Benefits

Complexity of classical workflow for gene-engineered T-cell production. Multiple manual steps are associated with the risk of high failure rates e.g. from handling errors, and result in high manufacturing costs.

Clinical manufacture of gene-modified T-cells currently is a complex process. It generally starts with obtaining the patient’s peripheral blood mononuclear cells (PBMC) by an initial leukapheresis step. The resulting PBMC are often enriched for T-cells and activated prior to gene-modification with viral or non-viral vectors. The modified T-cells are then expanded in order to reach the cell numbers required for treatment, after which the cells are rebuffered and/or cryopreserved prior to reinfusion. The cell product must be subjected to a number of quality control testing and has to meet all assay release criteria.

Each manipulation or addition of reagents to the cell preparation (e.g. washes, stimulation, transduction, feeding, sampling) creates a risk for error and for contamination that can lead to a failed production run. Moreover, these complex processes involve many different reagents, e.g. separation reagents, activation reagents, viral vectors, media, cytokines, different buffers etc. These reagents need to efficiently and stably work together as an integrated reagent system. Ideally, the process should become sufficiently robust enough to yield equivalent product quality independent of the patient and possibly the CAR used to modify the T-cells, assuming the transgene does not drastically impact the physiology of the expanding T-cell population. 

Because of these high technical demands, adoptive cellular therapy (ACT) using gene-modified T-cells has mainly been carried out by investigators who have developed their own manufacturing process for small-scale clinical trials while using the devices and infrastructure at hand. Hence standardized protocols are missing and anyone who has embarked on the task of manufacturing patient-specific ATMP for clinical use will admittedly agree that it is quite an undertaking: the cell-manufacturing process is labour intensive, as it comprises many (open) handling steps (e.g. density gradient cell processing, gene-modification, washing, feeding etc.) that run the risk of introducing operator error in the process, thereby reducing reproducibility of results. Successful cell manufacturing currently requires interventions from committed skilled operators who have undergone extensive training. Even then, the failure rate remains relatively high due to high skill and time demands on clean room personnel to make these complex products.

The high technological requirements of present manufacturing processes restrict its clinical use to a limited number of institutions worldwide. This in turn confines the number of runs and therefore the number of patients that can be served at any given time. As a result of the limited scalability and the complexity of the process, lack of automation and missing standardization the costs for manufacture of CAR T-cell products still remain high. Hence, the resulting commercial models are not competitive which presently impedes investment and consequently a broad dissemination of these promising therapies to the patients that need them. CARAT aims to reduce complexity of the process while increasing the levels of automation and standardization, thereby significantly reducing costs of manufacture.

In contrast to standard pharmaceutical drugs or immune-modulating agents, CAR T-cells, once infused in the patient, can assume “an independent life”. They will expand, secrete pro-inflammatory cytokines and mediate cytotoxicity upon target recognition. They will also widely home to tissues and possibly persist. Consequently, any associated toxicity can become uncontrolled and lead for instance to severe cytokine release syndrome with various levels of associated risks for the patient.

The severe cytokine release syndrome (CRS) is indeed a negative side effect of CAR T-cell therapy: CRS is associated with an uncontrolled response of the infused cellular product. Although the CRS can often successfully be controlled with Tocilizumab (anti-IL-6 antibodies) it would be preferable to directly regulate the level of response mediated by the CAR T-cells. In order to reach this, better methods to control the level and duration of the CAR T-cells response can and should be engineered. CARAT will tackle these issues through the development of innovative controllable CARs.

One of the major challenges for the translation of personalized cellular products into standard therapy is the fact that the production process is patient specific. Regulatory agencies are very familiar with drug manufacturing, but genetically engineered cellular products have special requirements. Although regulatory authorities are working hard to define optimal guidelines that can be harmonized and applied uniformly, the requirements for clinical manufacture of ATMP (advanced therapy medicinal products) still represent an enormous hurdle for translation into clinical routine. In Europe, the handling of genetically engineered ATMP is summarized in regulation 1394/2007 (§2)^[1]; in the USA, the regulations in Guidance for Human Somatic Cell Therapy and Gene Therapy must be followed^[2].

[1] REGULATION (EC) No 1394/2007 on advanced therapy medicinal products.

[2] GUIDANCE FOR INDUSTRY: FDA GUIDANCE FOR HUMAN SOMATIC CELL THERAPY AND GENE THERAPY. 1998.

CARAT relies on the CliniMACS® Cell Separation principle that has become an established technology since its introduction in 1997 for the development of GMP-compliant manufacturing processes for cellular products. In 2014 the CliniMACS System obtained approval by the U.S. Food and Drug Administration (FDA) for a specific treatment of Acute Myeloid Leukaemia (AML). To date, the CliniMACS System is the only technology worldwide for clinical enrichment of cellular products within a closed system.

The CliniMACS Prodigy® simplifies and standardizes conventional processes. In principle, the automated device is capable of performing all necessary steps from cell preparation, enrichment, washing, activation, transduction, expansion to final formulation and sampling in a closed sterile, single-use tubing set. Read more in Tools and Technologies.

The CliniMACS Prodigy will be adopted as a generic platform for integrated manufacturing of genetically engineered CAR T-cells with minimal user interaction. This fully automated approach will greatly simplify and improve robustness and reproducibility of the manufacturing procedure. Integration of enabling technologies from CARAT WP 1- 4 will finally result in the novel CARAT process illustrated in the figure below.

Essential steps of the CARAT process to manufacture CAR T-cells. A starting source of autologous cells is collected from the patient. Typically, the leukapheresis bag is connected to a closed sterile tubing set installed onto the CliniMACS Prodigy. T-cells or T-cell subsets are automatically selected using CliniMACS reagents. Enriched T-cells are then activated using the TransAct reagent in TexMACS medium in presence of supportive cytokines such as IL-2 or IL-7 and IL-15. The activated T-cells are gene-modified using a lentiviral vector and further expanded. After 10-12 days, the gene modified T-cells are washed and finally formulated for direct infusion into the patient or for cryopreservation. In process controls (IPC) and quality control (QC) are performed via use of the MACSQuant analyser.

The CliniMACS Prodigy is designed for saving time and costs: Integrated processes were refined to provide results in a shorter period of time and automation reduces manual handling steps. Thereby, personnel costs will be reduced and even expenses for GMP laboratories. As the CliniMACS Prodigy enables cell processing in a closed system, the clean room requirements will be reduced in comparison to processes with open handling steps, which will also positively impact manufacturing costs significantly. Moreover, automation with the CliniMACS Prodigy will allow for scale-up of manufacturing processes for commercial use, e.g. when hundreds to thousands of cell therapeutic doses per year are required to perform phase II/III clinical trials with the goal of FDA approval. In principle, the CliniMACS Prodigy is designed for device-based manufacturing, meaning one device is dedicated to the production of one patient product at a time (Kaiser et al.; 2015). Scale up could be achieved in centralized GMP units where multiple devices are operated in parallel. Such a unit-based production would preferentially be in organized areas where an operator could oversee several units at the same time. Partner 6, TrakCel, will develop validated tracking systems which are required to ensure logistic control of the material involved in a given manufacturing unit and during QC sampling.

Finally, instead of the centralized approach outlined above, CARAT aims at a future where gene-modified T-cells are manufactured cost-efficiently and localized at the point-of-care in a facility at the hospital. Such a decentralized mode for delivering cell therapy products to patients will certainly decrease the risks and costs, particularly the costs for transportation, personnel and GMP-manufacturing facilities.

Despite the clinical successes in treatment of hematologic malignancies, advanced strategies are needed to optimize CAR T-cell efficacy and to direct them e.g. towards poor-prognosis of solid tumours.

CARAT thus aims at enhancing clinical efficacy of input cells and establishing new generations of CAR molecules with improved efficacy and safety. Furthermore CARAT will develop new gene-delivery technologies leading to homogeneous, safer and more efficient cell products.

Finally, CARAT will develop new pre-clinical tools for monitoring CAR T-cell efficacy and for improving their therapeutic value.

Manufacture of ATMPs is not only technically challenging but also compliance with current regulations is regarded by many scientists as a major hurdle for translation of novel therapeutic concepts into clinical routine.

As CARAT will deliver gene-modified ATMPs, compliance with current regulations will be a crucial aspect affecting all steps of the cell manufacture workflow (see figure). Consequently, we designed a work package that focuses on regulatory and ethical issues (WP6) and will ensure that the CARART process fulfils GMP and other regulatory requirements. Read more about Work Packages.

Central role of regulations for ATMP manufacturing. Consequently, the entire CARAT work flow will be designed for regulatory compliance. Moreover, CARAT aims to reduce regulatory barriers also for other researchers e.g. by creating a ‘blue print’ IMPD.