Novel nanogel offers potential treatment for rheumatoid arthritis by trapping and degrading cfDNA

Abstract

Researchers have developed a novel nanogel that effectively traps and degrades cell-free DNA (cfDNA), a potential cause of rheumatoid arthritis (RA), according to a new study published in Proceedings of the National Academy of Sciences (PNAS).1 This breakthrough offers insights into the potential of nanomedicine for treating RA and various autoimmune diseases. Excessive levels of cfDNA in the serum and synovium have been identified as a causative factor of RA, participating in its pathogenesis. Scavenging cfDNA has been recognized as an effective therapeutic approach for RA. Cationic polymers, driven by static charge interactions, can neutralize anionic cfDNA and hinder the progression of joint inflammation in RA models. However, the development of biocompatible cationic materials for RA treatment is still in its early stages. “Innovative and effective strategies to locally and thoroughly scavenge cfDNA are urgently needed,” remarks Lingyun Sun, one of the corresponding authors of the study. Inspired by the biological functions of organelles, the nanogel is formed by a cationic peptide dendrimer that binds to negatively charged cfDNA and a nuclease enzyme that breaks down cfDNA into harmless fragments. The nanogel modulates key molecular pathways involved in RA pathogenesis, such as the toll-like receptor 9 (TLR9) signaling pathway, which is activated by cfDNA and triggers immune responses. “The nanogels with a positive charge on their surface can bind and scavenge anionic cfDNA through electronic interaction, inhibit the intracellular trafficking of cfDNA from the plasma membrane to the endolysosomes, and prevent it from reaching TLR9. By clearing cfDNA, the ligand for TLR9 is reduced, which in turn induces the expression of inflammatory genes,” explains Sun. The nanogel is based on a third-generation polylysine dendrimer (G3K) modified with 2,2′-bipyridine-4-carboxylic acid (BPY) groups on its periphery. The BPY groups enable cross-linking of the dendrimer via aromatic stacking interactions in an aqueous solution, forming a spherical nanogel with a diameter of about 200 nm. The nanogel has a high density of positive charges on its surface, enabling interaction with the negative charges of cfDNA. To enhance the cfDNA degradation capability of the nanogel, the researchers conjugated deoxyribonuclease I (DNase I), an enzyme that cleaves DNA strands, to the surface of the nanogel using active ester chemistry. The resulting nanogel, named DG3K10, exhibits high biocompatibility and stability, and rapidly targets inflamed joints where cfDNA accumulates. In vivo testing of the nanogel was conducted using two different models of RA: one induced by injecting CpG oligodeoxynucleotides (ODNs), synthetic DNA fragments that mimic bacterial DNA and activate TLR9, and another induced by immunizing mice with bovine type II collagen (collagen-induced arthritis [CIA]), which triggers an autoimmune response against the host's own cartilage. In both models, the nanogel demonstrated a remarkable ability to bind and degrade cfDNA in the serum and synovium of RA animals, as well as inhibit the TLR9-mediated inflammatory cascade in immune cells. The nanogel also exhibited fast and specific targeting to inflamed joints after intravenous injection, as confirmed by near-infrared fluorescence imaging. Moreover, the nanogel significantly alleviated clinical symptoms of RA, including paw swelling, joint temperature, bone mineral density, cartilage loss, and synovial inflammation. Compared with other cationic materials with different sizes and charge densities, or to methotrexate (MTX), a widely used anti-rheumatic drug, the nanogel demonstrated superior therapeutic efficacy and minimal toxicity. “The bioinspired nanogel represents a novel strategy for treating RA by locally scavenging cfDNA in inflamed joints. The nanogel could also be applied to other diseases where cfDNA plays a pathogenic role, such as systemic lupus erythematosus (SLE), sepsis, or cancer,” says Sun. The nanogel could be further modified with other functional molecules or enzymes to achieve multifunctional delivery and therapy. To further study the mechanisms underlying this association, the researchers performed mRNA sequencing (RNA-seq) on the synovial tissue of nanogel-treated CIA mice. The results showed that the therapeutic effects of DG3K10 on RA mice were associated with the TLR-9 signaling pathway, the myeloid differentiation factor 88/TNF receptor associated factor 6/nuclear factor kappa-B (MyD88-TRAF6-NFκB)-dependent cascade, cytosolic DNA-sensing pathways, and key cytokines like tumor necrosis factor. DG3K10 treatment effectively upregulated genes involved in immune regulation and downregulated genes involved in TLR9 activation and inflammatory cytokine production. “These findings deepen the understanding of the biomimicking strategy for RA treatment and shed new light on nanomedicine for curing various autoimmune diseases,” remarks Sun.