PI3K inhibitors

Question 1

What is the general substrate of PI3K

Answer 1

phosphatidyl-inositol

Question 2

Where can the inositol head group be phosphorylated

Answer 2

3,4,5 free OH groups

Question 3

Which OH groups cannot be phosphorylated on the inositol head group of PI

Answer 3

Positions 2 and 6

Nor 1 because it’s already bound to the phosphate group linking it to the DAG lipid anchor

Question 4

What determines where a phosphorylated PtdIns will localise intracellularly?

Answer 4

The positions in which it is phosphorylated

Question 5

How does PI3P concentration change in response to growth factor stimulation

Answer 5

Doesn’t change, considered to be constitutive lipid

Question 6

How do PIP3 and PI(3,4)P2 concentrations change after growth factor stimulation

Answer 6

Levels of the lipids rise rapidly following growth factor stimulation

Question 7

what is the result of a PtdIns being phosphorylated at the 3-OH position?

Answer 7

It’s no longer a substrate for phospholipase

Question 8

What kind of proteins do PIP3 and PI(3,4)P2 bind to

Answer 8

Proteins with a plextrin homology domain which is found in protein kinases, adaptor proteins and regulators of small GTPases

Question 9

What protein domains do PI(3)P bind to? and where are these found?

Answer 9

FYVE - Mostly in proteins involved in membrane trafficking

PX - in proteins that regulate NADPH oxidase complex, endocytic trafficking

Question 10

What are the four members of the class I PI3K isoforms

Answer 10

p110alpha/PIK3CA, p110beta/delta/gamma

Question 11

What are the 3 members of the class II PI3Ks

Answer 11

C2alpha, C2beta, C2gamma

Question 12

What is the class III PI3K

Answer 12

vps34

Question 13

What is the substrate of the class I PI3Ks and what do they produce from it

Answer 13

PI(4,5)P2 is the substrate and it forms PIP3

Question 14

How does PIP3 bind to downstream protein substrates

Answer 14

Via the proteins PH domain

Question 15

What is the substrate of class II/III PI3Ks

Answer 15

PI and converts it to PI(3)P which binds its target proteins by FYVE and PX domains.

Question 16

Why is there a delay in the production of PI(3,4)P2 compared to PI(3,4,5)P3 following growth factor stimulation?

Answer 16

PIP3 is produced first, then through the action of 5-phosphatase PIP3 is used to make PI(3,4)P2

Question 17

What are the class 1A PI3Ks

Answer 17

p110alpha, beta and delta

Question 18

what is the class 1B isoform of PI3K

Answer 18

p110 gamma

Question 19

How are class 1A PI3Ks activated?

Answer 19

By tyrosine kinase receptors, the SH2 domain of the regulatory subunit on each isoform binds to the phosphorylated kinase domain on the tyrosine kinase
Also activated by Ras/Rac/cdc42

Question 20

How are class 1B PI3Ks activated?

Answer 20

By GPCRs, also by Ras/Rac/cdc42

Question 21

What is the lipid phosphatase that regulates the production of PIP3

Answer 21

PTEN

Question 22

Which position on PIP3 is dephosphorylated by PTEN

Answer 22

At the 3 position

Question 23

Can PI3K signalling cause treatment resistance

Answer 23

Yes, following hormonal therapy, chemotherapy, radiotherapy and targeted agents

Question 24

Which parts of the PI3K pathway often become mutated in cancer

Answer 24

Oncogenic tyrosine kinases and Ras
PIK3CA/p110 alpha frequently mutated in cancers
Akt/PKB is activated as a result of mutation or upstream PI3K activation

Question 25

What are the hotspots in which PIK3CA becomes mutated in solid tumours?

Answer 25

The helical domain and the kinase domain.

Question 26

What kind of mutations are found in PIK3CA in segmental overgrowth syndromes?

Answer 26

Mosaic mutations in PIK3CA during embryonic development

Question 27

Why does the inhibition of mTORC1 using rapamycin actually increase the amount of PI3K signalling

Answer 27

Because: Downstream of mTORC1 is s6K1, a kinase which inhibits IRS-1 which is responsible for PI3K activation. By inhibiting mTORC1, this negative feedback loop no longer functions and PI3K signalling is switched on and other targets of AKT can be targeted (not just mTOR)

Question 28

Can cells survive with a low amount of PI3K activity? and does this effect proliferation?

Answer 28

Cells can survive and proliferate normally with <10% of their class 1A PI3K activity

Question 29

What kind of PI3K inhibitor is Piqray?

Answer 29

A p110 alpha inhibitor.

Question 30

What combination therapy is used in ER+ breast cancer with Piqray and why is this the case?

Answer 30

p110 alpha inhibitors in ER+ breast cancer increases the expression of ER. Therefore, when combined with hormonal therapy there is a synergistic effect on tumour regression

Question 31

Where is p110 delta mainly expressed?

Answer 31

Leukocytes

Question 32

Does PI3K delta expression increase in leukemic cells compared to normal ones?

Answer 32

No, its present at high levels in both cell types

Question 33

In leukemia, is PI3K delta mutated?

Answer 33

Very rarely

Question 34

Which specific haem-malignancies are PI3Kdelta inhibitors effective in

Answer 34

B cell malignancies. Specifically B cell chronic lymphocytic leukaemia (CLL) and B cell follicular lymphoma

Question 35

Why is PI3Kdelta inhibition only effective in B cell malignancies?

Answer 35

In CLL, B-cells have a particular life cycle, going between the lymph nodes and bone marrow and then into circulation. To access the lymph nodes/bone marrow they respond to chemokines, to stay they resond to adhesion molecules, for co stimulation they respond to B-cell antigen receptors; all of which rely on PI3K delta. If this is blocked, the cells can no longer respond to the signals and accumulate in the circulation. In the circulation they are vulnerable to combination therapy (anti CD20 immunotherapy)

Question 36

Why are normal B cells dependent on PI3Kdelta for survival?

Answer 36

Signalling through adhesion molecules, chemokines, co-stimulatory molecules and B-cell antigen receptors all rely on PI3K delta for the modulation of these signals

Question 37

Are PI3K delta inhibitors responsible for the cytotoxic effect on B-cells

Answer 37

No, only make them vulnerable to the therapy used in combination

Question 38

What effect does PI3K delta inhibition have on different T cell populations and what effect does this have on solid tumours?

Answer 38

Greatly inhibits Tregs whilst only having a limited inhibition on effector T cells (Th and cytotoxic T cells) This tips the balance towards an active, anti-cancer response.