Longitudinal Single-Cell Transcriptomic Profiling Reveals Dynamic Immune Cell Alterations During Burosumab Therapy in X-Linked Hypophosphatemia.

X-linked hypophosphatemia (XLH) is a rare hereditary disorder characterized by PHEX gene mutations, elevated FGF23 levels, and impaired bone mineralization. Burosumab, a monoclonal antibody targeting FGF23, has demonstrated clinical efficacy; however, the immunological dynamics during treatment remain unexplored. This study employed longitudinal single-cell RNA sequencing (scRNA-seq) to characterize peripheral blood immune cell alterations across multiple treatment stages in pediatric XLH. We performed scRNA-seq on peripheral blood mononuclear cells from pediatric patients with XLH at five time points spanning pretreatment and burosumab therapy phases, along with healthy pediatric controls. A total of 93,112 cells were analyzed using comprehensive bioinformatic pipelines, including unsupervised clustering, pseudotime trajectory analysis, temporal gene expression profiling, and cell-cell communication inference. Eleven major immune cell populations were identified, with notable dynamic alterations in T cells and natural killer (NK) cell subtypes across treatment stages. The cellular proportion of T helper 2 (Th2) cells and regulatory T (Treg) cells were elevated before treatment and normalized during therapy, whereas T helper 17 (Th17) cells exhibited reciprocal patterns. Genes upregulated in Treg cells during early treatment were enriched in osteoclast differentiation pathway. Natural killer subtype 2 cells showed enrichment in osteoclast differentiation and interleukin-12 response pathways. Cell-cell communication analysis identified dynamic interactions among Th2 cells, Th17 cells, Treg cells, and NK cell subtypes mediated by KLRB1-CLEC2D and SELL-SELPLG ligand-receptor pairs. This longitudinal transcriptomic study provides the first comprehensive characterization of peripheral immune dynamics during burosumab therapy in XLH, offering new insights into the immunological mechanisms underlying treatment response.