Targeted co-delivery of methotrexate and rela sirna via folate liposomes for rheumatoid arthritis therapy

Abstract

Rheumatoid arthritis (RA) is a global healthcare challenge as it affects millions of people causing long-term disability with increased mortality rates. Specific features of RA are sustained synovial inflammation and progressive destruction of joints, which can be challenging for both patients and healthcare professionals. Besides the articular pathology, the extra-articular complications that lead to early deaths are significant. Patients develop higher risks of cardiovascular diseases due to systemic inflammation led by RA and also develop the risks of cancer and infections due to immune suppression by traditional therapies. So, even though RA seems like a disease that can merely disable the patients, it is safe to say that it can also pose as a silent killer due to these complications. While the existing therapeutic modalities, like disease-modifying anti-rheumatic drugs (DMARDs) such as Methotrexate (MTX), have shown effectiveness in reducing the symptoms of RA and slowing the disease progression, they are not without their major drawbacks. A key shortcoming is the incapacity to afford long-term therapeutic levels, leading to peaks in plasma concentrations (Cmax) and burst release of MTX in the bloodstream leading to less bioavailability at the inflamed tissues and more systemic accumulation. Moreover, traditional treatment modalities are frequently limited in addressing the immune-mediated component mechanisms underpinning RA. As in the case of MTX, it is the first line of therapy given to every patient, its inhibition of nucleotide synthesis leads to general dampening of the immune system and there is no specificity in terms of targeting the aggravated inflammation in joints. This emphasizes the need for ongoing research and advances in treatments that are based on a better understanding of RA pathophysiology and the translation of insights into therapeutic interventions. Hence, the current work focuses on a synergistic therapeutic strategy for RA management, using co-localization of folate-liposomal delivery of MTX and RELA siRNA. This targeted new approach not only relies on the co-presence of the well known DMARD MTX, with its anti-inflammatory and immunosuppressive properties, as well as of the RELA siRNA specifically targeting the NF-κB pathway implicated in the RA development's pathogenesis but also provides a precision therapy by using Folate liposomes. Folate receptors are overexpressed on inflammatory macrophages that are activated and these macrophages are one of the main mediators of synovial inflammation and joint destruction in RA, hence, they are a suitable target for drug delivery. The pursuit of this contemporary therapeutic strategy was tested through various in-vitro and in-vivo experiments. In vitro assessments demonstrated that the shape of the composite, liposome-folate complex is well stable and has a selective and sustained delivery of MTX and RELA siRNA to RAW264.7 macrophage cell line, commonly known to study macrophage biology and inflammation. Different functional assays suggested the ability of co-delivered therapeutics to bring the shift from inflammatory M1 to anti inflammatory M2 phenotypes, which are intermediate in RA. Moreover, Folate targeting liposomal co-delivery of MTX and RELA siRNA in the collagen-induced arthritis (CIA) rat model showed a significant drop in synovial inflammation and improvement in the animals' mobility with respect to the control groups, which showcased the enhanced efficacy of this nanoparticle mediated -co-delivery. Besides the fact that it is efficient in reducing the symptoms of RA and in preventing joint damage, the combination of MTX and RELA siRNA co-delivered via folate-liposomes has several advantages over conventional RA therapies. Firstly, by packing initially both curative agents through a separate delivery system, it managed the co-localized and sustained release of MTX and RELA siRNA thus evading possible systemic toxicity and side effects due to burst release in bloodstream. Another factor is that the usage of folate receptors in the targeting of effector macrophages will result in making the treatment more specific and more effective without causing many side effects in healthy tissues. Finally, the dual targeting mechanism, which aims to simultaneously act on several pathways involved in the RA pathogenesis, such as inflammation and immune dysregulation, could be the best way to obtain enhanced therapeutic outcomes compared to the monotherapy approaches. By this, we can propose to reduce the dose of MTX and obtain a more specific reduction in inflammation. We can target the following research gaps by hypothesized solutions: i. ii. iii. The combinatorial approach can enhance the NF-κB downregulation and target inflammation without general immune suppression. The off-target effects of MTX and its burst release due to high Cmax can be controlled by delivering the therapies in Folate liposomes. The dosage of single therapies can be reduced due to the synergistic effect of MTX and RELA siRNA. The next step for preclinical and clinical research will be to determine its safety and translation potential in treating RA.