Lesson 8 Evading Growth Supressors, Replicative Immortality

• Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell. 2011;144:646-74.

  
• TGF-β pathway

  1. Cancerous mutations in pathway components

  2. Anti‐growth signalling

  3. TGF‐

  4. P

  5. Smad

  6. Smad P

  7. Cell cycle inhibitors

  8. Transforming Growth Factor (TGF)‐β receptors

  9. P Smad

  
• Virtually all carcinomas (~ 85% of all cancers) are resistant to the growth-inhibitory effects of TGF-β.

  
• Common resistance mechanisms

  Deletion or inactivation of TGF-β receptors or Smads

  
• A third of ovarian cancers examined in one study had mutations in the TGFβ-receptor I gene, which result in loss of expression of the protein.

  
• normal


  • ovarian tumours
• Smad 4 deletions or mutations are associated with tumours that have invaded normal tissues.

  
• 97% slow-growing pancreatic tumours expressed Smad4, 44% of more aggressive pancreatic tumours did not.
• Pancreatic cancer symptoms

  • Jaundice
  • Light-coloured stools or dark urine
  • Pain in abdomen and/or back
  • Weight loss for no known reason
  • Loss of appetite
  • Fatigue
• All vague: often correct diagnosis takes time, By diagnosis many pancreatic cancers have metastasised

  
• Pancreas function


  • Produces enzymes to break down food
  • Produces hormones insulin and glucagon to regulate blood glucose
• Pancreatic cancer is the 11th most common, 3rd most lethal
• Animal model of pancreatic cancer

  • Mouse mutants to examine genetic changes associated with pancreatic cancer
  • >90% of pancreatic carcinomas have activating K-RAS mutations
  • 55% (more aggressive tumours) bear deletions/mutations in SMAD4

  
• Animal model of pancreatic cancer

  1. Generated mice expressing an activated KRas gene in pancreatic cells during embryogenesis

  2. Crossed those mice to animals lacking Smad4

  3. Observed pancreatic cancer formation in the mice with different genotypes

  4. Mice with both mutations developed pancreatic cancer faster than those with only one

  
• Very difficult to overcome loss-of-function alterations, Mutations in TGF-β receptors or Smads tend to be gene deletions or frame-shift mutations, Can't design drugs to enhance activity of proteins that aren't made

  
• Gene therapies might be needed to re-introduce the wild type gene (or correct the mutation) so the missing protein can be made, would need most cancer cells to be altered…impossible?

  
• Targeted therapies to restore growth suppression pathway? No
• Hayflick limit on proliferation

  • Normal cells do not divide forever
  • PDL = population doubling limit

  
• Telomeres


  • The ends of chromosomes
  • Cell divides and chromosomes are replicated, ends (50-100bp) not replicated, the telomeres are eroded, chromosomes get shorter
  • Eventually, "interior" DNA damaged and/or chromosomes fuse
  • Cells stop dividing: senescence, apoptosis
  • Number of times cells divide before this: "Hayflick limit" (45-65 for somatic human cells)

  
• Telomerase

  • Restores telomere length
  • Adds DNA after every cell cycle, restores chromosome length
  • Expressed in embryonic stem cells, germ cells but not normal somatic cells
  • Cancer cells re-activate telomerase: they can bypass the Hayflick limit, replicate indefinitely
• Telomerase composition

  • TERT: a reverse transcriptase protein enzyme
  • TERC: an RNA that provides the template for reverse transcription
  • These associate with scaffolding proteins to form a complex
• TERT promoter was more frequently mutated (C146T, C124T) in tumours than any other non-coding sequence
• Cells with wild type sequence downregulated TERT expression and telomerase activity upon differentiation, but those with promoter mutations retained expression and activity
• Melanomas with mutations in TERT promoter and Ras or Braf


  Have worse prognosis
• Imetelstat


  • A lipid-conjugated oligonucleotide that is complementary to and binds with high affinity to the RNA template of telomerase
  • In vitro: blocked telomerase activity and promoted senescence
  • In mice implanted with lung cancer cells expressing luciferase: reduced tumour growth

  
• • Imetalstat was assessed in a randomised controlled trial, in pre-treated patients with non-small cell lung cancer
  • Slight benefit of Imetalstat improvement in survival rates: not statistically significant
  • Seemed more effective in patients with short telomeres: not statistically significant

  
• Limitation of targeting telomerase: Drugs like imetalstat only stop telomere restoration, cells will continue to divide until telomeres are short enough to cause cell death/ senescence, too slow!