Nit2/ω-amidase as a potential non-invasive diagnostic marker for moyamoya disease

Moyamoya (MM) is a progressive arteriopathy of the internal carotid artery and its branches that can lead to stroke and can be treated with surgical revascularization. The enzyme Nit2/ω-amidase (NIT2) is involved in glutaminase II and methionine salvage pathways. Recent studies have shown correlations between these metabolic pathways and MM arteriopathy, specifically increases in L-methionine, NO metabolites, and homocysteine. Here, we present data demonstrating that elevated levels of NIT2 correlate with the presence of MM, suggesting that it may have potential utility as a non-invasive biomarker for this condition.

Urine (n = 58) and blood plasma (n = 29) samples were collected from MM patients (aged 0-19) who were undergoing surgical revascularization and compared to samples collected from age- and sex-matched healthy controls (n = 23). The samples were then analyzed using Olink Explore 3072 proteomic proximity extension assays (PEA) to determine expression levels across ~ 3000 validated protein assays, revealing NIT2 as significantly elevated in MM patients compared to controls. We then conducted a secondary analysis of NIT2 using enzyme-linked immunosorbent assay (ELISA) for independent validation. Results were further analyzed for salient radiographic and clinical features, including Suzuki grade, bilateral vs. unilateral disease, radiographic stroke, and transient ischemic attack (TIA).

ANOVA analysis of the MM urine and plasma samples showed a statistically significant increase in Nit2/ω-amidase (NIT2) compared to the control samples, with MM patients exhibiting 1.6-2.3-fold increases in plasma expression (p = 0.04) and 4-11-fold increases in urinary expression (p ≤ 0.001). Higher levels of NIT2 in the urine were associated with moyamoya, bilateral disease, higher grade, stroke, and TIA.

NIT2 levels are significantly elevated in MM patients compared to matched controls, and higher levels correlate with measures of more severe disease. These novel data, coupled with the association of this molecule with metabolic pathways impacted by MM, support further investigation into NIT2 as a putative biomarker for MM.