molecular biology of neoplasia I - lecture notes - julia

Question 1

what is the somatic mutation hypothesis of cancer?

Answer 1

• cell growth, differentiation, and survival are under genetic control
• malignant transformation is a result of accumulated mutations in specific classes of genes
• tumor mass results from the clonal expansion of a single progenitor cell that has incurred the genetic damage
• genetic damage is in the aspets of cell growth that are under genetic control

Question 2

what are the three classes of genes involved in cancer? (list)

Answer 2

1. growth promoters
2. growth suppressors
3. caretakers

Question 3

what are growth promoters?

Answer 3

• one of the three classes of genes involved in cancer
• typical mutations activate the encoded protein
• strucutral mutation in primary amino acid sequence results in increased expression levels

Question 4

what are growth suppressors?

Answer 4

• one of the classes of genes involved in cancer
• typical mutations inactivate the encoded protein
• a structural mutation of the primary amino acid sequence increases expression levels

Question 5

what are caretaker genes?

Answer 5

• one of the types of genes involved in cancer
• generally ensure the stability of the gene (ie proteins involved in gene repair)
• neither promote nor inhibit
• when repair genes are inactivated, likelihood of mutation in oncogene or tumor suppressor gene increases

Question 6

what are the six classic phenotypic hallmarks of cancer? (classic because two more have been added since)

Answer 6

1. dysregulation of cell proliferation by constitutive activation of growth-stimulatory pathways or independence of proliferation signals - via alterations to oncogenes
2. insensitivity to growth inhibitory signals or loss of growth inhibition pathways - via alterations to tumor suppressor genes
3. evasion of apoptosis
4. limitless replicative potential - cells normally have a limited number of replications that they can undergo - tumor cells must get around that
5. angiogenesis - to allow for the tumor to get nutrients to grow
6. invasion and metastasis

Question 7

what are “landscaper genes”?

Answer 7

• required for the malignant phenotype
• “looking outward”
• code for proteins involved in angiogenesis, cell-cell and cell-matrix adhesion, and proteolytic enzymes required for invasion

Question 8

what is protoconcogene versus an oncogene?

Answer 8

• protooncogene = gene that encodes a protein that mediates or stimulates cell proliferation
• oncogene = inappropriately activated protooncogene, either by mutation or aberrant expression (over-expression or ectopic expression)

Question 9

what are the types of proteins encoded by oncogenes? (8) give some examples of each type

Answer 9

1. cell surface receptors
   - such as PDGF receptor, EGF receptor, M-CSF receptor
   - these are often growth factor receptors actibng via tyrosine specific protein kinase activity
2. GTP-binding proteins
   - membrane associated second messengers
   - like ras proteins
3. membrane/cytoskelton tyrosine-specific protein kinases
   - such as src
4. cytoplasmic tyrosine-specific protein kinases
   - transmit signals from the membrane associated protooncogenes
   - such as fes
5. steroid-type growth factor receptors
   - such as thyroid hormone receptor (erb-4)
6. serine/threonine specific protein kinases
   - such as raf
7. growth factors
   - extracellular - act on membrane receptors
   - such as EGF, PDGF, M-CSF
8. nuclear proteins
   - myc, fox, jun

Question 10

give an example of how mutations in genes for growth factors can lead to neoplasm

Answer 10

• c-sis encodes platelet derived growth factor (PDGF)
• v-sis is the oncogenic form (c-sis is therefore a protooncogene)
• v-sis can be picked up by a virus during transformation => transforming virus
• they didn’t explain how the mutated PDGF affects cell growth differently…

Question 11

how can mutations in the genes for growth factor receptors allow for neoplasm?

Answer 11

• in transmembrane receptor tyrosine kinases
• oncogenic versions of them are overactive
• most often the EGF-R family involved
• c-erbB1 gene = oncogene => overexpression - this is found in 80% of squamous cell carcinomas of the lung
• c-erbB2 gene = oncogene for HER2-neu receptor = estrogen receptor - amplified in some breast, ovary, lung, stomach cancers

Question 12

what signal transduction molecules can be involved in neoplasm development? (3)

Answer 12

• non-receptor protein tyrosine kinases such as src
• cytoplasmic serine/theronin kinases such as raf
• gtp-binding proteins such as ras

Question 13

how are gtp-binding proteins involved in development of neoplasm? what is an example of a commonly altered gene?

Answer 13

• play a critical role in signal transduction from RTKs
• mutation in ras gene is most common abnormality of dormant oncogenes in human tumors
• Ki-ras is involved in lung, ovarian, colon and pancreatic cancers
• N-ras is involved in leukemias

Question 14

what is the difference between qualitiative and quantitative activation of proto-oncogenes?

Answer 14

• both are result of dominant, gain-of-function mutations
• qualitative => changes in structure of gene (loss of regulatory elements) => abnormal gene product (oncoprotein) => aberrant function
• quantitative => upregulation or ectopic expression of a structurally normal growth-promoting protein
• example: breast cancer cells often produce excess cyclin D and E

Question 15

review: what is the pathway of ras signaling?

Answer 15

1. growth factor binds to growth factor receptor => activation
2. activates adaptor proteins
3. ras undergoes GDP-GTP exchange
4. activates raf
5. activates MEK
6. activates MAP-kinase
7. activates SRF (transcription factor)

ras is inactivated by an intrinsic GTPase activity

Question 16

how is ras signaling different when ras is mutated?

Answer 16

• normally, ras signal is self-limited by an intrinsic GTPase activity (hydrolysis of GTP)
• if mutant, this process is blocked - ras remains active => continuous activation of map kinase pathway

Question 17

what are common mutations seen in ras?

Answer 17

• all ones that diminish its GTPase activity
• point mutations
• c-K-ras codon 12 often has a GGC –> GTC => gly –> val
• this inactivates the inherent GTPase activity of ras
• commonly seen in pancreatic and colon carcinomas

Question 18

what are the mechanisms of oncogene activation? (2)

Answer 18

1. gene amplification (normal protein, but greatly overproduced)
2. chromosome rearrangement
   - can have a nearby strong enhancer that causes the normal protein to be overproduced
   - can fusion of an actively transcribed gene to another gene that results in overproduction of the otehr gene (CML)

Question 19

how is HER2-neu involved in cancer? (what does it correlate with? how is it used in treatement?)

Answer 19

• amplified in 20-25% of breast carcinomas
• correlates with presence of nodal metastasis and high grade
• also correlates with short survival in node positive patients
• gene that encodes it = c-erbB2 (the protein is also sometimes called ErbB-2…confusing)
• treatment = antibody directed against extracellular domain of HER2-neu = specific antagonist of ligand-growth factor receptor interaction = first drug developed against a specific oncogene to achieve FAD approval

Question 20

how does the drug gleevec work?

Answer 20

• for CML
• in CML, ATP must bind to the BCR/ABL protein so that the protein can add tyrosine to the substrate
• gleevec (aka sti571) binds intot he ATP binding site, preventing ATP from binding and thereby inactivating BCR/ABL

Question 21

what are tumor suppressor genes?

Answer 21

• genes that inhibit cell proliferation
• inactivation or loss therefore leads to tumor development by eliminating negative regulator signals

Question 22

what is the two-hit hypothesis? how does it relate to the differences in heritable and non-heritable cancers?

Answer 22

• says that, for cancer to develop, have to get deletion or mutation of both alleles of a specific gene
• developed to explain retinoblastoma, which is highly heritable, whereas only about 1 in 30,000 normal people develop tumor
• inheritane of a single mutant gene greatly increases risk of developing tumor in an autosomal dominant fashion
• argues that those with hereditary Rb already have one mutation in one allele of their Rb gene (this is the first hit) and then they only have to develop the “second hit” = mutation in remaining Rb gene
• however, those without the hereditary Rb have to get two hits which is really unlikely

Question 23

what is loss of heterozygosity? how is it detected?

Answer 23

• loss of one of the two alleles for a gene
• detected with PCR
• normally DNA sequences are slightly different in various regions
• if one copy of the gene is deleted, there will only be one band on the PCR
• prevalence differs at different positions within the genome - more prevalent at certain “hot spots”
• common in tumor tissue

Question 24

what are the possible ways to lose heterozygosity? (6)

Answer 24

recessive mutation since both copies need to be eliminated

1. nondisjunction
2. nondisjunction and duplication
3. mitotic recombination
4. gene conversion
5. deletion
6. point mutation

Question 25

what is the role of the Rb protein? how is it involved in cancer development?

Answer 25

• primary regulatory protein of the G1/S phase transition
• inhibits passage through the restriction point by repressing transcription of a number of genes involved in cell cycle progression and DNA synthesis
• mutation inactivates it => removal of negative regulator of cell cycle progression

Question 26

how are cyclin-dependent kinase inhibitors involved in cancer?

Answer 26

• central to malignant transformation - at least on of the four key regulators of cell cycle is altered in most human malignancies
• these 4 are:
• p16
• cyclin D
• CDK4
• pRb

Question 27

what is p53? what is the consequence of the loss of p53?

Answer 27

• transcription factor responsible for cell cycle arrest and indcution of apoptosis upon DNA damage
• loss results in increased mutation frequencies and general genome instability
• involved in up to 50% of cancers
• dominant negative mutation (where mutation in one allele prevents function of the other allele)

Question 28

review: what is the pathway by which p53 acts?

Answer 28

• dna damage => induction of p53 levels and transactivation ability (due to posttranslational modifications)
• activates transcription of CDK-inhibitor p21
• p21 blocks cell cycle progression in the G1 and G2 phases
• this allows time for the damaged DNA to be fixed
• if the damage can’t be repaired, it triggers apoptosis by activating transcription of pro-apoptotic genes (bax)

Question 29

how does HPV cause cancer? (what do the virally encoded proteins do?)

Answer 29

• E6 binds to and promotes degradation of p53
• E7 binds pRb and displaces E2F transcription factors

Question 30

what is xeroderma pigmentosum? what causes it?

Answer 30

• sensitivity to sunlight with the development of carcinomas at an early age
• normal skin fibroblasts can repair UV damage to DNA by inserting new bases
• patients have defective excision of pyrimidine dimers - can’t repair damage
• example of a problem with a caretaker gene

Question 31

what is heriditary non-polyposis colon cancer syndrome? what causes it? how is it identified?

Answer 31

• family carcinomas of the colon affecting predominantly the cecum and proximal colon
• due to mutant mismatch repair genes (hMSH2, hMLH1, hPMS2, hPMS2)
• results in decreased capacity to correct errors made during DNA replication, esp errors in repetitive nucleotide sequences
• results in replication error (can be detected by microsatellite instability = tandem repeats of one ot six nucleotides scattered through the genome - same in every tissue and fixed for life - but get contractions and expansions of these repeats in tumor)