Molecular Biology of Neoplasms

1. Molecular Pathogenesis /
Mechanisms Of Carcinogenesis
• Dr.Pandey,U.S.
• Professor & HOD
• Deptt. Of Pathology,
• GOVT. MEDICAL COLLEGE,
• BETTIAH-845438(West
Champaran,Bihar)

12. Hallmarks Of Molecular Carcinogrnesis

15. • Proteins within the cell control the cell cycle
– Signals affecting critical checkpoints determine
whether the cell will divide (cyclins, kinases)
Checkpoints Monitor Cell Cycle Progression
G1 checkpoint
M checkpoint G2 checkpoint
Control
system
Figure 8.9A

17. Hallmarks of Cancer
1. Self sufficiency in growth factors
2. Insensitivity to growth-inhibitory signals
3. Evasion of apoptosis
4. Defect in DNA Repair
5. Limitless replicative potential by Telomerase
6. Sustained angiogenesis
7. Ability to invade and metastasize
8. Evasion in Immune Surveillance
9. Role of micro-RNA (mRNA)
10. Epigenetic Changes
TEN (10)- fundamental changes of cell Physiology:

18. 18
Hall Marks Of Cancer:10-Fundamental Changes of Cell Physiology
CHRONIC INFLAMMATION
1.Self-sufficiency in growth signals: Tumors have the capacity to proliferate without
external stimuli, usually as a consequence of oncogene activation.
2. Insensitivity to growth-inhibitory signals: Tumors may not respond to molecules that
are inhibitory to the proliferation of normal cells such as transforming growth
factor-β (TGF-β), and direct inhibitors of cyclin-dependent kinases.
3.Defects in DNA repair: Tumors may fail to repair DNA damage caused by carcinogens
or unregulated cellular proliferation.
4.Evasion of Apoptosis : Tumors may be resistant to programmed Cell Death,as a
Consequence of inactivation of p53 or other changes.
5. Limitless replicative potential: (By TELOMERASE Activity)Tumor cells have
unrestricted proliferative capacity, associated with maintenance of telomere
length and function.
6. Sustained angiogenesis: Tumors are not able to grow without formation of a
vascular supply, which is induced by various factors, the most important being
vascular endothelial growth factor (VEGF).
7. Ability to invade and metastasize: Tumor metastases are the cause of the vast
majority of cancer death & depend on processes that are intrisic to the Cell or
are initiated by the signals from the tissue environment.
8.Evasion of Immune Surveillance:
9.Role OF micro-RNA(mRNA)
10.Epigenetic Changes

19. The control of cell division
>> is critical to normal tissue structure and function. It is regulated
by a complex interplay of many genes that control the cell
cycle,with DNA replication (S phase) and mitosis as major
checkpoints
>>The cell cycle is tightly regulated to minimise
transmission of genetic damage to subsequent cellgenerations
>>Progression through the cell cycle is primarily controlled by
cyclins,associated kinases and their inhibitors. Retinoblastoma
(RB) and p53 are major suppressor genes involved in the G1/S
checkpoint control
>>Cancer may be perceived as the consequence of loss of cell cycle
control and progressive genetic instability

20. Cell Cycle Regulatory Genes
Commonly Altered IN Human Cancers

22. Some acronyms!
• Myc
• Sis
• Erb
• Src
• Ras
• Yes
• Abl
• Fos
• jun
Myelocytomatosis
Simian sarcoma
Erythroblastoma
Rous sarcoma virus
Rat sarcoma
2 viruses Y73 & ESH sarcoma, isolated from a
chicken owned by Mr. Esh
Abelson murine leukaemia virus
Finkel biskis jinkins reilly mouse sarcoma
junana

24. 1.Oncogenes: are Cancer causing genes,
& are mutations of NORMAL genes ( PROTO-oncogenes )
• Genes known as proto-oncogenes code for
proteins that stimulate cell division
• mutated forms, called oncogenes, cause
stimulatory proteins to be overactive, with the
result that cells proliferate excessively
• gain of function mutations

25. ONCOGENES

26. Oncogenes ( Contd. )

27. Activation of proto-oncogenes
 1.Point mutations
 2.Gene Amplification
 3.Translocation
 4.Chrosomal Rearrangements: -Altered Regulation
-Fusion Gene
 5Transduction: transfer of genetic material by mediators
Transduction into Retrovirus(Viral Oncogene or v-onc )
 6.Loss of degradation signals
 7.Viral insertation

28. 2.Tumor Suppressor Genes( TSGs)
Definition – Tumor suppressor genes are the normal
genes that regulates the growth of cells.
• Loss of function of tumor suppressor genes can lead
to cancer.
• When mutated, the mutant allele behaves as a
recessive; that is, as long as the cell contains one
normal allele, tumor suppression continues.
• (Oncogenes, on contrast, behave as dominant; one
mutant or overly-active allele can predispose the cell
to tumor formation).

33. TSGs
 Divided into 2 categories –
 Caretaker genes – maintains the integrity of
genome by repairing DNA.
e.g. BRCA1, BRCA2 ,p53.
 Gatekeeper genes - inhibits the proliferation
of cell with damaged DNA and Apoptosis.
e.g. p53, RB, APC ,NF1 , CDH1( E-cadherin).

35. Mechanism of action of TSGs
1) Suppression of cell division – it is the main
mechanism adopted by most of the TSGs.
e.g. RB, APC, p53, p21 & p27.
2)DNA damage repair - capable of fixing DNA
damage including DNA mismatch. e.g. p53,
BRCA, MSH2(mutS homolog2), MLH1(mutL
homolog1), ATM (ataxia telangiectasia mutated
gene).

36. 3)Induction of Apoptosis -
Apoptosis maintains normal homeostasis and
suppresses cancer. e. g. p53, APC, PTEN
4) Inhibition of metastasis-
Tumor suppressors that can inhibit metastasis
e.g. metastin , breast cancer metastasis
suppressor1 ( BCMS 1), CDH1
 Some TSG e.g. p53 may utilize more than one
mechanism for their tumor suppressing
function.

37. Loss Of Heterozygosity (LOH)
• Because tumor suppressor genes are recessive .
Cells that contain one normal and one mutated gene
— are heterozygous — still behave normally.
• However, there are several mechanisms which can
cause a cell to lose its normal gene and thus be
predisposed to develop into a tumor. These may
result in a "loss of heterozygosity" or "LOH".

38. Mechanisms of LOH:
1. Deletion of
– the normal allele;
– the chromosome arm containing the normal allele;
– the entire chromosome containing the normal allele
(resulting in aneuploidy).
2. Loss of the chromosome containing the normal allele
followed by duplication of the chromosome containing
the mutated allele.
3. Mitotic recombination. The study of tumor suppressor
genes revealed that crossing over with genetic
recombination — occasionally occurs in mitosis .
Function of tumor suppressor can also be blocked by
methylation of their promoter region.

39. 3.Genomic Instability Of Cancer Cells-Enabler Of
Malignancy : By Defect in DNA - Repair

43. 4.EVASION OF APOPTOSIS

44. 4.Evasion Of Apoptosis (Contd.)
ESCAPING CELL DEATH BY APOPTOSIS: GENES REGULATING
APOPTOSIS AND CANCER
Another mechanism of tumour growth is by escaping cell death by
apoptosis. Apoptosis in normal cell is guided by cell death receptor,
CD95, resulting in DNA damage. Besides, there is role of some other
pro-apoptotic factors (BAD, BAX, BID and p53) and apoptosis-
inhibitors (BCL2, BCL-X).
In cancer cells, the function of apoptosis is interfered due to
mutations in the above genes which regulate apoptosis in the
normal cell. For example:
a) BCL2 gene seen in normal lymphocytes, but its mutant form with
characteristic translocation (t14;18) (q32;q21) was first described
in B-cell lymphoma. It is also seen in many other human cancers
such as that of breast, thyroid and prostate.
b) CD95 receptors are depleted in hepatocellular carcinoma and
hence the tumour cells escape apoptosis.

45. 5.Limitless Replicative Potential By Telomerase Activity
TELOMERES AND TELOMERASE IN CANCER
After each mitosis (cell doubling) there is progressive
shortening of telomeres which are the terminal tips of
chromosomes. Telomerase is the RNA enzyme that helps
in repair of such damage to DNA and maintains normal
telomere length in successive cell divisions.
Cancer cells in most malignancies have markedly
upregulated telomerase enzyme, and hence telomere
length is maintained. Thus, cancer cells avoid ageing,
mitosis does not slow down or cease, thereby
immortalising the cancer cells.

46. 5.Limitless Replicative Potential by Telomerase Activity

48. 6. Sustained Perfusion Of Cancer:VASCULARISATION

49. 6. Sustained Perfusion Of Cancer Cells :VASCULARISATION:Cancer cells
can not thrive&metastasise without neovascularisation,which supplies them nourishment &
oxogen: Vascularisation occurs by 2-process –i).Angiogenesis & ,ii) other modes

51. ii)Other Modes of Tumour Vascularisation

52. 7.Ability To Invade & Metastasize

54. 7.MECHANISM AND BIOLOGY OF LOCAL INVASION AND METASTASIS: (Contd.)
The process of local invasion and distant spread by lymphatic and haematogenous routes (together called
lymphovascular spread) discussed above involves passage through barriers before gaining access to the
vascular lumen.
1. Aggressive clonal proliferation and angiogenesis :The first step in the spread of cancer cells is
the development of rapidly proliferating clone of cancer cells. This is explained on the basis of tumour
heterogeneity.
2. Tumour cell loosening : In epithelial cancers, there is either loss or inactivation of E-cadherin and
also other CAMs of immunoglobulin superfamily, all of which results in loosening of cancer cells.
3. Tumour cell-ECM interaction : Loosened cancer cells are now attached to ECM proteins, mainly
laminin and fibronectin.
4. Degradation of ECM :Tumour cells overexpress proteases and matrix-degrading enzymes,
metalloproteinases (e.g. collagenases and gelatinase), while the inhibitors of metalloproteinases are
decreased.
5. Entry of tumour cells into capillary lumen : The tumour cells after degrading the basement
membrane are ready to migrate into lumen of capillaries or venules : by
( a) AMF: Autocrine Motility Factor
( b) Cleavage Products Of Matrix Components formed following degradation of ECM.
6. Thrombus formation : The tumour cells protruding in the lumen of the capillary are now covered
with constituents of the circulating blood and form the thrombus.
7. Extravasation of tumour cells:Tumour cells in the circulation (capillaries, venules, lymphatics) may
mechanically block these vascular channels and attach to vascular endothelium and then extravasate to
the extravascular space.
8. Survival and growth of metastatic deposit :The extravasated malignant cells on lodgement in the
right environment grow further under the influence of growth factors (PDGF,FGF,TGFbeta,&VEGF)
produced by host tissues, tumour cells and by cleavage products of matrix components. The metastatic
Deposits grow further if the host immune defence mechanism fails to eliminate it.

55. Invasion-Metastasis Cascade
AggressiveClonalProliferation-PrimT
TUMOR CELL LOOSENING
ADHESION OF Tumor Cells
DEGRADATION OF E C M
TRANSFORMATION
GROWTH
BM INVASION ( Local Invasion )
ANGIOGENESIS
INTRAVASATION
EMBOLIZATION (Circulating Tumor
Cells)
ADHESION
EXTRAVASATION ,ANGIOGENESIS
Micrometastasis,Colonisation
(METASTATIC GROWTH )
etc.

56. 8.Escape Of Immune Surveillence or
Evasion of Host Defence

59. 9.Role OF micro-RNA(mRNA): ONCOMIRS
microRNAs (miRs) are small noncoding,single stranded RNAs,
approximately 22 nucleotides in length, that mediate sequence-
specific inhibition of messenger RNA (mRNA) translation through
the action of the RNA-induced silencing complex (RISC).

60. MICRO-RNAs IN CANCER: ONCOMIRS
..Normally, microRNAs function as the post-translational gene regulators of cell
proliferation, differentiation and survival.
..In cancer, microRNAs have an oncogenic role in initiation and progression and
are termed as oncogenic microRNAs, abbreviated as oncomiRs. These
oncogenic microRNAs influence various cellular processes in cancer such as
control of proliferation, cell cycle regulation, apoptosis, differentiation,
metastasis and metabolism.
9.Role OF micro-RNA(mRNA): ONCOMIRS (Contd .)
MicroRNAs (miRNAs) are small noncoding single-stranded RNAs that are incorporated into
silencing complexes and can mediate post-transcriptional gene silencing. Deletions of
miRNA sequences can drive oncogene expression, whereas overactivity can inhibit tumor
suppressor gene function. Thus miRNA-200 is important for invasion and metastasis,
whereas miRNA-155 upregulates genes that promote proliferation, including MYC.
Conversely, deletions of miRNA-15 and miRNA-1- lead to the upregulation of antiapoptotic
BCL-2 in chronic lymphocytic leukemias.Long intervening noncoding RNAs (lincRNAs) can
regulate the activity of chromatin “writers” that modify histones and thereby control gene
expression; other noncoding RNA species also have roles in post-transcriptional gene
silencing or affect the maturation of ribosomes.

61. Posttranslational modification of histones (e.g., acetylation) and
DNA methylation—without changes in the primary DNA sequence
—can influence gene expression including silencing tumor
suppressor genes (e.g., p14ARF in GI cancers and p16INK4a in
various malignancies). Therapeutic strategies to demethylate
selected DNA sequences may be efficacious in these cases.
10.EPIGENETIC - CHANGES
Epigenetics= Relating to Epigenesis.
Epigenesis is development or Regulation of Expression of
Gene Activity without Alteration of Genetic Structure.
or Epigenesis is Heritable-Chemical-Modification of D.N.A. or
Chromatin that does NOT ALTER D.N.A.SEQUENCE (Robin)
“Epigenetics” refers to factors other than the sequence of DNA that
regulate gene expression (and, thereby, cellular phenotype).
Epigenetic Change shows Carcinogens act on Activators or
Suppresssor of Genes & NOT On GENES THEMSELVES

64. Stromal Microenvironment & Carciogenesis
• Cancers are Heterogenous Collections,including
malignant cells, as well as non-cancerous inflamm
-atory & stromal cells.
Increasingly it is recocnised that while
tumours clearly influence their sorrounding
stroma (e.g.,by inducing neovascularisation &
desmoplastic matrix),the host stromal cells,ECM
,& inflammation can also modulate(i.e.,inhibit &
augment)tumour growth.

66. Knudson’s two hit hypothesis
• To account for the sporadic and familial occurrence
of retinoblastoma ( Knudson in1971) & Adenoma-
Ca.Sequence in Colorectal Ca. states-
• Two mutation( hits) are required with RB gene
located on Ch 13q14 for development of
retinoblastoma.
• In Familial cases :- a child inherit a defective copy of
RB gene (1st hit) the other copy is normal.
Retinoblastoma develops when the normal copy
undergo somatic mutation( 2nd hit).
• In Sporadic cases :- both normal RB alleles are lost
by somatic mutation in one of the retinoblast.

68. Adenoma- Carcinoma Sequence

70. Warburg Effect
• Growth -Promoting Metabolic Changes :
 1. Excessive Nutrient Acquisition
 2. Altered Metabolic Pathways
 3.Oncometabolites in Tumerogenesis

72. THANKS………….

73. Theories/ Mechanisms Of Cancer Oncogenesis

74. THEORIES/ MECHANISMS OF CANCER ONCOGENESIS (CONTD.)
4. Immune Surveillance Theory: This hypothesis suggests that an immune
competent host mounts an attack pn developing tumour cells so as to destroy them
while an immune-incompetent host fails to do so.
5.Epigenetic Theory: According to the epigenetic theory ,the carcinogenic agents
act on activators or suppressors of genes and not on the genes themselves and
result in the abnormal expression of genes. Errors in epigenetic processes which
may appear
in cancer are in DNA methylation and histone modification.

75. resulting in mutations
Theories/Mechanisms Of Cancer Oncogenesis (Contd.)

76. Theories/Mechanisms Of Cancer Oncogenesis (Contd.)

77. Molecular Basis Of Multistep Carcinogenesis
No single genetic alteration is sufficient to induce cancers in vivo. A variety of controls influenced by
multiple categories of genes oncogenes,tumor suppressor genes, apoptosis-regulating
genes,senescence-modulating genes—must be lost for the emergence of cancer cells.
This situation is exemplified by the colon adenoma-tocarcinoma sequence (Fig.); the
evolution of benign adenomas to carcinomas is marked by increasing and additive effects
of mutations,affecting K-RAS(6p11.12),APC(5q21),SMAD 2,& SMAD 4 genes on 18q. The
accumulation of mutations with increasing genetic instability can be promoted by loss of
p53(17p13), DNA repair genes, or both.
Over time, tumors acquire additional changes that result in greater malignant
potential (e.g., accelerated growth, invasiveness,angiogenesis, and the ability to
form distant metastases) .

79. Molecular Basis Of Multistep Carcinogenesis (Contd.)
• Genes that regulate cellular proliferation in a more or less tissue specific
manner,such as APC(5q21),NF-1(17q11),& RB(13q14), are called GATEKEEPER-
genes, whereas those that regulate genomic stability(DNA repair genes) are
called CARETAKER-genes ( MSH2-2p22;MLH1-3p21;PMS1-2q31-33; PMS2-
7p22,Xerpoderma Pigmentosa-7 Nucleotide Excision Repair-genes likeXPV-genes-
9q22 ;ATM-genes -11q22-23 ).These latter genes regulate the likelihood of a
particular cell developing a mutator phenotype ( cells unusually susceptible to
addition mutations) resulting in mutations in GATEKEEPER-genes.
Despite the fact that tumors are initially monoclonal in origin, by the time they
become clinically evident, they are extremely heterogeneous.

80. Flow chart depicting scheme of Mol,Basis of Cancer

81. CHARLES C COLTON 1820, Lacon1:322
The greatest fool may ask more than the wisest man can answer

90. Flow chart depicting scheme of Mol,Basis of Cancer