Abstract Background Senescence is recognized as a critical barrier to epithelial regeneration and a driver of abnormal remodeling within the lung microenvironment. During early lung development, premature or dysregulated senescence can disrupt epithelial-stromal plasticity and contribute to long-term pulmonary dysfunction. However, the heterogeneity of senescent cell states and their responsiveness to therapy across developmental stages remains poorly understood. In this study, we applied a senotyping approach integrating multi-omic profiling and functional assays to define age-specific patterns of senescence and to evaluate how these differences influence the response to senolytic and MYC-targeted therapies. Methods Human pediatric lung tissue samples from the BRINDL repository were processed for single-nucleus RNA sequencing (snRNA-seq) using 10x Genomics technology to characterize plasticity markers defined here as the transcriptional flexibility cell cell-specific way to transition between progenitor and differentiated states, and senescence-associated gene expression (SASP). Samples were grouped into preterm, term, childhood, and adolescent, to map age-dependent differences in senescence programs. Gene network inference from snRNA-seq data revealed MYC as a recurrently activated transcriptional regulator in senescent epithelial and stromal niches showing senescence signatures (CDKN1A, CDKN2A, SERPINE1). To functionally validate these findings, precision-cut lung slices (PCLS) and air-liquid interface (ALI) airway cultures derived from 2-5 yrs and 15-16 yrs donors were irradiated and treated with senolytic Fisetin or the MYC inhibitor JQ1. Functional readouts, including epithelial barrier integrity (TEER), SASP cytokines, and apoptosis markers, were measured to evaluate how MYC modulation influences senescence and epithelial repair within the developing lung microenvironment. Results snRNA-seq revealed age-dependent differences in senescence and stress-response pathways, with MYC and KLF4 activation linked to epithelial plasticity and early stress signaling, in alveolar type 1 and type 2 cells and stromal cells. In early developmental models (2-5 years), treatment with Fisetin and JQ1 led to reduced SASP cytokine levels (IL-6, IL-8), decreased apoptosis, and improved epithelial barrier integrity. In contrast, adolescent models (15-16 years) showed opposite responses, with increased expression of senescence markers and pro-inflammatory cytokines (IL-1β, CCL2), along with epithelial disruption and matrix remodeling. These findings suggest that MYC-targeted and senolytic therapies may have stage-specific and potentially opposing effects, highlighting the need to consider developmental context in designing senescence-directed lung interventions. Conclusions Our findings identify MYC as an age-dependent regulator of senescence and show that senolytic and MYC-targeted treatments can have divergent effects during lung development. These results emphasize the importance of senotyping-guided, age-specific strategies when designing senescence-targeted interventions for the developmental lung diseases. This abstract is funded by: NIH
Shaikh et al. (Fri,) studied this question.