Flashcards in Molecular Basis of Carcinogenesis (Exam 3) Deck (37)
Name some of the properties of a malignant, or transformed, cell (11 total)
1. Loss of contact inhibition/high saturation density
2. Immortalization (avoid apoptosis)
3. Ability to grow w/o attachment to solid substrate
4. Reduced requirement for mitogenic growth factors (constitutive activation of pathways) and inability to halt cell cycle in the absence of growth factors
5. Altered morphology (little/no cytoplasm; rounded)
6. Invasive, that is capable of outgrowth into neighboring normal tissues to extend the boundaries of the tumor.
7. Metastatic, that is capable of shedding cells that can drift through the circulatory system and proliferate at other sites in the body.
8. Increased transport of glucose (lower O2 requirement--ferment sugar in place of oxidative phosphorylation)
9. De-differentiated, that is, lack many of the specialized structures and functions of the tissue in which they grow.
10. Sustained angiogenesis
11. Clonal in origin (derived from a single cell)
What differentiates a benign tumor from a malignant one?
Benign tumors have lost cell controls (are immortal), but are not invasive or metastatic.
What are two critical types of genes in which mutations are likely to cause cancer?
1. Oncogenes, which normally stimulate cellular proliferation (analogous to the “gas pedal” of your car), are activated.
2. Anti-oncogenes or tumor suppressors, which normally inhibit cellular proliferation (analogous to the “brake pedal” of your car), are inactivated.
What is the Knudson theory, and how does this apply to Loss of Heterozygosity? What is LOH?
The Knudson "two hit" theory is that two mutations must occur to knock out both copies of a gene. Refers to a tumor suppressor like Rb. This is why sporadic retinoblastoma is so much less less common than hereditary; two mutations in the same spot is much more rare, and takes longer. People born with a mutation are at a much higher risk because only one recombination event is required to turn off the brakes. LOH occurs as an "inappropriate" crossing over during mitosis between non-homologous chromosomes. Odds are 50% that heterozygosity is maintained, 50% that one cell gets both mutations and one gets none, in which case heterozygosity is lost.
Examples of cancers that are inherited as autosomal dominant disorders are (4):
1. Familial Adenomatous Polyposis (FAP-APC gene)
2. Familial Retinoblastoma (RB gene)
3. Familial Breast and Ovarian Cancer (BRCA1 and BRCA2 genes)
4. Wilms tumor syndromes
Examples of cancers that are inherited as autosomal recessive disorders are (4):
1. Xeroderma pigmentosa (XP genes)
2. Ataxia-telangiectasia (AT gene)
3. Bloom’s syndrome
4. Fanconi’s congenital aplastic anemia (FA genes)
This is the hallmark of a antioncogene or tumor suppressor gene.
In cases of "inherited" retinoblastoma (i.e. when there was a parent and other family members who also had the disease), the DNA from normal tissue of the patient or from other unaffected family members often shows a defect in the retinoblastoma gene, but has one normal copy of the gene per cell.
+++++++ In these patients it appears that normal, nonmalignant retinal cells, are heterozygous for the retinoblastoma gene, but the tumor cells have descended as a clone from a single cell that has acquired homozygosity for the retinoblastoma susceptibility gene. ++++
The Rb protein is ______ in rapidly proliferating cells at ____ of the cell cycle, but is ____ in non-proliferating cells in ____ of the cell cycle. ______ are responsible for phosphorylating Rb, allowing it to proceed from G1 to S phase.
hyperphosphorylated; S or G2; hypophosphorylated; G0 or G1; CDK (Cyclin Dependent Kinases);
HPV encodes two proteins, ____ which targets Rb, and ___ which targets p53.
____ encodes a T antigen (Large T) that targets ____ and ____.
SV40; Rb and p53
HeLa cells were isolated from a cervical carcinoma and have been growing in culture for over 60 years. These cells express HPV _____
E7 and E6 protein (E6 inhibits p53, another important tumor suppressor). If E7 and E6 expression is blocked, the cells return to normal phenotype. This bodes well for therapy as affecting just two proteins can have a drastic effect.
Kaposi Sarcoma encodes a ____, which binds ___ to push cells into S phase. Often seen in patients who have ____.
cyclin; CDK; AIDS
Rb appears to have a dominant inheritance pattern. We know, however, that a cell must be homozygous for the mutation to become tumorigenic. Explain.
What is inherited in a dominant fashion is the susceptibility to retinoblastoma. People who are heterozygotes for retinoblastoma have only one normal RB gene in each cell of their body including the cells of the retina. These cells will regulate their proliferation normally and will be non-malignant. However, loss of the single, normal RB gene by any number of events will produce a tumor. Thus if one cell among the millions of retinal cells has no RB protein, it will lose the ability to regulate its proliferation, grow out of control, thereby generating a clone of cells, which will become a malignant tumor. Thus people who are heterozygotes for the RB gene are likely to develop the disease and will pass on the defective gene to 1/2 of their children, so it appears to be autosomal dominant in its inheritance.
Persons who survive inherited retinoblastoma have an increased risk for developing a second neoplasm, which is typically mesenchymal in origin, for example,______. Cells of these tumors are also defective in RB function.
Lets go over CML one more time. Called the ______ chromosome, this involves a translocation between chromosomes ___ and ____. This creates a novel gene product called the _____, which is a ______. This protein is constitutively activated, and inhibits p27, an inhibitor of the cell cycle. The drug for this is _____ aka _____.
Philadelphia; 9/22; Bcr-Abl; protein kinase; Imantinib; Gleevec. (We conclude that Bcr-Abl promotes cell cycle progression and activation of cyclin-dependent kinases by interfering with the regulation of the cell cycle inhibitory protein p27.)
What is FAP? What gene encodes it? This disease appears to have autosomal dominant inheritance, but is actaully another example of _____.
FAP (Familial Adenomatous Polyposis); Encoded by the APC gene; Loss of heterozygosity.
[Like RB, FAP is inherited in an autosomal dominant fashion, in which patients that inherit one defective APC gene will be at higher risk (90% will develop colon cancer by age 50) to develop colon cancer. Cancer develops when the wild-type gene is lost by LOH in cells in adenomatous polyps of the colon during the first 20 years. Thus, these benign adenomatous polyps may become malignant by LOH.]
Describe the Wnt signaling pathway and how a mutation in the gene encoding APC disrupts normal function to produce cancer.
WNT binds Frizzled, APC complex is bound, which frees Beta-catenin to relocate to nucleus and transcribe c-myc. W/o APC, Beta-catenin is perpetually in the nucleus.
From lecture notes: The APC gene encodes a cytoplasmic protein that regulates the localization of the Beta-catenin protein. Beta-catenin is kept at the plasma membrane by being bound to E-cadherin in normal cells. The APC protein causes the degradation of any unbound and free Beta-catenin in the cytoplasm. When the APC is lost in FAP patients, Beta-catenin goes to the nucleus to produce transcription of oncogenes like c-myc. Thus, loss of APC tumor suppressor causes an overexpression of the c-myc oncogene, resulting in cancer!
In what two ways does p54 suppress tumorgenesis?
p53 restrains tumor formation by two different mechanisms. In the first, p53activates the p21 Cdk inhibitor gene in response to DNA damage and stress. Loss of p53in cells prevents the p21 gene from being transcribed, leading to the increased activity of the multiple Cdks normally turned off by p21 and resulting in increased cell proliferation.
A second way in which p53 restrains tumor formation is by inducing apoptosis.
Why are mutations in BRCA1/BRCA2 a big deal?
BRCA1 and BRCA2 function as “caretaker” genes, like p53, which serve to maintain genomic integrity. The gene products encoded by BRCA1 and BRCA2 are nuclear proteins that co-localize with RAD-51 at sites of DNA damage, and play a role in:
1. homologous recombination repair of double-stranded breaks
2. There is also evidence that BRCA1 and BRCA2 interact with the p53-mediated DNA damage checkpoint
3. Loss of BRCA1 or BRCA2 leads to the accumulation of other genetic defects, which can then lead to cancer formation.
In addition to their roles in DNA repair, BRCA1 and BRCA2 have been implicated in a variety of cellular processes, including DNA synthesis, regulation of gene transcription (similar to p53, one target of BRCA1 transcriptional activation is the Cdk inhibitor p21), cell cycle checkpoint control, centrosome duplication and ubiquitination.
Inherited mutations in BRCA1/BRCA2 display what inheritance pattern?
The p53 gene is a very, important cancer predisposing gene because mutant p53 is found in about _____% of all cancers!
The p53 gene has several important properties, which explain why it is one of the most important genes in human cancer (3):
1. p53 protein acts as a transcription factor important for the expression of genes, which prevent cells from replicating damaged or foreign DNA. p53 is also required for apoptosis, in which cells commit suicide if their DNA is damaged beyond repair. In p53 defective cells, damaged DNA is replicated, thereby producing additional mutations including chromosomal rearrangements, which can lead to cancer. In this manner, p53 acts as a “guardian of the genome”.
2. Certain p53 point mutations are found more frequently than others in human cancers. These mutations are called “hotspots”. For example, mutational “hotspots” produce alterations in amino acids 248 or 273 of p53 in all human cancers. Some “hotspots” are unique to specific cancers. For example, an alteration in amino-acid 157 of p53 is found mainly in lung cancer and is the result of the mutagenic chemicals found in cigarette smoke
3. p53 interferes with some viral infections. Adenovirus and HPV have oncogenes that act by inactivating p53, (for example, Adenovirus E1B and HPV E6 proteins). Remember that these viruses also inactivate RB protein. In fact, destruction of both RB and p53 either by cellular mutations or viruses is a major route to cancer.
Why was p53 originally thought to be an oncogene?
One mutant subunit (of tetramer) will inactivate 3wt subunits. Tumors then, have both wt and mutant p53, and it was thought that the presence of wt indicated that p53 was an oncogene (remember that Rb was the model, and 100% of Rb in a tumor will be mutant b/c of clonal nature of cells.) The mutant p53 protein is MORE stable than wt, so is virtually impossible to get wt/wt/wt/wt.
Definition of a proto-oncogene (aka c-onc):
A host gene that is homologous to an oncogene that is found in a virus but which can induce transformation only after being altered (such as mutation or a change of context such as coming under the control of a highly active promotor). It usually encodes a protein that functions in DNA replication or growth control at some stage of the normal development of the organism.
[Once retroviral oncogenes had been discovered, a surprising observation was made: Unlike the situation with DNA virus oncogenes which are true viral genes, there are homologs of all retrovirus oncogenes in cells that are not infected by a retrovirus. These cellular homologs are often genes involved in growth control and development/differentiation (as might be expected)]
[Note that c-oncs are not identical to their corresponding v-oncs. It appears that the virus has picked up a cellular growth controlling or differentiation gene and, after the gene was acquired by the virus, it has been subject to mutation.]
Characteristics of cellular proto-oncogenes
1) These are typical cellular genes with typical control sequences. As with most eucaryotic genes, most have introns (while retroviral oncogenes - v-oncs - do not)
2) They show normal Mendelian inheritance because they are normal genes, essential to the functions of the cell.
3) As with all genes in the eukaryotic genome, they are always at same place in genome (cf. what would be expected of endogenous retroviruses that had, over time, become incorporated into the cellular genome)
4) There are no LTR sequences (v-oncs always are in an LTR context)
5) Viral oncogenes are most like the c-onc of the animal from which the virus is thought to have acquired the gene. Thus, v-src of RSV is more like chicken src than human src. Note that v-onc was long ago acquired accidentally by the virus from the genome of a previous host cell
6) Cellular oncogenes are expressed by the cell at some period in the life of the cell, often when the cell is growing, replicating and differentiating normally. They are usually proteins that are involved in growth control.
7) Cellular oncogenes are highly conserved
If v-onc and c-onc are so alike, why does the v-onc introduced by a virus cause havoc in the cell?
This is due to differences in the genes, mutations that have occurred in the gene once it was picked up by the virus. Such changes include:
1. Amino acid substitutions or deletions which result in altered translation products
2. Many v-onc proteins are fusion proteins translated from a v-onc that is a hybrid gene of a c-onc and a viral gene.
3. V-oncs are inserted into the host genome along with LTRs which contain promotors/enhancers. This is likely to result in over expression of a gene that we know is probably involved in control of DNA transcription and replication!
We find an _____ in the viral genome of many retroviruses that transform cells to neoplasia. It should be emphasized that the oncogene in RNA tumor viruses is _____ (necessasy/unnecessary) for viral replication. It is an additional gene that gives the virus its capacity to transform the host cell.
Definition of virally-induced transformation:
ONCOGENE (onc); not necessary
Definition: Changes in a cell that result from integration of viral genetic sequences into the cellular genome that confer on the infected cell certain properties of neoplasia.
Rous sarcoma virus (RSV): added gene was called src (for sarcoma). src is not needed for viral replication. It is an extra gene to those (gag/pol/env) necessary for the continued reproduction of the virus. Deletions/mutations in src ___
abolish transformation and tumor promotion but the virus is still capable of other functions. RSV is unusual in that it has managed to retain its whole genome of gag/pol/env.
[many oncogenic viruses have lost genes in substituting cellular dna, and cannot complete replication without a helper virus]
The retroviral RNA genome consists of two identical strands held together by a tRNA molecule. The ___ gene codes for internal virion proteins, the ___ gene for virus membrane glycoproteins, and the __ gene for a virus polymerase
gag; env; pol
[When these genes are the only ones present the virus does not cause tumors, but replicates through an intermediate proviral DNA and integrates into the host cell genome. It can be transmitted as an integrated provirus through somatic or sex cells as a cellular gene. ]
Retroviruses that contain a ___- segment also have the ability to rapidly transform appropriate infected cells to the malignant phenotype. They therefore rapidly induce tumors after infection.
Retroviruses with v-onc genes (examples, all animals for some damn reason)
v-src - The oncogene of Rous Sarcoma Virus. This caused fibrosarcomas in certain birds.
v-erb - The oncogene of avian erythroblastosis virus causes erythroblastosis in chickens. Two related genes, erb-A and erb-B have been identified.
v-ab - The oncogene found in Abelson leukemia virus from mice.
v-myc - This gene is usually fused with a portion of the gag gene. It appears that this gene is capable of eliciting neoplastic transformation of cells.
How do viral oncogenes work?
The products of oncogenes mimic hormones or growth stimulating factors either by
1. resembling natural hormones or
2. by affecting the structure of the cell surface receptors for these hormones. These altered receptors then send signals to the cell nucleus in an unregulated manner to affect growth.
When and why do we care about n-myc?
N-myc is a gene expressed in neuroblastomas. The level of expression of N-myc is a very important diagnostic factor. Patients with 10 copies, 90% are dead within 1.5 years.
Are c-onc genes that cause spontaneous malignancies involved in any way with retroviruses?
No. The proto-onc or c-onc genes that are involved in spontaneous malignancies that have nothing to do with a retrovirus. Thus, a change in gene regulation or gene structure due to a cellular mutation could activate these genes to cause a malignancy. Either quantitative changes (too much protein) or qualitative changes (overactive or unregulated protein) in the proto-oncogene are responsible for these effects.
Analysis of DNA from human bladder cancer cells has shown that c-ras genes have been point mutated. These mutations produce a ras protein that is unregulated and is always “on”. Detection of these ras mutations indicates a ____ prognosis.
The HER2/neu oncogene (also called erbB2), which encodes a integral membrane protein kinase (see v-erb-B) is amplified in about 20% of breast cancers. Higher levels of amplification correlate with a____ prognosis.