MAnufacturing of TUmour-REactive Natural Killer cells

Project Information

MATURE-NK

Grant agreement ID: 765104

DOI

10.3030/765104

Project closed

EC signature date 18 May 2018

Start date 1 June 2018

End date 30 November 2022

Funded under

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Total cost

€ 3 347 488,66

EU contribution

€ 3 347 488,66

Coordinated by

MEDIZINISCHE HOCHSCHULE HANNOVER
Germany

Periodic Reporting for period 2 - MATURE-NK (MAnufacturing of TUmour-REactive Natural Killer cells)

Reporting period: 2020-06-01 to 2022-11-30

Summary of the context and overall objectives of the project

Standard therapies of leukemia include remission chemotherapy and hematopoietic stem cell transplantation. More recently, immunotherapies with checkpoint inhibitors and chimeric antigen receptor (CAR)-modified cytotoxic T lymphocytes have evoked striking successes. Despite these important improvements, significant fractions of leukemia patients are either unresponsive or will relapse from these treatments. Therapy options for most solid cancer types are even less promising.
Natural killer (NK) cell-based therapies are emerging as novel attractive therapeutic approaches to improve the efficacy of cancer treatment and to overcome resistance to current therapies of leukemia and solid cancers. NK cells are a type of lymphocytes and form an important part of the innate immune system with the ability to kill harmful cells. Whereas T lymphocytes can have toxic side effects, NK cell usually are well tolerated.
The main objective of the MATURE-NK project was to further develop and improve technology and reagents for NK cell-based immunotherapies. This included improving selection of tumor-reactive NK cell subtypes and developing technology for enhancing their anti-tumor cytotoxicity as well as manufacturing of therapeutic NK cells for clinical NK cell infusions.
The project was designed to combine expert laboratories in basic, clinical and company-related research and development in the NK cell field. Thirteen early stage researchers performed the research work as part of their PhD theses and were trained to become experts for the growing area of cellular immunotherapies.
In conclusion, the project has achieved significant progress in NK subtype selection, modification and manufacturing of therapeutic NK cells that will impact on the further development of NK cell-mediated cancer therapies. This includes improvements in selection of NK cell donors, tumor-reactive NK subsets and amplification of NK reactivity by CAR-modification, checkpoint inhibitors and NK cell engagers. Exemplary therapies were successfully tested in animal cancer models, GMP-compliant manufacturing of CAR-modified NK cells was improved and an exemplary protocol for a clinical leukemia trial drafted. The results have been described in numerous scientific manuscripts and in a priority application filed in 2022 by an industrial partner.

Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far

A major focus was in the first part to improve selection, modulation and modification of therapeutic NK cells. The following major results were achieved:
1) Improved phenotypic and genotypic selection of NK cell donors were developed including refined KIR/MHC combinations for donor NK cell selection by a new metric and new antibody reagents.
2) Improved tumor-reactive NK subsets were analyzed supporting the efficacy of adaptive NK cells and of NK cells generated from induced pluripotent stem cells (iPSC) for an off-the-shelf strategy.
3) Significant progress in amplification of NK cell functions was achieved:
i) Promising CAR modifications were employed in head and neck squamous cell cancer. A series of second generation CARs with improved functionality against leukemia cells was tested resulting in a priority application.
ii) A "checkpoint" anti-NKG2A antibody was evaluated that potentiated NK cell-mediated anti-leukemia activity. Novel NK cell engagers co-engaging activating receptors efficiently boosted NK cells from transplanted leukemia patients. These data were basis of a press release.
iii) Genetic knock-outs of HIF-1α or SMAD4 have been tested to increase the resistance of NK cells to hypoxia and the immunosuppressive activity of TGF-β. Increased persistence and activity could be achieved.
4) Clinical NK cell-based trials have been analyzed and support a focus on preventing inhibitory interactions, improving persistence and off-the-shelf donor strategies.

In the second part of the project procedures were evaluated in preclinical models, manufacturing processes were improved and a clinical protocol for an NK cell-infusion trial was drafted:
1) Two novel human immune system (HIS) mouse models were developed, one with expression of human Fc receptors and another with autologous human leukemia. These are important for preclinical evaluation of therapeutic approaches. The adaptive NK subset was shown to possess strong anti-tumor functionality in a human acute myeloid leukemia (AML) mouse model. A novel tetravalent NK cell engager combining an activating receptor and a tumor antigen was successfully tested in a mouse model of human hepatocellular cancer.
2) GMP-compliant manufacturing processes incorporating CAR-modifications were improved. For peripheral blood NK cells a fully automated process was designed. As proof-of-concept, CAR-NK cells generated were tested in a preclinical model for human AML with positive results. A process for cord blood stem cell-derived NK is also available.
Finally, an exemplary clinical trial protocol for a CAR-NK cell therapy for AML was drafted.

The results of this project have so far been described in 16 publications with ESRs as first or co-authors, 13 additional publications are in different stages of submission or final preparation (all with acknowledgement of MATURE-NK). Further 12 publications with principal investigators as authors have been published. One priority application has been filed by an industrial partner. The results were disseminated to scientists, clinicians and companies at a "MATURE-NK" Satellite Meeting to the International NK2022 Conference and to a wider public via the website and an article in "EU Research" targeting a large relevant audience in government, the private sector and academia.

Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)

This project has achieved significant progress beyond the current state-of-the-art. Three developments should be especially important. First, adaptive NK cells based on their reduced checkpoint inhibition and iPSC-derived NK cells, because of the potential for generating off-the-shelf products, are promising candidates for NK cell infusion trials. Second, antibody-based reagents will be useful for NK cell checkpoint blockade and as NK cell engagers triggering activating receptors as shown in vitro with patient leukemia cells and in vivo in animal cancer models. Third, novel second generation CAR constructs in combination with fully automated NK cell manufacturing will lead to NK products with fortified anti-cancer reactivity. Combination therapies with these reagents and cells should be evaluated in clinical trials.
Aside the scientific impact, the project will have a major impact on future clinical trials of NK cell-mediated immunotherapies. We foresee improvements in regard of therapies of leukemia and possibly also solid cancers with corresponding impact on quality of life and life expectancy for relapsed leukemia and in the longer run also for other untreatable solid malignancies. Successful therapies will support the further growth of our industrial partners based on their proprietary reagents, cell therapy equipment and packages. Finally, the project provides an example for and has trained experts needed for the expected rapid increase in future cellular therapies.

Figure 1