Title: Next generation CAR19 studies
Funder: Department of Health and Social Care

Abstract: Chimeric Antigen Receptors (CARs)1 are chimeric proteins generated by fusing the antigen-binding region of a monoclonal antibody (mAb) to intracellular T-cell signalling domains. They graft the specificity of a mAb onto the potent effector mechanisms of a T-cell. Large numbers of CAR T-cells can be generated by isolating and transducing a patient s peripheral blood T-cells with an integrating vector coding for the CAR. After infusion, these cells can home, extravasate and penetrate a solid core of tumour, proliferate8 and persist9. Recent clinical data described in neuroblastoma10 and more dramatically in CD19 CAR2&ndash;5 targeting for treatment of B-cell malignancies (CAR19) have shown unprecedented responses in patients with refractory disease. CAR therapy is already a disruptive force in the treatment of cancer and looks set to soon find routine application in salvage of refractory B-cell malignancy. Despite the promising clinical data, many challenges remain before CAR19 can be more widely adopted in the NHS, even for the narrow indication of refractory B-cell malignancy. Severe immediate and late toxicity have been evident which include cytokine storms, delayed macrophage activating syndromes, with CAR designs associated with better responses typically causing more toxicity11. Permanent depletion of the B-cell compartment can be poorly tolerated in some patients. Large-scale manufacturing of retroviral and lentiviral vectors remains largely an unsolved problem12. It is also unclear what the most cost-effective manufacturing model of bespoke CAR T-cell products for each patient will be. We have developed a CAR19 strategy which overcomes the above challenges with a view to wider deployment of CAR19 beyond experimental clinical studies. We have developed a CD19 CAR based on a different CD19 binder characterized at UCL. This CAR transmits a 41BB activation signal which appears to be the optimal format for B-cell malignancies. We believe this CAR is at least as potent as those tested in the US. With safety in mind, we have generated a sort-suicide gene termed RQR8. This is a small type-I membrane protein which contains two copies of the Rituximab binding epitopes from CD20 which allows selective destruction of CAR T-cells by administration of Rituximab. We also have addressed manufacturing problems: UCL has pioneered retroviral and lentiviral pseudotyping with the RD114 envelope. This envelope allows generation of stable, high-titer vectors particularly effective at transducing T-cells. Using RD114 pseudotyping, we have generated means of generating large amounts of virus and purifying it. In addition, we have developed a means of incorporating T-cell activation domains in the lentiviral capsid this allowing one reagent to both activate and transduce T-cells. These new technologies allowing us overcome the problem of large-scale vector manufacture and efficient T-cell transduction. Using this stimulating and transducing lentiviral vector, we have developed a simple 10-day process using disposable bioreactors to expand and transduce T-cells at scale sufficient for therapeutic application. The simplicity and relatively small amount of handling of this methodology should make it transferrable to most accredited haematopoietic stem cell processing labs present in all large centres in the UK, providing a cost-effective solution to CAR T-cell manufacture. We believe we can deliver a CAR19 programme which will be highly competitive both technically but particularly in terms of scalability and cost-effectiveness. Our immediate aim is to implement two large phase I studies in acute and chronic B-cell malignancies and show feasibility of our approach and safety of the therapeutic. We plan to demonstrate CAR therapy is feasible using typical NHS and UK infrastructure. We will determine cost-effectiveness of our programme. Our longer term vision is that we can provide a viable national strategy for CAR19 within the NHS, and a model for application of CAR therapy to other cancers.

Lay abstract: Despite much advancements many patients with leukaemia and lymphoma die of their disease. Some patients relapse and conventional treatments i.e. chemotherapy and radiotherapy no longer work; other patients never achieve remission despite intensive chemotherapy and radiotherapy. For some patients with certain types of leukaemias and lymphomas, we know that cure is impossible with conventional treatments. Even for those cured of their cancer, chemotherapy and radiotherapy is highly toxic, and can leave patients with long-term effects on their health. Treating cancer by engineering the patient s own immune system to attack it is a new radically different form of cancer treatement. We propose a strategy where this form of treatment can be brought to NHS patients. T-cells are a type of white-blood cell which form part of our immune system. The role of T-cells in our body is to detect and kill any cells which have become infected with a virus. T-cells are incredibly powerful machines, able to precisely hunt down and kill infected cells. They also can divide and survive in our bodies indefinitely, continuously patrolling for recurrence of the infection. Since cancer cells develop from normal cells in our body, T-cells rarely recognize them as something they need to attack. Researchers have long tried to coax these powerful cells to kill cancerous tissue. We believe that the best way of doing this is by genetic engineering. T-cells can be easily taken from a blood sample and grown in the laboratory. Using genetic engineering tools developed originally for gene-therapy, these T-cells can be &ldquo;re-programmed&rdquo; in the laboratory so that a patient s own T-cells recognize and kill cancer cells. This can be achieved by introducing a gene for an artificial protein called a chimeric antigen receptor, or CAR for short, into a patient s T-cells. After being modified in the laboratory, these T-cells can be returned back to the patient in a drip. The CAR directs them to now attack cancer cells with the same vigour and determination as they would naturally attack a virus infection. These CAR T-cells can divide and live in the patient, constantly hunting for any cancer cells preventing relapse. Because of the way they kill cancer cells and how long they can survive in a patient, we expect the long-term effects on health will be very different and favourable compared with chemotherapy and radiotherapy. Recent clinical studies in the US leave little doubt that CAR T-cell therapy will be game-changer in cancer therapy in many types of leukaemia and lymphoma. In UCLH we have a programme of CAR T-cell studies. We have developed a powerful but safe CAR gene-therapy vector against leukaemia and lymphoma. We would like to develop and test this in a number of clinical studies in lymphoma and leukaemia. Substantive funding from i4i would allow us to run an efficient programme quickly giving access to NHS patients from our referral base access to CAR therapy. The long-term goal of this project is to provide a timely and cost-effective strategy for CAR therapy across the NHS.

Research activity codes: 6.2 CELLULAR AND GENE THERAPIES
Health categories: CANCER AND NEOPLASMS