The Evaluation of the Therapeutic Efficacy and Side Effects of a Macromolecular Dexamethasone Prodrug in the Collagen-Induced Arthritis Mouse Model

Abstract

Purpose: To investigate the efficacy and safety of N-(2-hydroxypropyl) methacrylamide (HPMA) copolymer-dexamethasone conjugate (P-Dex) in the collagen-induced arthritis (CIA) mouse model.

Methods: HPMA copolymer labeled with a near infrared fluorescence (NIRF) dye was administered to mice with CIA to validate its passive targeting to inflamed joints and utility as a drug carrier system. The CIA mice were treated with P-Dex, dexamethasone (Dex) or saline and the therapeutic efficacy and skeletal toxicity evaluated using clinical scoring and micro-computed tomography (μ-CT).

Results: The NIRF signal of the HPMA copolymer localized to arthritic joints consistent with its passive targeting to sites of inflammation. While the CIA mice responded more rapidly to P-Dex compared to Dex, the final clinical score and endpoint μ-CT analyses of localized bone erosions indicated that both single dose P-Dex and dose equivalent daily Dex led to comparable clinical efficacy after 30 days. μ-CT analysis of the proximal tibial metaphyses showed that P-Dex treatment was associated with significantly higher BMD and BV/TV compared to Dex and the saline control, consistent with reduced glucocorticoid (GC) skeletal toxicity.

Conclusion: These results validate the therapeutic efficacy of P-Dex in the CIA mouse model. P-Dex treatment averted the adverse effects of GC's on systemic bone loss, supporting its utility in clinical development for the management of rheumatoid arthritis.

Keywords: ELVIS; HPMA copolymer; collagen-induced arthritis; glucocorticoid; prodrug.

Figure 1

Clinical scores of the P-Dex, Dex and saline treated groups of mice with CIA and healthy controls. Scores for all four paws were summed for each mouse. Error bars have been omitted for clarity of presentation.

Figure 2

The effect of P-Dex, Dex, saline on paw swelling in the CIA mice.

Figure 3

Representative in vivo NIRF imaging of mice at different time points post-injection of IRDye CW 800-labeled HPMA conjugate (P-IRDye). The mouse had one arthritic limb (left hind) and three non-arthritic limbs. Persistent NIR fluorescence signals were detected only in arthritic limb, consistent with targeting and retention of the polymeric carrier in the arthritic joint.

Figure 4

Micro-CT analysis of the calcaneus in CIA mice after different treatments. Graphical representation of the BMD (a) and BV/TV (b) values among all the treatment groups. Density changes among all the treatment groups are as visualized in images from Data viewer in all three planes i.e. coronal, transaxial and saggital planes (c), using color-coded bar. Blue color with higher number represents bone with higher density.

Figure 5

Micro-CT analysis of the calcaneus in CIA mice from the different treatment groups. 3D-reconstructed images of the calcaneus (a). Note the eroded bone surface in the untreated group, which is reverted back to normal in treatment groups. 3D representation of the trabecular bone analyzed inside the calcaneus (b).

Figure 6

Micro-CT analysis of the calcaneus in CIA mice from the different treatment groups. (a), (b) and (c) show the differences among groups for BS/TV, Tb.N, and Tb.Sp respectively. (d) Images from CT-Vox between the groups. Note the increased porosity, eroded surface and exaggerated periosteal reaction of the untreated group. (e) 3D coronal cuts using CT-Vol, to have a visual representation of bony architecture inside the calcaneus.

Figure 7

Comparison of BMD and BV/TV measurement of calcaneus and proximal tibial metaphysis (PTM) in four groups. *Significantly different from healthy control group, p < 0.05; **Significantly different from saline group, p < 0.05; *** Significantly different from free Dex group, p < 0.05.