Proteasome inhibition promotes Foxn1 expression in thymic epithelial cells and induces thymic regeneration in mice

The thymus plays a critical role in sustaining T-cell immunity, although its function is highly vulnerable to acute injury and physiologically declines with age, resulting in compromised immune responses. Impaired thymic function represents a major clinical challenge, particularly in settings of immunosuppression associated with cancer therapy and aging. Yet, effective strategies to rejuvenate the thymus remain limited. To explore novel regenerative approaches, we focused on FOXN1, a master regulator of thymic epithelial cell (TEC) development and function. By developing a custom screening platform, we tested a library of FDA-approved compounds for their ability to induce FOXN1 in TECs. Proteasome inhibition emerged as a potent and previously unrecognized mechanism for upregulating FOXN1 in both murine and human primary TECs. Among the hits identified in the screening, the antiparasitic drug nitazoxanide (NTZ) stood out for its proteasome inhibitory activity and for inducing Foxn1 expression while preserving cell viability, unlike other proteasome inhibitors. Mechanistically, NTZ-induced proteasome inhibition triggered endoplasmic reticulum stress (ER) and the adaptive unfolded protein response (UPR), ultimately engaging autophagy in TECs. In this context, the induction of autophagy acted as a compensatory mechanism to support cell survival in response to proteasome inhibition. Notably, when administered in mice, NTZ significantly accelerated functional thymic recovery after radiation-induced damage, promoting restoration of thymic architecture and cellularity of both stromal and hematopoietic compartments without disrupting physiological T-cell selection or tolerance mechanisms. Consistent with our in vitro findings, NTZ treatment induced Foxn1 and its downstream targets in TECs in vivo and conferred protection to TECs following irradiation. These findings uncover proteasome inhibition and, more broadly, modulation of ER stress and UPR pathways as a previously unrecognized mechanism regulating Foxn1 expression and position NTZ as a promising pharmacological strategy to enhance immunity in patients experiencing T-cell deficiencies due to cancer-related immunosuppression, infections, and age-related thymic atrophy.