Urinary catheter blockage is a frequent problem for long-term catheterized patients, who are often referred to as ‘blockers’. Encrustation of the catheter is generally caused by urease-positive microorganisms, such as Proteus mirabilis [1]. Urease metabolizes urea to ammonia, resulting in alkaline urine, causing precipitation of struvite and apatite crystals, and these encrustations then lead to catheter blockage [1]. Long-term catheterized patients often have asymptomatic infections present within the bladder, therefore the prescription of antibiotics is generally discouraged. Here, we examine the use of urease inhibitors as an anti-virulence mechanism to prevent catheter blockage. Acetohydroxamic acid (AHA) is a urease-inhibitor and is prescribed for the treatment of frequent catheter blockage, it is not licensed in the UK or EU and is rarely used owing to its toxic side effect. Urease inhibitors have been well-examined in the literature, often for use in agriculture. However, other than AHA, none have been licensed as a therapeutic for catheter blockage. Our study aims to identify new and improved urease inhibitors which can be prescribed to long-term urinary catheter users who are frequent blockers.

Computational modelling docking experiments were used to dock small molecules onto the target-site using the crystal structure of the enzyme, urease from Sporosarcina pasteruii (PDB: 4UBP) (Fig.1). Docking was carried out using Cresset Flare software. Compounds were designed around the structures of known inhibitors: 2-mercaptoacetamide (2-MA), thiourea, and the natural flavonoid, quercetin [2,3]. Analysis of the docking score and the contacts of the compound to the urease crystal structure were used to rank the compounds. Compounds were filtered by Lipinski’s rules and the ability to purchase the compounds, the potency of the compound was assessed in vitro using the Berthelot assay against purified Canavalia ensiformis urease and a whole-cell assay of P. mirabilis.

From the virtual screening of high-ranking compounds, A5, A6, and A11, were identified and purchased. The ability of these compounds to inhibit urease was assessed using in vitro urease activity assays, with comparisons to known inhibitors (Fig. 2). Inhibitory concentration 50 (IC50) describes the ability of the compound to reduce the enzyme activity by 50%, IC50 can be used to determine the potency of the compounds.

Analysis of the IC50 showed that A11 appears to be 500-fold more potent than AHA against purified C. ensiformis urease (Fig. 2A). The whole-cell P. mirabilis assay informs on the ability of compounds to cross the bacterial membrane and access the intracellular urease. Results appear to show A11 to be 50-fold more potent than AHA (Fig. 2B).

We have demonstrated the small molecule, A11, as a potential therapeutic to treat recurrent catheter blockages. Future work is required to assess A11’s ability to prevent urinary catheter blockage using in vitro models of a catheterized urinary tract.

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Continence 2S2 (2022) 100465
DOI: 10.1016/j.cont.2022.100465