cells, due to the e-
selectin. Then, following the immune system, these white blood cells travel to the cancer and
the trail will attack the cancer cells. In experiments using mice over 75% of cancer cells died or
were dying after 2 hours of treatment.

TARGETED THERAPY
Small molecule drugs and monoclonal antibodies. May starve cancer of hormones it needs to
grow, bind to antigens on cancer cells, mark cancer cells to they are recognised by the immune
 system, may inhibit angiogenesis, may inhibit growth signals, may trigger apoptosis, may carry
toxins to cancer cells, may deliver cell-killing substances.

E.g. Herceptin/ Trastuzumab for HER2 breast cancer. HER2 gene causes expression of HER2
receptor which dimerises to activate a tyrosine kinase and a cascade with mitogenic effect.
Herceptin is a monoclonal antibody which has two antigen-binding regions, specific to the HER2
receptor, which will bind to its extracellular portion thus preventing tyrosine kinase activation.
Herceptin will also recruit immune effector cells to upregulate the immune response against
cancer cells.

e.g. Gleevec/ Imatinib for chronic myelogenous leukaemia and acute lymphocytic leukaemia.
Prevents tyrosine kinase autophosphorylation.

e.g. venetoclax : is a BH3 mimetic. In short, Bax is a pro apoptotic protein (by its triggering of
cytochrome C release), and Bax is regulated by Bcl2, which thus downregulates apoptosis. In B
cell lymphoma, Bcl2 may be overexpressed, causing apoptosis evasion. Bcl2 itself is inhibited by
BH3 proteins (e.g. Noxa, Puma). By synthesising a BH3 mimetic: venetoclax, we are able to
induce apoptosis in cancerous cells.

Detection

Detection of cancers is becoming increasingly important because of tumour heterogeneity. This is

cancerous tumours’ variety in gene pool and variation in exact DNA which means they are differently
prone to different drugs, and the later cancer is caught the more likely it is to have strains resistant
to certain treatments.
The current methods for cancer testing are medical procedures, protein biomarkers and medical
imaging. However each has drawbacks, medical procedures being very invasive and requiring much
infrastructure, protein biomarkers having a low specificity and a high rate of false positives and
medical imaging causing radiation damage and only being effective for certain populations. Lately a
new technique has been developed, that will soon hopefully be commercialized and popularized, as
not only does it not have any of the above disadvantages but also has earlier detection than any
other test. For example, in a lung cancer patient the blood test discovered the cancerous recurrence
100 days earlier than conventional methods. Furthermore, this is not an extremely complicated
process, but begins with a very simple blood test. The DNA is extracted from the blood, and a
molecular library is prepared from this. Then next-generation sequencing and computational
analysis is used to identify cancerous cells.

However, we do not want to become too precise.

suppress CD4􏰁 and CD8􏰁 T cells by their uptake of ar- ginine and high intracellular level of arginase that
depletes their surroundings of arginine, an essential amino acid for T cell ac- tivation (42, 45, 74). MDSC-
produced ROS and peroxynitrite inhibit CD8􏰁 T cells by catalyzing the nitration of the TCR and thereby
preventing T cell-peptide-MHC interactions

MDSC also block T cell ac- tivation by depriving the environment of cysteine, an amino acid that is essential for
T cell activation. T cells lack the enzyme to convert methionine to cysteine and the membrane trans- porter to
import cystine, which could be reduced intracellularly to cysteine, and therefore must obtain their cysteine from
ex- tracellular sources

Metastatic dissemination may occur remarkably early however cancer cells may be
maladapted to their new microenvironments/ suppressed by immune system/ wrong
resources, only an environmental change or mutation will result in their growth into
tumours in and of themselves.

Age-related
Supportive stroma is key
Inflammation : growth factors that sustain proliferative signaling, survival factors that limit
cell death, proangiogenic factors, extracellular matrix-modifying enzymes that facilitate
angiogenesis, invasion, and metastasis, and inductive signals that lead to activation of EMT
and other hallmark-facilitating programs

inflammatory cells can release chemicals, notably reactive oxygen species, that are actively mutagenic for nearby cancer cells, accelerating
their genetic evolution toward states of heightened malignancy

Cancer stem cells CSC

**Hence tumours rapidly become genetically heterogeneous as mutations (unless fatal and
cytotoxic) are left more unchecked. This submits tumours not only to population dynamics
and natural selection – more proliferative and invasive cells having more success than their
relatives, but also makes cancer tumours once large extremely hard to treat, and susceptible
to drug resistance, as exemplified by recurrent tumours which are usually resistant to the
drugs employed in the remission of the former. **
n.b. operates as part of larger network with functional redundancy, mice with non-
functional Rb surprisingly free of abnormalities at least early in life. Similarly with TP53

Inflammation : growth factors that sustain proliferative signaling, survival factors that limit
cell death, proangiogenic factors, extracellular matrix-modifying enzymes that facilitate
angiogenesis, invasion, and metastasis, and inductive signals that lead to activation of EMT
and other hallmark-facilitating programs

inflammatory cells can release chemicals, notably reactive oxygen species, that are actively mutagenic for nearby cancer cells, accelerating
their genetic evolution toward states of heightened malignancy

Genetic alterations often occur in a preferred sequence but the total accumulation of
changes rather than order is responsible for determining tumour’s biologic properties
[Tumour suppressor genes not necessarily recessive at the cellular level]

natural selection

Investigators have identified somatic alterations present at various stages of colorectal

tumour formation
Alterations present in all or virtually all of the neoplastic cells studied
Alterations must provide cell with growth advantage, allowing it to outcompete other
neoplastic cells within the tumour
Or genetic alteration may have arisen coincidentally with another change that actually
provided the selective growth advantage
When the same mutations are present in many tumours, former more likely
Genetic alterations in oncogenes : ras gene mutation
When transfected in recipient cells, mutated ras confers neoplastic properties
50% colorectal carcinomas have ras
ras may be the initiating event in a subset of colorectal tumours
somatic activation by point mutation, as well as oncogenes activated by amplification or
rearrangement
amplified common, but rarer specific amplification
including neu c-myc c-myb in primary colorectal tumours
Oncogene rearrangements, other than a single trk, not observed in colorectal tumours

Allelic losses and tumour suppressor genes

Loss of chromosomal regions are frequent
Losses involve only one of the two parental chromosomes usually
Allelic losses interpreted as tumour suppressor gene regions
First detected cytogenetically, have studied more recently using probes that detect
restriction fragment length polymorphisms to determine whether one of the two parental
alleles is lost specifically in tumour DNA
Previous studies of allelic losses in tumours from patients with inherited predisposition to
particular tumour types
Inherited predisposition to colorectal tumour formation occurs in an autosomal dominant
syndrome, familial adenomatous polyposis
Linked locus mapped to chromosome 5q
Loss of large portion of chromosome 17p through chromosome loss or mitotic
recombination in more than 75% of colorectal carcinomas
Identified in some tumours, common region of loss contains the p53 gene
Mutations causing amino acid substitutions in the p53 gene product observed in remaining
p53 alleles of several colorectal cancers
Similar mutations in many remaining p53 alleles
APCC, HMPCC

Tumour suppressor genes hypothesised to act recessively at the cellular level: both
maternal and paternal copies of gene inactivated for growth-suppressive function to be
eliminated
Supported by molecular cloning and analysis of retinoblastoma gene
However not always
e.g. p53
predisposition syndromes usually from germline inactivation of one copy of a tumour
suppressor gene
double inactivation rare
singly mutated p53 genes in mice can cooperate with ras to transform primary rodent cells
in vitro
at cellular level, p53 may function as dominant negative taher than recessive mutation
explained perhaps in part by oligomerisation of p53 gene product
mutant p53 gene product may inactivate wt product by binding to it and preventing its
normal association with other cellular constituents
on this basis mutated p53 gene in colorectal tumour -> selective growth advantage
subsequent loss of p53 allele often associated with progression from adenoma to
carcinoma, and probably amplifies growth advantage of initial mutation
evidence in at least one colorectal tumour demonstrating postulated intermediate step

p53 heritable mutation Li-Fraumeni syndrome, sarcoma spectrum

many other allelic losses in colorectal tumours

median of four to five chromosomal arms suffering allelic losses
more losses and considerably worse prognosis than other patients
losses: contain suppressor genes, but also, when loss in a heterogenous fashion is that may
have no specific effect on phenotype but have arisen coincidentally with other genetic
alterations
or many tumour suppressor genes throughout genome

other somatic alterations

significant loss of methyl groups in DNA occurs very early in colorectal tumorigenesis
examination of DNA from even very small adenomas revealed that approximately one third
of DNA regions studied had lost methyl groups present in the DNA of normal colonic
mucosa: suggesting loss of 10-20 million methyl groups per cell
loss of DNA methylation has been shown to inhibit chromosome condensation, might lead
to mitotic nondisjunction -> loss or gain of chromosomes
epigenetic change of hypomethylation could contribute to instability of tumour cell genome,
and alter the rate at which genetic alterations including allelic losses occur

in addition to structural alterations of oncogenes and tumour suppressor genes noted

above, consistent differences in expression or activity of other genes/ gene products have
been noted in colorectal tumours
e.g. c-myc expressed at high levels in most colorectal carcinomas, especially those derived
from the descending colon
tyrosine kinase activities of proteins elevated in many
activities associated with several oncogenes
expression of glycoconjugates also increased in colorectal carcinomas
difficult to assess functionality and significance of altered levels: accompanying or
important?

Data from in situ DNA labelling of colonic epithelium : tumorigenesis preceded by

widespread cellular hyperproliferation
Initiation, promotion and progression in experimental models
And specific to cancers

VIRAL PRODUCTS ONCOGENICALLY INTERFERE WITH THE CELL CYCLE

Viruses may not tamper at the level of the genome, rather directly dysregulating cell signalling
pathways, those of the cell cycle. For example, the huma papilloma viruses HPV cause cervical
cancer by inactivating the Rb gene product
Oncogenic human papillomaviruses HPV, cervical cancer: inactivate Rb gene product
By E7 oncoproteins of HPV-16 and HPV-18 which competitively binding Rb, inhibiting binding its
normal physiological partners
And adenovirus E1A, SV40 T Ag

SURPLUS
APC, in Wnt signalling, signal attenuator “gatekeeper”
Highly conserved family of growth and developmental signals
Ligand-induced release of dishevelled inhibits the GSK-3beta kinase/APC/Axin complex from
degrading beta-catenin
Stabilised beta-catenin translocates from cytoplasm to nucleus and promotes transcription via Tcf-
Lef
Promotes cell growth/survival/migration, upregulates e.g. myc, cyclin D1
FAP familial adenomatous polyposis coli, 95% have colorectal polyps by the age of 35
Moving near immunoglobulin heavy chain promoter
Retinoblastoma (Rb), Chromosome 13, position 13q13.2, tumour suppressor gene which is
inactivated in cancers