Alkaptonuria in two Colombian patients: identification of HGD variants including a novel finding

Abstract Background: Alkaptonuria (AKU) is a rare autosomal recessive inborn error of metabolism caused by deficiency of homogentisate 1,2-dioxygenase (HGD), its deficiency results in homogentisic acid (HGA) accumulation, which oxidizes to form melanin-like pigments that deposit in connective tissues, leading to ochronosis and progressive multisystem complications. We describe two unrelated Colombian female patients with AKU, emphasizing clinical variability and novel genetic findings. Case reports: Case 1: Female with onset of symptoms at age 16, including scleral pigmentation, glaucoma, retinal detachment, and joint pain. Arthroscopy revealed ochronotic pigmentation and cystic changes in the glenohumeral cartilage. Lumbar MRI showed intervertebral disc narrowing and calcifications. Pigmented deposits were observed in the ears and sclera during adulthood. Whole-exome sequencing (WES) identified two HGD variants: c.164_166del (p.Thr55del) and c.774+69C>T, both classified as variants of uncertain significance (VUS). Case 2: Female with reddish urine noted perinatally and recurrent darkened urine episodes during infections. At age 8, she developed intermittent bilateral bone pain in knees and wrists, worsened by physical activity. No evident ochronosis was observed. Whole genome sequencing (WGS) revealed compound heterozygosity for HGD variants: c.808G>A (p.Gly270Arg), likely pathogenic, and c.774+69C>T (VUS). Conclusions: Both patients shared the intronic HGD variant c.774+69C>T in heterozygosity, a VUS located outside the canonical splice site, which limits its functional detection through WES. Its recurrence in two unrelated individuals—each with AKU-compatible symptoms and a second HGD variant—raises the possibility of pathogenic relevance. This deep intronic variant poses a diagnostic challenge, as WES has limitations in assessing the impact of non-coding variants on splicing regulation. In contrast, WGS enables a more accurate characterization of deep intronic regions, as demonstrated in the second case, where this same VUS was confirmed using that approach. This case report emphasizes the relevance and challenge of molecular diagnosis in AKU for confirmation, genetic counseling, and expanding understanding of the disorder’s genetic heterogeneity. Notably, we report a previously undescribed HGD variant in a Latin American population, underscoring the importance of genomic characterization in underrepresented groups.