The ALK inhibitor reported by Cephalon provides an example where macrocyclization imparted selectivity without introducing any apparent new interactions or obvious steric clashes, suggesting that differences in strain in the bound macrocycle ring are responsible. ALK is a member of the insulin receptor (IR) superfamily of receptor tyrosine kinases. The ALK gene can participate in gene fusions that form kinase proteins that are constitutively active and thus oncogenic. Avoiding inhibition of IR was considered important for a safe drug. Breslin et al. started with the acyclic 2,4-diaminopyrimidine inhibitor 44 (Figure ), which inhibited ALK in vitro with IC50 = 22 nM but was selective for ALK versus IR by a factor of only 3.6. This compound was shown by X-ray crystallography to bind ALK in a U-shaped geometry. Modeling of a less substituted scaffold based on 44 into the active site of ALK suggested that macrocyclization through introduction of a two-carbon linker would constrain the compound in the bound conformation. The resulting cyclic compound, 45, inhibited ALK with 7-fold improved potency in vitro, and displayed a ~20-fold increase in selectivity against IR (IC50(IR)/IC50(ALK) = 67). The authors attributed the increased selectivity to the conformationally constrained macrocycle disfavoring conformers that bind to IR, such that adopting the conformation required to occupy the active site of IR involves some strain. A relatively modest strain energy of 1.8 kcal/mol upon binding to IR versus ALK would be sufficient to achieve the 20-fold increase in selectivity that was observed. Optimization of 45 by adding or elaborating substituents led to compound 46, with further improved potency against ALK in cellular assays (IC50 = 10 nM), and >140-fold selectivity over IR. The inhibitor series that led to the CDK/FLT3/JAK2 inhibitor zotiraciclib, described above (Figure ), provides an example where macrocyclization imparted selectivity by exploiting a sequence difference between the kinase active sites. This difference stabilizes an interaction with residues at the active site of certain kinase targets that are not present in other family members.