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(1) Background: Refractory acute graft-versus-host disease (R-aGvHD) remains a leading cause of death after allogeneic stem cell transplantation. Survival rates of 15% after four years are currently achieved; deaths are only in part due to aGvHD itself, but mostly due to adverse effects of R-aGvHD treatment with immunosuppressive agents as these predispose patients to opportunistic infections and loss of graft-versus-leukemia surveillance resulting in relapse. Mesenchymal stromal cells (MSC) from different tissues and those generated by various protocols have been proposed as a remedy for R-aGvHD but the enthusiasm raised by initial reports has not been ubiquitously reproduced. (2) Methods: We previously reported on a unique MSC product, which was generated from pooled bone marrow mononuclear cells of multiple third-party donors. The products showed dose-to-dose equipotency and greater immunosuppressive capacity than individually expanded MSCs from the same donors. This product, MSC-FFM, has entered clinical routine in Germany where it is licensed with a national hospital exemption authorization. We previously reported satisfying initial clinical outcomes, which we are now updating. The data were collected in our post-approval pharmacovigilance program, i.e., this is not a clinical study and the data is high-level and non-monitored. (3) Results: Follow-up for 92 recipients of MSC-FFM was reported, 88 with GvHD ≥°III, one-third only steroid-refractory and two-thirds therapy resistant (refractory to steroids plus ≥2 additional lines of treatment). A median of three doses of MSC-FFM was administered without apparent toxicity. Overall response rates were 82% and 81% at the first and last evaluation, respectively. At six months, the estimated overall survival was 64%, while the cumulative incidence of death from underlying disease was 3%. (4) Conclusions: MSC-FFM promises to be a safe and efficient treatment for severe R-aGvHD.
The current treatment strategies for diseases are assessed on the basis of diagnosed phenotypic changes due to an accumulation of asymptomatic events in physiological processes. Since a diagnosis can only be established at advanced stages of the disease, mainly due to insufficient early detection possibilities of physiological disorders, doctors are forced to treat diseases rather than prevent them. Therefore, it is desirable to link future therapeutic interventions to the early detection of physiological changes. So-called sensor-effector systems are designed to recognise disease-specific biomarkers and coordinate the production and delivery of therapeutic factors in an autonomous and automated manner. Such approaches and their development are being researched and promoted by the discipline of synthetic biology, among others.
Against this background, this paper focuses on the in vitro design of cytokine-neutralizing sensor-effector cells designed for the potential treatment of recurrent autoimmune diseases, especially rheumatoid arthritis.
The precise control of inducible gene expression was successfully generated in human cells. At first, a NF-κB-dependent promoter was developed, based on HIV-1 derived DNA-binding motives. The activation of this triggerable promoter was investigated using several inducers including the physiologically important NF-κB inducers tumor necrosis factor alpha (TNFα) and interleukin 1 beta (IL-1β). The activation strength of the NF-κB-triggered promoter was doubled by integrating a non-coding RNA. The latter combined expressed RNA structures, which mimic DNA by double stranded RNAs and have been demonstrated to bind to p50 or p65 by previous publications. The sensitivity was investigated for TNFα and IL-1β. The detection limit and the EC50 values were in in the lower picomolar range. Besides the sensitivity, the reversibility and dynamic of the inducible system were characterized. Hereby a close correlation between pulse times and expression profile was shown.
The optimized NF-κB-dependent promoter was then coupled to established TNFα- and IL-1-blocking biologicals to develop sensor-effector systems with anti-inflammatory activity, and thus potential use against autoimmune diseases such as rheumatoid arthritis. The biologicals were differentiated between ligand-blocking and receptor-blocking biologicals and different variants were selected: Adalimumab, etanercept and anakinra. The non-coding RNA improved again the activation strength of NF-κB-dependent expressed biologicals, indicating its universal benefit. Furthermore, it was shown that the TNFα-induced expression of NF-κB-regulated TNFα-blocking biologics led to an extracellular negative feedback loop. Interestingly, the integration of the non-coding RNA and this negative feedback loop has increased the dynamics and reversibility of the NF-κB-regulated gene expression. The controllability of drug release can also be extended by the use of inhibitors of classical NF-κB signalling such as TPCA-1. The efficacy of the expressed biologicals was detected through neutralization of the cytokines using different experiments. For future in vivo trials, first alginate encapsulations of the cells were performed. Furthermore, the activation of NF-κB-dependent promoter was demonstrated using co-cultures with human plasma samples or using synovial liquids.
With this generated sensor-effector system we have developed self-adjusting cytokine neutralizer cells as a closed-loop delivery system for anit-inflammatory biologics.