Bio of Cancer 3

Cell genomes are under constant attack from endogenous biochemical processes

1. 10,000 depurinations/day/cell caused by H+ and OH- ions (loss of A or G) 2. 100-500 depyrimidinations/day/cell caused by H+ and OH- ions (loss of C or T) 3. Spontaneous Deamination- the removal of an amine group from a base - While spontaneous deamination of cytosine forms uracil, which is recognized and removed by DNA repair enzymes, deamination of 5-methylcytosine forms thymine. This conversion of a DNA base from cytosine (C) to thymine (T) can result in a point mutation. 4. Oxidation of bases by free radicals

Cells deploy a variety of defenses to protect DNA molecules from attack by mutagens

1. Melanocytes transfer their melanosomes to keratinocytes to protect the keratinocyte from UV light exposure. 2. Free radical scavengers and enzymes capable of converting free radicals to harmless chemicals - Superoxide dismutases -- converting the superoxide anion, O2- , to O2 or H2O2 - Catalases -- Converts H2O2 to H2O - Glutathionine-S-Transferase (GST) -- a "family" of enzymes that aids in detoxification by speeding up the linking of toxic compounds with glutathione (GSH), thus forming a less reactive substance

Activation of apoptosis by p53

1. P53 is a TF for Fas 2. P53 is a TF for IGFP-3, an inhibitor of insulin like growth factor (IGF-1/2) binding its receptor 3. P53 is a TF for Bax, a proapototic factor

Why should a cell commit suicide?

1. Programmed cell death is needed for proper development just as mitosis is. § formation of fingers and toes of the fetus § the sloughing of the inner lining of the endometrium at the start of menstruation § formation of the proper connections between neurons in the brain requires that surplus cells be eliminated 2. Programmed cell death is needed to destroy cells that represent a threat to the integrity of the organism. § cells infected with viruses § cells of the immune system which are self reactive § cells with DNA damage

Human cells are constructed to be highly resistant to immortalization and transformation 5 steps are involved

1. The mitogenic signaling pathway controlled by Ras 2. The cell cycle controlled by pRb 3. The alarm pathway controlled by p53 4. The telomere maintenance pathway controlled by hTert 5. The signaling pathways controlled by protein phosphatase 2a

Inherited defects in nucleotide-excision repair (NER), base-excision repair (BER), and mismatch repair (MMR) lead to specific cancer susceptibility syndromes

1. Xeroderma Pigmentosum- inherited defects in nucleotide excision repair 2. Hereditary nonpolyposis colorectal cancer ( HNPCC ) or Lynch syndrome is an autosomal dominant genetic condition that is associated with a high risk of colon cancer; it is an inherited defect in the eukaryotic homologs of the mismatch repair proteins mutS (hMSH2) and mutL (hMLH1)

Telomeres are complex molecular structures that are not easily replicated

6 bases 5' to 3' (TTAGGG)x (G rich) followed by a ss stranded G rich overhang

The karotype of cancer cells is often changed through alterations in chromosome number

Aneuploidy

What makes Bcl-2 a proto-oncogene?

B cells, like all activated lymphocytes, die a few days after doing their job; in this case, making Abs They die by apoptosis (programmed cell death) High levels of Bcl-2 protect these cells against apoptosis by suppressing the activation of caspases The translocation of Bcl-2 to chromosome 14 allows the constitutive expression of Bcl-2. Cancer results, not by increased proliferation, but by not dying on schedule Thus Bcl-2 works in direct opposition to p53; p53 induces apoptosis, bcl-2 prevents it.

A variety of other DNA repair defects confer increased cancer susceptibility through poorly understood mechanisms

BRCA1 is linked to breast and ovarian cancer and is located on 17q21. The BRAC1 protein is a component of a multi-subunit complex. Through this complex, BRCA1 repairs double-strand DNA breaks via homologous recombination. The BRCA2 protein interacts with single-strand DNA and the recombinase RAD51. Together with BRCA1 and a BRCA2 partner, PALPB2, BRCA2 completes DNA double strand breaks by recruiting RAD51 for the final steps of homologous recombination. The absence of BRCA-2 is also associated with a variety of chromosomal abnormalities; chromatid breaks (ctb) , tri-radials (tr) and quadri-radials (qr)

14q+ 18q- Translocation

Bcl-2 is a human proto-oncogene located on chromosome 18 Its product is an integral membrane protein located in the membranes of the ER, nuclear envelope and outer membrane of the mitochondria The gene was discovered as the translocated locus in a B cell lymphoma (bcl-2 was translocated from chromosome 18 to the immunoglobulin heavy-chain locus on chromosome 14) and is constitutively expressed Bcl-2 prevents apoptosis and is not your typical proto-oncogene - Prevents release of cytochrome c from mitochondria - Nothing to do with proliferation

What seems to be important is the ratio of anti-apoptotic Bcl-2 members to pro-apoptotic members and the dimerization of members:

Bcl-XL and Bcl-2 homodimers favor cell survival, preventing apoptosis by preventing cytochrome C release from the mitochondria Bcl-2 and Bcl-XL can also bind to Apaf-1 - A central checkpoint of apoptosis is the activation of Caspase-9. The BH4 domain of Bcl-2 and Bcl-XL can bind to the C terminal part of Apaf-1, thereby inhibiting the association of Caspase-9 with Apaf-1

Apoptosis as a morphological phenomenon

Blebs in the membrane Nucleus condenses into dense structures, pyknosis DNA is cut into different size pieces, called a DNA ladder, in two tumor cell lines when apoptosis is induced by expression of the pro-apoptotic Lats2 protein. Lats2-induces caspase-3- and caspase-9-dependent apoptosis. Fragmentation of the golgi Apoptotic nuclei

Models of Tumor Heterogeneity

Both models predict that only a few cells will actually be able to cause a tumor but the stochastic model predicts that it is random and present in any selected sub-population while the stem cell hypothesis predicts that a population can be selected and enriched

Histopathology provides evidence of multi-step tumor formation

Cancer is a multistep process during which cells acquire a series of mutations that eventually lead to unrestrained cell growth and division, inhibition of cell differentiation, and evasion of cell death. As cancerous tumors grow in size, they stimulate a process called angiogenesis, during which they acquire a new source of blood vessels that provides them with oxygen and nutrients. Eventually, tumor cells invade the surrounding tissue and spread to other parts of the body in a process called metastasis

Discovery of complementation groups in XP patients

Cell fusion experiments allowed for the discovery of 8 distinct genes, 7 of which, XPA to XPG, are part of nucleotide excision repair complex and XPV, which codes for DNA pol-h (eta) If two individuals have the same enzyme missing then they don't complement each other and they're still susceptible to UV radiation. But if you take two cells where one is missing A and one is missing F and you fuse the two cells à The A- cell will have a good F and the F- cell will have a good A à Those cells will be resistant to UV

The proliferation of cultured cells is also limited by the telomeres of their chromosomes

Cells which escape crisis are characterized by telomere stability and the reactivation of telomerase expression

Cancer development seems to follow the rules of

Darwinian evolution

To repair mistakes and mutations we have 4 important mechanisms

Direct repair: for endogenous and exogenous mutagens · Simplest; acts directly on damaged nucleotides, converting them back to their original structure · An alkylated guanine would pair with thymine; to fix this damage, an enzyme transfers the alkyl group to itself, restoring the guanine base to normal · Enzymes that perform this function: Ada in E.coli and MGMT in humans Excision Repair (ER) (BaseER and NucleotideER): for endogenous and exogenous mutagens · Removing damaged nucleotides and then filling in the gap · BaseER = Removal of a base from a single nucleotide · NucleotideER = Removal of an entire block of nucleotides Mismatch Repair: for errors made during replication · Detects the absence of proper base pairing; excise mismatched part of daughter strand & fill in the gap Nonhomologous end joining: replication errors, mutagens and damage caused by radiation · Ku proteins bind to broken DNA ends. · Ku proteins have an affinity for one another bringing them and the fragments in close proximity & the DNA fragments are joined by DNA ligase. · Often prone to errors (sequences flanking ds break often lost, mis-repair à hybrid structure)

A 2011 paper by Hanahan and Weinberg suggest the addition of two new emerging hallmarks and two new enabling characteristics

Emerging Hallmarks: 1. Deregulating cellular energetics 2. Avoiding immune destruction Enabling characteristics: 1. Genome instability 2. Tumor-promoting inflammation

Most human cancers develop over many decades of time

Epidemiological studies convincingly show that the incidence of most cancers increase dramatically with age (80% of cancers are diagnosed at age 55 or older).

Where DNA errors arise

Errors during DNA replication, but in this case repaired by Pol d's ability to proofread and repair in addition to replicate the DNA Spontaneous base changes DNA base changes due to mutagenic agents A break in the DNA caused by a clastogen (a mutagenic agent giving rise to or inducing disruption or breakages of chromosomes, leading to sections of the chromosome being deleted, added, or rearranged).

In reality, tumor growth is mixed, there is both death and division going on simultaneously. Selection for those cells best adapted at any given time.

Even in a rapidly growing tumor there will still be necrotic/apoptotic cells Survival of the fittest - competition for oxygen & nutrients

However, tumor instability becomes a major factor and selection may no longer be able to select for the most advantageous cell types

Evidence suggests that this chaos on the chromosomal level is not just a side effect of malignancy, but the direct cause and driving force of cancer

Human embryonic kidney (HEK) cells +/- Large T Ag of SV-40 Graph

HEK cells + wt LT becoming immortalized through taking out both Rb and p53 (one alone doesn't do it)

Multiple lines of evidence reveal that normal cells are resistant to transformation by a single mutated gene

Ras alone induces only epithelial hyperplasia in colonic epithelium (essentially normal cells found in increased numbers) or even an inactivated APC and an active ras produce hyperplasia Monozygotic Twin Studies: the distance between the two dots represents that the initial mutation was not alone sufficient to induce cancer.

Argument for stem cells as the cell of origin of cancer

Self-renewal is a key property of cancer cells. Stem cells already have the capacity to self-renew, therefore they would theoretically require fewer neoplastic changes to become fully transformed. Due to their longevity, there could be greater opportunity for genetic changes to accumulate in individual stem cells than in more mature progenitors with a limited lifespan.

Argument for combining the two by starting with stem cell but other changes occurring in the transit amplifying cells

Initial transforming events occurring in stem cells could cause an expansion of the stem cell pool and/or expansion of downstream proliferating progenitors. Statistically, secondary events are more likely to be accrued within these expanded pools of target cells, eventually resulting in full transformation. Thus, whereas a stem cell might be the initial target cell, depending on the consequences of initiating events a cell with a progenitor phenotype could actually be the penultimate precursor. The propensity for oncogenic events to induce changes that enable continued propagation as opposed to termination or differentiation could differ in stem cells versus downstream progenitors.

Skin carcinogenesis model

Initiation -> mutating the DNA Promotion -> causing those cells to proliferate Progression -> from a benign tumor to an aggressive metastatic invasive cancer

SIX Hallmarks of Cancer

Insensitivity to anti-growth signals Self-sufficiency in growth signals Evading apoptosis Sustained angiogenesis Limitless replicative potential Tissue invasion & metastasis

What happens in crisis

Lack of telomeres leads to end to end joining Repeated breakage, fusion, bridge formation

Inherited mutant alleles affecting p53 pathway predispose those affected to a variety of cancers

Li-Fraumeni syndrome (LFS) is an inherited familial predisposition to a wide range of certain, often rare, cancers due to a change in p53 Other multicancer families that resemble Li Fraumeni include mutations in Chk2 kinase (which phosphorylates p53) and those having mutations in the promoter of MDM2 causing over expression of MDM2.

Argument against stem cells as the origin of cancer

Stem cells are quiescent and divide less frequently than progenitors, and consequently are less likely to acquire mutations in the first place. Because stem cells are the key to tissue regeneration and the only cells capable of self-renewal, they might have evolved sophisticated inhibitory machinery to keep self-renewal in check as an anticancer defense.

Apoptosis, drug pumps and DNA replication mechanisms offer tissues other ways to minimize the accumulation of mutant stem cells

Stem cells may choose apoptosis over DNA repair which is not always 100% accurate or they may use highly effective DNA repair mechanisms. Stem cells express high levels of the the MDR gene product as seen by experiments using fluorescent dyes. They compared stem cells to differentiated cells for the ability to exclude a dye. Asymmetric DNA strand allocation - Keep conserved strand in stem cell population each time - The nonconserved strand will go on to the transit amplifying population and all the daughters after that

Bcl-2 and related proteins

Pro-survival (Bcl-2): Cells are resistant to apoptosis when present, cytochrome C remains in the mitochondria Pro-apoptotic: Bax binds bcl-2 and inactivates it, cells will be apoptotic when cytochrome c moves to cytoplasm, p53 is a TF for Bax

Human hTert and hTR at work extending the telomere DNA

hTERT provides 6 nt template region for extension

Cells which spontaneously transform after crisis have found a way to maintain their telomeres.

hTR RNA always present, just don't have protein product in mortal cells What the immortal cells are doing is turning on the telomerase enzyme itself

Nonmutagenic agents, including those favoring cell proliferation, make important contributions to tumorigenesis

TPA (also known as PMA), a potent tumor promoter: functions as diacylglycerol and activates PKC

The single stranded G rich strand is protected from degradation by

being base paired with the C rich strand

X-Rays

The energy of X-rays strips electrons from water molecules. The resulting free radicals proceed to generate reactive oxygen species (ROS) that create single and double strand breaks in the DNA double helix that can in turn lead to chromosome breaks if not repaired.

DNA repair is an important line of defense against mutations caused by exogenous carcinogens and by endogenous mechanisms. If not repaired before a cell replicates, they may contribute to carcinogenesis.

The genes responsible are a special group of TSG called caretaker genes in contrast to TSG like Rb and p53 which are gatekeeper genes.

Apoptosis vs. Necrosis

The main difference between apoptosis and necrosis is that apoptosis is a predefined cell suicide, where the cell actively destroys itself, maintaining a smooth functioning in the body whereas necrosis is an accidental cell death occurring due to the uncontrolled external factors in the external environment of the cell.

Some immortalized cells can maintain telomeres without telomerase

The mechanism is called alternative lengthening of telomeres (ALT)

A linear path of clonal succession oversimplifies the reality of cancer: intra-tumor heterogeneity

The prevailing dogma of cancer evolution is therefore one of ''gradualism'' in which acquisition of driver mutations occurs cumulatively over years to decades, resulting in incremental progression through increasingly malignant phenotypes.

Flow Cytometry PI Red F12 vs. Anexin FL1 Green

Top Left = Dead Cells Bottom Left = Healthy cells Top Right = Late apoptotic, Membrane no longer able to keep out PI Bottom Right = Early apoptotic, Membrane still intact but phosphatidylserine flipped out

Cancer cells need to become immortal in order to form tumors

When cells enter senescence as the result of telomere shortening, p53 is activated which is able to bind the unprotected end of the chromosome Activated p53 triggers growth arrest or cell death If p53, p16 or arf is missing or inactivated, then cell growth is not arrested nor is apoptosis induced. Cells may escape temporarily but the cells are said to be in "crisis." Lack of telomeres triggers chromosomal rearrangements which can be genetic suicide. To survive, the cell must reactivate telomerase or find another means to stabilize the telomeres (referred to as ALT, alternative lengthening of telomeres). Result: immortality

Use of flow cytometry to isolate stem cells from breast cancer tissue

Tumorigenicity was found in the CD24-, CD44+.

DNA adduct

a piece of DNA covalently bonded to a cancer-causing chemical.

Asymmetric and symmetric division of stem cells:

a. Asymmetric division where a stem cell gives rise to another stem cell and a transit amplifying cell (most common) b. Stem cells maintain the size of the population. If one is lost a transit amplifying cell can revert back or a stem cell can divide symmetrically to keep the number constant. c. A growing organ using symmetric growth followed by asymmetric growth for organ maintenance.

Exogenous alkylating agents

can add methyl groups to the bases which can lead to the loss of the base or a misread by DNA polymerase

UV radiation

causes pyrimidine dimers: covalent bonds between T-T, T-C and C-C. 71% are cyclobutane pyrimidine dimers (Thymidine Dimers) (A) while 24% are the pyrimidine 6-4 pyrimidinone photoproducts (B). When these pyrimidines are bound to each other that DNA can't be replicated or transcribed because those bases are not available to DNA polymerase

anoikis

cell death when cells can't bind to the extracellular matrix via their integrins. does not require p53

Telomerase

consists of two subunits, a reverse transcriptase, known as human telomerase reverse transcriptase (hTert) (p123) which maps to chromosome 5p and an RNA template known as telomerase associated RNA molecule (hTR) consisting of 451 ribonucleotides,

Following Cox-2 inhibitor celecoxib, mammary cancers induced by middle T of polyoma virus show

increased apoptosis and decreased proliferation

Senescence

is defined as cell cycle arrest following a limited number of cell divisions typically triggered by one or more dysfunctional telomeres. In contrast, cell lines from tumors do not show senescence; they continue to grow forever in tissue culture as long as they are properly cared for.

fungal toxin, aflatoxin B1 (AFB1),

is made by molds, largely Aspergillus flavus, that grow on peanuts and grains that have been improperly stored, notably in areas of high rainfall and humidity. is one of the most potent mutagenic substances by making a DNA adduct if not detoxified by epoxide hydrase or Glutathione S-transferase.

Cells accumulate genetic and epigenetic alterations as tumor progression proceeds: A possible five-hit scenario for colorectal cancer, showing the mutational events that correlate with each step in the adenoma-carcinoma sequence.

loss of APC --> DNA hypomethylation --> activation of K-ras (12p) --> loss of 18q TSG --> loss of p53

How liver inflammation can lead to liver cancer.

loss of mdr function chronic liver inflammation recruitment of inflammatory cells TNF-a activation of TNF-a receptor IKK phpsphorylates IKB NF-KB free to act as TF -TNF-a (more inflammation) -cyclin D, Myc (proliferation) -Bcl-X (protection from apoptosis) -COX-2 -> prostaglandin -> loss of contact inhibition, anchorage independent proliferation, increased proliferation

Noxa

mediates E2F-induced apoptosis

Apoptosis is a complex program that often depends on

mitochondria

In mammals, members of the Bcl-2 family act at the ____________ to control release of __________ _, which is required for the binding of _______-_ to _________ _______ __________ ______-_.

mitochondria cytochrome c caspase-9 Apoptosis Protease Activating Factor-1

5q- band 21 - APC

o 10% of all colon cancers have an inherited basis o Part of this 10% is due to FAP (familial adenomatous polyposis) o Inactivation of both APC alleles can be detected in most human intestinal tumors at very early stages of tumor development o Loss of heterozygosity (LOH) can occur by somatic mutation, loss of the entire chromosome or by somatic recombination of the good copy. o Patients with this genetic loss develop 1000s of polyps in their colon during their 20s and 30s o The gene responsible, APC (adenomatous polyposis coli) codes for a large protein consisting of 2840 amino acids o Its best understood function is to down regulate Beta-catenin, a transcription factor and a key effector of the Wnt signaling pathway.

Hypomethylation

o DNA methylation is a well-defined epigenetic mechanism involved in regulation of gene expression during development. In eukaryotes, methylation involves addition of a methyl group to the 5-carbon position of cytosine. In mammalian cells, this epigenetic modification of DNA primarily occurs on the cytosine of CpG dinucleotides and has been shown to play a major role in the transcriptional silencing of numerous genes. o Hypomethylation occurring in tumor generation would, therefore, allow for the expression of previously silenced genes or possibly chromosomal instability.

Bax expression is controlled by

p53

A classic cancer hallmark is the ability of malignant cells to

proliferate indefinitely

p21

responsible for actin stress fibers

Bcl-2 family members are regulated at the

transcriptional level

benzo[a] pyrenediolepoxide (BPDE)

ultimate carcinogen able to directly attack and form covalent adducts with DNA bases, which may then generate oncogenic mutations.

Benzo[a]pyrene

undergoes two successive oxidations mediated by cytochrome P450 enzymes (largely CYP1A1) and is converted to benzo[a] pyrenediolepoxide (BPDE).

By looking at what mutations took place in the DNA

you can sometimes figure out what the chemical was that caused those mutations

Inflammation can contribute to tumor growth by providing

§ growth factors that sustain proliferative signaling, § survival factors that limit cell death, § proangiogenic factors, § extracellular matrix-modifying enzymes that facilitate angiogenesis, invasion, and metastasis, - metalloproteases break down collage & fibronectin and allow tumor cells to escape primary site § inductive signals that lead to activation of epithelial to mesenchymal transition.

Both intrinsic and extrinsic apoptotic programs can lead to cell death

1. Apoptosis by internal signals pro-apoptotic signal -> release of cyt c from mt -> cyt c + Apaf-1 -> (procaspase-9 -> caspase-9) -> cyt c + Apaf-1 + caspase-9 = Apoptosome -> activation of executioner caspases -> cleavage of death substrates 2. 1. Apoptosis signaled by external signals - An example: FasL/Fas Cell death receptors: Binding of ligand to the Fas receptor induces apoptosis by direct activation of Fas associated death domain (FADD) and caspase-8. Fas ligand consists of three polypeptide chains, so its binding induces receptor trimerization. Caspase-8 bound to the receptor via FADD is then activated by autocleavage, leading to activation of downstream caspases and cell death. Two pathways can converge

The karotype of cancer cells is often changed through alterations in chromosome structure

1. Chromosome breaks 2. Chromosome translocations 3. Mutations in genes that influence mitotic spindle behavior can lead to chromosome instabilities

Cell genomes are under occasional attack from exogenous mutagens and their metabolism

1. Radiation - X-rays - UV irradiation 2. Chemicals - Occupational - Environmental - Dietary

Incipient cancer cells can escape crisis by expressing telomerase

85-90% of tumor cells show telomerase activity while most normal cells have very low levels of hTERT detected by the Trap assay

Normal stem cells within tissues

Don't need to divide as much Are partially shielded from toxic agents Have the ability to self-renew and also give rise to daughter cells that show a commitment to differentiate

The data suggest that the dynamic clonal diversification model better represents what happens in late tumors

Dynamic clonal diversification: One or two driver type mutations -> Once you have a rapidly proliferating pool -> mutations can happen in different cells and can all go different directions In a 2011 paper (Cell 144, 27-40, January 7, 2011) the authors cite several examples in which a more ''punctuated equilibrium'' evolutionary model may apply to development of cancers instead of the previously described gradualism. Genome-wide telomere attrition in somatic cells, for example, may generate naked DNA ends that act as a nidus for on-going genomic instability. End-to-end chromosome fusions resulting from telomere loss can lead to spiraling cycles of dsDNA breakage, aberrant repair and further chromosomal damage in both daughter cells. Iteration of this breakage and repair process can lead to extensive genomic remodeling in multiple competing subclones in only a few cell cycles. Under these scenarios, bursts of somatic mutation may accrue in relatively short periods of chronological time.

Small intestinal crypts

House stem cells that serve to constantly replenish epithelial cells that die and are lost from the villi. Protection of these stem cells is essential for long-term maintenance of the intestinal epithelium - the location of Paneth cells adjacent to stem cells suggests that they play a critical role in defending epithelial cell renewal - Mucus within crypts prevents chemicals from reaching the stem cells Cells btwn 5-20 = Transit amplifying cells divide every 12 hours, move up the villi, and eventually start to differentiate Enterocytes differentiate further & further, until they get to top of villi and undergo apoptosis (shed every 3rd or 4th day)

Telomerase plays a key role in the proliferation of human cancer cells

Human cancer cells do not need to be transfected with hTERT, have already turned on their own telomerase

Repair enzymes fix DNA that has been altered by mutagens

If the DNA is damaged, in the example below by ethylnitrosourea, resulting in an abnormal base, it is the job of repair enzymes, in this case O6-methylguanosine DNA methyltransferase (MGMT), to correctly restore it: sometimes referred to as direct (one-step) repair. Rather than reverse the change chemically, base excision repair (BER) and nucleotide excision repair (NER) can be used to remove a chemically altered base and replace it with the correct one. - Base Excision Repair (BER): targets chemically altered bases induced mainly by an endogenous mechanism. The first step of BER is carried out by a family of DNA-damage-specific glycosylases that flip the lesion outside of the helix and then cleave the base, an endonuclease cleaves the sugar-phosphate backbone. DNA polymerase β then replaces the nucleotides and ligase fills in the gaps. - Nucleotide excision repair (NER) is specific for helix-distorting lesions such as pyrimidine dimers and bulky DNA adducts induced by environmental agents which interfere with transcription and replication. The lesion, along with 24-32 other bases are excised by endonucleases and then DNA polymerases delta and eta, fill in the spaces with the help of Proliferating cell nuclear antigen (PCNA) which acts as a scaffold to recruit proteins involved in DNA replication, DNA repair, chromatin remodeling and epigenetics. Error prone repair or SOS repair is induced by single-stranded gaps and/or the presence of DNA degradation products. This system is capable of replication opposite thymidine dimers or apurinic sites and apparently does so by putting in most any base. It therefore causes a very broad spectrum of mutations, including duplications and deletions. This system is often the cause of mutations following either chemical or UV mutagenesis. Obviously such a repair system must be a desperate recourse for the cell, allowing replication past a region where the wild-type sequence has been "lost".

Other combinations of oncogenes / tumor suppressor genes

Ig Heavy chain enhancer ahead of myc genes § Placed in fertilized egg § B cell lymphomas arise after months of latency § Found other oncogenes activated in the tumors Large T Ag with insulin promotor § Placed in fertilized egg § Pancreatic hyperplasia seen in 50-70% § Pancreatic cancer seen in 1-2% of the mice

Transformation usually requires collaboration between two or more mutant genes

Myc gives immortality Ras gives altered morphology and anchorage independence Other combos: § src & myc § erbB & myc § ras & E1A, Large T or E7

Stem cells may or may not be targets of the mutagenesis that leads to cancer

One component of the cancer stem cell theory concerns how cancers arise. In order for a cell to become cancerous, it must undergo a significant number of essential changes in the DNA sequences that regulate the cell. The conventional cancer theory is that any cell in the body can undergo these changes and become a cancer cell. The stem cell theory is that cancer stem cells arise out of normal stem cells or the precursor cells that normal stem cells produce since the normal stem cells are the only cells that reproduce themselves. They are therefore around long enough to accumulate all the necessary changes to produce cancer. Self-renewal is a quality of stem cells that allows for the accumulation of mutations that can be passed on to daughter cells. A modification of the stem cell model is that a differentiated cell can acquire a mutation that reactivates a self-renewal program ("the induced pluripotent stem cell").

Tumor stem cells further complicate the Darwinian model of clonal succession and tumor progression

The ''cancer stem cell hypothesis.'' Two important related concepts of this hypothesis are that (a) tumors originate in either tissue stem cells or their immediate progeny through dysregulation of the normally tightly regulated process of self-renewal. (b) As a result of this, tumors contain a cellular subcomponent that retains key stem cell properties. These properties include self-renewal, which drives tumorigenesis, and aberrant differentiation that contributes to cellular heterogeneity "Recent studies provided evidence of the existence of a subpopulation of cells within a variety of tumor types with a tumorigenic potential that is lacking in the rest of the cells within these tumors. Traditionally, many cancer cells have been considered to have tumorigenic potential, even though no assay has yet demonstrated that a high percentage of single human cancer cells can form tumors. In contrast, the cancer stem-cell model has suggested that only small subpopulations of cancer cells have tumorigenic potential, based on experiments in which human cancer cells were xenotransplanted into NOD/SCID mice. For example, only one in a million (0.0001%) human melanoma cells is tumorigenic in NOD/SCID mice. Indeed, most human cancers have only rare (0.1%) tumorigenic/leukaemogenic cells (also called cancer-initiating cells or cancer stem cells) when transplanted into NOD/SCID or other highly immunocompromised mice"

p53 often ushers in the apoptotic death program

When the cell can't be repaired during cell cycle delay, p53 is capable of inducing programmed cell death, apoptosis.

18q- 21, Loss of 18q - Deleted in Colorectal Cancer (DCC) Tumor Suppressor Gene (TSG)

o Loss of heterozygosity occurs in about 70% of colorectal cancer and 50% of large benign adenomas. Early adenomas do not show this change. Among patients with stage II disease, the five-year survival rate was 93 percent in those whose tumor had no evidence of allelic loss of chromosome 18q and 54 percent in those with allelic loss; among patients with stage III disease, survival was 52 and 38 percent, respectively. o The gene products from 18q21.1 is thought to code for the netrin-1 receptor, binding to its ligand triggers signaling related to nervous system development. When not bound to netrin-1, the netrin-1 receptor acts as a tumor suppressor, which means that it keeps cells from growing and dividing too fast or in an uncontrolled way. Other studies suggest that when DCC is not bound to netrin-1, it triggers cell death (apoptosis).

12p activation - Ras

o The activation of KRas by point mutations in positions 12, 13 or 61. o Ras is an important signal transducer of growth factor signals o Oncogenic Ras stays bound to GTP and signals downstream molecules in the absence of growth factors o Found in 30-70% of all colon cancers

17p- - Immortalization depends on the loss of p53

o When cells enter senescence as the result of telomere shortening, p53 is activated when it binds the unprotected end of the chromosome. o Activated p53 triggers growth arrest or cell death o If p53 is missing or inactivated, then cell growth is not arrested nor is apoptosis induced. Cells may escape temporarily but the cells are said to be in "crisis." o Lack of telomeres triggers chromosomal rearrangements which can be genetic suicide. o To survive, the cell must reactivate telomerase or find another means to stabilize the telomeres (ALT). o Resulting in immortality