Cell permeability PROTACs

The next logical step in the development of the PROTAC technique as a viable therapeutic option was the design of a PROTAC scaffolding that would be independently cell permeable, negating the need for microinjection as the method for PROTAC delivery. A cell permeable PROTAC would also lessen the molecular biological manipulations necessary for PROTAC activity, emphasizing the capacity for the technique as both a tool for understanding protein function in the cell and as a potential therapeutic. 

In order to generate a cell permeable PROTAC, an alternative E3 ligase ligand was required. The IkBcx phosphopeptide functioned successfully to recruit the E3 ligase components, but the negatively charged... 

In order to ensure the cell permeability of the new PROTAC design, an eight amino acid polyarginine tag was included on the carboxy-terminus of the E3 ligase-targeting peptide sequence of the PROTAC. This significant positive charge serves to facilitate translocation of the PROTAC into cells via a mechanism that mimics the HIV Tat protein. ... 

Figure 3.10 Structure of PROTAC-4 (5). AP21998 binds the target protein FKBP(F36V), while the HIFla peptide recruits the von Hippel-Lindau E3 ligase (VHL). A polyarginine tag provides cell permeability.

Figure 3.11 PROTAC-4 is cell-permeable and mediates EGFP-FKBP degradation.

PROTAC-5 (6), the cell-permeable AR-targeting PROTAC, is shown in Figure 3.12, and is composed of DHT connected by a linker to the HIFla peptide with a C-terminal polyarginine tag. As with PROTAC-3, a HEK293... 

Figure 3.13 PROTAC-5 is cell-permeable and mediates GFP-AR degradation. One...

The success of the all-small molecule PROTAC-6 was an important step forward in the continued development of the PROTAC technique. Not only did it show the potential for significant variations in the structural design of the PROTAC, but also the flexibility in the choice of an E3 ligase target. In addition, it showed the successful improvement of PROTAC cell-permeability... 

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