Thread by @Biotech2k1

How to Build a Cell Therapy:

A look at the editing that goes into CAR-T and CAR-NK cell therapies.

1/ The first major challenge to any cell therapy is to overcome the rejection of the cells by host immunity. All cells have a MHC code on them that labels them as "self".

2/ When a person's MHC code does not match someone else, you can get the cells attacking the patient or the patients immune cells attacking the donor cells.

3/ There are 3 cells of the immune system that can target and kill other cells. To build a truly stealthy cell therapy, you have to overcome at least T cells and NK cells. The Macrophages are yet unproven if it makes any effect.

4/ If you are engineering a T cell, this can be done in 2 ways. The first is to knock out the T cell receptor (TCR). This will prevent the T cell from seeing the patient as foreign and attacking healthy tissues.

5/ It can be done in a 2 step process by inserting the CAR then knocking out the TCR, or it can be done by inserting the CAR into the TCR's location in the genome. This is a 1 step process that replaces the TCR with the CAR.

6/ There is some data for the benefits of inserting the CAR into the TCR locus, but it also lowers efficiency dramatically as its a very precise edit. Most TCR knockout is 98% efficient while insertion is more around 75% efficient.

7/ The next edit will be to knock out the T cells own MHC code. Since it is a cell, it displays its own MHC code of the person it belongs to. This can cause the recipients immune cells to kill it as foreign. Knocking out the MHC I is a key step.

8/ An even better step is to do a targeted insertion of the HLA-E type into that MHC locus. This not only removes the MHC so it won't cause T cells to reject it, it also uses the HLA-E to prevent NK cells from rejecting it.

9/ This single gene substitution has multiple big benefits by masking that cell from both the T cells and NK cells of the recipient. For NK cells they don't have a T cell receptor so they only need the insertion of a CAR and the replacement of their MHC code to HLA-E.

10/ The next major edits you will see for cell therapies are co-stimulation. This is a critical part of keeping these cells active in the patient so they can kill tumors longer. For a T cell, you will see things like CD-28 or 41BB. For the NK cells, you will see IL-15.

11/ Overcoming the immune system to prevent rejection and stimulating the cells to last longer and work harder are the basic 2 edits.

12/ The next big edit will be overcoming the tumor microenvironment (TME) of the patient. Most tumors exist because they evolve to avoid recognition by the immune system. The cell therapies need to be able to overcome these blockades.

13/ This can be done by administering PD-1 inhibitors along with the cell therapy, or they can just knock out the PD-1 receptor on the CAR-T cells to make them immune to PD-1 inhibition.

14/ Beyond these key edits, there are many others that can be disease specific. An example is knocking out CD38 receptors on cell therapies so they can work with drugs that target CD38 without harming the cell therapy. $FATE is using this approach for one of its therapies.

15/ When it comes to cell therapies, I am more in favor of NK cells as they have many functions that make them favorable as a CAR-NK then a CAR-T. The NK cell has stress ligands like NKG2D and KIR that allow it to detect stressed or damaged cells.

16/ This gives them the ability to detect cancer cells beyond just the CAR receptor. The NK cell also can be given and enhanced CD16 receptor so it works better with other antibodies for cancer treatments. This gives CAR-NK multiple shots on the tumor.

17/ The science here is still evolving, but iPSC derived CAR-NK cells with combination with already developed cancer antibodies hold big promise.