2What Is Cancer?• Cancer is a group of diseases caused by the uncontrolledmultiplication of abnormal cells in the body, a process calledneoplasia.• Abnormal new tissues called neoplasms are formed.• Neoplasms usually form masses called tumors that may bebenign (non cancerous) or malignant (cancerous).• Malignant or cancerous tumors grow rapidly,are invasive(to surrounding tissue) and metastatic (traveling viablood/lymph to invade distant tissues).• Cancers destroy healthy tissues causing loss of function anddeath.• Cancer is the 2nd major killer in populations of developedcountries & the leading cause of death in children 3-15 (US).• Cancers are genetic disorders caused by accumulation ofsomatic mutations (gene & chromosome) in a person’s cells.• Inherited mutations give a predisposition for certain cancers.
3Characteristics of Cancer Cells • Cancer cells are genetically altered via gene orchromosome mutations so:- lack normal controls over cell division or apoptosis.- may express inappropriate genes (e.g. for telomerase,enzyme that maintains length of DNA for continued division)- are genetically unstable due to loss of DNA repairmechanisms (so are more susceptible to radiation damagethan normal cells).• Divide excessively (proliferate) & indefinitely producingneoplasms.• Live indefinitely (do not show apoptosis).• Lose the normal attachment to other cells so becomemetastatic (travelling via blood/lymph to invade distantsites).• Secrete signals for angiogenesis (growth of blood vesselsinto tumor).
4Cancer Cells are Undifferentiated & Malignant undifferentiated tovarying degrees(even anaplastic, likestem cells) so divide& do not perform thenormal function ofmature cells.• The lessdifferentiated thecancer cell the moremalignant thecancer (the morerapidly growing is thetumor).
5What Causes Cancer?Inherited mutations in genes that affect cell cycle,DNA repair, or apoptosis: these mutations give agenetic predisposition for cancer.Somatic mutations to these same genes caused by:• Exposure to risk factors- environmental mutagens (carcinogenic chemicals,radiation)- hormones- weakening of immune system (as in AIDS).• Oncogenic (tumor) Virus infections- Epstein Barr virus (causes Burkitt lymphoma)- Human Papilloma Virus (causes cervical cancer).Tumor viruses transform human cells into cancer cells by:•Introducing viral cancer - causing oncogenes into host cell DNA•Causing Translocation and overexpression of host protooncogenes.
6Normal cell cycle is controlled by signal transduction: Growth factors bind to surface receptors on the cell; transmembrane proteins relay signals into the cell.Two types of growth factors:Growth factors stimulate cell division.Growth-inhibiting factors inhibit cell division.Healthy cells divide only when growth factor and growth-inhibiting factor balance favors cell division.Cancer cells divide without constraint(e.g., mutations in growth and growth-inhibiting factor genes).
7Regulation of cell division by signal transduction.
8The Cell Cycle Oncogenes G2 (cell growth) M (mitosis) S (synthesis) DNA repairgenesREPAIRS AHEADG1G0 (resting)Tumor suppressor genes
9Cell cycleCDKp53 is known as the ‘ guardian of the genome ‘
10CONTROL of CELL CYCLE Differentiated The Cell has 3 major Cells enter G0Cells enter“checkpoints” that arenon dividingG1sensitive to signals, thecheckpointG0 phase-G1, the G2 and the Mcheckpoints.• If a cell does notG1pass G1 checkpointControlit enters a non-systemSdividing GO Phase.• Most somatic cellsG2are in GO.M• Some cells in GO(e.g. liver cells) canreenter cell cycle ifM checkpointneeded.G2 checkpointIn cancer cells genes that control cell cycle have mutatedso cells divide excessively producing neoplasms.
11Cancer and genes:Three classes of genes are frequently mutated in cancer:Proto-oncogenes ( oncogenes)Tumor suppressor genesMutator genes
12Proto-oncogenes oncogenes: Proto-oncgenes are genes that possess normal gene products and stimulate normal cell development.OncogenesOncogenes arise from mutant proto-oncogenes.Oncogenes are more active than normal or active at inappropriate times and stimulate unregulated cell proliferation.Some tumor viruses that infect cells possess oncogenes:RNA tumor viruses = possess viral oncogenes (derived form cellular proto-oncogenes) capable of transforming cells to a cancerous state.DNA tumor viruses = another class of tumor viruses; do not carry oncogenes, but induce cancer by activity of viral gene products on the cell (no transformation per se).
13Types & effects of different types of mutations: Point mutations: occur in protein coding or controlling sequences.Deletion: frameshifts may lead to defective proteins.Gene amplification: random over-replication of small segments of DNA results in extra copies (up-regulates cell growth).Mutator genes:Mutator gene increases spontaneous mutation rate of other genes.Mutator gene products are involved in DNA replication and repair; mutations make the cell error prone.
14Proteins Control Cell Division The cell cycle is controlled by proteinsfrom inside & outside the cell.• Intracellular Cyclins and Cyclin Dependent Kinases(CDKs) control the checkpoints.• Hormones or extracellular proteins from other cells(called Growth Factors) signal target cell to divide.- Hormones (e.g. Growth Hormone) or Growth Factorsbind to receptor proteins of target cell membrane.- This triggers a molecular signaling pathway.- A series of linked proteins activate Cyclin- CDKs whichAllows Cells to Pass Cell Cycle Checkpoints & divide.
15How Growth Factors Trigger Cell Division or HormonePlasmamembraneRelay•G1 checkpoint preventsProteins opendamaged DNA from replicatingcheckpoints•Checkpoint controlled byReceptorCyclin - CDKproteinSignalTransductionControlPathwayG1SsystemMG2
16Tumor Suppressor Proteins Inhibit Cell Division & Prevent Cancer Tumor suppressor proteins are proteins thatbind to checkpoint proteins to stop the cellcycle & prevent cell division if DNA is damaged.• Tumor suppressor proteins stop division ofmutated cells until mistakes in DNA arerepaired by enzymes.• TS proteins keep most mutations from beingpassed on to daughter cells & developing intocancer.• If the genes for TS proteins mutate the brakeon cell division is removed cancers may result.• Two important TS proteins are the p53 protein& the RB protein.
17The p53 Tumor Suppressor Protein The p53 tumor suppressor protein is activatedwhen DNA is damaged. The p53 gene is called the“guardian angel of the genome”P53 proteinactivatesInternalgenes forsignallingpathwayproteins that•Prevent cellentering Sphase•Repair DNA•CauseDNA repairapoptosis (ifApoptosisDNA isCell cannot enter S phaseirreparable)
18Gene Mutations That Cause Cancer Mutations in 4 types of genes cause Cancer• Proto - oncogenes: genes that code for normal proteinsused in cell division–Growth factors–Membrane Receptors for Growth Factors–Signaling Proteins (e.g. ras proto- oncogenemutates in 30% of cancers).• Tumor Suppressor genes: gene that code for proteins thathelp prevent uncontrolled cell division by blocking key steps(e.g. DNA replication).- Retinoblastoma susceptibilty (RB) gene- p53 gene mutates in >50% of cancers.• DNA Repair genes• Genes for Apoptosis
19How Carcinogens Cause Cancer Inactivation of DNA Repair Genes Oncogenesof Genes forTumorApoptosisSuppressorGenes
20Oncogenes Are Mutated Proto-oncogenes A cell can acquire a cancer - causing oncogenefrom•A virus•A mutation in a proto-oncogene.Oncogenes still code for the proteins needed forcell division but they cause cancer by producing– Too much of the protein– An abnormally active protein, e.g. protein thatactivates division by itself– Protein that is made when it is not needed– Protein that should be made by a different (i.e.dividing) cell.
21Cancer causing Mutations •Proto-oncogenes form active oncogenes by- being misplaced (e.g. by translocation) to a site wherethe gene is continually expressed resulting inoverproduction of a protein that stimulates cell division(e.g. in Chronic Myeloid Leukemia)- By mutating to a form that is over expressed.•Cancer causing Mutations in Tumor Suppressor genesinactivate the genes so normal protein product is not formed.MutatedoncogeneTumorSuppressorgeneneoplasm
22Over Stimulation Hyperactive of Cell relay protein Division by GrowthStimulationMembranefactorof CellReceptorHyperactiveDivision byrelay proteinOncogeneNormal product(product ofof ras generas oncogene)Relayissues signalsproteinson its own.Transcription factor(activated)DNATranscriptionProtein thatstimulatesTranslationcell division
23Cancer from Mutation of Tumor Supressor Gene Tumor-suppressor gene Mutated tumor-suppressor geneCancerNormalfromProteinMutationDefective,preventsof TumornonfunctioningcellSupressorproteindivisionif DNA isGeneProteindamagedabsent(cell divisionCell divisionnot blocked)allowed if DNArepairedMutations accumulatein cancer cells
24Multiple Genetic Changes Cause Cancer Cancers result from a series of genetic changes in a cell lineage• Inherited (germline) cancers begin with an inherited cancersusceptibility mutation in every cell that is passed on tooffspring.• Inherited cancers may follow a dominant pattern, e.g. InheritedRetinoblastoma caused by a mutation in the Rb tumorsupressor gene increases cancer risk 10,000 x.• However, Inherited cancers need at least one more somaticmutation for cancer to develop (“2 hit hypothesis for cancercausation”) .• Sporadic cancers are caused solely by somatic mutationsoccurring in certain body cells so are not passed on tooffspring.• Accumulation of somatic mutations in a cell over timeeventually leads to uncontrolled cell division and cancer.• Therefore sporadic cancers tend to appear much later in life thaninherited cancers.
25Accumulation of Mutations Cause Cancer • inheritance of a germ cell mutation acts as a riskfactor for cancers by reducing the number ofsomatic mutations required to cause cancer.• Early mutations show up in all subsequent stagesof a cancer.Normal1423Chromosomes mutationmutationsmutationsmutationsNormal cellMalignant cell
26Oncogenes Normal genes (regulate cell growth) 1st mutation (leads to accelerated cell division)1 mutation sufficient for role in cancer development
27Tumor Suppressor Genes Normal genes (prevent cancer)1st mutation(susceptible carrier)2nd mutation or loss (leads to cancer)
28The Two-Hit Hypothesis First hitFirst hit in germline of childSecond hit(tumor)
29Pathogenesis of Colon Cancer Colon Cancer is usuallySporadic & develops in a seriesof steps caused by a series ofsomatic cell mutationsLoss ofRemoval of polypstumor-prevents cancersuppressorColon wall gene (e.g. APC)Small benignNormal colongrowth (polyp)epithelial cells= adenomaAPC: adenomatous polypodsis coli
30Pathogenesis of Colon Cancer -2 Activation ofLoss ofCampbell, & Reeceras oncogenetumor-Biology figsuppressorgene p53Loss ofAdditionaltumor-mutationssuppressorMalignant tumorLarger benignSmall benign gene(adenocarcinoma)growth (adenoma)growth (polyp)Environmental Risk factors for Colon Cancer• Low fiber diet• Smoked meats (contain heterocyclic aromatic amines that areconverted to mutagens in the liver)• Low intake of fruits & vegetables (antioxidants)• Low intake of cruciferous vegetables (a chemical in brocolli,brussels sprouts, cabbage activates enzymes that blockformation of mutagens)
31Genetic Abnormalities Associated With Hematologic Malignancies A- Point MutaionMutaions within the RAS oncogenes or P53 tumor- suppressor gene are common in many haempoietic malignancies. The point mutation may involve several base pairs. In 35% of cases of AML the nucleophosmin gene shows an insertion of 4 base pairs.B- TranslocationIncludes two main mechanisms:1- Fusion of parts of two genes to generate a chimeric fusion gene that codes a novel fusion protein. Ex: BCR- ABL in t(9; 22) in chronic myeloid leukaemia.2- Overexpression of a normal cellular gene. Ex: overexpression of BCL-2 in the t(14; 18) translocation of follicular lymphoma or MYC gene in Burkitt,s lymphoma.
32D- Duplication or amplification E- Epigenetic alterations C- DeletionsMay involve a small part of a chromosome, the short or long arm or the entire chromosome. Losses most commonly affect chromosomes 5, 6, 7, 11, 20 and Y. The critical event is probably loss of a tumor suppressor gene.D- Duplication or amplificationGains are common in chromosomes 8, 12, 19, 21 and Y. It is not a common feature in haematologic malignancy but has been described involving the MLL gene.E- Epigenetic alterationsMeans alterations in the mechanism by which genes are transcribed and are stably inherited with each cell division so they are passed on as the malignant cell divides. The most important mechanisms are methylation of cytosine residues in DNA and enzymatic alterations such as acetylation or methylation of the histone protein that package DNA.
33Prostate Cancer• Prostate Cancer is the most common cancer among men (esp.>65 yrs, African - Americans) & 2nd in cancer deaths in men.• Risk factors include increasing age, race, family history, fat diet,male hormones over many years.• Adenocarcinoma occurs in periphery of prostate gland.• Metastasises to lungs, bones (bone pain often first symptom asearly stage of primary tumor may be asymptomatic).• Tumors are graded from 1 (well differentiated cells) to 5 (leastdifferentiated cells, high malignancy).• Manifestations (some similar to Benign Prostatic Hyperplasia):- changes in voiding pattern, dysuria, hematuria,– from metastasis low back pain from bone, wt loss, anemia, &shortness of breath.• Screening (important for early asymptomatic cancers) includes:- digital rectal exam (palpation of prostate by DRE detectsnodular lump)- transrectal ultrasonography (measures prostate vol: moresensitive than DRE)- PSA blood test
34PSA Test for Prostate Cancer • Tumor cells express abnormal genes so form abnormalproteins (antigens) so can act as tumor markers.• Prostate Specific Antigen (PSA) a glycoproteinreleased by prostate gland into the blood identified as amarker of prostate cancer in• PSA is highly specific to prostate gland but not specific toprostate cancer. Elevated blood PSA can also occur withnon cancerous conditions (i.e. false positives occur withprostatitis or with benign prostatic hyperplasia BPH)• The need to treat stage 1 cancers detected by PSA test iscontroversial: stage 1 tumors are asymptomatic, not detectedby digital exam & present in 80% of men over 80 yrs.• Treatment of Prostate Cancer includes surgery,radiation & hormonal manipulation (e.g. androgen inhibitors) ifexpected survival >10yrs, otherwise “watchful waiting” ispreferred in elderly patients.• PSA test is used to assess treatment (correlates withprostate size & cancer stage).
35Etiology of Breast Cancer Breast cancer is most common cancer in women & 2nd mostcommon in cancer deaths in women (after lung cancer).Risk Factors for Breast Cancer• Prolonged exposures to estrogens (early menarche & latemenopause). Breast cancers that are estrogen receptor +ve aretreated with drugs (e.g. tamoxifen) that bind to these receptors.• Late Childbearing (having first child after age 30)• Breasts with a high proportion of lobular (milk producing)and ductal tissue density.• Not breast feeding babies increases post menopausal BC.• Exposure to radiation.• High alcohol consumption.• Family History of BC & Genetic Predisposition in 5-20% ofcases (inheriting mutated breast cancer susceptibilitygenes, BRCA-1 or BRCA-2).
36Genetics of Breast Cancer • % of breast cancers are Familial .• Most involve mutations in 2 Tumor Suppressor genesinvolved in DNA repair so are used as genetic markers.• Both genes also increase the risk of ovarian cancer.- Breast Cancer Susceptibility Gene 1 (BRCA1) onchromosome 17- Breast Cancer Susceptibility Gene 2 (BRCA2) onchromosome 13.• Mutated HER-2/neu Gene (Human Epidermal GrowthFactor Receptor 2 gene is an Oncogene for a protein thatstimulates cell division & occurs in % of BreastCancers.• Her-2/neu breast cancers strike early in adulthood & spreadquickly.• Herceptin is a monoclonal antibody based drug that bindsto Her-2/neu receptors & blocks cell division in tumors.
37Pathogenesis of Breast Cancer Tumor (usuallyadenocarcinomain milk ducts)Campbell, & Reece,Biology, figGlandulartissueNote: lumpectomy a possible treatment in early stagesA tumor grows from aCancer cells invadesingle cancer cell.neighboring tissue.
38Pathogenesis of Breast Cancer -2 LymphvesselBloodvesselCancer cellMetastatic tumor• Cancer cells spread viaSmall percentagelymph & blood to otherof metastisised cancerparts of the body.cells may survive and• “Sentinel lymph node ”biopsy determines ifestablish a new tumor inacancer has spread & ifanother part of the body.further lymph noderemoval required.
39Diagnostic methods used to study malignant cells 1- Karyotype analysis: It is a direct morphological analysis of chromosomes from tumor cells under the microscope.2- Fluorescent in situ hybridization analysisFISH analysis involves the use of fluorescent- labelled genetic probes which hybridize to specific parts of the genome. This can detect extra copies of genetic material or reveal chromosomal translocation.3- Southern blot analysis: It involves extraction of DNA from leukaemic cells followed by restriction enzyme digestion, gel electrophoresis and transfer by blotting to a suitable membrane. The DNA is then hybridized to a probe complementary to the gene of interest.4- Polymerase chain reaction: Can be performed on blood or bone marrow for a number of specific translocations such as t(9; 22) and t(15; 17). It is very sensitive and can detect one abnormal cell in one million normal cells. It is of great value to diagnose minimal residual disease.
405- DNA microarray:Allows rapid and comprehensive analysis of cellular transcription by hybridizing labelled cellular mRNA to DNA probes which are immobilized on a solid support. This approach can rapidly determine mRNA expression from a large number of genes and may be used to determine the mRNA expression pattern of different leukaemia or lymphoma subtypes.6- Flow cytometry: Normal cells each have a characteristic profile but malignant cells often express an aberrant phenotype that can be useful in allowing their detection.7- Immunohistochemistry: Antibodies can be used to stain tissue sections with fluorescent markers.Value of using these methods:a- Initial diagnosis.b- Establishing treatment protocol.c- Monitoring response to therapy.
41Cytogenetics • Cytogenetics is the original cancer genetic test used to identify abnormal mutatedchromosomes by karyotype analysis.• Cytogenetics identified the Philadelphiachromosome resulting from a translocationerror in chromosome forming an oncogenefor chronic myelogenous leukemia (CML) in1960.• Karyotype Analysis is done by culturing tumorcells, arresting them in metaphase & spreadingchromosomes via use of hypotonic solutions.• Chromosomes are stained and interpreted by acytogeneticist.• Process may take weeks (as many tumors do notgrow in vitro).