Introduction to the Molecular Biology of Cancer - XY3121 Molecular Medicine - PDF

XY3121: Molecular Medicine Introduction to the Molecular Biology of Cancer Dr Peter Oatley [email protected]  Cell Cycle Checkpoints Learning Outcomes Part 1: Cell Cycle Checkpoints...

XY3121: Molecular Medicine Introduction to the Molecular Biology of Cancer Dr Peter Oatley [email protected]  Cell Cycle Checkpoints Learning Outcomes Part 1: Cell Cycle Checkpoints Recall the cell cycle phases Understand the function of Cdk and Cyclins in the cell cycle Know the function of the three main checkpoints in the cell cycle and their regulation  Cell Cycle Checkpoints and Cancer Lecture Structure Part 1: Cell Cycle Checkpoints Organ/body size regulation The Cell Cycle - Recap - Checkpoints - Cyclins & Cyclin Regulation - G0 Checkpoint 1-3 Part 2: Cancer  Cell Cycle Checkpoints The Cell Cycle - Organ/body size regulation Organ/body size is determined by balance of: 1) Cell growth 2) Cell division 3) Cell Survival - All tightly regulated by cell signalling External Factors affecting cell signalling 1) Mitogens - Stimulate cell division - Trigger a G1/S-Cdk activity wave that overcomes cell cycle blocks 2) Growth factors - Stimulate cell growth Note: Some molecules - Promoting synthesis of proteins and other macromolecules fit in more than one 3) Survival factors - Promote cell survival category. - Suppressing programmed cell death (apoptosis) factors  Cell Cycle Checkpoints The Cell Cycle - Recap Mitosis - The process of cell division - Accurate chromosomal DNA segregation → Produce two daughter cells - Regulated by the cell-cycle control system Phases: G1 - Cell growth S-Phase - DNA Synthesis Interphase G2 - Cell growth M-Phase - Mitosis and Cytokinesis  Cell Cycle Checkpoints The Cell Cycle - Recap Mitosis - The process of cell division M-Phase: Mitosis - Prophase, Prometaphase, Metaphase, Anaphase, Telophase Cytokinesis - Two daughter cells created (Includes Telophase) Molecular Biology of the Cell Cell Cycle Checkpoints DNA cell  Cell Cycle Checkpoints The Cell Cycle - Checkpoints 1) End of G1 (G1/S Transition) - Before committing to cell-cycle entry & DNA replication 2) G2/M transition - Early mitotic events that lead to chromosome alignment on the mitotic spindle 3) Metaphase-to-anaphase transition - Sister-chromatid separation  Cell Cycle Checkpoints The Cell Cycle - Checkpoints Checkpoints regulated by Cyclin-Cdk complex activity  Cell Cycle Checkpoints The Cell Cycle - Checkpoints Checkpoints regulated by Cyclin-Cdk complex activity  Cell Cycle Checkpoints The Cell Cycle - Cyclins Cyclin induced Cdk autophosphorylation and Cdk activation Active Cdk-Cyclin complex phosphorylates substrates Pollard & Thomas: Cell Biology  Regulation of Cyclin-dependent kinases Cell Cycle Checkpoints Kinase Activation Inactivation The Cell Cycle - Cyclin Regulation Bind inhibitors Phosphorylation Ubiquitination Pollard & Thomas: Cell Biology  Cell Cycle Checkpoints The Cell Cycle - G0 Most Cells in the body are in G0 - Nondividing state - Disassembled cell-cycle control system - Expression of Cdks genes is permanently turned off in some cells (E.g. Neurones) - Others can be reactivated to G1 by Mitogens Checkpoint 1 G1→ S  Cell Cycle Checkpoints Checkpoint 1 - G1→ S Mitogens stimulate cell cycle entry - RTK/Ras/MAPK Signalling leads to G1-Cdk Activation - G1-Cdk inactivates Retinoblastoma (Rb) protein → Active E2F protein released (and bind DNA) → Active G1/S-cyclin → Active S-cyclin → DNA Synthesis (S-Phase)  Cell Cycle Checkpoints Checkpoint 1 - G1→ S Regulated Proteolysis Decreases G1/S-cyclins SCF complex (Skp, Cullin, F-box) - Ubiquitin ligase family member - Activity changes depending on the F-Box component - Regulated by substrate phosphorylation SCF catalyses the ubiquitylation of: 1) CKI proteins - in late G1 → Release inhibition of S-Cdk and DNA replication 2) G1/S-cyclins - destruction in early S phase  Cell Cycle Checkpoints Checkpoint 1 - G1→ S DNA Damage Blocks Cell Division - Process requires p53 - p53 normally ubiquitinated by Mdm2 - DNA damage is detected (See XY1110 Genetics) → Activates kinase cascade → p53 phosphorylated (no longer Ubiquitinated) → Inactivation/reduction of Mdm2 (not shown) - Stable p53 binds DNA (Tetramer) → CDKN1A gene expression → p21 protein → + Many more - p21 protein inhibits G1/S-cyclin and S-cyclin → Cell Cycle arrest or apoptosis Note: Short telomers can also activate p53 → Limit replication of cells Checkpoint 2 G2/M  Cell Cycle Checkpoints Checkpoint 2 - G2/M Active M-Cdk Promotes…. Requires active M-Cdk - Assembly of the mitotic spindle M-Cdk regulated by phosphorylation balanced: - Chromosome condensation - CAK - Cdk activating kinase - Sister chromatid attachment - Wee1 - Cdk inhibitory kinase - Nuclear envelope breakdown - Cdc25 - Phosphatase - Actin cytoskeleton and Golgi rearrangement M-Cdk induces positive feedback to produce more active M-Cdk  Checkpoint 3 Metaphase-to-anaphase S-cyclin  Cell Cycle Checkpoints Checkpoint 3 - Metaphase-to-anaphase Regulated Proteolysis Triggers Metaphase-to-Anaphase Transition Anaphase promoting complex or cyclosome (APC/C) - Ubiquitin ligase family member - Activity changes depending on associations: - Cdc20 in mid-mitosis - Cdh1 from late mitosis through early G1 S-cyclin  Cell Cycle Checkpoints Checkpoint 3 - Metaphase-to-anaphase Regulated Proteolysis Triggers Metaphase-to-Anaphase Transition Anaphase promoting complex or cyclosome (APC/C) - Ubiquitin ligase family member APC/C catalyses the ubiquitylation of: A) Securin → Separase released to separate sister-chromatids B) S-cyclin and M-cyclin - Inactivates Cdks → Cdk targets dephosphorylated → Mitosis Completion APC/C remains active in G1 → period of Cdk inactivity APC/C is turned off for the next cell cycle  Cell Cycle Checkpoints and Cancer Summary Cell Cycle Checkpoints The cell cycle: G1 - Cell growth S-Phase - DNA Synthesis G2 - Interphase M-Phase - Mitosis and Cytokinesis Checkpoint 1) End of G1 (G1/S Transition) - Signalling → Retinoblastoma (Rb) releases E2F protein → Active G1/S-cyclin and S-cyclin→ DNA Synthesis (S-Phase) - DNA Damage Blocks Cell Division via p53 - SCF complex (Skp, Cullin, F-box) Checkpoint 2) G2/M transition - Active M-Cdk Checkpoint 3) Metaphase-to-anaphase transition Anaphase promoting complex or cyclosome (APC/C)  Cell Cycle Checkpoints Recommended reading Molecular Biology of the Cell 6th Edition Chapter 17; 965-979, 992-994 Questions? Break  Cancer Introduction Learning Outcomes Part 2: Cancer Understand the key features of cancers Identify different cancer stages/grades from a cancer’s features and how the name of a cancer relates to its origin Have examples of cancer risk factors and their involvement in cancer genesis Recall the common terms used to describe genes involved in cancer development Know genes commonly associated with cancer development and classify them from their function  Cancer Introduction Lecture Structure Part 2: Cancer Cancer - Types & NHS Facts - Features - Metastasis - Risk Factors & Development Cancer Critical Genes - Definitions - Specific gene examples Diagnosis Cancer Introduction  Cancer Introduction Cancer - Types Cancer Incidence Increase with Age 1 in 2 people will develop some form of cancer during their lifetime. >200 types of cancer grhosp.on.ca  Cancer Introduction Cancer - NHS Facts Most common types of cancer in the UK: Smoking and Lung Cancer Deaths - Breast cancer - Lung cancer - Prostate cancer - Bowel cancer Reducing your risk of cancer - Healthy eating - Regular exercise - Not smoking Key symptoms - Lump suddenly appears on body - Unexplained bleeding - Changes to bowel habits  Cancer Introduction Cancer - Features of Cancers What general features can you recall?  Cancer Introduction Cancer - Features of Cancers Multifactorial Genetic Disorder ~300 cancer critical genes - Diverse functions In a typical lifetime, every single gene is likely to have undergone 1010 separate mutations. Molecular Biology of the Cell  Cancer Introduction The Warburg effect Cancer - Features of Cancers Multifactorial Genetic Disorder Types of Genetic mutations: - Increase growth signal response - Metabolism shift - Insensitive to antigrowth signals - Evade apoptosis - Genetically unstable - Increase replication potential - Angiogenesis - Tissue invasion and metastasis Molecular Biology of the Cell  Molecular Biology of the Cell Cancer Introduction Cancer - Features of Cancers Benign/Malignant Non-cancerous Cancerous Benign tumours Malignant tumours “Cancers" - Remain localised at their site of origin - Connective/Muscle tissue: Sarcomas - Generally amenable to surgical removal - Bone marrow WB stem cells: Leukemias - Patient generally survives - Lymphocytes (and precursors): Lymphomas Exceptions: Vulnerable locations (e.g. brain) - Epithelial cells: Carcinomas  Molecular Biology of the Cell Cancer Introduction Cancer - Features of Cancers Malignant tumours “Cancers" - Connective/Muscle tissue: Sarcomas - Bone marrow, white blood stem cells: Leukemias - Lymphocytes (and precursors): Lymphomas - Epidermal cells: Carcinomas Carcinomas further subdivisions: - Squamous cell carcinoma - Resemble stratified squamous epithelium - Basal cell carcinoma - Transitional cell carcinoma - Kidney/Ureter - Adenocarcinoma - Grow in a glandular pattern. Tissue of origin is often added as a descriptor: Example 1: Renal cell adenocarcinoma Example 2: Bronchogenic squamous cell carcinoma Cancer - Metastasis Metastasis: The process by which cancer cells spread to other parts of the body. i.e. Primary tumours can form secondary tumours. Cell survival rate: 1:1,000-1,000,000 Molecular Biology of the Cell  Cancer Introduction Cancer - NHS Facts Cancer stages Stage 0 - Cancer is where it started (in situ) and hasn't spread Stage 1 - Cancer is small and hasn't spread anywhere else Stage 2 - Cancer has grown, but hasn't spread Stage 3 - Cancer is larger and may have spread to the surrounding tissues and/or lymph nodes Stage 4 - Cancer has spread to at least 1 other body organ. TNM staging system T Tumour size, with numbers 1 to 4 (1 for small, 4 for large) N Lymph nodes, with numbers 0 to 3 (0 means no lymph nodes have cancer, 3 means many do) M Metastases, with numbers 0 or 1 (0 means it has not spread, 1 means it has)  Cancer Introduction Cancer - Risk Factors Genetic and Lifestyle - Up to 50% of cancers could be prevented by a change in lifestyle - Viral and bacterial infections contribute significantly to human cancers Papillomavirus - Cervical cancers Hepatitis-B and -C - Liver cancers Pathophysiology: The Biologic Basis for Disease in Adults and Children  Cancer Introduction Time Cancer - Development Accumulation of Factors - Germline mutations - Maternal stress - Excess energy/material sources - Increase cell proliferation - Carcinogens - DNA damage - Failed DNA repair - Failed Apoptosis - Changes in gene expression & cell behaviour Pathophysiology: The Biologic Basis for Disease in Adults and Children Molecular Biology of Cancer  Molecular Biology of the Cell Cancer Introduction Cancer Critical Genes - Definitions Proto-oncogenes - Normal genes that help cells grow/divide/stay alive. Oncogenes - Mutated proto-oncogenes that have the potential to cause cancer. i.e. Gain of function/activity. Mutations often have a dominant effect (Single mutations)  Cancer Introduction Cancer Critical Genes - Definitions Proto-oncogenes - Normal genes that help cells grow/divide/stay alive. Oncogenes - Mutated proto-oncogenes that have the potential to cause cancer. i.e. Gain of function/activity. Mutations often have a dominant effect (Single mutations) Tumour suppressor genes - Normal genes that regulate cell division, DNA repair and apoptosis. Loss of function can lead to cancer. Mutations often have a recessive effect (Requires multiple mutations) Molecular Biology of Cancer Cancer Critical Genes - Retinoblastoma (Rb)  Cancer Introduction Cancer Critical Genes - Retinoblastoma (RB) Retinoblastoma Protein (pRb) - Childhood tumour develops from neural precursor cells in the immature retina - 1:20,000 Children  Cancer Introduction Cancer Critical Genes - Retinoblastoma (RB) Retinoblastoma Protein (pRb) - Childhood tumour develops from neural precursor cells in the immature retina - 1:20,000 Children - RB gene - Regulates initiation of the cell-division cycle (See part 1) - Loss of function leads to unregulated entry into S-Phase Molecular Biology of Cancer Cancer Critical Genes – RAB (Ras protein)  Cancer Introduction Cancer Critical Genes - RAB (Ras protein) Epidermal Growth Factor (EGF) Receptor Signalling - Receptor Tyrosine kinase regulates epithelial tissue development and homeostasis Classic Ras/Raf/MAPK signalling pathway….. Grb2 → Sos → Ras → Raf → Mek → Erk Oncogene Product Note: Many genes for downstream signalling proteins are also oncogenic. E.g.1 Ras GTPase activity increase. E.g.2 Myc overexpression in B cell lymphocytes.  Cancer Introduction Cancer Critical Genes - RAB (Ras protein) Ras - GTPase Protein - Part of the RTK/Ras/PI3K Pathway - RTK linked with Ras and Rho family of monomeric GTPases Molecular Biology of the Cell  Cancer Introduction Cancer Critical Genes - RAB (Ras protein) Ras - GTPase Protein - RTK linked with Ras and Rho family of monomeric GTPases Classic Ras/Raf/MAPK signalling pathway….. Grb2 → Sos → Ras → Raf → Mek → Erk MAP Kinase (Erk) has many targets Scaffold Protein Scaffold Proteins - help prevent cross-talk between Parallel MAP Kinase Modules Molecular Biology of the Cell  Cancer Introduction Note: Several anticancer drugs target the Ras-MAP-kinase signalling pathway Molecular Biology of the Cell  Cancer Introduction Cancer Critical Genes - RAB (Ras protein) RTK/PI3K Pathway - PI3K also stimulated by Ras via RTK/Ras/PI3K Active Akt → Inhibit Apoptosis → Cancer Cell Survival Molecular Biology of the Cell  Cancer Introduction Cancer Critical Genes - RAB (Ras protein) RTK/PI3K Pathway - PI3K also stimulated by Ras via RTK/Ras/PI3K Active Akt → Inhibit Apoptosis → Cancer Cell Survival Molecular Biology of the Cell  Cancer Introduction Cancer Critical Genes - RAB (Ras protein) PI3K/Akt/mTOR Pathway - Transmit signals for cell growth/division - Increases metabolism - Cell division driven with both: 1) Mitogens (e.g. glucose) 2) Growth factors (e.g. insulin and insulin-like growth factors) - Activation of the pathways can lead to abnormal cell growth Example 1: Type 2 Diabetes – High insulin Example 2: Active PI3K Molecular Biology of the Cell Molecular Biology of Cancer Cancer Critical Genes - TP53  Cancer Introduction Cancer Critical Genes - TP53 p53 Protein - Very little p53 protein under normal conditions (rapidly degraded) - Regulates cell survival - Maintains DNA stability - p53 protein tetramers bind DNA - TP53 mutated in ~50% of cancers P P P P53 P Note: Cancer cells are ineffective in DNA repair - Radiotherapy effects these cells more! Cofactor Recruitment Note: CDKN1A gene encodes p21 protein - p21 inhibits Cdk complexes required for S phase progression Borrero & El-Deiry (2021) BBA - Reviews on Cancer, 1876:188556  Cancer Introduction Cancer Critical Genes - TP53 p53 Protein Tumour incidence in p53-deficient mice - Very little p53 protein under normal conditions (rapidly degraded) - Regulates cell survival - Maintains DNA stability - p53 protein tetramers bind DNA - Mutated in ~50% of cancers - p53 not essential for normal development but… Donehower (1996) Seminars in Cancer Biology 7:269-278  Cancer Introduction Cancer Critical Genes - TP53 p53 Protein - Current drug development Restore TP53 expression Restore p53 Conformation Prevent mutant p53 signal interruption Reduce p53 degradation Borrero & El-Deiry (2021) BBA - Reviews on Cancer, 1876:188556 Cancer Diagnosis  Cancer Introduction Cancer - Diagnosis Various symptoms Blood Tests - Antigens and other Biomarkers - Circulating tumour cells (CTCs) - number and genetic analysis MRI/CT - Computerized Axial Tomography (CAT) - X-ray 3D tomography image stack - Magnetic Resonance Imaging (MRI) Biopsy - Cellular observation: Mitosis, Necrosis, Immune cells - Genetic: DNA sequencing, protein expression/activity (Biomarkers)  Cancer Introduction Cancer - Diagnosis Biopsy - Cellular observation: Mitosis, Necrosis, Immune cells Dividing cells Dividing cells Cells in all the Cells move confined to throughout the epithelial layers through the basal layer lower third are proliferating basal lamina  Cancer Introduction Cancer - Diagnosis Biopsy - Cellular observation: Mitosis, Necrosis, Immune cells Cancer grades - What the cells look like under a microscope. Grade 1 - Cancer cells that resemble normal cells and aren't growing rapidly Grade 2 - Cancer cells that don't look like normal cells and are growing faster than normal cells Grade 3 - Cancer cells that look abnormal and may grow or spread more aggressively  Cancer Introduction Summary Part 2: Cancer Cancer - Types & NHS Facts - Features - Metastasis - Risk Factors & Development Cancer Critical Genes - Definitions - Specific gene examples Diagnosis  Introduction to Cell Biology Recommended reading Molecular Biology of the Cell 6th Edition Chapter 20; 1091-1130 Questions?