T Cell Therapeutics

Question 1

What is preparation of CAR T cells?

Answer 1

Taking T cells out of someone’s body
Purify and magnetically store for CD4 and CD8 T cells
Transfect a vector into those T cells that express chimeric antigen receptor - transduce them normally with a lentivirus
Infused back into patient to attack cell it is binding to

Question 2

What is a chimeric antigen receptor?

Answer 2

Normally a single chain Fv (a single antibody heavy and light chain joined together into a single protein) linked onto a membrane component to embed into the membrane , then link to set of activating domains

Question 3

What activating domains did first generation CAR-T cells have?

Answer 3

ITAM related to ZAP70 which is related to signalling from T cell receptor

Question 4

What do new generation CAR T cells have as activating domains?

Answer 4

Have additional co stimulatory domains as ZAP 70 on its own wasn’t sufficient

Question 5

What were the first approvals for CAR-T cells all targeting in 2017?

Answer 5

Blood based tumours not solid tumours

Question 6

What is the black box warning for CAR-T cells?

Answer 6

Severe risk of side effects e.g. cytokine release syndrome, neurotoxicity

Question 7

Why do CAR-T cells have high success rates?

Answer 7

They hang around so if any cancer cells escape first wave, they are likely to be targeted later on

Question 8

How can we prevent cytokine release syndrome?

Answer 8

Have antibdoies that can mop up high levels of TNF alpha, IL-1 beta and IL-6 etc.

Question 9

What has been done to help the issue of balancing survival v exhaustion?

Answer 9

Changes made in costimualtory molecules, can change the way T cells behave

Question 10

What is the problem with viral integration site

Answer 10

Can affect other genes, and proliferation. Lentiviruses are less problematic than other retroviruses but can still have issues

Question 11

What is the problem with memory CAR-T cells?

Answer 11

Want some cells in effector memory pool that can be reactivated in future for sustaining CAR-T survival in patients long term

Question 12

What does stimulatory domain 41BB do?

Answer 12

Slower expansion of cells, means they are more durable

Question 13

What does stimulatory molecule CD28 do?

Answer 13

Fast expansion, less durable

Question 14

What is the issue with resistance around CAR-T cells?

Answer 14

Avoiding resistance is an issue

Question 15

What does CD40L accessory molecule do?

Answer 15

Facilitates T cells memory formation and induces increased immunogenicity on tumour cells

Question 16

What does ICOSL do?

Answer 16

Enhances PI3K signalling pathwya and upregualted the expression of Bcl2

Question 17

What does IL-7 do in CAR-T cells

Answer 17

Inhibits T cells apoptosis and exhaustion

Question 18

what is the problem with CAR-T cells getting to the site?

Answer 18

Normal T cells get to the site thorugh other cells producing chemokines, then once it is there has peptides and MHC that it recognises to become activated
Tumours might not necessarily do this, therefore how you get T cells into a tumour, is by finding a reason for the T cell to want to go into the tissue in the first place

Question 19

What does IL-12 do in CAR-T cells?

Answer 19

Mitigates Treg-suppression and reprogram tumour-assocaited macrophages and dendritic cells

Question 20

What is anakinra?

Answer 20

Il-1 receptor antagonist

Question 21

What is tociluzimab?

Answer 21

IL-6 receptor blocker

Question 22

What is lenzilumab?

Answer 22

GM-CSF binder

Question 23

What does CD28 costimualory molecule do?

Answer 23

Facilitates full and sustained T cell activation, growth and survival

Question 24

What does 4-1BB co stimulatory molecule do?

Answer 24

Be assocaited with more anti-apoptotic proteins and reduced T cells exhaustion

Question 25

what does ICOS co stimulatory molecule do?

Answer 25

Presents the characteristics of Th17 cells with increased expression of Il-17A, IL-17F, and IL22 following antigen recognition

Question 26

What is the problem with genetic loss of antigen?

Answer 26

If tumour cells start to lose the target molecule that you are working with then CAR-T cells can no longer see those tumour cells

Question 27

What is the problem with CAR transduction of tumour?

Answer 27

Because you normally take these T cells out the patient themselves, if you have put some vector into tumour cells, then you can have CAR tumour cells whihc can mask and block off ability of CAR T cells to see the antigen

Question 28

What is trogocytosis?

Answer 28

T cell can itself remove antigen, means T cell cna interact with tumour cell but instead of killing it, can have antigen transfer to surface of T cell and this decreases antigen on tumour cell and makes it difficult to see how T cells can target required cells

Question 29

What is the problem with loss of death receptor signalling on tumour cells?

Answer 29

Mutaitons in cells can take away ability to respond to cell death triggers, lose ability to kill tumour cell well

Question 30

What is the problem with immunosuppressive tumour microenvironment?

Answer 30

If you have a solid tumour, often find environment is immunosuppressive. Don’t turn on and activate T cells and other immune cell to cause damage, if anything it is the other way round to try and stop them

Question 31

How can we improve CAr-T cell performance?

Answer 31

Making small alginate polymer, porous but contains lentiviruses, some cytokines and everything it needs. So when T cells went in there, they would get ransfected by virus and stimulated. Can all be done in vivo

Question 32

What is good about alginate polymer?

Answer 32

Doesn’t produce all cells at once which reduces risk of things likecytokine release syndrome

Question 33

What do you have to ensure with alginate polymer?

Answer 33

Make sure vectors only express in T cells under certain conditions, this reduces likelihood of your tumour cells expressing chimeric antigen receptor

Question 34

How can you improve CAR-T cells for solid tumours?

Answer 34

Have to start with T cells that have already infiltrated that tumour so need to take a biopsy and isolate specific T cells. Find those which are cytotoxic So instead of taking T cells at random out of bloodstream, take ones specifically found in the tumour

Question 35

What are some of the ways in which next-generation CAr-T cells can be improved?

Answer 35

Bivalent CAR/ LINK CAR - two single chain Fvs together, can make things wider in specific or so that you have to link both to activate it, making it more specific SynNotch - first target is primer, second recognises after priming SNIP CAR - being able to turn off CAr T cells HLAs - can get immune system to have a blind spot to that particular cell expressing HLA Tethered - have tethered cytokine by peptide linker, can actually stimulate receptors on that CAR-T cell

Question 36

What are benefits of CAR-NK cells?

Answer 36

Good tumour infiltrating capability Can kill if they recognise antibdoies, cytokines, and through perforins and granzymes Kill other cells first without being primed Reduces risk of graft v host disease CD19 works just as well on an NK cell as it would on T cell