#5599 FEBUXOSTAT IN COMBINATION WITH ANAKINRA OR PH NEUTRALIZATION IMPROVE THE OUTCOMES IN HYPERURICEMIA-RELATED URIC ACID CRYSTALLURIA IN MICE
Authors: Ehreiser Louisa , LI Qiubo , LI LI , Anders Juliane , Steiger Stefanie 
Published: June 2023

Abstract

Background and Aims
Currently, it is unknown whether CKD patients with hyperuricemia (HU)-related uric acid (UA) crystalluria, also known as chronic UA nephropathy, would benefit from targeting HU, UA crystallization, and/or inflammation to slow down the progressive decline of kidney function. Although large multi-center RCTs with xanthine oxidase inhibitors (XOIs) such as allopurinol and two Mendelian randomization studies have disproven a causal link between HU and CKD progression, one cannot rule out that XOIs may still represent a viable therapeutic approach for patients with HU-related UA crystalluria in order to prevent CKD progression. This issue requires clarification. To address this, we used our well-established CKD mouse model of HU with UA crystalluria and tested numerous therapeutic approaches including XOI and anakinra.

Method
Alb-creERT2;Glut9lox/lox (male and female) mice were injected with tamoxifen and placed on an acidogenic diet with the purine inosine to induce HU and crystalluria-related CKD. After chronic UA nephropathy was established on day 14, mice were treated either 1) with the XOIs allopurinol or febuxostat, 2) with sodium bicarbonate supplementation to neutralize urinary pH to prevent UA crystallization, or 3) with anakinra to inhibit IL-1β-driven inflammation, or 4) a combination of the aforementioned therapies for 2 weeks. Saline was used as control. On day 0, 14, and 28, GFR (as primary endpoint) as well as serum UA levels and urinary UA crystals (as secondary endpoints) were determined. At the end of the study, we quantified kidney injury, immune cell infiltration, inflammation, and interstitial fibrosis using histological analysis, ELISA, RT-qPCR, and flow cytometry.

Results
Therapy with XOIs but not with sodium bicarbonate and anakinra reduced serum UA levels in mice with HU and UA crystalluria. Treating mice with febuxostat improved kidney function (increase in GFR) by preventing kidney injury, UA crystalluria, UA crystal granuloma formation, and interstitial fibrosis compared with the control group, while allopurinol worsened the outcomes of chronic UA nephropathy. On the other hand, sodium bicarbonate supplementation had no effects on serum UA levels, kidney inflammation, immune cell infiltration and interstitial fibrosis but improved kidney function due to less urinary UA crystal deposition (neutralization of urine pH) and granuloma formation as compared with saline-treated mice. As expected, anakinra did not slow down the progression of chronic UA nephropathy as noticed by a similar GFR decline compared with the control group. Although, anakinra reduced the number of infiltrating immune cells and the mRNA expression of inflammatory mediators in the kidney as well as the serum IL-1β concentrations, we observed more interstitial fibrosis. In combination, allopurinol with anakinra did not improve the outcomes of chronic UA nephropathy despite the inhibitory effect of anakinra on the inflammatory response because allopurinol triggered tubulointerstitial nephritis and vasodilation. Interestingly, both combination therapies febuxostat with anakinra and febuxostat with sodium bicarbonate were protective by improving kidney function and preventing UA crystal deposition, kidney injury, inflammation, interstitial fibrosis and UA crystal granuloma formation compared with the single therapies.

Conclusion
Our interventional study is the first pre-clinical study, which reveals that the XOIs allopurinol and febuxostat have differential effects on the outcomes of HU-related UA crystalluria. While allopurinol contributes to the progression of CKD by causing tubulointerstitial nephritis in mice, febuxostat rather seems to be renoprotective similar to urinary pH neutralization with sodium bicarbonate. We identified both combination therapies febuxostat with anakinra or sodium bicarbonate as most beneficial in order to prevent CKD progression in mice. Thus, febuxostat in combination with IL-1β inhibition or prevention of urinary UA crystallization may represent therapeutic approaches for patients with chronic UA nephropathy.