Elsayyad, Nihal Mohamed ElmahdyGomaa, ImanSalem, Mohamed AAmer, RehamEl-Laithy, Hanan M2022-10-012022-10-012022-09https://doi.org/10.1016/j.lfs.2022.121001https://bit.ly/3URjGXvAims: This study aims at formulating combined delivery of Risedronate sodium (RIS) and Vitamin D3 (VITD3) for augmented therapeutic outcome against osteoporosis (OP) using deep lung targeted PAMAM-G5-NH2 dendrimers to minimize RIS gastrointestinal side effects and enhance both drugs bioavailability through absorption from the alveoli directly to the blood. Methods: RIS-PAMAM-G5-NH2, VITD3-PAMAM-G5-NH2, and RIS/VITD3-PAMAM-G5-NH2 were prepared and evaluated in vitro for particle size (PS), zeta potential (ZP), %loading efficiency (%LE), morphology and FTIR. The efficacy of the RIS/VITD3-PAMAM-G5-NH2 compared to oral RIS was evaluated in OP-induced rats by comparing serum calcium, phosphorus, and computed bone mineral density (BMD) pre- and post-treatment. Additionally, a comprehensive metabolomics and molecular pathways approach was applied to find serum potential biomarkers for diagnosis and to evaluate the efficacy of inhaled RIS/VITD3-PAMAM-G5-NH2. Key findings: RIS/VITD3-PAMAM-G5-NH2 was successfully prepared with a %LE of 92.4±6.7% (RIS) and 83.2±4.4% (VIT-D3) and a PS of 252.8±34.1 adequate deep lung delivery. RIS/VITD3- PAMAM-G5-NH2 inhalation therapy was able to restore serum calcium, phosphorus, and BMD close to normal levels after 21 days of treatment in OP-induced rats. The WNT-signalling pathway and changes in the metabolite levels recovered to approximately normal levels upon treatment. Moreover, histone acetylation of the WNT-1 gene and miR-148a-3p interference proved to play a role in the regulation of the WNT-signalling pathway during OP progression and treatment. Significance: Pulmonary delivery of RIS/VITD3-PAMAM-G5-NH2 offers superior treatment for OP treatment compared to the oral route. Molecular and Metabolic pathways offer a key indicator of OP diagnosis and progression.en-USOsteoporosisRisedronate SodiumVitamin DPAMAM DendrimersMolecular PathwaysMetabolomicsArticlehttps://doi.org/10.1016/j.lfs.2022.121001