Unit 8 - Resistance Mechanisms

Question 1

Around what percentage of ER+ patients have A) de novo resistance and B) acquired resistance (in early patients)?

Answer 1

30% for both

Question 2

Describe 4 mechanisms for anti-hormone resistance

Answer 2

Pharmacogenetic – CYP2D6 SNPs can affect the response – those with poor enzyme activity have poorer response – this enzyme carries out the conversion of tamoxifen to endoxifen the more potent inhibitor of ER

Alterations in cell cycle/apoptosis
Increased expression of cyclin D1 (amplified in 60% of luminal B cancers) – to prevent G1 arrest
Increased AKT signalling can lead to mis-localisation of p21/p27 as well as changes in apoptotic elements (Increase BCL-2 and decreased Bad)
Also upregulation of GRP78 can regulate autophagy to prevent cell death

ER negativity/loss
Around 15-25% go onto lose ER – has been linked to EGFR/HER2 hyperactivation as well as PI3K/MAPK signalling and mutations in FOXA1.

Epigenetic events
40% of TNBC have DNA methylation of ER promoters and miRNA-206 up-regulation. This can also be due to other tumour suppressor genes like RB/PTEN.
Additionally histone deacetylation can result in decreased expression of some genes – and inhibitors of HDACs can actually recover ER in ER- cells.

Question 3

Describe ways that ER activity can be lost

Answer 3

1. Mutations in ER are seen in 25% of patients with acquired anti-hormone resistance – most in helix 12 (Y537S) which lead to constitutive activation via the AF-2 domain.
   As well as splice variants
2. Deregulation of ER coregulators – Increased expression of FOXA1 or mutation of NCoR (Or inhibition at phosphorylation level)
3. Deregulation of growth factor signalling (entire pathway GFs + kinases)– which can promote proliferation

Question 4

Describe the ways that growth factors signalling can promote anti-hormone resistance

Answer 4

• Promote Nuclear transcription factors like AP-1 promoting non-classical genomic signalling at AP-1 responsive elements
• Can phosphorylate AF-1 at S118 or its co-activators to promote ligand independent genomic signalling
• Growth factor pathway can also promote its own target elements to promote proliferation
• Can increase non-genomic signalling by hyperactivation of molecules like Src

Question 5

Discuss the various mechanisms that have been investigated in the treatment of anti-hormone resistance.

Answer 5

1. Targeting signalling pathways
   HER2 and EGFR are commonly upregulated in these cells and can help provide receptor cross-talk mechanisms which drive resistance – so use of herceptin/gefitinib have been investigatd – clinical trials mixed
2. Co-treatment of anti-hormones and RTK inhibitors
   Given the up-regulation of these molecules is an early event in resistance (due to ER inhibiting their expression) suggestion that this might be effective – trials also inconclusive as many RTKs can contribute to signal cross-talk
3. Targeting mTOR
   Around 70% of breast cancer patients have PI3K pathways hyperactivation (PI3KCA or PTEN) which relates to poor prognosis – molecules like Everolimus (as well as Buparlisib and apelisib) can be used alongside anti-hormones like exemestane
4. Targeting CDKs
   CDK4/6 hyperactivation (or loss of INK4) found in about 20% of anti-hormone resistant patients so use of Palbociclib prevents its activity (Rb phosp and E2F transcription) – can be used alongside anti-hormones in either 1st line or 2nd line therapy.
   Also ribociclib and albemaciclib

Question 6

What is the disadvantage of Palbociclib in treating anti-hormone resistant breast cancer?

Answer 6

Resistance occurs within about 2-3 years in most patients by:

• CDK6 hyperactivation
• Cyclin E amplification
• RB loss/mutation
• P53 loss
• Akt mutation/hyperactivation
• RTK/RAS amplification

Question 7

How does Everolimus work?

Answer 7

Binds to FKPB12 via a methyl group and this complex blocks Mtorc1/2 kinase activity blocking both protein synthesis and AKT – which may also play a role in blocking cross-talk with ER
Is an allosteric irreversible inhibitor

Question 8

What mechanisms do melanoma cells become resistant to vemurafenib treatment?

Answer 8

Upstream mechanisms – hyperactivation of RTKs (IGFR/EGFR/PDGFR) or their ligands
Downstream mechanisms – BRAF amplification/alternative splicing, MEK/RAS mutations, CRAF hyperactivation, COT/MLK1-4 hyperactivation (MEK/ERK) or NF1 inactivation
Alternative pathways – Cyclin D1 hyperactivation, PTEN deletions, PI3K/Akt hyperactivation
Low levels of Jarid1b due to microphthalmia-associated transcription factor (MTIF) which result in slow cycling, chemo-resistant cells

Question 9

What mechanisms are there to treat vemurafenib resistant patients

Answer 9

• Immunotherapy like ipilimumab/nivolumab/pembrolizumab
• Combinations of trametinib(MEK inhibitor) or pan-Raf/src inhibitors with vemurafenib
• Change dosing regime to ensure a proportion of the tumour population is sensitive to therapy