Application of combined gene and cell therapy within an implantable therapeutic device for the treatment of severe hemophilia A

Periodic Reporting for period 2 - HemAcure (Application of combined gene and cell therapy within an implantable therapeutic device for the treatment of severe hemophilia A)

Okres sprawozdawczy: 2017-05-01 do 2018-10-31

The long-term goal of HemAcure was to develop and refine tools and technologies for a novel ex vivo prepared cell-based therapy to treat the bleeding disease, haemophilia A (a genetic deficiency in clotting factor VIII (FVIII)) that ultimately lead to improved life quality of the patients.
Within the 3-year project 2 main objectives were pursued: a) providing preclinical data demonstrating proof of concept for the indication haemophilia A, and b) development of a cell therapy product manufacturing process according to Good Manufacturing Practice (GMP)-compliant regulatory requirements.
The concept comprises using the patient’s own, autologous blood outgrowth endothelial cells (BOECs). These cells are genetically modified to introduce functional copies of the FVIII gene, allowing the cells to produce and secrete functional clotting factor VIII protein into the blood stream. To ensure the highest safety standards, the procedure is performed ex-vivo and guided by continuous quality testing throughout the complete process. After cell expansion, they are transplanted into a proprietary, GMP manufactured pre-vascularised medical device (Cell Pouch™), implanted under the skin.
All planned tasks of the program were successfully completed. UKW set up a standardized operating procedure (SOP) for the isolation steps and expansion of BOECs. UPO achieved high efficiency in transduction and genetic repair of the BOEC genome. UNILO analysed the cells after the performed large scale expansion and could demonstrate their safety. Based on the SOP in cell isolation, transduction and cell expansion it could be shown that a therapeutic dose of FVIII can be achieved consistently at a cell production cost that is feasible from a clinical perspective. SERC produced cGMP Cell Pouches™ for implantation into the haemophilia model. The transduced and expanded FVIII BOECs were successfully transplanted into the Cell Pouch™ allowing the cells to engraft into a supportive natural tissue environment and showed survival within the device. IMS/GRU, responsible for GMP-compliance of the production process for the BOECs completed the necessary risk assessments based on the product requirements of the Advanced Therapy Medicinal Product (ATMP). Quality gate protocols between the Work Packages were installed to achieve reliable quality of the deliveries and checklists for regulatory requirements were created building the foundation for ATMP development under GMP-compliance.

We developed tools and technologies to enable the establishment of a combined ATMP, composed of therapeutic cells with the implantable medical device. Our first-time right concept based on the quality-by-design approach enabled us to identify sources of variability affecting our processes, setup measurements to control them enabling us to generate a streamlined, safe and cost-effective GMP conform process that will be translatable into the clinical setting.
Several manufacturing protocols were generated by UKW to establish a GMP compliant method for cell isolation and cultivation of BOECs using animal component free coated vessels and defined medium for cell growth and expansion. The production and purification of recombinant lentiviruses carrying the genetic code for the human FVIII gene was established. Lentiviruses are capable of effectively introducing functional copies of FVIII into the BOECs enabling them to produce functional clotting factor VIII. These steps were implemented into GMP conditions by UKW and IMS/GRU.
During and after expansion of the corrected BOECs at UNILO,the cells were analysed for their safety profile, functionality and senescence state to evaluate and define release criteria for the therapeutic cell product for future human clinical studies. As a first in vivo preliminary test, UPO conducted an initial test on BOECs in the haemophilia A mouse model showing that corrected BOECs injected interperitoneally released therapeutic levels of FVIII into the blood stream. After conducting surgical implantation, dosing and additional supporting proof of concept studies, SERC demonstrated that the cells met the defined release criteria, transplanted the corrected BOECs into the Cell Pouch™, demonstrating survival of cells for the three-month period. Cell PouchTM transplanted cells were analysed at various time points for cell survival, safety and clotting factor VIII production. For corrected BOECs, it was indicated that the cells were able to produce FVIII within the Cell Pouch™. However dosing optimization with larger sample size is required, but importantly, FVIII released from BOECs within the Cell PouchTM could be detected in blood and an improvement in clotting was shown possible.The implantation and transplantation studies of the scalable, pre vascularized, removable Cell Pouch™ will be used in developing an Instructions For Use document suitable for use in future human clinical studies.
Different sets of design and manufacturing protocols were generated, based on current European GMP regulations leading to the preparation of a common technical document (CTD) for an ATMP, composed of therapeutic cells and an implantable medical device, a combined Advanced or Gene Therapeutic Medicinal Product. With this document the first step is achieved to develop an ATMP medical product, which can be evaluated in a clinical phase I/II study.
Within the project duration, the team was present on various international Haemophilia conferences and attended Venture Capitalist events. Independent of the occasion the concept of HemAcure met with great interest. Currently we are evaluating the best way presenting the results to the scientific community.

Within this project we linked several technologies from different sectors and streamlined them in GMP-compliant process right from the start of the design and process development. We applied systematic quality engineering approaches, as they permitted us to identify the sources of and the control of variation, an issue that is at the core of current GMP.
This enabled us
• to have the finished HemAcure product consistently meet its predefined characteristics from the start
• to facilitate an immediate and easy transfer regarding patient safety from pre-clinical to clinical steps
• to avoid significant increase of production cost due to unplanned corrective action during scale-up
• to provide a solid basis for continuous improvement of the product and the process during its lifecycle
The economic impact can be measured by new jobs created throughout the project and scientists could train and develop their skills. The challenging application of systematic quality engineering approach for designing an advanced medicinal therapeutic product process, allowed all contributors to collect an enormous knowhow resulting in highly-skilled workers with ideal career opportunities.
The structure of HemAcure represents a good example how complex and technically advanced biomedical challenges can be handled by combining efforts from multiple perspectives. The project provides a solid basis for a future medical device, which can ease the burdens of patients affected by severe haemophilia A. Regular infusions with recombinant FVIII would no longer be necessary and the life quality of patients would immediately increase. Also, the European Healthcare system would profit of such a treatment, especially as the developed ATMP could apply to a variety of different applications.

Graphical overview of the interplay of all HemAcure work packages and the future vision of HemAcure

Periodic Reporting for period 2 - HemAcure (Application of combined gene and cell therapy within an implantable therapeutic device for the treatment of severe hemophilia A)

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