The potential of senescent cell clearance to extend healthspan and delay the onset of age-related diseases has driven the development of innovative senotherapeutics. Thus, niosome-based gene therapy has emerged as a promising strategy to overcome the off-target limitations of first-generation senolytics, senomorphics, and senoreverts. In the present study, we investigated two new niosome formulations for genetically modifying primary umbilical cord stroma senescent MSCs (UC-MSCs) in a Palbociclib-induced senescent model, using non-senescent UC-MSCs as a control. These niosomes were formulated by fixing the cationic lipid composition (1,2-di-O-octadecenyl-3 trimethylammonium propane, DOTMA, D) and varying the non-ionic surfactants (polysorbate 20, P20, or polysorbate 80, P80) and the helper lipids (chloroquine, CQ, or cholesterol, CH) (DP20CQ and DP80CH, respectively) at two mole ratios (1:2:1 and 1:2:2 for DP20CQ; 1:2:2 and 1:2:4 for DP80CH). Results on transfection revealed that the DP20CQ formulations outperformed those with DP80CH in both UC-MSCs phenotypes from four different donors. Notably, DP20CQ niosomes prepared at 10/1 and 15/1 DOTMA/DNA ratios achieved the highest transgene expression levels in senescent UC-MSCs, surpassing even the commercial reagent Lipofectamine. Further, DP20CQ formulations exhibited adequate physicochemical properties for transfection and promoted a marked endosomal escape, particularly at the DOTMA/DNA ratio 10/1, without compromising cell viability. These findings underscore the value of niosomes as robust gene delivery vehicles for senescent cells in therapeutic antiaging strategies.
