Molecular, Cellular and Genetic Basis of Cancer

MOLECULAR, CELLULAR AND GENETIC BASIS OF CANCER https://cdn.the-scientist.com/assets/articleNo/70781/aImg/48671/cancer-cells-articlel.webp Dr Katherine Karakoula A.Karakoula@wlv.ac.uk Learning outcomes ➢ Understand the concepts of the molecular, cellular and genetic basis of cancer. ➢ Recognise the role of ▪ proto-oncogenes ▪ oncogenes and ▪ tumour suppressor genes in the development of cancer. ➢ Other factors that could cause cancer. ➢ Recognise important deregulated pathways and associated genes involved in the development of cancer. Cellular Basis of Cancer • Cancer is characterized by abnormal and uncontrolled growth • In normal tissues, the rates of new cell growth and cell death are kept in balance • In cancer, this balance is disrupted • This disruption can result from 1) uncontrolled cell growth or 2) loss of a cell's ability to undergo apoptosis What is the molecular basis of cancer? CANCER: GENETIC DISEASE • Mutations in genes result in altered proteins • During cell division • External agents • Random event • Most cancers result from mutations in somatic cells • Some cancers are caused by mutations in germline cells 5 Malignant versus Benign Tumors Benign or low grade (I or II) tumours: • Slow growing • Less likely to come back • Do not spread to other parts of the brain or spinal cord Malignant or high grade (III or IV) tumours: • Life-threatening and fast growing • Likely to come back • Capable of spreading by invasion and metastasis https://teachmephysiology.com/wp-content/uploads/2017/03/cellcycle-1.png The cell cycle is a four-stage process. The stages G1, S, and G2 make up interphase, which accounts for the span between cell divisions. Cancer is Caused by Nonlethal Genetic Mutations Affecting Certain Genes. ➢ Proto-oncogenes ➢ Oncogenes ➢ Tumor suppressors What is a proto-oncogene? Genes are made of sequences of DNA that contain the information necessary for the cells to function and grow properly. Genes contain instructions (codes) that tell a cell to make a specific type of protein. Each protein has a specialized function in the body. A proto-oncogene is a normal gene found in the cell. Each one is responsible for making a protein involved in cell growth, division, regulation of programmed cell death (apoptosis), inhibition of cell differentiation. Essential for cell growth and development and for maintaining healthy tissues and organs. Most of the time, these genes work the way they are supposed to, but sometimes things go wrong. Can proto-oncogenes cause cancer? ➢ A proto-oncogene can’t cause cancer unless a mutation occurs in the gene that turns it into an oncogene. ➢ Everyone has proto-oncogenes in their body. In fact, proto-oncogenes are necessary for our survival. ➢ When a mutation occurs in a proto-oncogene, it becomes permanently turned on (activated). This is called a gain-of-function mutation .The gene will then start to make too much of the proteins that code for cell growth. Cell growth occurs uncontrollably. Uncontrollable cell growth leads to cancer. ➢ These mutations are also considered dominant mutations. This means that only one copy of the gene needs to be mutated in order to start cancer. There are at least three different types of gain-of-function mutations that can cause a proto-oncogene to become an oncogene: ➢ Point mutation. This mutation alters, inserts, or deletes only one or a few nucleotides in a gene sequence, in effect activating the protooncogene. ➢ Gene amplification. This mutation leads to extra copies of the gene. ➢ Chromosomal translocation. This is when the gene is relocated to a new chromosomal site that leads to higher expression. Most of the mutations that cause cancer are acquired, not inherited. This means that you aren’t born with the gene error. Instead, the change happens at some point during a person’s life. Proto-oncogene activation Examples of proto-oncogenes Ras The first proto-oncogene to be shown to turn into an oncogene is called Ras. Ras encodes an intracellular signal-transduction protein. In a normal cell, Ras is transiently activated and recruits Raf, to activate the MAP-kinase pathway to transmit growth-promoting signals to the nucleus. When Ras is mutated, it leads to continuous stimulation of cells without any external trigger causing an uncontrolled growth-promoting signal. Most cases of panceatic cancer have a point mutation in the Ras gene. Many cases of lung, colon, and thyroid tumours have also been found to have a mutation in Ras. HER2 Another well-known proto-oncogene is HER2. This gene makes protein receptors that are involved in the growth and division of cells in the breast. Many people with breast cancer have a gene amplification mutation in their HER2 gene, also known as HER2-positive breast cancer. Myc The Myc gene is associated with a type of cancer called Burkitt’s lymphoma. It occurs when a chromosomal translocation moves a gene enhancer sequence near the Myc proto-oncogene. Cyclin D Cyclin D is another proto-oncogene. Its normal job is to make a protein called Rb tumor suppressor protein inactive. In some cancers, like tumours of the parathyroid gland, Cyclin D is activated due to a mutation. As a result, it can no longer do its job of making the tumour suppressor protein inactive. This in turn causes uncontrolled cell growth. What are oncogenes? A mutated (changed) form of a type of gene called a proto-oncogene, which is involved in normal cell growth and division. When a proto-oncogene is changed so that too many copies are made or it becomes more active than normal, it is called an oncogene. Oncogenes may cause normal cells to become cancer cells and grow in the body. Mutations that lead to the conversion of proto-oncogenes to oncogenes usually occur during a person’s lifetime and are not inherited from a parent. Oncogene mutations are usually a gain of function and are dominant. They regulate cell proliferation, growth, and differentiation, as well as control of the cell cycle and apoptosis. Currently major molecular targets for anti-cancer drug design Selected oncogenes and associated cancers Category / Protein Proto-oncogene Function Growth Factors PDGF (β chain) SIS Fibroblast growth HST-1 factors INT-2 Mode of Activation Overexpression Overexpression Amplification TGFα Overexpression ERB-B1 ERB-B2 PDGF-R Overexpression Amplification Overexpression KIT Point Mutation Transforming growth factor α Growth Factors Receptors EGF-receptor family PDGF receptor Receptor for stem cell (steel) factor Associated Cancer Astrocytoma, osteosarcoma Stomach cancer Bladder & breast cancer Melanoma Astrocytomas Hepatocellular carcinomas SCC of the lung, gliomas Breast and ovarian cancers Gliomas Gastrointestinal stromal tumours Proteins Involved in Signal Transduction GTP-binding K-RAS H-RAS N-RAS Non-receptor tyrosine kinase RAS signal transduction WNT signal transduction Nuclear Regulatory Proteins Transcriptional activators Cell-Cycle Regulators Cyclins ABL BRAF β-catenin Point mutation Colon, lung, pancreatic tumours Point mutation Bladder & kidney Point mutation tumours Melanoma, leukaemia, Translocation lymphoma Point mutation CML, ALL Point Melanomas mutation/Overexpression Hepatoblastomas & HCC C-MYC Translocation N-MYC L-MYC Amplification Amplification CYCLIN D Translocation Amplification CYCLIN E Overexpression Amplification or Point mutation CDK4 Burkitt lymphoma Neuroblastoma, small cell carcinoma of lung SCC of the lung Mantle cell lymphoma Breast & oesophageal cancers Breast cancer Glioblastoma, melanoma, sarcoma Cyclin-dependent kinase Adapted from Table 7-6, Kumar V, Abbas AK, Fausto N, Aster J. Robbins & Cotran pathologic basis of disease, 8th edition. Elsevier; 2010 Tumour Suppressor Genes (TSGs) ➢ Tumor suppressors: genes normally arrest cell division (negative regulators of cell cycle entry or progression - arrest the cell cycle if DNA is damaged or chromosomes are abnormal). ➢ TSG encode proteins that are receptors for secreted hormones that function to inhibit cell proliferation ➢ TSG negative regulators of growth signalling pathways (e.g. APC or PTEN) ➢ TSG gene mutations are usually a loss of function and recessive. The transformation of a normal cell to a cancer cell is accompanied by the loss of function of one or more tumour suppressor genes and both gene copies must be defective in order to promote tumour development. Examples of tumour suppressor genes Inherited Disease Gene Protein function APC Negative regulator of the signalling pathway Adenomatous polyposis coli (APC) BRCA1, BRCA2 Components of DNA repair systems Familial breast and ovarian Spontaneous breast cancers cancer CDH1 E-cadherin, a cell Hereditary adhesion diffuse gastric molecule cancer Many epithelial cancers CDKN2A INK4a, inhibitor of cyclindependent kinase Cdk4 Some esophageal and pancreatic cancers MEN1 Transcription Multiple factor and protein endocrine kinase neoplasia Some familial melanomas Spontaneous Tumours Most colon cancers Many metastatic cancers NF1 PTEN RB SMAD4 TP53 TSC1, TSC2 VHL WT1 Neurofibromin, Neurofibromatosis Ras-GTPase type 1 activation Negative regulator of PI3K growth Cowden disease signalling pathway Repression of transcription factor E2F Retinoblastoma, osteosarcoma Signal transducer in Juvenile polyposis TGF-signalling Transcription Li-Fraumeni factor; guardian syndrome of the genome' Some tumours of neural crest origin 30%-50% of spontaneous cancers Retinoblastoma, sarcomas, several carcinomas Colon and pancreatic cancers Most frequently mutated in human cancers Inhibitor of Tuberous Rare mTOR sclerosis von HippelUbiquitin Lindau Many renal cell carcinomas ligase disease Transcriptio Wilms Some leukaemias n factor tumour Important deregulated pathways in cancer p53/MDM2/p14ARF pathway (decreased apoptosis, increased proliferation) p16INK4a/CDK4/RB1 pathway (loss of cell cycle control) EGFR/PTEN/Akt/mTOR pathway (increased proliferation, cell migration + invasion) Platelet Derived Growth Factor (PDGF) + Receptor (increased proliferation) Other chromosome abnormalities p53/MDM2/p14ARF pathway (decreased apoptosis, increased proliferation) p53 tumour suppressor gene • located on chromosome 17p13.1 • highly conserved gene containing 11 exons encoding 53kDa protein • involved in many human tumours eg colon, breast, bladder, brain tumours etc. In >50% of all human tumours the p53 pathway is aberrant. Inactivation of the p53 protein renders it unable to signal and activate the cell’s apoptotic machinery resulting in increased survival of cancer cells. The p53 protein controls cell division through regulation of transcription of other genes p53 induces transcription of: MDM2 p21 Gadd45 CD45 Bax p53 mutations are most commonly in DNA binding motifs preventing transcription control G:C➔A:T at CpG sites cell cycle DNA repair apoptosis p53 prevents transcription of: p14 ARF Bcl-2 cell cycle apoptosis p53 protein is induced in response to DNA damage functional p53 DNA damage accumulation of p53 in nucleus G1 arrest DNA repair apoptosis inactivated p53 DNA damage no G1 arrest replication of damaged DNA genomic instability p14ARF acts as a growth suppressor -TSG • p14ARF protein encoded by CDKN2A locus on 9p (alternative reading frame to p16INK4a) • hypermethylated or homozygously deleted in cancers • p14ARF binds to MDM2 and inhibits MDM2 degradation of p53 • p14ARF expression negatively regulated by p53 p53 p14ARF mdm2 Loss of normal TP53 function from alterations in either TP53, MDM2 or p14ARF The p16INK4a/CDK4/RB1 pathway CDK4 and CDK6 genes: act as oncogenes and promote cell division; both encode cyclin dependant kinases which form complexes with cyclin D to phosphorylate the RB protein 28 Carcinogenesis The main stages of carcinogenesis The complete process of transformation, invasion and metastasis. carcinogenesis: malignant of neighbouring tissues Each stage comprises a series of genetic alteration of cells affecting specific genes that are involved in regulating cell properties relevant for the individual stage The Hallmarks of Cancer • Normal cells can only ° The Hallmarks of Cancer: Douglas Hanahan and Robert A. Weinberg (2000) Cell 100, 57–70 divide for a predetermined number of times before reaching senescence (irreversible inhibition of replication). Replicating Senescent Cell immortalisation and tumourigenesis Immortalisation is the acquisition of an infinite lifespan. Normal mammalian somatic cells proliferate a limited number of times before undergoing senescence. Senescent cells may remain metabolically active even though they have permanently ceased proliferation. Immortalisation: essential step for the malignant transformation of normal cells. Telomerase, the enzyme responsible for maintaining telomeres at the ends of chromosomes. Unlike normal cells that lack detectable levels of telomerase activity, approximately 90% of human tumours consist of cells that contain an active telomerase enzyme. Cellular principles of invasion and metastasis • Cancer cells spread to distant sites in the body via the blood stream/lymphatics. • Metastatic cells: less adhesive than normal cells and are able to degrade and penetrate tissue barriers such as the extracellular matrix (ECM) of surrounding connective tissue and the basement membrane of blood vessels. • After gaining access to the systemic circulation they can invade normal tissue at various sites in the body forming secondary colonies. Valastyan, S. and Weinberg, RA. Cell 2011 The invasion - metastasis cascade involves: ▪ Acquisition of local invasiveness ▪ Invasion of the cell into blood/ lymph vessels (intravasation) ▪ Transport through the blood/lymph vessels to distant tissue sites ▪ Escape of the cancer cells from circulation (extravasation) ▪ Ability to adapt to the local tissue environment and to proliferate Signs for Genomic Changes in Cancer • Changes in chromosome numbers - Aneuploidy (the presence of an abnormal number of chromosomes in a cell i.e. 45 or 47 instead of 46), various cancers i.e. prostate, neuroblastoma, brain, colon, etc. • Chromosomal changes - Increase in DNA copy number - Loss in chromosomal (bladder cancer, endometrial cancer, brain tumours, cervical cancer) Genetic instability of tumour cells Genetic analysis of solid tumours revealed the presence of a high degree of genetic abnormalities, such as aneuploidy, chromosome translocations etc. This is likely due to the lack of active p53 protein, and the ability of cancer cells to avoid cell death through apoptosis. Other mechanisms may also play a part here, e.g. mitotic defects that result in chromosome miss-segregation chromosomal instability (CIN) is widespread in cancer cells from epithelial origin. The best example is translocation between chromosomes 9 and 22 [t( 9;22)], i.e. Philadelphia (Ph) chromosome, in chronic myeloid leukemia (CML), resulting in the translocation of proto-oncogene ABL at 9q34 to BCR on chromosome 22 Alteration of genetic mechanisms in cancer Human karyotype is diploid hence 1 allele mutation in a TSG is not enough to cause cancer. Loss of heterozygosity (LOH): cross chromosomal event that results in loss of the entire gene and the surrounding chromosomal region -TSGs. Microsatellite instability (MIN): This is a phenomenon often seen in colorectal cancer cells with defective DNA mismatch repair system, e.g. in hereditary nonpolyposis colorectal cancer (HNPCC). Microsatellites are regions of repetitive DNA sequences in the genome that are prone to shortening or extension if the mismatch repair enzymes are defective. Chromosomal anomalies cause various disorders Anomalies that affect autosomes (the 22 paired chromosomes that are alike in males and females) and the 2 sex chromosomes (X and Y). 2 major categories: Numerical abnormalities Trisomy (an extra chromosome) Monosomy (a missing chromosome) Structural abnormalities Translocations (a whole chromosome or segments chromosomes inappropriately join with other chromosomes) Deletions and duplications of various parts of chromosomes of If these alterations activate or inactivate genes that result in a proliferative advantage over normal cells, then a cancer may develop. Replication of damaged DNA can lead to chromosome abnormalities The process begins with accidental DNA damage in a cell that lacks functional p53 protein. Adapted from the Molecular Biology of the Cell. 4th edition., Alberts B, Johnson A, Lewis J, et al., New York: Garland Science; 2002. Epigenetic events Epigenetics is the study of stable changes in cell function that do not involve alterations in the DNA sequence. Hypermethylation - transcriptional silencing that can be inherited by daughter cells following cell division – TSGs activation. Causes of cancer Mutations and cancer Cancer development is based on the accumulation of somatic mutations over lifetime. Germ line mutations are typically not involved, but in very rare cases of inherited cancer predisposition, they are contributing to disease progression. The basal mutation rate is low in humans but it may be enhanced through exposure to chemical mutagens radiation tumour viruses. Chemical mutagens : can modify DNA through alkylation or deamination of DNA bases, or through intercalation between base pairs and formation of DNA adducts (e.g. aromatic hydrocarbons). Examples of alkylating agents: Ethyl methyl sulfate, nitrogen mustards, mitomycin, methyl methane sulfonate (MMS), diethyl sulfate, and nitrosoguanidine DNA-intercalating agents: acridine orange, ethidium bromide (EtBr), proflavin, daunorubicin etc. Oxidative damage may also affect DNA integrity. Radiation Ultraviolet radiation may induce skin cancer (eg, basal and squamous cell carcinoma, melanoma) by damaging DNA - formation of thymidine dimers, which may escape excision and re-synthesis of a normal DNA strand. Patients with inherent defects in DNA repair (eg, xeroderma pigmentosum) or immune suppression by drugs or an underlying disease are particularly prone to skin cancers due to ultraviolet exposure. Accidental/occupational ionizing radiation & exposure to therapeutic irradiation may lead to leukaemia, breast cancer, sarcomas and other solid cancers years after exposure. Drugs ▪ Estrogen, oral contraceptive: may slightly increase the risk of breast cancer, but this risk decreases over time. ▪ Estrogen alone used for hormone replacement therapy: increases the risk of uterine cancer. ▪ Combined estrogen and progesterone: increase risks of breast and ovarian cancer similar to estrogen alone. ▪ Long-term use of anabolic steroids: increase the risk of liver cancer and accelerate prostate cancer. ▪ Treatment of cancer with chemotherapy drugs alone or with radiation therapy increases the risk of developing a second cancer, as do immunosuppressive drugs given for organ transplantation. The cancers most commonly increased due to immunosuppressive drugs are myelodysplastic syndrome and acute myeloid leukemia. Dietary substances ➢ Certain substances consumed in the diet can increase the risk of cancer. ➢ A diet high in fat: increased risk of colon, breast, and possibly prostate cancer. ➢ Large alcohol consumption: hgher risk of developing head and neck and esophageal cancer. ➢ A diet high in smoked and pickled foods or in meats cooked at a high temperature increases the risk of developing stomach cancer. ➢ People who are overweight or obese have a higher risk of cancer of many cancer types, particularly cancers of the breast, endometrium, colon, kidney, and esophagus. Physical factors: Chronic skin, lung, gastrointestinal, or thyroid inflammation may predispose to development of cancer. For example, patients with long-standing inflammatory bowel disease (ulcerative colitis) have an increased risk of colorectal carcinoma. Sunlight and tanning light exposure increases the risk of skin cancers and melanoma. Immune Disorders Immune system dysfunction as a result of inherited genetic mutation, acquired disorders, aging, or immune-suppressing drugs interferes with normal immune surveillance of early tumours and results in higher rates of cancer. Known cancer-associated immune disorders include ▪ Ataxia-telangiectasia (acute lymphocytic leukemia, brain tumours, gastric cancer) ▪ Wiscott-Aldrich syndrome (lymphoma) ▪ X-linked agammaglobulinemia (lymphoma) Immune deficiency resulting from immune-suppressing drugs or HIV infection (lymphoma, kidney cancer, melanoma, cervical cancer, head and neck cancer, Kaposi sarcoma) Autoimmune disorders such as systemic lupus erythematosus, rheumatoid arthritis, and B-cell lymphoma Viral causes of cancer Oncogenic viruses, which contain viral oncogenes in their genomes that are in most cases derived from cellular proto-oncogenes. They tend to use one of the following mechanisms to stimulate proliferation of their host cells: ▪ Insertion of a strong promoter in the vicinity of a host cell proto-oncogene ▪ Expression of proteins that neutralise host cell tumour suppressor proteins ▪ Expression of proteins that prevent or delay apoptosis Characteristics of viral carcinogenesis include: ▪ Tumour viruses often establish persistent infections in the human host ▪ Host factors are important determinants of virus-induced carcinogenesis Viruses are rarely complete carcinogens; they require additional factors to fully activate carcinogenesis. Human tumour viruses Virus (Group) Associated Human Cancer DNA VIRUSES Papilloma virus family Human papilloma virus (HPV) (various subtypes) Genital tumours, squamous cell carcinoma Herpes virus family Human herpes virus 8 (HHV8) Epstein-Barr virus (EBV) Kaposi sarcoma Burkitt's lymphoma, Hodgkin's disease, Nasopharyngeal carcinoma Hepadenavirus family Hepatitis B virus Hepatocellular carcinoma RNA VIRUSES Retrovirus family Human T-cell leukaemia virus Human immunodeficiency virus Flavivirus family Hepatitis C virus Adult T-cell leukaemia AIDS-related malignancies Hepatocellular carcinoma Adapted from Table 43-1, Brooks GF, Carroll KC, Butel JS, Morse SA. Jawetz, Melnick & Adelberg’s medical microbiology, 24th ed. McGraw-Hill, 2007) Bacteria may also cause cancer. Helicobacter pylori infection increases the risk of several kinds of cancer (gastric adenocarcinoma, gastric lymphoma, mucosa-associated lymphoid tissue [MALT] lymphoma). Parasites of some types can lead to cancer. Schistosoma haematobium causes chronic inflammation and fibrosis of the bladder, which may lead to cancer. Opisthorchis sinensis has been linked to carcinoma of the pancreas and bile ducts. The importance of DNA repair systems DNA repair systems: fix damage that may have occurred to the DNA. Principal DNA repair mechanisms include: mismatch repair, base and nucleotide excision repair, repair of depurinated sites and repair of double-strand breaks. In hereditary cancers, characterized either by microsatellite instability or chromosomal instability, the underlying basis for the genomic instability is mutations in DNA repair genes compared to sporadic (non-hereditary) cancers where genomic instability, at least at the early stages of cancer development, is not due to mutations in DNA repair genes or mitotic checkpoint genes. Inherited diseases caused by DNA repair defects Disease Protein Affected Affected Function Manifestation Bloom syndrome topoisomerase III alpha, RMI1 and RMI2 genes Recombination repair? Immunodeficiency, cancer susceptibility, chromosome breaks Breast cancer susceptibility BRCA1, BRCA2; proteins of DNA repair complexes Homology-directed DNA repair Breast and ovarian cancer Cockayne syndrome Nucleotide excision repair protein CSA Transcription-coupled nucleotide excision repair Poor growth, early senility, neurological degeneration Fanconi anemia ANCA, FANCC or FANCG Repair of DNA crosslinks? Anaemia, leukaemia, chromosome breakage Hereditary nonpolyposis colon cancer (HNPCC) Proteins of mismatch repair - MLH1, MSH2, MSH6, and PMS2 Post-replication mismatch repair Cancer susceptibility Nijmegen breakage syndrome Activator of nuclear protein kinases, NBS1 Signalling for DNA double-strand break repair Growth retardation, immunodeficiency, cancers Werner syndrome DNA helicase and exonuclease Unknown Premature aging, short telomeres Xeroderma pigmentosum Nucleotide excision repair proteins XPD and XPB Genome-wide nucleotide excision repair Cutaneous photosensitivity Adapted from Table 9.1, Meisenberg G, Simmons WH. Principles of medical biochemistry, 3rd ed. Mosby/Elsevier, 2012) References and further reading Chapter 20. Cancer. In: Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P. Molecular biology of the cell, 5th edition. New York: Garland Science; 2007 , Available from: http://www.garlandscience.com/product/isbn/9780815341055. Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation Cell 2011 Mar 4;144(5):646-74 [Abstract available at http://www.ncbi.nlm.nih.gov/pubmed/21376230]. Karp G. Chapter 16. Cancer. In: Karp G. Cell and molecular biology: concepts and experiments, 6th edition. Australia: Wiley; 2010, Available from: http://au.wiley.com/WileyCDA/WileyTitle/productCd-EHEP000329.html. Stricker TP, Kumar V. Chapter 7. Neoplasia. In: Kumar V, Abbas AK, Fausto N, Aster JC, Perkins JA. Robbins and Cotran pathologic basis of disease, 8th ed. USA: Saunders; 2010, Available from: http://www.us.elsevierhealth.com/pathology/robbins-cotran-pathologic-basis-of-diseasehardcover/9781416031215/. Weinberg RA. The biology of cancer, 2nd ed. USA: Garland Science; 2013, Available from: http://www.garlandscience.com/product/isbn/9780815342205. Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P. Molecular biology of the cell, 4th ed. New York: Garland Science/Taylor & Francis LLC; 2002, Hanahan D, Weinberg RA. Hallmarks of cancer: 74, http://www.ncbi.nlm.nih.gov/pubmed/21376230. the next generation. Cell 2011;144(5):646-