Arginine

Question 1

What are CARs?

Answer 1

Chimeric antigen receptors they are artificially designed protein receptors that recognise specific antigens on cancer cells and activate immune cells (e.g. T cells) to directly recognise and destroy tumour cells. CAR is built of TCR and antibody

Question 2

How are CAR T cells made?

Answer 2

1. Collect blood from a patients
2. Isolate T cells
3. Transduce T cells (viral gene transfer)

Question 3

What is mechanism of action of CAR T cells?

Answer 3

for example in neuroblastoma, tumours express GD2 on the surface and CAR T cells engineered to recognise GD2 will activate and release perforin and granzymes, creating pores in tumour cells which results in tumour killing

Question 4

What makes CAR T cells clinical trial successful?

Answer 4

• Cells need to be detectable by flow cytometry
• CAR T cells need to persist for weeks
• Response correlates with cancer clearance
  E.g. CAR T cells for acute lymphoblastic leukaemia
  (Lee 2015)

Question 5

Are CAR T cells working for all cancers?

Answer 5

No, for solid cancers it is harder than for blood cancers. E.g. for clinical trial for glioblastoma, anti-GD2 CAR T cells showed low efficacy with majority of patients not responding to treatment (Louis 2011)

Question 6

Are CAR T cells successful in all blood cancers?

Answer 6

No, it is not successful in acute myeloid leukaemia as CAR T cells fail to recognise CD33. These CAR T cells can’t be detected by flow cytometry and not many patients benefit from this therapy (Wang 2015)

Question 7

Why did standard CAR T cells fail?

Answer 7

• They fail to divide
• TME is very suppressive e.g. no IL-2 for proliferation of T cells, presence of Tregs, MDSCs, macrophages etc, competition for nutrients e.g. glucose or arginine

Question 8

How tumours compete for arginine?

Answer 8

Tumours consume a lot of arginine because of high intrinsic expression of enzyme arginasee which is main enzyme which metabolise arginine

Question 9

What is neuroblastoma?

Answer 9

It is a paediatric cancer and it is the most common extra cranial solid cancer in children that has one of the lowest survival rates of childhood cancers.

Question 10

What is arginine?

Answer 10

It is an amino acid needed for T cell proliferation and cell viability

Question 11

How does arginine act?

Answer 11

1. Arginine binds to two arginine receptors (CAT-1 and another unknown receptor)
2. Inside cells, arginine is metabolised by arginase 1 and arginase 2 in urea cycle into ornithine which is important for cell proliferation (arginasee 1 is a cytoplasmic enzyme and arginase 2 is mitochondrial enzyme)
3. Arginine can also be metabolised by NOS and converted into citrulline
4. Tumour cells and T cells don’t express ‘recycle’ enzymes; OTC and ASS-1 so they are dependent only on arginine from the environment and without arginine they cannot divide

Question 12

Neuroblastoma and arginase

Answer 12

Neuroblastoma has a high expression of arginase 2 and show high arginase activity and is able to metabolise a lot of arginine

Question 13

Neuroblastoma and lab of Carmela De Santo

Answer 13

They look into improving CAR T cells in treatment of neuroblastoma by:
- Inducing expression of recycle enzymes OTC and ASS-1 by CAR-T cells so that they can synthesise their own arginine
- Re-engineering CAR T cells to compete for arginine metabolism by expressing higher amount of arginase 1 and arginase 2
- CAR T cells adaptation to other amino acids (cysteine and tryptophan)
All of the above improvements lead to increased proliferation of CAR T cells and stronger anti-tumour response.