L10: The Hallmarks of Cancer PDF

**L10: The Hallmarks of Cancer** A diagram of a brain Description automatically generated **\ **The Hallmarks of Cancer framework, published in 2000, identifies key features that drive cancer. These include: 1. Sustained Proliferative Signaling 2. Evasion of Anti-Proliferative Signaling 3. R...

**L10: The Hallmarks of Cancer** A diagram of a brain Description automatically generated **\ **The Hallmarks of Cancer framework, published in 2000, identifies key features that drive cancer. These include: 1. Sustained Proliferative Signaling 2. Evasion of Anti-Proliferative Signaling 3. Resisting Cell Death (Evasion of Apoptosis) 4. Enabling Replicative Immortality 5. Activation of Invasion and Metastasis 6. Angiogenesis ### **1. Sustained Proliferative Signaling** - In normal cells, division is tightly controlled by mitogenic signaling molecules and receptors. - Cancer cells bypass these controls by producing their own growth signals, becoming more sensitive to normal signals, or mutating receptors to be independent of external growth factors. - Example: HER2 amplification in metastatic breast cancer---amplification of the HER2 gene on chromosome 17 leads to overexpression of the HER2 receptor, making the tumor responsive to targeted therapy (Herceptin), which inhibits the receptor\'s signaling cascade, 'sensitising' the tumour to human epidermal growth factor. - Amplification status of Her2 graded into 4 categories. There needs to be on average twice as many genomic copies of the HER2 gene as there are copies of chromosome 17, for patients to benefit from Herceptin treatment. ### **2. Evasion growth suppressors** - Tumor suppressor genes (TSGs) normally prevent excessive cell growth by regulating the cell cycle. In cancer, these genes are often mutated or silenced e.g. methylation of promoter region. - Key Genes: TP53 (on chromosome 17) and RB (on chromosome 13) are examples of critical TSGs involved in growth suppression. - Example: Retinoblastoma occurs when RB is inactivated, allowing uncontrolled cell cycle progression. RB usually governs G1/S and G2/M transitions via complexes with E2F. Most cancers find a way to impair RB function. ### **3. Resisting Cell Death (Evasion of Apoptosis)** - Apoptosis is a programmed cell death mechanism that removes damaged or unneeded cells. Cancer cells often resist apoptosis by altering apoptotic pathways. - Regulators of Apoptosis: The Bcl2 family of proteins is frequently involved in cancer, with Bcl2 acting as an anti-apoptotic protein. - Example: In B-cell lymphoma, BCL2 is overexpressed due to a translocation with the IGH gene, preventing apoptosis and promoting tumorigenesis. - TP53 (pro-apoptotic), mutated in roughly 50% of all cancers ### **4. Enabling Replicative Immortality** - Cancer cells bypass cellular senescence (growth arrest entered when telomeres become too short) and crisis (genomic instability due to shortened telomeres) to maintain indefinite growth. - Telomerase Activation: Telomerase is often reactivated in cancer cells, preventing telomere shortening and extending the replicative lifespan of cells. - Challenge in Treatment: Telomerase inhibition is a potential therapy due to near universal association with malignant cells and lack of expression in normal cells, but its effects are slow to manifest, and the enzyme\'s 3D structure remains elusive, complicating drug development. ### **5. Activation of Invasion and Metastasis** - Cancer cells gain the ability to invade surrounding tissues and spread to distant sites through the blood or lymphatic system. This process involves changes in cell adhesion and motility. - Genes normally involved in cell migration, e.g. during early embryonic development. Are upregulated. - Key Proteins Involved: E-cadherin (normally involved in epithelial cell adhesion) is often downregulated in metastatic cancers, while N-cadherin (involved in neuronal and mesenchymal cell migration) is upregulated in high-grade carcinomas. ### **6. Angiogenesis** - Tumors need blood vessels to supply nutrients and oxygen to sustain their growth. Cancer cells can induce angiogenesis (the formation of new blood vessels). - Key Genes Involved: RAS and MYC are critical in angiogenesis and also play roles in cell growth, survival, and metastasis. - Example: Activating mutations in RAS are found in many cancers, such as pancreatic, bowel, and lung cancers. RAS interacts with Her2.  **Emerging Hallmarks of Cancer\ In addition to the six hallmarks, other capabilities have been identified as playing significant roles in cancer:** - Deregulating Cellular Energetics: Cancer cells often reprogram their metabolism to support their rapid growth. - Avoiding Immune Destruction: Cancer cells can evade immune surveillance, which is an area of active therapeutic research. - Promoting Inflammation & Loss of Genome Stability: These features contribute to cancer progression. ### **Clonal Expansion and Clonal Evolution** - Cancer involves the accumulation of genetic mutations that provide a competitive advantage. These mutations lead to clonal expansion, where the most competitive cells dominate. Given the genomic instability of such clones, they are likely to acquire additional abnormalities leading to further evolution of that clone. - Clonal Evolution: Tumors evolve as clones acquire further mutations, leading to greater genetic heterogeneity, which is a hallmark of aggressive cancers. - Mutation rates in cancer cell lines are higher due to a variety of reasons: deregulation of DNA damage repair pathways, disturbance of chromosome disjunction events, sensitivity to genotoxic agents (mutagens, carcinogens, teratogens)... ### **BRCA1 and BRCA2 Mutations** - BRCA1 and BRCA2 are tumor suppressor genes that play a crucial role in repairing DNA double-strand breaks via homologous recombination. Mutations in these genes greatly increase the risk of breast, ovarian, and other cancers. - Inheritance of Mutations: Inherited mutations in BRCA1 or BRCA2 significantly increase the lifetime risk of developing breast cancer (50-80%) and ovarian cancer (30-50%). - BRCA1/2 genes both function in homologous recombination, a highly accurate repair pathway for DNA DSBs, particularly important for breaks formed during DNA replication. So far on this module we have explored the cyto-genomics of pre-natal diagnosis, and touched on some of the most common genetic diseases affecting this community of patients such as infertility, miscarriage, and severe genetic diseases caused by large genomic imbalances or chromosomal rearrangements. You then heard from Duncan Baker, who provided an overview of common molecular genomic diagnostic technologies and some examples of rare diseases for which these technologies are most suited for diagnostic insight. Although individually rare, there are many of these type of diseases, so collectively they are quite common. It is estimated that between 3.5-5.9% of the population have or are likely to develop a rare disease in their lifetime, this equates to approximately 300 million individuals globally. In the final 6 lectures of this module we will look beyond cases of inherited disease, and focus on the genetics of cancer, which in the majority of cases is acquired during one\'s lifetime. To start off therefore, in this lecture, we will summarise a landmark review - "The Hallmarks of Cancer" - which provides a framework to categorise and make sense of the causative abnormalities that also drive and progress instances of cancer. I have made this review available on google classroom to help you further your understanding of content covered in this lecture. I have also provided other papers, which explore individual examples of important genes covered by this lecture and the review. I do not expect you to read all of this material for the exam, but being able to discuss one or a few of these examples in more depth will help you to demonstrate evidence of further reading and depth of understanding in the exam. Although cancer can affect individuals of any age, it is far more common in later life, particularly from the sixth decade onwards. New cases of cancer are estimated to affect approximately 600-700 individuals per 100,000 [[(Cancer Research UK)]](https://www.cancerresearchuk.org/health-professional/cancer-statistics/incidence/all-cancers-combined#heading-One), with an associated mortality rate of roughly 160 per 100,000, that is slightly higher for the male population, and lower for females. (NIH\_NCI, 2020-US population). Breast, lung, prostate, colon, skin and bladder are among the most common cancers, followed by the haematological malignancies that affect the bone marrow, lymphatic system an the blood. These diseases are characterised by clonal heterogeneity, meaning that the disease is composed of a complex mixture of multiple cell lines/clones some of which is classified as apparently normal tissue, which is thought to be recruited by the cancer to sustain its growth and proliferation. In 2000, The Hallmarks of Cancer was published to provide a simplified and accessible framework to categorise and make sense of the huge variety of genetic abnormalities observed in cancer. These include: 1. Sustained proliferative signalling 2. Evasion of anti-proliferative signalling 3. Resisting cell death 4. Enabling replicative immortality 5. Activation invasion and metastasis and, 6. Angiogenesis This lecture will summarise each of these important features of cancer and explore in more detail some common examples, which also serve as diagnostic and prognostic markers of disease The first of these hallmarks is sustained proliferative signalling. Under normal circumstances, the decision by a cell to divide is tightly regulated. In general the process is dependent on the release of mitogenic signalling molecules, the detection of the presence of these signals by cell surface receptors, and signalling cascades inside the cell, that bring about a response to the singal and result in the growth and division of the cell. Cancerous tissues find a way around this regulation - and it's apparent that they do this in a number of different ways. For example, cancers are able to upregulate and release their own mitogenic signalling molecules, or stimulate the normal tissues around them to to do the same. Alternatively cancerous cells can become more sensitive to the growth factors, so that they are able to grow and divide out of control even in the presence of normal levels of mitogenic signalling molecule. In addition to this, from receptor, phosphorylation cascade to transcription factor - activation mutations may be acquired by cancerous cells, which completely uncouple them from the dependance on outside factors. An example of one of these is shown in the slide. Upregulation and overexpression of the The Human Epidermal Growth Factor Receptor 2 (Her2) on the surface of malignant cells is observed in cases of metastatic breast cancer. This upregulation is caused by amplification of the HER2 gene, on chromosome 17. Amplification above a certain threshold qualifies the patient for treatment with a monoclonal antibody called Herceptin, which specifically binds to the receptor and inhibits intracellular signalling cascade, thus "sensitizing" the tumour to human epidermal growth factor. The amplification status of Her2 is graded into 4 categories, shown on the right of the slide. Sections of 4 different breast tumours are shown. Nuclei appear a bluish purple colour as the chromatin has been stained using a dye similar to leishmans (used in G-Banding). You should also be able to see a brownish precipitate that seems to have localised to the surface of cell membranes. This precipitate is a product that has been generated by the enzyme "horseradish peroxidase", has been specifically conjugated to the Her2 protein via a complex of antibodies. Treatment with Herceptin is not curative, but it does help to slow down the rate disease progression and metastasis to provide patients with extended life expectancy. However, to qualify for this treatment the HER2 gene must be amplified above a certain threshold. This forms the cornerstone of diagnostics for this type of tumour. The goal is to identify patients who are expected to respond to herceptin based on the amplification status of the HER2 gene. Patients who show an obvious and massive over expression of Her2 (as seen at the bottom of the slide (classified 3+) -- are prescribed Herceptin. Those classified as zero do not show any evidence of overexpression. The staining you can see is therefore thought to represent normal expression levels of this protein. These patients, and those with low levels of amplification (+1) are not offered herceptin, as they do not show sufficient response to the drug. Borderline cases need to be investigated by more quantitative and precise means, and are promptly sent for cytogenomic analysis for further investigation by FISH. You can see the FISH results for HER2 amplification of the right of the slide for 4 different borderline patients. [[The HER2 gene has been painted red. The gene is located on chromosome 17, the centromere of which has been painted green]](https://www.ogt.com/products/product-search/cytocell-her2-erbb2-amplification/). Normal cells should therefore have 2 green signals and 2 red signals. The borderline patient shown top left is therefore negative for HER2 amplification, as there appears to only be 2 copies of the gene. In this case -- herceptin would not be recommended. Top right, there is evidence of amplification, but the ratio is approximately 1.5, they would not be offered herceptin. Only patients who have a red to green signal ratio of 2 or more are expected to benefit from herceptin treatment. In other words there needs to be on average twice as many genomic copies of the HER2 gene as there are copies of chromosome 17. The tumours at the bottom of the slide -- clearly show evidence of massive HER2 amplification, way above the required threshold ratio of 2. These patients would qualify for treatment with herceptin. Herceptin prevents dimerisation of Her2, which suppresses the downstream proliferative signalling of this receptor by the Ras signalling cascade. Another way for cancers to uncouple from normal growth and proliferative regulation, is to become desensitised to growth suppressors. The genes involved in normal growth and proliferation suppressions are commonly referred to as tumour suppressor genes. In cancer, their function is commonly lost via mutation, transcription suppressions e.g. via methylation of a promoter region, by deletion or altered regulation of genes that regulate TS expression. TP53 on chromosome 17, and RB on 13 are the two most prominent examples of this in cancer. The example shown on the right of the slide, is that of retinoblastoma. This gene is downregulated, deleted or inactivated in most cancers. This protein has a crucial role in cell cycle progression in normal cells. RB forms a complex with E2F, which essentially functions to downregulate the transcription of genes involved in G1/S and G2/M transitions - thus loss of RB is a major way for cancer cells to avoid this growth suppression. Evasion of apoptosis is also a common hallmark of cancer. The components that regulate apoptosis are clustered into those involved in an intrinsic pathway that sense and transduce signals from inside the cell, and those involved in a extrinsic pathway, which receive and process extracellular signals. The genes involved in sensing apoptotic signals are called REGULATORS. Intrinsic and extrinsic signals are carried by apoptotic triggers, which either promote (TS) or suppress (oncogene) the decision for a cell to apoptose. The Bcl2 family of proteins are a good example to remember for this class of abnormality as the are commonly affected in cancer. The triggers regulate the apoptotic effectors, which are a family of proteases (caspases) and once activities, will progressively dismantle the cell. TP53 is another very good example to remember - as this gene is mutated in around 50% of all cancers. This slide shows diagnostic examples of both genes that I have just mentioned. TP53 would be classified as a regulator protein and is involved in sensing DNA damage, hypoxia and oncogene overexpression. The gene is a negative regulator of apoptosis, and is commonly lost in cancer. When lost it is commonly associated with poor prognosis. For example, MDS - a type of precancerous disorder of the bone marrow, can if left untreated to progress and transform into an aggressive form of acute myeloid leukaemia. The loss of TP53 via deletion (shown top right) and mutation/inactivation is common - and a marker of likely progression. In the example shown, a normal 17 is shown at the bottom of the image - and an abnormal 17, where the short arm has been lost via structural rearrangement, is missing the TP53 gene. BCL2 has anti-apoptotic activity. Variants of this gene that result in over activation or inappropriate expression are commonly associated with tumourigenesis. In the example shown (bottom right of the slide), the BCL2 genehas been fused (via translocation) to IGH. This rearrangement couples the expression of BCL2 to the IGH promoter - thus resulting in overexpression of BCL2. This abnormality is a common marker and is frequently diagnostic of B-cell neoplasms, which are a type of lymphoma. Normal cells have a replicative lifespan - but to create a tumour mass it is widely accepted that cells need to be immortal. To achieve this hallmark, cells must overcome senescence and crisis. Senescence is an irreversible non-proliferative state, which is entered when telomeres become too short. You should be aware that telomeres are protective caps at the end of each chromosome arm, consisting over long sequences of a hexanucleotide repeats. Each round of DNA replication results in the progressive shortening of telomere caps, because he normal replication apparatus is not able to copy all the way to the end. Over many rounds of DNA replication/cell division - telomeres become so short that the protective structures at the end are no longer formed - resulting in chromosome-chromosome fusions. These structures are unstable, and their presence in the cell triggers crisis - which leads to apoptosis. Cancer cells have been found to avoid senescence by either activating the expression of the enzyme telomerase (which permits replication to the very end of telomeres, and thus prevents telomere shortening) - or by upregulating recombination based telomere maintenance mechanisms. Telomerase is a very promising therapeutic target, due to its near universal association with malignant cells, and lack of expression in non-malignant/normal cells. But - the lack of a 3D structure of the enzyme has hampered efforts to target this protein with inhibitors. Moreover, once telomerase has been inhibited (hypothetically) - it's expected for there to be a long delay before senescence/crisis initiation... So although this protein shows the hallmarks of a potential target - the therapeutic impact would likely be far slower than what would be perceived as clinically desirable. In order for a small malignant mass of cells to proliferate and grow into a life threatening tumour - the cells must find a way to sequester enough nutrients to support the enhanced metabolism in that tissue. This is done by stimulating the growth of new blood vessels. There are many genes involved in angiogenesis - but there are two genes in particular that I'd like you to remember. These are RAS and MYC. The reason why these genes are important ,particularly for cancer diagnostics, is because they feature in many other hallmarks, and are common to many cancers. In the example shown - activating mutations in RAS are associated with around 70% of Pancreatic cancers, around 60% of Bowel cancers, and 30% of melanomas, lung and cervical cancers - to name but a few. In addition to angiogenesis, RAS is also involved in cell growth and proliferation (e.g. via interaction with Her2 which we considered earlier, survival, cell migration and invasion). A trait acquired by late stage cancers, is an ability to spread to other tissues and parts of the body, that are remote to the initial lesion. This process is called invasion and metastasis. Cells that have acquired this ability are commonly characterised by the depletion, inhibition or loss of key products involved in cell-cell interactions or cell-ECM interaction. In addition - the genes that are normally involved in cell migration, for example during early embryonic development - are also found to be upregulated. E and N Cadherins are two good examples to remember. E-Cadherin normally acts to help assemble epithelial cell sheets, and is commonly depleted in metastatic cancers. Conversely, N-Cadherin, which is normally expressed in migrating neurons and mesenchymal cells during embryonic development - is often upregulated and high grade carcinomas. The formation of metastases follows a gradual 6 stage process, beginning with local invasion to juxtaposed tissues, followed by entry into the blood or lymphatic system. Malignant cells are the transported through the vascular system until they escape into a distant tissue, where they must form a small node, that grows into a larger malignant tumour mass. Since the original publication of the Hallmarks of Cancer in 2000 - there have been other cellular capabilities that have emerged as key features of cancerous growth. These hallmarks include the deregulation of cellular energetics to support the continuous growth and proliferation of a malignant mass. Avoiding destruction by the immune system - which will surely lead to very exciting therapeutic targets in the future - in fact some exist already. THe promotion of inflammation and the loss of genome stability. You do not need to know all of these in detail for this module - but you are welcome to engage in further reading to learn more. THE EMERGING HALLMARKS WILL BE TREATED AS FURTHER READING IN THE EXAM Ultimately the acquisition of multiple hallmarks is dependant on the accumulation of a critical mass of genomic alterations, that uncouple the cell from vital cellular pathways. Mutations in these cells occur randomly, and only those that confer an advantage to the host end up featuring as major biomarkers in the cancer. This principle leads on to two important features of tumourigenic behaviour - Clonal expansion and clonal evolution. Clonal expansion is the term used to describe the propagation of a particularly competitive cell line within a tumour. Given the inherent genomic instability of such a clone - these cells are likely to acquire additional abnormalities, which if they confer an additional advantage - leads to the evolution of that clone. This tumours are often charaterised as highly heterogeneous populations of cells, some belonging to major clones, others belonging to minor clones - which may be dwindling in number due to a lack of competitiveness - or growing in number as they have newly acquired a competitive edge. Cancers that have gained and inability to stability maintain their genome are thus empowered with the potential to colonise the initial site, and develop all the hallmarks needed to mature into high grade metastatic disease. Mutation rates in many cancer cell lines are higher due to a variety of reasons, including increased sensitivity to genotoxic agents, deregulation of DNA damage sensing components and DNA damage repair pathways, and disturbance of chromosome disjunction events. The example given concerns two of the best known genes in cancer studies. Mutation/inactivation of the breast cancer susceptibility genes BRCA1 and BRCA2 (both TSs) confers the greatest risk for the development of breast and ovarian cancer. Inheritance of either inactivated gene, leads to a high risk of early onset BC - and a lifetime risk of 50-80% BC, and 30-50% OC. Other cancers are also predisposed such as prostate, stomach and pancreatic. The BRCA1/2 genes both function in HR, which is a high fidelity (i.e. highly accurate) repair pathway for DNA DSBs. ALthough there are several different break repair pathways, HR is particularly important for breaks formed during DNA replication - which features heavily of course in highly proliferative tissues or cancer.