Rapamycin-activated Caspase-9 (RapaCasp9)

Engineered FKBP–FRB–caspase-9 fusion proteins, commonly known as Rapamycin-activated Caspase-9 (RapaCasp9), are synthetic safety switches designed to provide control over adoptive cell therapies, such as Chimeric Antigen Receptor (CAR) T-cell therapy (Stavrou et al., 2018). These proteins consist of a truncated human Caspase-9, which lacks the endogenous Caspase Activation and Recruitment Domain (CARD), fused to two dimerization domains: the FK506-binding protein (FKBP12) and the FKBP-rapamycin binding (FRB) domain of the mammalian target of rapamycin (mTOR) (Stavrou et al., 2018; Straathof et al., 2005). The system is activated by the administration of rapamycin (sirolimus) or its analogs, which function as molecular glues to induce the heterodimerization of the FKBP and FRB domains (Stavrou et al., 2018). This dimerization brings the fused Caspase-9 catalytic domains into close proximity, leading to their auto-activation and the subsequent initiation of the intrinsic apoptotic pathway through the activation of executioner caspases like Caspase-3 (Di Stasi et al., 2011; Gargett & Brown, 2014). This technology serves as a critical safety mechanism to mitigate severe adverse effects, such as cytokine release syndrome (CRS) or graft-versus-host disease (GvHD), by allowing for the rapid and selective elimination of the engineered cells in vivo (Di Stasi et al., 2011). The use of rapamycin as an inducer is particularly advantageous due to its established clinical profile and ability to cross the blood-brain barrier, though its systemic immunosuppressive effects must be considered (Stavrou et al., 2018).

Other names

iCasp9 (rapamycin-inducible)FKBP-FRB-Casp9Inducible Caspase-9Engineered FKBP–FRB–caspase-9 fusion proteinsRapamycin-inducible Caspase-9

Rapamycin-induced heterodimerization of the FKBP and FRB domains brings the fused Caspase-9 catalytic domains into close proximity, leading to their auto-activation and the subsequent initiation of the intrinsic apoptotic pathway.