Date: 8th September 2021
Rheumatoid arthritis (RA) is a chronic disease affecting as much as 1% of the worldwide population, and with no known cure, modifying antirheumatic biologic drugs have revolutionised the treatment of this autoimmune disease. However, ~40% of patients fail to respond to treatment, and drugs that suppress the immune system can have substantial adverse effects. Now, researchers have designed genetically engineered stem cells to create a synthetic gene circuit that senses changing levels of endogenous inflammatory cytokines to trigger a proportional therapeutic response of biologic drug release. The engineered cells reduced inflammation and prevented bone erosion caused by rheumatoid arthritis.
Antirheumatic biologic drugs aim to suppress the immune system and are usually designed to target several inflammatory cytokines and pathways, such as interleukin-1 (IL-1), IL-6, and tumor necrosis factor–α (TNF-α). However, whilst the severity of RA fluctuates over time, the drugs do not, and are administered continuously at high concentrations, often predisposing patients to substantial adverse effects (AEs), such as increased risk of infection. Specific therapeutic strategies developed to sense and respond to endogenous levels of inflammatory mediators could meet these unmet clinical needs, and overcome challenges such as AEs, but are so far lacking.
Now, researchers at Washington University School of Medicine in St. Louis, US, led by Farshid Guilak have engineered cells into cartilaginous constructs that exhibited rapid activation and recovery in response to inflammation. These cells were engineered into bioartificial implants and significantly mitigated RA disease severity, preventing increased pain sensitivity and bone erosion.
To start, the team used CRISPR-Cas9 genome engineering to create a self-regulating synthetic gene circuit in induced pluripotent stem cells (iPSCs). They inserted the gene for IL-1Ra (Il1rn) (or luciferase (Luc) as a reporter control) at the Ccl2 locus, creating a self-regulating gene circuit that transcribed Il1rn in response to inflammatory activation of Ccl2 – dubbed as SMART cartilage cells (Stem cells Modified for Autonomous Regenerative Therapy). Cell-based injection of the genome-engineered Luc-iPSCs were able to sense and respond to inflammation in vivo in a model for murine model of arthritis, showing increased luminescence intensity concordant with increased clinical score in older mice, as disease progression occurred.
However, the team wanted to create a stable platform for the SMART cells, so they used a previously designed cell-based implant. They seeded the cells onto 3D woven scaffolds and chondrogenically differentiated them over 21 days into chondrocyte-like cells that produced a proteoglycan-rich matrix. This ensured minimal migration of the cells out of the target area, and the platform did not required vasculature or innervation for long term survival.
The tissue-engineered bioartificial implants containing SMART cells were then implanted into arthritic mice. Reporter gene activation showed the implant constructs were responsiveness in vivo, and cell survival in a chronic inflammatory environment was seen up to 5 weeks after implantation. Next, the team assessed the activation and reactivation dynamics of this genetically engineered drug delivery platform. Here, they saw that induced flares up of RA were concomitant with clinical scores and increased reporter luminescence, which waned as flare up subsided. Furthermore, these constructs were able to be reactivated after a week-long washout period. Overall suggesting that the in vivo on-off kinetics were proportional to endogenous inflammatory signals.
However, could the implants mitigate arthritis symptoms? Here, the team showed that animals receiving SMART cell implants demonstrated significant amelioration of arthritis severity, including 40% reductions in both clinical scores and ankle thickness, compared to control animals. They also had lower inflammation and had reduced cartilage degradation and proteoglycan loss than the control cohort, this was accompanied by a significant decrease in mechanical pain sensitivity.
Micro–computed tomography of the bone structure also showed that mice receiving the SMART implants were completely protected from bone erosions that are characteristic of inflammatory arthritis, and had a significant reduction in their inflammatory cytokine profiles as compared to control animals.
Conclusions and future applications
The team here have combined the principles of synthetic biology and tissue engineering to develop a genome-engineered implantable drug delivery system that could automatically sense and respond to inflammatory cytokines to produce therapeutic levels of anticytokine biologic drugs in a self-regulating manner. The implants exhibited extended viability and therapeutic function in vivo and could be repeatedly reactivated by inflammatory challenges. The anticyotkine therapy alleviated pain sensitivity and bone erosion, a feat not achievable by current clinically available disease-modifying drugs.
Cell-based therapies are rapidly evolving, and are a new frontier in medicine – providing new precisely engineered platforms for highly controlled, long term drug delivery. Other genetically controlled smart cells that are stimuli-responsive and function under physiological conditions are also being investigated. However, the work presented here represents the next development in the journey by creating a niche for them to reside. By combining genome engineering of therapeutic cells with tissue engineering, creating sense and respond SMART cells within a bioimplant, the team have created a stable implantable drug delivery system that could likely overcome the burden of repeated administered injections and reduce the risk of AEs. Although, its long term use is yet to be assessed for safety and efficacy, this will be addressed in the near future. Interestingly, similar implantable niches are being explored to monitor autoimmune disease and effectiveness of treatments as they reflect aspects of the immune status.
The team are now interrogating other biomaterials and synthetic gene circuits for further tuning of the cellular responses in other chronic inflammatory disease models. Advances in biofabrication are currently revealing a range of new biomaterials for example biomimetic hydrogels, nanoengineered bioinks and self-healing gels which are now able to support tissue construction and may aide the development of the implants. The team eventually hope to engineer the platform to manufacture more than one drug in respond to different inflammation triggers, increasing its potential applications.
For more information please see the press release at Washington University School of Medicine in St. Louis
Choi et al. (2021) A genome-engineered bioartificial implant for autoregulated anticytokine drug delivery. Science Advances 7, 36
https://doi.org/10.1126/sciadv.abj1414
