Biology, 2nd semester exam, cancer

The p53 exerts its effects in two ways.

(1) It induces the transcription of p21, which inhibits the CDK/cyclin-mediated phosphorylation of Rb required for the cell to transition to the S phase. (2) If DNA damage inflicted upon the cell cannot be successfully repaired, p63 mediated the transcription of genes implicated in the process of apoptosis.

At least seven different regulatory systems must be perturbed in order for a normal cell to grow as a cancer.

(1) Push the gas pedal: Cancer cells continue dividing in situations in which normal cells would quietly wait for a special chemical signal (2) No break Conversely, tumor cells must ignore "stop dividing" commands that are sent out by the adjacent tissues (3) No suicide (4) Attraction of blood vessels Then they have to persuade nearby blood vessels to build the infrastructure they need to thrive. (5) Immortality The great majority of cells in tumors quickly die of their genetic defects reproduce indefinitely if the tumor is to grow. The survivors do so in part by manipulating their telomeres, gene-free complexes of DNA and protein that protect the ends of each chromosome. (6) Invasion Tumors: the ability to invade nearby tissue and then metastasize to distant parts of the body that gives cancer its lethal character. (7) Evasion of immune system detection The anti-tumor activity of immune system rapidly eliminates cells with uncontrolled growth. Successful tumor cells therefore have to block these mechanisms.

There are two mechanisms by which proto-oncogenes can be converted to cellular oncogenes:

(1) Quantitative (regulatory): Tumor formation is induced by an increase in the absolute number of proto-oncogene products (resulted in enhanced gene expression) or by its production in inappropriate cell types (2) Qualitative (structural): Conversion from proto-oncogene to transforming gene (c-onc) with changes in the nucleotide sequence (by e.g. point mutation) which are responsible for the acquisition of the new properties leading to an increase of the stimulatory effect of oncoproteins on cell growth.

There are six types of proteins involved in the transduction of a growth factor signal:

(1) growth factors, (2) growth factor receptors, (3) intracellular signal transducers, (4) nuclear transcription factors which elicit the mitogenic effect through the regulation of gene expression, (5) cell cycle regulators, and (6) many types of microRNAs.

The mutations that activate these oncogenes are either:

(1) structural mutations that lead to the constitutive activity of a protein without an incoming signal (e.g., the protein kinases or Ras) or (2) regulatory mutations that lead to the expression of the gene at an elevated level or at the wrong place and time (e.g., growth factors or transcription factors).

Viral infections that can cause cancer:

- Epstein-Barr - HPV (human papilloma virus) - Rous sarcoma virus - Helicobacter pylori - Hepatitis B virus - Kaposi's sarcoma-associated herpesvirus

NF-kB provides...

...a molecular link between inflammation and cancer. Oncogenic activation of the NF-kB gene has also been identified in human tumors, including: - multiple myeloma, - acute lymphocyte leukemia, - prostate, and - breast cancers.

When the proto-oncogene is mutated or overregulated, it is called...

...an oncogene and results in unregulated cell growth and transformation. At the cellular level, only one mutation in a single allele is enough to trigger an oncogenic role in cancer development. The chance that such a mutation will occur increases as a person ages.

Steps of metastasis

1. Migration, 2. intravasation (the invasion of a cancer through the basal membrane and into blood vessel), 3. transport, 4. extravasation (cancer cells exiting the capillaries and entering organs), and 5. metastatic colonization . The first step (migration) and the last step (metastatic colonization) have been demonstrated to be rate limiting and their rate dictates the overall metastatic ability of the cancer.

Loss-of-function mutation:

A mutation that results in reduced or abolished protein function.

Pre-metastatic niche,

A site of future metastasis that is altered in preparation for the arrival of tumor cells. Haematopoietic precursor cells in the bone marrow expressing VEGFR1 (vascular endothelial growth factor receptor 1) appear to home in on specific sites before the tumour cells get there, paving the way for wandering metastatic cells by forming niches where they can locate and multiply.

Following further examination, a fundamental principle of cancer biology was revealed:

Almost all known oncogenes are altered forms of normal genes or proto-oncogenes.

Cell cycle regulators

Alteration in the normal function of these molecules often results in unchecked cell growth. - cycline D Increased expression of this regulator of the G1 S transition is commonly found in tumors. - CDK4 This regulator is among the most commonly altered gene in this class of genes. - CDK inhibitors: p16, p21, p27

Xeroderma pigmentosum (XP) genes.

Among eight genes so far identified as responsible for XP, XPA through XPG are involved in nucleotide excision repair of DNA damage induced by UV as well as various chemical carcinogens. If tumor suppressor genes (e.g. p53) are affected, the result may be cancer. XP is characterized by sun sensitivity, ocular involvement, and greater than 1000-fold increased risk of cutaneous and ocular neoplasms. The median age of onset of non-melanoma skin cancer is before age ten years

Ubiquitin ligases

Are enzymes that attach a small peptide called ubiquitin to proteins, flagging it for proteolysis in proteosomes.

Sarcomas

Cancers derived from mesoderm cells (e.g. bone, muscle)

miRNAs

Constitute a new class of oncogenes. MicroRNAs (miRNAs) are an abundant class of small noncoding RNAs that function as negative gene regulators. miRNA deregulation is involved in the initiation and progression of human cancer;

EGFR receptor

Contains an extracellular ligand-binding domain, a single transmembrane domain, and a cytoplasmic protein tyrosine kinase domain. Getting the signal from a growth factor outside the cell to inside the nucleus where gene expression is regulated requires several steps: - binding of the growth factor to the receptor, - receptor dimerization, - autophosphorylation, - activation of intracellular transducers (including RAS) - and a cascade of serine/threonine kinases, and - regulation of transcription factors for gene expression

Gene therapy of tumors 1b. Delivery of pro-apoptotic genes

Delivery of p53 gene to a p53-defected tumor cells can help to trigger the apoptotic pathway, that is, to kill the tumor cells.

Same allele, different locus, different phenotype

Different mutations in the same gene can result in different phenotypes. A good example is the RET proto-oncogene. Germline mutations of RET lead to multiple endocrine neoplasia (MEN) type 2.

Non-receptor tyrosine kinases are another important group of signaling molecules.

Genes that code for - cytoplasmic tyrosine kinases such as SRC, - serine/ threonine kinases such as RAF and MAPK, and - nuclear kinases such as ABL, can also undergo oncogenic activation

Penetrance.

If all carriers of a certain dominant allele in a population do not express the trait (same genotypes/different phenotypes), the gene is said to have incomplete penetrance.

Autosomal Recessive Inheritance

In autosomal recessive inheritance, two copies of the allele are required for the trait to be expressed. Carriers of one disease allele will not develop the illness, and several generations may be unaffected, leading to the appearance of skipped generations. Males and females are equally affected. If both parents carry one copy of the recessive allele, one in four offspring, on average, will express the trait

Loss of heterozygosity gene

In hereditary cancer syndromes, individuals are called heterozygous (having one or more dissimilar gene pairs). This is because they start life with a mutation (inherited from one of the parents) in one of the alleles linked to cancer susceptibility, but it is balanced by a normal counterpart. These individuals are predisposed to cancer because all their cells have already sustained the first hit to cancer-linked genes. If the critically needed normal suppressor gene that balances this germline mutation is lost at some time during an individual's life, a condition called loss of heterozygosity occurs.

Helicobacter pylori

Is a bacterium capable of inducing chronic inflammation in the stomach and initiating carcinogenesis. However, only less than 1% of those infected will develop gastric cancer. It is a multi-step process where inflammation is considered the initial and required step in the process. Helicobacter pylori and its role in gastritis and peptic ulcer disease (NOT in cancer formation!).

Kaposi's sarcoma-associated herpesvirus (KSHV; or human herpesvirus 8; HHV8)

Is a co-factor of Kaposi's sarcoma, a cancer of skin and connective tissues. Kaposi's sarcoma is associated frequently with AIDS patients. KSHV is thought to contribute to tumorigenesis through several mechanisms. Some viral products induce the production of cytokines and growth factors that can influence uninfected neighboring cells

Human papilloma virus (HPV)

Is a doublestranded DNA virus that infects the epithelial cells of skin and mucosa. The epithelial surfaces include all areas covered by skin and/or mucosa such as the mouth, throat, tongue, tonsils, vagina, penis, and anus. Transmission of the virus occurs when these areas come into contact with a virus, allowing it to transfer between epithelial cells. HPV plays an important role in the cause of cervical cancer.

Epstein-Barr virus (EBV)

Is a gamma-herpesvirus, and encodes several viral proteins that affect host gene expression. EBV has been shown to act as a causative factor in B-cell lymphoproliferative diseases and probably nasopharyngeal carcinoma. However, it is thought to be only a contributing factor for a type of cancer called Burkitt's lymphoma (it acts along with malaria to generate this cancer).

An oncogene

Is a gene that has the potential to cause cancer. In tumor cells, they are often mutated or expressed at high levels

A proto-oncogene

Is a normal gene that can become an oncogene due to mutations or increased expression. The resultant protein encoded by an oncogene is termed oncoprotein. Proto-oncogenes code for proteins that help to regulate cell growth and differentiation. Proto-oncogenes are often involved in signal transduction and execution of mitogenic signals, usually through their protein products. Upon activation, a proto-oncogene (or its product) becomes a tumor-inducing agent, an oncogene.

Platelet-derived growth factor (PDGF)

Is a polypeptide that is normally important for extracellular matrix production, but overexpression may result in proliferation as a result of autocrine stimulation. Moreover, PDGF is a component of a wound response and its normal role is to stimulate epithelial cells around the wound edge to proliferate and repair the damage.

Ubiquitination

Is a post-translational modification (an addition to a protein after it has been made) where ubiquitin is attached to a substrate protein. The addition of ubiquitin can affect proteins in many ways: - It can signal for their degradation via the proteasome, - alter their cellular location, - affect their activity, and - promote or prevent protein interactions.

NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells)

Is a protein complex that controls transcription of DNA. NF-κB is found in almost all animal cell types and is involved in cellular responses to stimuli such as stress, cytokines, free radicals, ultraviolet irradiation, oxidized LDL, and bacterial or viral antigens. NF-κB plays a key role in regulating the immune response to infection (κ light chains are critical components of immunoglobulins). Incorrect regulation of NF-κB has been linked to: - cancer, - inflammatory, and autoimmune diseases, - septic shock, - viral infection, and - improper immune development.

a transcription factor (sometimes called a sequence-specific DNA-binding factor)

Is a protein that binds to specific DNA sequences, thereby controlling the rate of transcription of genetic information from DNA to messenger RNA. Transcription factors perform this function alone or with other proteins in a complex, by promoting (as an activator), or blocking (as a repressor) the recruitment of RNA polymerase (the enzyme that performs the transcription of genetic information from DNA to RNA) to specific genes

A fibroblast

Is a type of cell that synthesizes the extracellular matrix and collagen, the structural framework (stroma) for animal tissues, and plays a critical role in wound healing. Fibroblasts are the most common cells of connective tissue in animals.

Viral oncogenes / oncovirus

Is a virus that can cause cancer

Mdm2

Is an important negative regulator of the p53 tumor suppressor. Mdm2 protein functions both as an E3 ubiquitin ligase that recognizes the N-terminal trans-activation domain (TAD) of the p53 tumor suppressor and an inhibitor of p53 transcriptional activation.

Tumor protein 53 (p53; encoded by the TP53 gene in human)

Is induced when DNA is damaged or in the case of other stresses such as hypoxia and cell cycle abnormalities.

Carcinogenesis or oncogenesis or tumorigenesis

Is the actual formation of a cancer, whereby normal cells are transformed into cancer cells. The process is characterized by a progression of changes at the cellular, genetic, and epigenetic level that ultimately reprogram a cell to undergo uncontrolled cell division, thereby forming a malignant mass.

Retinoblastoma (Rb)

Is the firstly identified tumor suppressor gene. Children with hereditary retinoblastoma inherit a single defective copy of the RB gene, sometimes seen as a small deletion on chromosome 13. They develop retinal tumors early in life and generally in both eyes.

Ras

Is the most commonly mutated oncogene in cancer, with 10-20% of tumors harboring mutations in Ras. Ras mutations are found in large number of tumors of the colon, pancreas, and thyroid. In the inactive state, Ras binds GDP

Fibroblast growth factors (FGFs)

Make up a large family of polypeptide growth hormones normally expressed during proliferation of cells required for normal wound healing, but overexpression can lead to tumor formation

Different locus, different allele, same phenotype

Many cancer susceptibility syndromes are genetically heterogeneous (a mixture), which means that different mutations (genotypes) can be expressed as the same phenotype (e.g., cancer). These different mutations may be located within the same gene but at different locations (locus heterogeneity) or on different genes altogether (allelic heterogeneity).

Tumor suppressor genes

Most cancer susceptibility genes are tumor suppressor genes. Tumor suppressor genes are just one type of the many genes malfunctioning in cancer. These genes, under normal circumstances, suppress cell growth. Some do so by encoding transcription factors for other genes needed to slow growth. For example, the protein product of the suppressor gene TP53 is called p53 protein. It binds directly to DNA and leads to the expression of genes that inhibit cell growth or trigger cell death. Other tumor suppressor genes code for proteins that help control the cell cycle. Both copies of a tumor suppressor gene must be lost or mutated for cancer to occur

Oncogenes

Most cancers have mutations in proto-oncogenes, the normal genes involved in the regulation of controlled cell growth. These genes encode proteins that function as growth factors, growth factor receptors, signal-relaying molecules, and nuclear transcription factors (proteins that bind to genes to start transcription).

Autosomal Dominant Inheritance

Most hereditary cancer syndromes are inherited in autosomal dominant fashion. Dominant inheritance occurs when only one copy of an allele is required for a particular trait to be expressed (phenotype). In autosomal dominant inheritance, multiple generations express the traits, with no skipped generations

Function of proto oncogenes

Most proto-oncogenes encode proteins that participate in signal transduction pathways through which signal to divide (or not divide) are relayed from outside the cell to the regulatory machinery within

Regulation of p53 protein by MDM2

Normally, the level of p53 protein in a cell is low. The activity of p53 in a cell is regulated at the level of protein degradation, not at the level of expression of the p53 gene. The MDM2 protein, a ubiquitin ligase, is its main regulator.

Rous sarcoma virus (retrovirus of chicken)

Oncogenes were first discovered as viral (v-) oncogenes in retroviruses that cause tumors

Growth factors (GFs)

Overexpression of these polypeptide growth hormones can elicit (in cells normally in the G0 phase) an aberrant transition from G0 to G1, with subsequent uncontrolled growth.

The retinoblastoma protein (Rb)

Prevents the cell from replicating damaged DNA by preventing its progression along the cell cycle through G1 into S. Rb binds and inhibits transcription factors of the E2F family, As long as E2F is inactivated, the cell remains stalled in the G1 phase.

Steps of The multiple-hit hypothesis

Progression toward an adenocarcinoma begins with the activation of the K-ras (Kristen rat sarcoma) oncogene and the deletion of p53 gene. Deletion of the DDC (deleted in colorectal carcinoma) gene initiates cell surface changes and further mutations promotes metastasis. The specific mutations described are characteristic, but can occur in any order.

Gene therapy of tumors 1a. Down-regulation of proto-oncogenes

RNA interference is used to down-regulate the overexpression of proto-oncogenes.

Epidermal growth factor (EGF)

Signaling serves as an important model for how a signal from an extracellular growth factor can be transduced through a cell, regulate gene expression, and trigger cell proliferation. The EGF receptor (EGFR) is a tyrosine kinase receptor

Holes in the two-hit hypothesis

So, according to the two-hit hypothesis, tumor suppressors act recessive at the phenotypic level (both alleles must be mutated/lost for cancer to develop). However, it turned out that this hypothesis is not true for the inherited form of retinoblastoma itself, because it is actually inherited in an autosomal dominant fashion.

The role of inflammation in the cancer formation

Some viral and bacterial infections induce a chronic inflammatory response that contributes to the process of carcinogenesis. Inflammation is a host defense mechanism against infectious agents and injury, as it is a consequence of wound healing. Under normal conditions it is highly regulated and short lived: such acute inflammation typically resolves itself with the help of anti-inflammatory factors. By contrast, recent evidence suggests that it is lingering, chronic inflammation that plays an important role in causing cancer.

The multiple-hit hypothesis

The classical example of the principle that several consecutive mutations are needed for the development of most tumor

TUMOR-SUPPRESSOR GENES

The human body has mechanisms exerted by tumor suppressor genes that normally 'police' the processes that regulate cell numbers and ensure that new cells receive DNA that has been precisely replicated. Many tumor suppressor gene products act as stop signs to uncontrolled growth and therefore may inhibit the cell cycle, promote differentiation, or trigger apoptosis. Both alleles of a tumor suppressor gene must lose their function before a tumor develops.

Signal transduction proteins

The next level at which defects in cell growth and development can occur is at the level of downstream signal transduction proteins.

Growth factor receptors (GFRs)

The normal binding of GFs to their receptors results in signal transduction in response to receptor dimerization. Several GFRs have been identified that are capable of activation, even in the absence of specific ligand. Many of these receptors have intracellular domains that function as tyrosine kinases.

P53: fix or kill!

The p53 gene is a tumor-suppressor and a pro-apoptotic gene. The p53 protein prevents a cell from completing the cell cycle if (1) its DNA is damaged or (2) the cell has suffered other types of damage. When the damage is minor, p53 halts the cell cycle — hence cell division — until the damage is repaired. When the damage is major and cannot be repaired, p53 triggers the cell to commit suicide by apoptosis

Metastasis

The process by which tumor cells migrate from a primary site to other parts of the body is called metastasis. Metastasis is the fundamental difference between a benign and malignant growth and represents the major clinical problem of cancer.

K-ras oncogene

The protein product of the normal KRAS gene performs an essential function in normal tissue signaling, and the mutation of a KRAS gene is an essential step in the development of many cancers. Like other members of the Ras family, the KRAS protein is a GTPase and is an early player in many signal transduction pathways

BRCA1 and BRCA2 genes are very different, at least some functions are interrelated.

The proteins made by both genes are essential for repairing damaged DNA.

Two major classes of genes are thought to be involved in the formation of tumors,

The protooncogenes and tumor suppressor genes.

Hunting for oncogenes (1) Discovery of c-src proto-oncogene.

The src gene (v-src) was isolated from the genome of Rous sarcoma virus (RSV) followed by radiolabeling and hybridization with the chicken genome. Hybridization signals in Southern blot analysis indicate the presence of chicken genes (v-src) homologous to that of virus oncogene.

Epidermal growth factor receptors (EGFRs):

There are at least three members of the family of tyrosine kinase receptors, erbB-1, erbB-2, and erbB-3, which when mutated, lead to errant signaling and growth in the absence of cognate ligand. Overexpression of erg b-1 (also known as HER2; human epidermal growth factor receptor 2) is associated with breast cancers.

Cell surface molecules

There are numerous cell surface molecules that antagonize normal cell growth. (a) Transforming growth factor (TGF)-β receptor mediates its inhibitory effects by stimulating the production of CDK-Is. (b) Protein product deleted in colon carcinoma (DCC) regulates cell growth through the integration of signals from the cellular environment

Hepatitis B virus (HBV)

There is a strong association between hepatitis B virus infection and liver cancer (hepatocellular carcinoma). Host-viral interactions evoke an immune response that results in liver necrosis, inflammation, and regeneration.

Intracellular signal transduction proteins

These molecules possess an antagonistic role to the actions of intracellular proto-oncogenes

Transcription factors

Transcription must be considered as an ultimate target of oncogene action. Many oncogenes act directly as transcription factors. The transcription factors are expressed at very low levels at quiescent cells but are quickly turned on when cells receive signal to divide and are rapidly translocated to the nucleus to mediate gene transcription.

Hunting for oncogenes (2) Cloning of a human oncogene.

Transfer of DNA from human bladder tumor cells to cultured mouse cells yielded transformed cells. In the process the transformed cells acquired an activated human oncogene, and approximately a million base pairs of irrelevant human DNA from the tumor cells. Human DNA can be detected in mouse cells that by using a radiolabelled probe for the Alu family of repetitive sequences, which is present every few thousand base pairs in the human genome (but not in the mouse DNA). Therefore, an Alu probe hybridizes to many restriction fragments in a Southern blot of DNA from a primary transfectant

Gene therapy of tumors 1e. Delivery of angiogenesis inhibitor genes to tumors

When tumors reach the size of a pea, they require the formation of microcapillaries for efficient feeding. Delivery of antiangiogenesis factors to the tumors can cause the starvation and eventually death of the tumor.

Traditional anti-tumor therapies

Whole-body therapies: - chemotherapy, - radiotherapy Local therapies: - surgery, - chemotherapy, - radiation therapy, - local heat therapy

Loss of function of one allele predisposes the cell to tumor development.

With a germinal mutation, all cells are predisposed. The tumor arises after loss of function of the second allele. When somatic mutation occurs in a single cell, loss of function of both alleles rarely affects the same cell. But with a germ cells mutation, loss of function of the second allele is frequent, since all cells carry the first mutation, i.e., are predisposed.

E7 protein

acts as the primary transforming protein. E7 competes for retinoblastoma protein (pRb) binding, freeing the transcription factor E2F to transactivate its targets (genes encoding enzymes involved in DNA replication), thus pushing the cell cycle forwards.

A typical retrovirus contains

an RNA genome that codes for three genes or groups of genes: - gag (group-specific antigen), - pol (polymerase), and - env (coat protein, envelope). As with all genes of higher organisms, a cellular oncogene (c-onc) consists of exons and introns with defined structure and sequence, as in the gene src (the name is derived from sarcoma, a tumor that is induced by a mutation in this gene). The virus may contain parts of the cellular oncogenes (c-src). This is designated viral oncogene (v-src)

Epigenetic factors

are mechanisms outside the gene such as a cell's exposure to carcinogens or hormones, or genetic variations that modify a gene or its protein by - methylation, - demethylation, - phosphorylation, or - dephosphorylation. These factors can alter what is ultimately expressed; they can change a phenotype. For example, hormone and reproductive factors may influence the penetrance of certain cancer-linked mutations

Gene therapy of tumors 1d. Delivery of immunomodulatory genes (interleukins, tumor necrosis factors) to tumor cells

can trigger the recognition of tumor cells by the immune system.

adenocarcinomas

cancers of glandular tissue (e.g. breast)

carcinoma

cancers that occur in epithelial cells are classified as carcinoma

Two-hit hypothesis

early onset at multiple sites in the body of an inherited form of cancer called hereditary retinoblastoma. Inheriting one germline copy of a damaged gene present in every cell in the body was not sufficient to enable this cancer to develop. A second hit (or loss) to the good copy in the gene pair could occur somatically, though, producing cancer. This hypothesis predicted that the chances for a germline mutation carrier to get a second somatic mutation at any of multiple sites in his/her body cells was much greater than the chances for a noncarrier to get two hits in the same cell.

Women with an abnormal BRCA1 or BRCA2 gene

have up to an 60% risk of developing breast cancer by age 90; increased risk of developing ovarian cancer is about 55%

Viral E6 protein

in association with host E6 associated protein, which has ubiquitin ligase activity, act to ubiquitinate p53, leading to its proteasomal degradation.

Individuals with sporadic retinoblastoma,

in contrast, inherit two normal RB alleles each of which has undergone a somatic loss-of function mutation in a single retinal cell. Because this is an unlikely occurrence, sporadic retinoblastoma is rare, develops late in life, and usually affects only one eye

Loss of heterozygosity (LOH)

is a gross chromosomal event that results in loss of the entire gene and the surrounding chromosomal region.

Gene therapy of tumors 1c. Suicide therapy

is based on the delivery of the tk (thymidine kinase) gene of herpes simplex virus to tumor cells. Addition of acyclovir (an anti-herpes virus drug) converts acyclovir to toxic product, which in turn arrests the cell cycle.

Certain variations of the BRCA1 gene...

lead to an increased risk for breast cancer.

chronic inflammation, in the absence of an infectious agent,

leads to an increased risk of cancer. A key mediator of the inflammatory response is the transcription factor NF-kB. It is induced by several cell types, such as macrophages, target cells of inflammation, and cancer cells. In addition to inflammation, NF-kB has other downstream effects that contribute to tumorigenesis, such as the: - inhibition of apoptosis and - the promotion of metastasis and angiogenesis.

Serine/threonine kinases

phosphorylate serine and threonine residues.

Tyrosine kinases

phosphorylate tyrosine residues in target proteins.

Gain-of-function mutations,

which are much less common, confer an abnormal activity on a protein.