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Flashcards in CANCERRRRR Deck (33):

. Describe at least five different properties of malignant cancer cells

Phenotype properties:
Unresponsive to normal signals for proliferation control.
De-differentiated (lack specialized structures of tissues in which they grow).
Invasive (can grow out into neighboring tissues).
Metastatic (can shed into circulation and proliferate elsewhere in the body).
Generally it's the metastasis that's fatal in cancers.
Clonal origin (derived from a single cell).
Increased transport of glucose.
Lack of contact inhibition (will grow over each other).
Immortality (ability to grow indefinitely).
Can grow without an attachment to a solid substrate.


2 . Describe the multi‐step process for carcinogenesis, and discuss the relative importance of heredity and
the environment and why early events may include mutations in DNA repair genes. Hint 6 steps.

Not considered to be an inherited disease (not inherited as a single Mendelian gene).
On the other hand, cancer susceptibility genes are certainly heritable (see Knudson below).
Carcinogenesis is characterized by the accumulation of many genetic alterations or mutations, particularly over a long period of time-- thus age is strongly associated with cancer, as are environmental factors that produce high rates of mutations.
If DNA repair genes are damaged early on, the rate at which you accumulate DNA mutations - since you can't repair them as well - goes up markedly.
Multi-step process for cancer:
(1) Normal cell
(2) Increased proliferation: With a mutation or two, an immortalized cell (see below for examples).
(3 + 4) Early/progressive neoplasia: With a few more mutations, get abnormal growth patterns.
(5) Carcinoma: A full-on tumor.
(6) Metastasis: A tumor that's spreading through the circulatory system.
I think the point he's trying to make here is that you need a fairly wide assortment of mutations (though order isn't important) to result in a cancer:
Turn on oncogene or make oncogene protein much more active
Turn off tumor suppressor genes (both cell cycle regulatory and DNA repair genes)
Turn off apoptotic genes and turn on anti-apoptotic genes.


What types of genes are usually mutated in tumor initiation? Describe the effect on cellular proliferation that the product of these genes has

Either activated oncogenes (proliferation genes) or silenced tumor suppressor genes.
Oncogenes: accelerate proliferation. Tumor suppressors: slow down proliferation.


What type of cytogenetic abnormalities are associated with malignancy? You should be able to give at least two different examples

Translocations of chromosomes, deletions on chromosomes:
Can activate oncogenes (for example, by putting an extremely active promoter upstream of one).
Can inactivate tumor suppressor genes (for example, by translocating another gene into the middle of the tumor suppressor sequence)
Notice that this is kind of a silly point to make, at least on its face. Anything that can bring a promoter nearby an oncogene (like pretty much any chromosomal rearrangement) or anything that can interrupt transcription or promotion of a tumor suppressor gene (likewise) can be associated with malignancy.


Give at least two examples of events that can produce loss of heterozygosity and how they support
Knudson’s theory.

LOH = Loss Of Heterozygosity.
Means you inherit a heterozygous state (say, for a working tumor suppressor gene), but convert to (negative) homozygosity at some point.
Loss of this heterozygosity means you lose the one working copy of the gene that you have, through:
-mitotic recombination
-chromosome loss
-and/or environmental factors


What is knudsons theory

If you're heterozygous, you have one "strike" against you through your genes (one copy of suppressor gene knocked out in your parents' passed-on DNA). If you have one more "strike" (ie, exposure to UV light causes an unrepaired mutation in the other copy of that gene), you're unable to produce that tumor suppressor gene product at all-- which leads, potentially, to cancer.
Example: Familial retinoblastoma vs acquired retinoblastoma (in children vs. adults, bilateral vs. unilateral)-- former is easier to acquire since it only requires one mutation event.


Are cancers associated with both dominant and recessive syndromes? You should be able to give a different example of each type.

Familial retinoblastoma: A recessive disease but inherited in a dominant way: need both tumor suppressor (RB) genes knocked out to show a phenotype (thus recessive), but everyone who inherits heterozygosity winds up with the disease (thus dominant) due to LOH problems.
Notice that this inheritance pattern shows a vertical pedigree (looks like autosomal dominant).
Sporadic retinoblastoma: Mutations in both tumor suppressor genes needed to knock out function- thus not strongly inherited (not sure how he's tying this to the LO, as somatic sporadic mutations shouldn't be inherited at all)


Describe how the RB (retinoblastoma) gene was first identified. You should be able to describe the important cytogenetic and molecular evidence.

Cytogenetic analysis of cells from retinoblastomas showed that the region around
chromosome 13q14 often had an abnormal structure.


What are the properties of the protein product of the RB gene? List at least three biochemical properties

[Notice that the RB protein is a universal protein-- not just found in the retina.]
An inhibitor of the cell cycle that prevents proliferation.
Normally hypophosphorylated (little PO4) to prevent proliferation.
Hyperphosphorylated by CDKs (cyclin-dependent kinases) to be turned off.
When turned off, allows normal cell proliferation.
When kept off or inhibited, allows unchecked cell proliferation and carcinogenesis.
General note: RB protein works by binding to a variety of transcription factors


Describe how the RB protein functions during the cell cycle and why it is important in cancer. You should be able to give an explanation of how the loss of RB may produce a malignancy

As mentioned, functions to block G1 moving to S (replication) phase. Without RB, a cell has no 'brake' on its proliferation.
Notice that there are certain tissues that are particularly susceptible to the loss of RB-- that is, losing RB there has a particularly acute effect. Example is, obviously, the retina.


What is the hallmark of a tumor suppressor gene or anti-oncogene? You should be able to use the RB gene as an example

Prevents cells from proliferating by controlling cell cycle.


How were oncogenes discovered? You should be able to describe the method with at least three different examples

Discovered in oncogenic retroviruses (specifically Rous Sarcoma Virus in chickens). With one particular viral gene segment (v-onc), tumors are rapidly induced in the infected cells after infection; without it, integration into the host genome occurs without activation of oncogenes.
Examples: v-src, v-erb, v-myc, etc. Watch for the v- at the beginning of it. (as opposed to c-myc, which is an endogenous oncogene in the human genome.)
Often oncogenes mimic growth factor receptors to achieve their nefarious ends.
Method: take cells, put them in agar, watch for proliferation. Normal cells won't be able to proliferate (no anchorage to grow on)-- infected cells will proliferate regardless of anchorage (see characteristics of cancer cells, above).


What functions do the protein products of viral oncogenes perform? You should be able to give at least four examples of oncogenes of known function

RB protein is a target of some tumor viruses (eg. human papilloma virus), which produce proteins which inactivate RB (and/or p53) in the cell in which the virus has taken up residence.
This is a common theme: RB and/or p53 inactivation by viral proteins allowing rapid, unchecked cell proliferation.
Notice Kaposi's sarcoma [HIV/AIDS] is also caused by RB/p53-inactivating proteins.
Notice also that retroviral-induced cancers (ie, resultant from viral reverse-transcription of their oncogenes into human DNA) are very rare in humans. Our viruses tend to just inactivate p53 and RB rather than encode oncogenes themselves



As mentioned, the viral src, erb, myc, etc, sequences. Notice that viral copies of oncogenes tend to be more powerful effects than their endogenous counterparts.
v-src: phosphorylates various tyrosine residues in other proteins (similar to ABL in humans).
v-erb: mimics epidermal growth factor receptor (unregulated).
v-sis: mimics platelet-derived growth factor (unregulated).
Endogenous oncogenes are marked c-onc:
Notice that c-onc genes are part of normal functioning of human cells; therapy can't target all c-onc, just their overexpression.
c-onc genes need to undergo mutation before they become carcinogenic.



APC gene product: keeps beta-catenin outside the nucleus; without APC, beta-catenin goes to nucleus and begins uncontrolled transcription of oncogenes (like c-myc).
LOH in APC produces familial adenomatous polyposis (FAP), which leads to colon polyps and, eventually, metastatic colon cancer.
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 Ecadherin
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!


BRCA1 (breast-cancer gene 1)

BRCA1 (breast-cancer gene 1): Its gene product forms the scaffold for protein assembly that "checks up on" the cell cycle to make sure that the DNA has replicated faithfully. When it's knocked out, the check on the cell cycle is removed.
Can either be familial (LOH as above) or sporadic.
Note that in sporadic cases, the mutation can be on other (unspecified) genes that regulate or have an effect on BRCA's expression



p53 you may recall from Li-Fraumeni syndrome-- mutant or inactivated p53 is found in ~50% of all cancers.
Protein involved in DNA mutation repair at "checkpoint" in cell cycle.
Notice that mutations in p53 can be "dominant-negative;" that is, one bad copy of p53 can inactivate the other, good copy of p53.
Along with RB, one of the two major gene products knocked out by oncoviruses to produce cancers


Why are oncogenes useful as molecular markers in prognosis? You should be able to give at least two examples of oncogenes that are currently being used and also include the evidence of why these are good markers.

The level of expression of oncogenes tends to correlate with the rapidity of the progress of the cancer.
One example: The level of expression of the N-myc gene is used in prognosis analysis for neuroblastoma.
Another example: Increased expression of the HER2/neu gene correlates with poor prognosis in breast cancer.


targeting a few drugs that target them.

Herceptin: drug antibody therapy against the HER2/erb2 oncogene product.
Small molecules: able to inhibit action of cancerous proteins by, among other pathways, binding to their active sites. Example is Gleevac, an ATP analogue, that inhibits ABL tyrosine kinase in patients with BRC-ABL translocation on the Philadelphia chromosome.
(BRC-ABL: translocation causing leukemia; resistant to radiation therapy. ABL is an ATP-dependent tyrosine protein kinase that PO4s various other proteins.)
Gleevac = specific to tyrosine kinase proteins: specific fit to their ATP-binding pocket.
Notice can use combined oncogene targeting therapy and radiation therapy: former makes the cancer more susceptible to the radiation.


Targeting tumor repressor genes:

You can inject RB directly into RB-negative tumors
You can use drugs that only kill cells with p53 deficiencies
You can use drugs which correct the mutant conformation of dominant-negative p53 proteins (see above).


what is the location of the RB gene?

chromosome 13q14


What do hela cells express?

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.


2 . Describe why p53 was originally incorrectly thought to be an oncogene.

The p53 gene was initially found to be an oncogene, in that certain p53 mutant
genes were dominant to the wild-type gene in producing cellular transformation. Later studies
indicated that this gene behaved as a classic tumor suppressor in Li-Fraumeni syndrome. The
explanation was found by showing that the oncogenic p53 mutations produce a mutant p53
protein that can bind the wild-type p53 protein and inactivate it. These “dominant-negative” p53
mutations can be viewed as “spoilers” or “monkey wrenches”.


4 . Describe the cellular function of the p53 protein.

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”.


5 . Recognize that oncogenic viruses make proteins to inactivate both Rb and p53.

p53 interferes with the life cycle of many human viruses including
Adenovirus and HPV (human papilloma virus). The viruses 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.


Discuss how oncogenes were discovered, describing at least three different examples of the method
used in the discovery.
2 . Discuss the functions of protein products of viral oncogenes, including at least four examples of
oncogenes of known function.
3 . Describe why oncogenes are useful as molecular markers in prognosis and discuss at least two
examples of oncogenes that are currently being used, including the evidence of why these are good
4 . Differentiate between oncogenes and tumor suppressor genes and describe the function of these two
types of cancer genes and how mutations in them may combine to produce cancers.

Retroviruses are RNA containing membrane enclosed viruses that bud from the
cell membrane of infected cells and which usually do not kill the infected cell.
2. The RNA genome consists of two identical strands held together by a tRNA
molecule. The gag gene codes for internal virion proteins, the env gene for virus membrane
glycoproteins, and the pol gene for a virus polymerase (See Fig. 1). When these genes are the
only ones present the virus does not cause tumors, but replicates through an intermediateproviral 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 v-onc segment also have the ability to rapidly
transform appropriate infected cells to the malignant phenotype. They therefore rapidly induce
tumors after infection. A few examples are:

a. v-src - The oncogene of Rous Sarcoma Virus. This caused
fibrosarcomas in certain birds.
b. v-erb - The oncogene of avian erythroblastosis virus causes
erythroblastosis in chickens. Two related genes, erb-A and erb-B have been identified.
c. v-abl - The oncogene found in Abelson leukemia virus from mice.
d. 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.


5 . Describe two examples of how molecular, genomic, and clinical information (Bioinformatics) about a
patient's cancer are being used for targeted therapy and for "personalized medicine" in cancer.

rIn an example of this type of procedure, a
“heat map” is created in which changes in gene copy number are correlated with tumor grade.
The “heat map” is created by hybridization of the tumor DNA to “gene chip” containing all human
genomic DNA sequences as molecular probes. Red color indicates increases and green
indicates decreases. It is called a “heat map” because it resembles graphical representation of
hot and cold areas of the earth.
In breast cancer, this type of analysis is already being used in the clinic. The Breast cancer
Xpress Chip measures the expression of 123 genes known to be altered in breast cancers.
This includes tumor suppressors such as BRCA1 and p53 and oncogenes such as estrogen
receptor (ER) and erbB2. The information is then used for diagnosis, prognosis and therapy. For
example, patients with high erbB2 are treated with Herceptin and those with high ER levels are
treated with tamoxifen (estrogen antagonist).
With the advent of whole genome sequencing, cancer therapy is being designed on a
personal basis depending on the type of mutations (oncogene, tumor suppressor, etc.) in the
tumor and the known therapeutic responses to these lesions (see Dancey et al., 2012).


Cellular Transformation is assayed by:

a. Tumor formation in animals after administration of the oncogenic virus.
b. Transformation of cell morphology and growth regulation after infection of
cultured cells in vitro. Foci of infected cells developing as transformed clones can be seen and


. The protein (pp60v-src)

. The protein (pp60v-src) coded by the v-src gene is a membrane bound protein
kinase that phosphorylates tyrosine residues in several different proteins. Theses proteins then
change the properties of the cells by affecting gene expression.



v-erb-B codes for a protein that is similar in structure to the cell surface receptor
for epidermal growth factor (EGFR). This raises the possibility that this protein has growth
stimulating properties like EGFR. This receptor is a member of a family of related proteins that
(like pp60v-src) exhibit tyrosine-specific protein kinase activity



v-abl codes for a protein kinase that phosphorylates tyrosine residues on other
proteins. It is similar to the human cellular gene (c-ABL) that is found in the BCR-ABL
translocation in the Philadelphia chromosome and is overexpressed in BCR-ABL CML.


Properties of c-onc genes

1. Although some have nearly the same DNA sequences as the cognate v-onc
gene most are quite different. Examples:
a. c-src has a different carboxy-terminal amino acid sequence that v-src and
has numerous introns that do not exist in v-src.
b. c-myc also has many introns not present in v-myc, although the coding
sequences are nearly identical (7 amino acid changes).
2. Thus, if v-onc genes originated from c-onc genes, substantial rearrangements
occurred during or after the capture (at least with many such oncogenes).
3. A few c-onc genes ligated to retroviral promoters can transform normal cells
when introduced via DNA mediated transformation (examples: c-ras). Other c-onc genes do not
directly transform normal cells.
4. Analysis of DNA from human bladder cancer cells has shown that c-ras genes
have been point mutated, supporting the qualitative model above. Mutations are found in
either codon 12 or codon 61 of the ras gene product. These mutations produce a ras protein
that is unregulated and is always “on”. Detection of these ras mutations indicates a poor
5. Gene amplification of c-onc genes has also been detected in some cancers and
is very useful for prognosis. For example, N-myc, a member of the c-myc family of oncogenes,
is found amplified in neuroblastoma. 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 poor prognosis. This supports the
quantitative model above. Note: Drugs have been designed that inhibit HER2/neu/erbB2
(see below).
6. Translocations of c-onc genes have been observed in certain human cancers,
and also indicate a poor prognosis. The translocation results in inappropriate and high level
expression of the oncogene (See Burkitt lymphoma and the Philadelphia chromosome (BCRABL
translocation) as discussed in the previous lecture). Note: Drugs have been designed
that inhibit ABL tyrosine kinase that is overexpressed in the BCR-ABL translocation (see
7. It is clear that several genes must simultaneously be changed to transform a
normal cell to a malignant one. Mutations in both tumor suppressors and oncogenes are
needed. The best evidence for this idea has been found in colon cancer. Loss of growth
Molecules to Medicine: Cell Physiology 17-11-2015 /10-11/ Robert A. Sclafani
regulation and an increased mutation rate coupled to loss of programmed cell death (apoptosis)
can be particularly deadly.