Cellular immunotherapy for B cell cancers shows promise
Dr Bahram 'Bob' Valamehr - Fate Therapeutics Inc., San Diego, USA
As we heard, CAR T therapy has changed the way we think about cancer treatment, however, the process is complex. As you can see here, you start with an impaired starting material, there are complex logistics, time to delivery to the patient is lengthy, the cost is high and the limited availability creates a challenge for multi-dosing. At the same time, when you engineer the product not every cell is engineered and not every engineered cell is pristine.
At Fate Therapeutics we have been working on using IPSCs as a starting material. These are pluripotent cells that are maintained in a petri dish, have the unlimited self-renewal ability and can at any time become any of the 200 cell types in the body. We’ve taken advantage of this product, using it at the single cell level to do precise engineering to create master cell banks to create uniform composition of cell material and have extensive characterisation to be able to create clonal cell lines to make homogeneous cell products.
This is done by the scheme you see here. We reprogram, engineer and we drop single cells into a 96-well plate, allow that population to expand, it becomes a continuous culture. The culture maintains its pluripotency, genomic stability and now you have the ability to do very precise engineering. As you can see in the middle, you can drop several genes then into a targeted locus and end up with a pure population of cells carrying those genes.
In another strategy you can knock out various genes, here we targeted beta-2 microglobulin in a biallelic manner at the single cell level, got rid of the expression of B2M and hence HLA class 1 expression. So now we have a pure product that is knocked out.
We take that engineered product and we induce differentiation into haematopoiesis. Here we go from an IPSC to a CD34 positive haematopoietic progenitor cell and continue that differentiation into NK cells and T-cells. Here we illustrate in NK cells. You can see that nicely the population becomes homogeneous for CD56 positive which is a pan-marker for NK cells. Other activating receptors on NK cells also come up, as you can see here NKp30. The process is highly scalable, one million IPSCs gives you 1E12 NK cells and that output is continuous. You can every single time start this process and deliver 1E12 NK cells. At the global gene expression level these NK cells look very similar to their primary counterpart and this process has been transferred to now a manufacturing process where it’s done in multiple sites – our collaborators at the University of Minnesota and internally at Fate.
The product is released based on the fact that it’s pure for haematopoietic cells and NK cells, has high viability that now allows you to have a homogeneous high quality product that’s low cost, each dose is $2,500, is thawed [?] and directly infused, there is no processing needed. So it becomes a true administered off the shelf product in an out-patient setting analogous to pharmaceutical drug product development.
This process, as I mentioned, starts with an IPSC population, is engineered, single cells are selected and a master cell bank is fully characterised and allows for production of T- and NK cells in an off the shelf manner. Using this process we have now started three clinical investigations. First is FT500 which is the first IPSC derived product cleared for clinical investigation in the US. Twelve patients have been dosed, a combined 60 doses have been given to those twelve patients and so far the product has been safe and well-tolerated.
FT596 is the first engineered product, it’s the first engineered IPSC product in clinical investigation in the world to the best of our belief. The first patient with the lowest dose has showed anti-leukaemia effect and also persistence in the bone marrow. FT596 is a multi-factor engineered product which I will talk about.
FT596 consists of three anti-tumour modalities. It’s the first cancer immunotherapy product to be engineered with three active anti-tumour modalities. It consists of hnCD16, a high affinity non-cleavable CD16 which binds to antibodies and elicits antibody dependent cellular cytotoxicity. This is a key action of general killing by NK cells and we’ve enhanced that feature. So now we can combine FT596 with any monoclonal antibody to elicit enhanced ADCC. We’ve also introduced a CAR that has been calibrated to NK biology targeting the CD19 antigen and finally we’ve added IL-15 receptor fusion to support persistence and reduce the dependency for exogenous cytokine support.
Because the clone was selected to have one copy of each product you can see on the far right hand side, sorry the top should say CD16, the orange should say CAR 19 and the bottom should say IL-15 receptor fusion, you can see that each are expressed homogeneously and this is very challenging if you want to engineer NK cells or even T-cells to have this profile. So compared to primary NK cells you can see such uniform differential.
To show what step manner these engineering attributes do we created a co-culture assay which consists of cancer cells that are CD19 positive, CD20 positive. So these are antigens on B-cells that are usually targeted, as we heard earlier. We also co-cultured with a CD19 negative, CD20 positive cancer cell line. When we introduced a CAR-19 you can see you can effectively eliminate the CD19 positive target cells and when you add the IL-15 it is completely eliminated because IL-15 enhanced persistence and durability of the product.
With utilising the hnCD16 we add rituximab, the anti-CD20 antibody, and cleared the second half of the product that you could argue mimics antigen escape. We used CD20 here, we could have used CD22, CD123 or other monoclonal antibodies that target B-cell malignancies.
This product in vivo is highly efficacious as a monotherapy targeting leukaemia. It has the ability to control tumour in an extensive period. In combination, going after lymphoma, you can see where rituximab fails to control the tumour burden in combination with FT596 it does a great job maintaining durability and control of the tumour. Finally in the allogeneic system you can see that compared to CAR T, FT596 as a monotherapy is comparable if not better.
So my final data slide I will recap what this combinational strategy is. Here we used Raji lymphoma cell line, that’s CD19 positive, CD20 positive, and compared FT596 to five different primary CAR T products. As you can see here in this low effector target ratio, so an in vitro stress test where only 0.321 ratio of the effector cells compared to the target, you can see that FT596 is comparable to primary CAR T. However, when you add Rituxan you can see that you get a more deeper, durable response.
In a different format, trying to mimic antigen escape, we knocked out CD19 from the Raji cell line and created CD19 negative, CD20 positive cells and then introduced a high capacity test to look at effector function. As you can see here, primary CAR T and FT596 alone failed to control the tumour elimination over a long term period, however in combination with Rituxan FT596 was able to eliminate the tumour load in the in vitro culture.
So this is what’s being presented at ASH for Fate Therapeutics specific to CAR programmes. As I mentioned, FT596 is publication number 301 presented later today. We also have a CAR BCMA in combination with Darzalex, a product where we’ve knocked out CD38 to eliminate fracture sites. So this is a true NK product that can sustain ADCC as well as CAR BCMA activity. This is publication number 3214, IND to be followed the second half of next year. And excitingly we are also working on a first off the shelf CAR T product where we’re utilising some of the top engineering attributes that are coming out of Sloan Kettering and out of Dr Michel Sadelain’s lab to create a first off the shelf CAR T product and that will be filed in a few months and that’s publication number 4434. Thank you.