Future improvements to the CAN methodology have the potential to dramatically streamline CPS projects. For example, CAN currently requires relatively high peptide concentrations to efficiently perform SPAAC (typically >50 μM) before NCL. Although we limited our demonstration to aqueous conditions with denaturant, it is worth noting that SPAAC is much more versatile than NCL since SPAAC can be performed in solvent mixtures more conducive for solubilizing peptides, such as MeCN/H2O that is compatible with being directly lyophilized. 107-108 Furthermore, unlike NCL, SPAAC does not suffer from competition with thioester hydrolysis. Future iterations of CAN may employ faster bioorthogonal chemistries for linking the peptides, as long as the chemistries are SPPS compatible. In fact, such bioorthogonal reactions that are also mutually orthogonal to DBCO and azide SPAAC would potentially enable multiple ligations to be performed in one pot via a CAN strategy. Such a combination would greatly increase yields in CPS by eliminating the need for intermediate HPLC purifications after each ligation. Finally, CAN parameters can be incorporated into an improved version of our "Automated Ligator" (Aligator) program, to predict segments that will benefit most from CAN.