XDH Humanized Mouse Model for Hyperuricemia and Gout Drug Research

Hyperuricemia, a metabolic disorder characterized by elevated uric acid levels in the blood, is a growing global health concern. It significantly increases the risk of gout—an inflammatory arthritis caused by urate crystal deposition in the joints—as well as kidney stones and cardiovascular complications. The rising prevalence of hyperuricemia and gout underscores the urgent need for innovative research models to facilitate drug development and therapeutic advancements.

To address this need, Cyagen has developed the B6-hXDH humanized mouse model, which incorporates the full-length human XDH gene while eliminating the native mouse Xdh gene. This model closely mimics human uric acid metabolism, making it an invaluable tool for preclinical studies targeting xanthine oxidase inhibitors (XOIs) and next-generation therapeutics for hyperuricemia and gout.

Discover how our B6-hXDH mouse model is poised to advance disease research and novel treatment discoveries.

Hyperuricemia and Gout: A Growing Global Health Concern

Hyperuricemia (HU) is a metabolic disorder characterized by abnormally elevated levels of uric acid in the blood. As the final product of purine metabolism in the body, excessive uric acid can crystallize into urate deposits in the joints, leading to gouty arthritis, or accumulate in the kidneys, forming stones. During a gout attack, patients often experience intense joint pain, redness, swelling, and fever.^[1] Long-term hyperuricemia also increases the risk of kidney disease, hypertension, and cardiovascular conditions.

The global prevalence of gout and hyperuricemia has been rising, making them among the most common chronic diseases after the "three highs" (hypertension, hyperlipidemia, and hyperglycemia). Currently, the prevalence of hyperuricemia worldwide ranges from 2.6% to 36%, with trends showing increasing incidence among younger populations - while the condition demonstrates a higher prevalence with increasing age.^[2]

Figure 1. Disease progression stages of gout induced by hyperuricemia.^[3]

Disease Progression and Treatment of Hyperuricemia and Gout

Gout is a common chronic inflammatory disease caused by the deposition of monosodium urate (MSU) crystals in both joint and non-joint structures. Elevated serum uric acid levels (hyperuricemia) are the primary risk factor for MSU crystal formation and gout onset. Clinically, hyperuricemia is associated with recurrent acute gouty arthritis attacks, tophus formation, chronic tophaceous arthritis, joint deformities, and kidney-related complications include chronic interstitial nephritis and urate nephrolithiasis, a type of kidney stone disease.^[4-5]

As of 2020, over 1.1 billion individuals worldwide were affected by hyperuricemia and gout, with projections estimating that the number of patients in China alone is expected to reach 200 million and 43.25 million, respectively, by 2024. The increasing patient population is driving continuous demand for effective treatments, with the global hyperuricemia and gout drug market expected to experience sustained long-term growth.

Since hyperuricemia is closely related to uric acid metabolism, current treatments primarily aim to manage the condition by reducing uric acid production or enhancing its excretion. Xanthine oxidoreductase (XOR), a key enzyme regulating uric acid synthesis, serves as an important therapeutic target.^[6]

Figure 2. Certain diseases, purine-rich diets, and alcohol metabolism can increase uric acid (UA) production.^[6]

Xanthine Oxidoreductase and Uric Acid Synthesis

Xanthine oxidoreductase (XOR) is a molybdenum-containing enzyme that exists in two interconvertible forms: xanthine dehydrogenase (XDH) in its reduced state and xanthine oxidase (XO) in its oxidized state. XDH can be converted into XO through reversible thiol oxidation or irreversible proteolytic modification.^[7] In its reduced state, XDH catalyzes the conversion of hypoxanthine to xanthine and subsequently to uric acid, producing NADH in the process. In its oxidized state, XO catalyzes the oxidation of xanthine to uric acid and hydrogen peroxide.

As a pivotal regulator of purine metabolism, XOR acts as a key regulatory point by catalyzing the oxidation of hypoxanthine to xanthine and further to uric acid - thereby controlling uric acid production.^[8-9] Inhibiting XOR activity can reduce serum uric acid levels, preventing urate crystal deposition in joints and tissues. This alleviates gout symptoms and reduces the risk of complications such as kidney stones, making XOR inhibition a key therapeutic strategy for hyperuricemia and gout. Xanthine oxidase inhibitors (XOIs), such as allopurinol and febuxostat, reduce uric acid production by inhibiting xanthine oxidase and have become essential treatments for hyperuricemia and gout.^[10]

Figure 3. Pathway of uric acid synthesis catalyzed by the xanthine oxidoreductase (XOR) system.^[9]

XDH Humanized Mouse Model for Drug Research

Despite their clinical use, XOIs like allopurinol and febuxostat present safety concerns. Allopurinol is associated with severe allergic reactions, while febuxostat carries an FDA black box warning due to cardiovascular risks, limiting its widespread use. As a result, there is a pressing need to develop safer alternatives, with small interfering RNA (siRNA)-based therapies emerging as a promising approach.

The XDH gene encodes both xanthine dehydrogenase (XDH) and xanthine oxidase (XO). However, genetic and protein differences between humans and mice present challenges for testing siRNA-based therapies targeting the human XDH gene or its mRNA. To address this, Cyagen has developed the B6-hXDH humanized mouse model (Catalog No. C001586), in which the murine Xdh gene has been replaced in situ with the full-length human XDH gene, including upstream and downstream untranslated regions (UTRs) and introns. This model provides an advanced platform for evaluating novel XOIs and RNA-based therapeutics to facilitate their transition into clinical trials.

Model Validation Data: Expression & Biochemistry Results

Gene expression analysis results indicate that B6-hXDH mice successfully express the human XDH gene, while the mouse Xdh gene is no longer expressed. The relative expression levels of the human XDH gene across tissues are similar to those of the native murine gene.

Figure 4. Comparison of human XDH gene and mouse Xdh gene expression across tissues in wild-type (WT) and B6-hXDH mice.

Protein Expression Analysis

Western blot results confirm that B6-hXDH mice successfully express the human xanthine oxidase protein in the liver and kidneys.

Figure 5. Protein expression analysis of human xanthine oxidase in the liver and kidney of wild-type (WT) and B6-hXDH mice.
Note: The antibody used for detection cross-reacts with both human and mouse xanthine oxidase, resulting in the appearance of human xanthine oxidase bands in WT mice during Western blot analysis.

Serum Uric Acid and Blood Urea Nitrogen Levels

Blood biochemical analysis results indicate that the serum uric acid (UA) levels in B6-hXDH mice are slightly lower than those in wild-type mice, with more pronounced differences observed in male mice. No significant differences were observed in the serum blood urea nitrogen (BUN) levels between B6-hXDH and wild-type mice.

Figure 6. Comparison of serum uric acid (UA) and blood urea nitrogen (BUN) levels between wild-type (WT) and B6-hXDH mice.

Conclusion

The B6-hXDH humanized mouse model (Catalog No. C001586) features an in situ replacement of the murine Xdh gene with the complete human XDH sequence, including non-coding regulatory regions. These mice express human XDH gene and xanthine oxidase (XO) protein in a pattern similar to the endogenous mouse gene. This model accurately replicates human-like XDH gene and protein expression patterns, making them particularly suitable for studying the pathophysiology of hyperuricemia and gout and an ideal preclinical platform for developing novel xanthine oxidase inhibitors and innovative small RNA-based therapies.

Cyagen Metabolic Disease Models

Cyagen collaborates extensively with leading pharmaceutical companies, biotechnology firms, and academic research institutions worldwide to develop a comprehensive range of metabolic disease models. Our gene modeling experts have developed disease models related to metabolic conditions such as liver disease, obesity, diabetes, hyperuricemia, and atherosclerosis, accelerating research and drug discovery efforts in these fields.

Recommended Models for Metabolic and Cardiovascular Diseases

References
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[2]Huang Yuchai, Lü Yongman. Scientific Understanding and Standardized Management of Hyperuricemia [J]. Chinese Journal of Health Management, 2023, 17(4): 316-319.
[3]Verywell Health. (n.d.). Hyperuricemia (high uric acid). Retrieved January 2, 2025, from https://www.verywellhealth.com/hyperuricemia-high-uric-acid-189838
[4]Dalbeth N, Choi HK, Joosten LAB, Khanna PP, Matsuo H, Perez-Ruiz F, Stamp LK. Gout. Nat Rev Dis Primers. 2019 Sep 26;5(1):69.
[5]Dalbeth N, Gosling AL, Gaffo A, Abhishek A. Gout. Lancet. 2021 May 15;397(10287):1843-1855. doi: 10.1016/S0140-6736(21)00569-9. Epub 2021 Mar 30. Erratum in: Lancet. 2021 May 15;397(10287):1808.
[6]Du L, Zong Y, Li H, Wang Q, Xie L, Yang B, Pang Y, Zhang C, Zhong Z, Gao J. Hyperuricemia and its related diseases: mechanisms and advances in therapy. Signal Transduct Target Ther. 2024 Aug 28;9(1):212.
[7]Cicero AFG, Fogacci F, Kuwabara M, Borghi C. Therapeutic Strategies for the Treatment of Chronic Hyperuricemia: An Evidence-Based Update. Medicina (Kaunas). 2021 Jan 10;57(1):58.
[8]Furuhashi M. New insights into purine metabolism in metabolic diseases: role of xanthine oxidoreductase activity. Am J Physiol Endocrinol Metab. 2020 Nov 1;319(5):E827-E834. 
[9]Bortolotti M, Polito L, Battelli MG, Bolognesi A. Xanthine oxidoreductase: One enzyme for multiple physiological tasks. Redox Biol. 2021 May;41:101882.
[10]Lee Y, Hwang J, Desai SH, Li X, Jenkins C, Kopp JB, Winkler CA, Cho SK. Efficacy of Xanthine Oxidase Inhibitors in Lowering Serum Uric Acid in Chronic Kidney Disease: A Systematic Review and Meta-Analysis. J Clin Med. 2022 Apr 27;11(9):2468.