MSOP-1004 Cancer Biology Lectures 5 & 6 2024 PDF

MSOP-1004 Biomedical Sciences & Therapeutics: Cancer Biology – Lectures 5 & 6 Dr Simon Scott  BRCA1 & 2 ‘breast cancer genes’ The BReast CAncer susceptibility genes: BRCA1 (chromosome 17) & BRCA2 (chr13) Autosomal recessive inheritance Some families show v. increased risk of b...

MSOP-1004 Biomedical Sciences & Therapeutics: Cancer Biology – Lectures 5 & 6 Dr Simon Scott  BRCA1 & 2 ‘breast cancer genes’ The BReast CAncer susceptibility genes: BRCA1 (chromosome 17) & BRCA2 (chr13) Autosomal recessive inheritance Some families show v. increased risk of breast/ovarian cancer due to inherited mutations Individual lifetime risk of cancer: 50-80% for breast cancer (5-7% of overall cases – ~95% non-inherited/ sporadic), 30-50% ovarian cancer. Also increases inherited prostate, pancreas & colon cancer  BRCA1 & 2 BRCA proteins involved in DNA repair (ds breaks), DNA recombination, regulating cell cycle, and probably inhibition of oestrogen receptor gene (suppressing signalling) Truncated proteins often produced (loss of function) In some sporadic cases, epigenetics can reduce gene expression (mentioned in PHAM1054)  PTEN A phosphatase is encoded by the PTEN (phosphatase and tensin) gene. PTEN enzyme dephosphorylates the membrane lipid PIP3 (phosphatidyl-inositol-3 phosphate) → PIP2, antagonising the PI3 kinase that phosphorylates PIP2 → PIP3 stimulating cell prolif/inhibiting apoptosis Loss of PTEN activity → PI3 kinase pathway ON → unregulated cell proliferation → carcinogenesis Epidermal Growth Factor PTEN PIP2 PIP3 PI3K AKT + mTOR protein kinases EGF receptor Proliferation Apoptosis PTEN, Cowden syndrome & cancer Germline PTEN mutation linked to Cowden syndrome (~1:20,000 people) PTEN on chromosome 10 Autosomal dominant inheritance – dominant negative effect (PTEN dimer) – (like Huntington’s disease – PHAM1054) Growths on skin and mucous membranes Predisposition to cancers: prostate, breast, glioblastoma, thyroid, uterus, kidney & melanoma 70% of prostate cancers have mutant PTEN N.B. not all phosphatases are TS or kinases oncogenic Harmatomas – pre-cancerous skin/intestinal growths in Cowden Syndrome  p53 and cell cycle Normally, p53 is continuously synthesised and rapidly degraded after binding to Mdm2 If DNA is damaged by chemicals/radiation, both p53 & Mdm2 become phosphorylated and can’t bind together p53 now accumulates and initiates expression of >50 genes (p53 is a transcription factor). Binds as a tetramer to responsive elements in gene promoters Mutations in p53 alleles have a dominant negative effect as mutant p53 subunits in the tetramer interfere with function of the whole complex  N.B. Estimated that 20,000 bases of DNA are modified/damaged by endogenous ROS in a single cell!  p53 and cell cycle p53-mediated p21 protein production inhibits Cyclin D/Cdk4 and ‘arrests’ cell cycle at G1/S boundary/‘checkpoint’, giving time for enzymes to repair DNA damage before cell division Similarly p53-mediated p21 inhibits cyclin- cdk complexes at G2/M checkpoint Mutant p53 often unable to bind to target gene promoter - can’t initiate cell cycle arrest  p53 and checkpoint control DNA damage phosphorylation See also Fig 6.5 in P P Mdm2 p53 Pecorino textbook promoter P21 CyclinD Cdk4 p21 G1 S P53 acting as transcription factor  …via Rb protein P53 acting as transcription factor Rb1 is NOT phosphorylated → cell cycle arrest at G1/S checkpoint  P53, DNA repair and angiogenesis p21 also binds proliferating cell nuclear antigen (PCNA) inhibiting in DNA replication p53 also directly initiates transcription of DNA repair enzyme gene (e.g. XP) mutant p53 initiates transcription poorly p53 also initiates transcription of the thrombospondin gene – inhibitor of angiogenesis  p53 and apoptosis p53 is pivotal in the cell’s decision to undergo cell cycle arrest/DNA repair OR apoptosis p53 transcriptionally regulates several mediators of apoptosis (see Table 6.2 in Pecorino textbook) After extensive DNA damage the cell ‘commits suicide’ rather than producing mutations. But if lack of functional p53 → poor apoptosis Mitochondrial pro-apoptotic proteins are controlled by p53. These release cytochrome c which activates the ‘apoptosome’ → Stimulation of intrinsic apoptosis  p53 and apoptosis p53 induces several genes involved in apoptosis For example, p53 controls the BAX/Bcl2 apoptotic pathway. DNA damage causes p53 phosphorylation leading to expression of pro-apoptotic BAX protein… p53 also drives expression of transmembrane receptor genes such as Fas – extrinsic apoptosis stimulation In both cases a series of enzymes called caspases are involved in a cascade which leads to apoptotic cell death e.g. Fas p53 and apoptosis DNA damage phosphorylation p53 P P Mdm2 promoter BAX BAX Bcl2 BAX Apoptosis Bcl2 No apoptosis p53 – Responses to DNA damage p53 downstream effects - some examples See also Fig 6.3, 6.6 & 6.7 in textbook  P53 mutations 75% of p53 gene mutations are ‘mis-sense’ mutations leading to a single amino acid substitution. Most of these are in the DNA binding domain Mutations in p53 pathway proteins (e.g. Mdm2, Bax) also often found in tumours. Mutations in the p53 gene can be inherited if they are present in germ-line cells. This gives offspring a genetic predisposition to developing cancers (as they have a genetic instability/mutator phenotype)  Li-Fraumeni syndrome Inherited p53 mutations are characteristic of Li-Fraumeni (cancer) syndrome (LFS), an autosomal dominant condition (chromosome 17) > 70% of LFS patients have germline p53 mutation (other de novo, or other genes) → 25x increased risk of cancer by 50 yrs old Individuals with LFS usually carry one functional p53 allele (i.e. heterozygous) Unlike most tumour suppressors, some specific p53 mutations have a dominant negative effect. The mutant p53 binds to wild-type p53 (tetramer) making complex nonfunctional Figure shows p53 tetramer bound to DNA and red ‘hotspots’ of mutation seen in cancer – mostly affect DNA binding domains See also Fig 6.4 in Pecorino textbook  p53 mutations Some mutations can lead to reduced p53 expression/ tumour suppression – haploinsufficiency (one wt allele, but doesn’t produce enough protein) Thus p53 tumourogenesis is usually not due to a loss of hetereozygosity (LOH) – an exception to most TS genes p53 null ‘knockout’ mice develop tumours when young LFS can also be caused by mutations in other genes involved in the ‘p53 pathway’ (e.g. CHEK2) - e.g. proteins involved in controlling cell cycle checkpoints  ‘Knockout’ mice Hetero/homozygous ‘knockout’ mice have been used to study impact of gene mutations in many genes LFS inheritance HPV & cancer: Rb/E7 N.B. Not sex linked Virus interference with TS proteins Some oncogenic viruses express proteins that inhibit function of TS proteins as part of their replication Human papillomavirus (HPV) E7 inactivates Rb → overrides G1/S checkpoint. HPV E6 inactivates p53 → prevents apoptosis of infected cells E6 & E7 promote p53/ Rb degradation Both contribute to cell proliferation: Warts (benign), cervical cancer (malignant) Amino acid 72 in p53 gene is polymorphic: usually proline or arginine. Arginine more susceptible to E6 degradation of p53 → higher mutation rate. With both Arg alleles: 7x cervical cancer incidence Virus interference with TS proteins Also see Fig 6.9 in Pecorino textbook HPV E6 interference with p53  Cancer Treatment Cancer formation (carcinogenesis) and its molecular basis is complex – treatment solution isn’t easy either! ‘Conventional’ treatments: surgery, radiotherapy & chemotherapy still predominate However, research into molecular mechanisms of cancer is leading to new technologies and drugs. New combination therapies often more effective  Principles of Cancer Therapies First treatment was surgically excision of tumour mass Difficult for more inaccessible tumours. Note that any cells remaining can potentially ‘repopulate’ by clonal expansion. Also tumour may have metastasised to unknown secondary sites Surgery still used by often in combination with radiotherapy and/or chemotherapy These newer therapies aim to be cytostatic (prevent proliferation) and cytotoxic (kill cells). Principles of Cancer Therapy Radiotherapy and most chemotherapies aim to cause DNA damage → cell death (necrosis, apoptosis). Most tumour cells grow in a fast, uncontrolled way - impaired cell cycle arrest and DNA repair, so more cytotoxicity. However, fast growing normal cells often affected (e.g. hair follicles, gut epithelium) in systemic treatment Some new drugs target angiogenesis, preventing neovasculature formation → tumour necrosis  Tumour Cell Sensitivity Not all cells equally sensitive to therapy. This hetereogeneity can be due to cell type (e.g. proliferation rate, DNA repair efficacy), development of resistance (i.e. gene mutations in tumour) or location in tumour (e.g. distance from blood vessel for drug delivery, hypoxia)  Tumour Cell Sensitivity Drugs designed to have maximum efficacy with minimum side-effects The therapeutic index (TI) indicates this – the difference between minimum effective dose (MED) and maximum tolerated dose (MTD) Greater therapeutic index, less side effects (most drugs) Most chemotherapies MaxTE chemo X used near MTD Effect aspirin X MinE TI See Pecorino Fig 1.6 Ch1 MED Dose MTD  Chemotherapeutic Agents Drugs that target DNA, RNA and proteins, to damage DNA/disrupt normal cell cycle → apoptosis (if enough damage) Side effects on rapidly proliferating normal cells (e.g. alopecia, bone marrow depletion). Can disrupt treatment regimen Novel, more efficacious drugs with less side effects desired Knowledge of molecular differences between neoplastic and normal cells can aid drug design  Clinical Trials Drug testing is multiple stages; cell culture, animal models, clinical trials (Phase I-III) Phase I (n