KRAS/NRAS

Question 1

What molecular technique does the RAS service use?

Answer 1

Pyrosequencing

Question 2

What sample type does the RAS service require?

Answer 2

FFPE blocks which we cut

Or sections that we cut ourselves

Question 3

What is the problem with FFPE? Why does it work well with Pyro?

Answer 3

Cross linked DNA - cross links form between bases which interfere with PCR
DNA becomes heavily fragmented

Works well with Pyro because it is only sequencing short reads of DNA

Question 4

What type of referrals does our RAS service mainly receive?

Answer 4

Referrals from patients with metastatic colorectal cancer which expresses EGFR

Question 5

What is EGFR?

What is it’s significance in colorectal cancer?

Answer 5

Receptor tyrosine kinase that transmits a growth-inducing signal to cells

It is upregulated in colorectal cancer

Ligands that activate EGFR are usually tightly controlled, however in a tumour micro-environment these ligands are increased and lead to upregulation of EGFR

Drugs such a cetuximab target this TK and block its activity

Question 6

What is the significance of RAS mutations with regards to EGFR therapy.

Answer 6

Cetuximab won’t work in patients that have RAS mutations as these mutations cause constitutive activation of the pathway regardless of EGFR

NICE guidelines only recommend Cetuximab in wild type RAS patients

Question 7

What are the RAS genes that we test? What codons do we look at?

Answer 7

NRAS and KRAS

codons 12, 13 and 61 in both

Question 8

What type of mutations can be present? What do these mean?

Answer 8

Base change can cause:
transition - pyrimidine swapped for a pyrimidine
transversion - pyrimidine swapped for a purine

Question 9

Name a common mutation we see in KRAS.

Answer 9

c. 35G>T p.(Gly12Val)

c. 35G>A p.(Gly12Asp)

Question 10

What is the sensitivity of the RAS assay?

Answer 10

Change in peak height - 10%

New peak can be down to 5%