What is the primary difference between cell lines/in vitro reserach?
Cell lines can be transformed while primary cells can't be transformed through a
single mutuation
Mutations that activate _____ oncogenes not sufficient to cause cancer in primary
cells
1-2
What is an example of a insufficient mutation which makes it not cause cancer?
Mutated/activiated K-Ras
What do mutation resistance rely on? (2)
DNA stability and tissue organization
How does DNA stability impact mutations (2)?
- DNA is fixed/unchanged
- Very robust defense mechanism against cancer
How does tissue organization help prevent mutation accumulation? (2)
1) low cell division rate of stem cells protecting against DNA instability
2) stem cells are not located in parts of tissue exposed to carcinogens and
mutagens
What is important about mutations in the transit-amplifying cells?
These cells are already scheduled to die at some point, likely won't persist long
enough to become neoplastic
Mutations can be caused by ______ and _____ sources
Exogenous and endogenous
What are some exogenous sources of DNA mutations?
Mutagenic agents (chemical & physical)
What are some endgenous sources of DNA mutations? (3)
1) DNA replication by DNA polymerase during S phase
2) Spontaneous chemical alternations of nucleotide precursors that become
embedded into the DNA
3)Mutagenic agents
Damage from endogenous sources much ______ per day than amount of damage
caused by exogenous mutagenic agents in most tissues
Higher!
What do defenses do cells have against mutagens? (2)
Physical and chemical
What physical defenses do cells have against mutagens?
,Preventing penetration of mutagen into tissue
e.g. Melanin or keratinocytes
What are some chemical defenses against DNA damage? (5)
- ROS mitigation
- GST-mediated detoxification of carcinogens (linked to glutathione)
- Mismatched repair
- Dealkylating repair enzymes
- Excision repair enzymes (BER/ NER)
What are repairs to fix specific lesions in DNA? (3)
1. Base Excision
2. Nucleotide Excision
3. Error prone
What is base excision repair (BER)?
Repairs lesions caused by endogenous sources (ROS or depurination)
How does base excision repair work? (5)
1) DNA glycosylases recognizes abnormal bases
2) The covalent bond to deoxyribose gets cleaved (has trouble recognizing T
because it's in normal DNA)
3) AP endonuclease cleaves
Deoxyribosephosphate
4) DNA polymerase adds individual nucleotide or longer stretch of nucleotides
5) DNA ligase seals the repaired DNA
What is nucleotide excision repair (NER)?
Repairs lesions from exogenous sources by recognizing abnormal bases and
cleaving DNA around the damage.
How does nucleotide excision repair work? (4)
1) large protein complex recognizes the abnormal base
2) cleaves the DNA fragment around the helix distortion
3)Polymerase fills in the gap with DNA replication machinery (PCNA/RPA)
4) DNA ligase seals up repaired DNA
How are nucleotide excision repair and p53 related?
P53 can activate nucleotide excision repair to facilitate global repair of the
genome.
What type of polymerases are used for error prone repair?
High-fidelity polymerases with low error rates
What is error-prone repair?
A mechanism that attempts to repair DNA lesions under non-ideal
circumstances using error-prone polymerases.
Why are error prone polymerases used compared to high fidelity polymerases?
,Can add nucleotides to a growing DNA strand even when the base on
complementary strand is missing or DNA adduct is attached
Why is error-prone repair a gamble cell?
There's a risk incurring more mutations with error prone repair vs cell death due
to failed DNA replication
Inherited Defects in __________ Cause Susceptibility To Specific Cancer Types
DNA Repair Pathways
What are some inherited defects that can cause suspectibility to cancer? (3)
1) Xeroderma pigmentosum (XP)
2) Hereditary non-polyposis colon cancer (HNPCC)
3) BRCA1/2
What type of repair does XP use?
NER
What condition is characterized by extreme sensitivity to UV radiation?
Xeroderma pigmentosum (XP)
What are the characteristics of XP?
Infants burn with minimal exposure, many freckles, parchment like skin
What is the risk of skin cancer for individuals with Xeroderma pigmentosum
before age 20?
2000-fold increased risk compared to the general population.
At what age do XP patients typically present with skin cancer?
Around 10 years old, compared to around 60 years old in the general population.
What causes Xeroderma pigmentosum?
Inherited defects in one of eight genes (7 are NER, 1 is error-prone polymerase)
XP individuals are at relatively no increased risk for cancers of internal organs --
why is this important?
Suggests that UV radiation has been a significant mutagen for most of human
evolution, in response made a DNA repair system to fix damage caused by this is
singular mutation
Hereditary non-polyposis colon cancer (HNPCC) is a
Familial cancer syndrome
What is the lifetime risk of developing colon cancer for individuals with hereditary
non-polyposis colon cancer (HNPCC)?
80% lifetime risk
What types of cancers are individuals with HNPCC at increased risk for? (5)
Brain, endometrial, stomach, ovarian, and urinary tract carcinomas.
What is the mechanism of developing HNPCC? (5)
Loss of APC --> DNA hypomethylation --> activation of K-RAS --> loss of 18q TSG
--> loss of p53
How quickly can adenomas progress to carcinomas in HNPCC patients?
, In 2-3 years, compared to 8-10 years for other types of colon cancer
What genetic defects are associated with hereditary non-polyposis colon cancer?
Mismatch repair (MMR) genes
What happens to mismatch repair genes in HNPCC?
Inherit one defective allele, undergo loss of heterozygosity (LOS) to lose
remaining WT copy of allele
Why is the loss of heterozygity in MMR important?
High mutation rates in mircosatellite repeats of genes on TGF-B --> causing
unchecked cell cycles
What is important about TGF-β receptor and mutations?
If TGF-β signaling is lost, would be advantageous for cell that
wants to undergo unchecked cell cycles
What is the role of BRCA1 and BRCA2 genes?
They help maintain genomic integrity through DNA repair
How do BRCA1/BRCA 2 maintain genomic integirty?
DNA repair proteins!
Large complexes form with other proteins in the nucleus to repair ionizing
radiation
What is the role of BRCA1 and BRCA2 genes thought to be previously?
TSG -- regulating proliferation, survivial and differnation
What is the risk of developing breast cancer for carriers of mutant BRCA1 or
BRCA2 by age 70?
50-70% risk.
What type of DNA repair do BRCA1 and BRCA2 assist with?
Homology-directed DNA repair of double-strand breaks.
What do dsdna breaks require and how? (2)
Repair machinery to access undamaged sequence on sister chromatid
1) sister chromatid is the template
2) unwinds sister chromatid
What happens to homologous repair in cells lacking BRCA1 and/or BRCA2
function?
It is compromised, leading to inability to repair double-strand DNA breaks.
If BRCA1/2 are so important to homologous repair of dsdna breaks,why don't
more cancers have defects in these genes?
We don't know, still under investigation
BRCA2 somatic mutations can be associated with these carincomas outside of
breast and ovarian cancer? (2)
Colon and prostate
Cell lines can be transformed while primary cells can't be transformed through a
single mutuation
Mutations that activate _____ oncogenes not sufficient to cause cancer in primary
cells
1-2
What is an example of a insufficient mutation which makes it not cause cancer?
Mutated/activiated K-Ras
What do mutation resistance rely on? (2)
DNA stability and tissue organization
How does DNA stability impact mutations (2)?
- DNA is fixed/unchanged
- Very robust defense mechanism against cancer
How does tissue organization help prevent mutation accumulation? (2)
1) low cell division rate of stem cells protecting against DNA instability
2) stem cells are not located in parts of tissue exposed to carcinogens and
mutagens
What is important about mutations in the transit-amplifying cells?
These cells are already scheduled to die at some point, likely won't persist long
enough to become neoplastic
Mutations can be caused by ______ and _____ sources
Exogenous and endogenous
What are some exogenous sources of DNA mutations?
Mutagenic agents (chemical & physical)
What are some endgenous sources of DNA mutations? (3)
1) DNA replication by DNA polymerase during S phase
2) Spontaneous chemical alternations of nucleotide precursors that become
embedded into the DNA
3)Mutagenic agents
Damage from endogenous sources much ______ per day than amount of damage
caused by exogenous mutagenic agents in most tissues
Higher!
What do defenses do cells have against mutagens? (2)
Physical and chemical
What physical defenses do cells have against mutagens?
,Preventing penetration of mutagen into tissue
e.g. Melanin or keratinocytes
What are some chemical defenses against DNA damage? (5)
- ROS mitigation
- GST-mediated detoxification of carcinogens (linked to glutathione)
- Mismatched repair
- Dealkylating repair enzymes
- Excision repair enzymes (BER/ NER)
What are repairs to fix specific lesions in DNA? (3)
1. Base Excision
2. Nucleotide Excision
3. Error prone
What is base excision repair (BER)?
Repairs lesions caused by endogenous sources (ROS or depurination)
How does base excision repair work? (5)
1) DNA glycosylases recognizes abnormal bases
2) The covalent bond to deoxyribose gets cleaved (has trouble recognizing T
because it's in normal DNA)
3) AP endonuclease cleaves
Deoxyribosephosphate
4) DNA polymerase adds individual nucleotide or longer stretch of nucleotides
5) DNA ligase seals the repaired DNA
What is nucleotide excision repair (NER)?
Repairs lesions from exogenous sources by recognizing abnormal bases and
cleaving DNA around the damage.
How does nucleotide excision repair work? (4)
1) large protein complex recognizes the abnormal base
2) cleaves the DNA fragment around the helix distortion
3)Polymerase fills in the gap with DNA replication machinery (PCNA/RPA)
4) DNA ligase seals up repaired DNA
How are nucleotide excision repair and p53 related?
P53 can activate nucleotide excision repair to facilitate global repair of the
genome.
What type of polymerases are used for error prone repair?
High-fidelity polymerases with low error rates
What is error-prone repair?
A mechanism that attempts to repair DNA lesions under non-ideal
circumstances using error-prone polymerases.
Why are error prone polymerases used compared to high fidelity polymerases?
,Can add nucleotides to a growing DNA strand even when the base on
complementary strand is missing or DNA adduct is attached
Why is error-prone repair a gamble cell?
There's a risk incurring more mutations with error prone repair vs cell death due
to failed DNA replication
Inherited Defects in __________ Cause Susceptibility To Specific Cancer Types
DNA Repair Pathways
What are some inherited defects that can cause suspectibility to cancer? (3)
1) Xeroderma pigmentosum (XP)
2) Hereditary non-polyposis colon cancer (HNPCC)
3) BRCA1/2
What type of repair does XP use?
NER
What condition is characterized by extreme sensitivity to UV radiation?
Xeroderma pigmentosum (XP)
What are the characteristics of XP?
Infants burn with minimal exposure, many freckles, parchment like skin
What is the risk of skin cancer for individuals with Xeroderma pigmentosum
before age 20?
2000-fold increased risk compared to the general population.
At what age do XP patients typically present with skin cancer?
Around 10 years old, compared to around 60 years old in the general population.
What causes Xeroderma pigmentosum?
Inherited defects in one of eight genes (7 are NER, 1 is error-prone polymerase)
XP individuals are at relatively no increased risk for cancers of internal organs --
why is this important?
Suggests that UV radiation has been a significant mutagen for most of human
evolution, in response made a DNA repair system to fix damage caused by this is
singular mutation
Hereditary non-polyposis colon cancer (HNPCC) is a
Familial cancer syndrome
What is the lifetime risk of developing colon cancer for individuals with hereditary
non-polyposis colon cancer (HNPCC)?
80% lifetime risk
What types of cancers are individuals with HNPCC at increased risk for? (5)
Brain, endometrial, stomach, ovarian, and urinary tract carcinomas.
What is the mechanism of developing HNPCC? (5)
Loss of APC --> DNA hypomethylation --> activation of K-RAS --> loss of 18q TSG
--> loss of p53
How quickly can adenomas progress to carcinomas in HNPCC patients?
, In 2-3 years, compared to 8-10 years for other types of colon cancer
What genetic defects are associated with hereditary non-polyposis colon cancer?
Mismatch repair (MMR) genes
What happens to mismatch repair genes in HNPCC?
Inherit one defective allele, undergo loss of heterozygosity (LOS) to lose
remaining WT copy of allele
Why is the loss of heterozygity in MMR important?
High mutation rates in mircosatellite repeats of genes on TGF-B --> causing
unchecked cell cycles
What is important about TGF-β receptor and mutations?
If TGF-β signaling is lost, would be advantageous for cell that
wants to undergo unchecked cell cycles
What is the role of BRCA1 and BRCA2 genes?
They help maintain genomic integrity through DNA repair
How do BRCA1/BRCA 2 maintain genomic integirty?
DNA repair proteins!
Large complexes form with other proteins in the nucleus to repair ionizing
radiation
What is the role of BRCA1 and BRCA2 genes thought to be previously?
TSG -- regulating proliferation, survivial and differnation
What is the risk of developing breast cancer for carriers of mutant BRCA1 or
BRCA2 by age 70?
50-70% risk.
What type of DNA repair do BRCA1 and BRCA2 assist with?
Homology-directed DNA repair of double-strand breaks.
What do dsdna breaks require and how? (2)
Repair machinery to access undamaged sequence on sister chromatid
1) sister chromatid is the template
2) unwinds sister chromatid
What happens to homologous repair in cells lacking BRCA1 and/or BRCA2
function?
It is compromised, leading to inability to repair double-strand DNA breaks.
If BRCA1/2 are so important to homologous repair of dsdna breaks,why don't
more cancers have defects in these genes?
We don't know, still under investigation
BRCA2 somatic mutations can be associated with these carincomas outside of
breast and ovarian cancer? (2)
Colon and prostate
