with good selectivity against several other kinases of this subset and in the kinome.
CC-90003 (69) [124], a structurally related covalent ERK1/2 inhibitor from
Celgene, was investigated in phase I clinical trials (NCT02313012) which were,
however, terminated in 2016 due to lack of response, an inappropriate PK-profile,
and neurotoxicity [125]. Analogous probe 70 (see also PDB: 5LCJ) enabled imaging
of ERK1/2 in living cells by employing trans-cyclooctene (TCO)/tetrazine (Tz) click
chemistry [126].
Covalent TAK1 targeting was accomplished by Gray and co-workers with
closely related inhibitors such as 71a (IC 50 ¼ 5.1 nM) and 71b (IC 50 ¼ 50 nM;
k inact /K I ¼ 6.9 Â 10
3 M
À1 s
À1 ) shown in Fig. 25b. Inhibitor 71b hit multiple kinases
in a panel among those being 10 kinases with a cysteine at the D1 position. However,
both compounds possessed a moderate selectivity against MEK1 and ERK2 also
sharing this cysteine. Interestingly, replacement of the acrylamide warhead by a
shorter chloroacetamide moiety led to a strong increase in selectivity against the
latter two kinases, which was accompanied by a rearrangement of the DFG motif in
TAK as observed by X-ray crystallography (see, for example, the PDB structures
5JH6 vs. 5E7R).
Various warhead chemistries were evaluated by Yang et al. to covalently trap
GSK-3β [127]. Out of their series, one of the most promising inhibitors was
fluoroacetamide 72 (Fig. 25c; IC 50 ¼ 17 nM). Labeling of Cys199 was suggested
by experiments with a fluorescent competitor probe and by digestion/MS using an
acrylamide-based analog. In contrast, two derivatives equipped with
2-chloropyridine moieties as S N Ar-type warheads proved to be reversible inhibitors
albeit with similar biochemical potency. Another interesting study was conducted by
Wissner et al. who installed two independent warheads on a quinazoline scaffold to
generate dual irreversible inhibitors of EGFR and vascular endothelial growth factor
receptor 2 (VEGFR2, also termed KDR) as exemplified by compound 73
[128]. While the 4-(dimethylamino)crotonamide residue was directed toward the
aforementioned F2 cysteine in EGFR, the substituted benzoquinone moiety was
meant to target Cys1045 located at the D1 position of VEGFR2, presumably via a
radical-based reductive addition mechanism. Very recently, α-cyanoacrylamide 73
has been described as dual GSK-3β/CK-1δ inhibitor [129]. While being only
moderately potent, the compound covalently modified GSK-3β as shown by X-ray
crystallography (PDB ID: 6H0U) while no covalent interaction can be assumed for
CK-1δ, which is devoid of cysteine moieties in the active site.
2.8 Development of Inhibitors Targeting Remote Cysteines or
Inactive Conformations
Remote cysteines are located outside of the regular kinase domain. Such cysteines
may also be amenable to covalent targeting with ATP pocket ligands, provided that
the cysteine of interest is positioned in spatial proximity to ATP binding cleft. A first
76
M. Gehringer
CC-90003 (69) [124], a structurally related covalent ERK1/2 inhibitor from
Celgene, was investigated in phase I clinical trials (NCT02313012) which were,
however, terminated in 2016 due to lack of response, an inappropriate PK-profile,
and neurotoxicity [125]. Analogous probe 70 (see also PDB: 5LCJ) enabled imaging
of ERK1/2 in living cells by employing trans-cyclooctene (TCO)/tetrazine (Tz) click
chemistry [126].
Covalent TAK1 targeting was accomplished by Gray and co-workers with
closely related inhibitors such as 71a (IC 50 ¼ 5.1 nM) and 71b (IC 50 ¼ 50 nM;
k inact /K I ¼ 6.9 Â 10
3 M
À1 s
À1 ) shown in Fig. 25b. Inhibitor 71b hit multiple kinases
in a panel among those being 10 kinases with a cysteine at the D1 position. However,
both compounds possessed a moderate selectivity against MEK1 and ERK2 also
sharing this cysteine. Interestingly, replacement of the acrylamide warhead by a
shorter chloroacetamide moiety led to a strong increase in selectivity against the
latter two kinases, which was accompanied by a rearrangement of the DFG motif in
TAK as observed by X-ray crystallography (see, for example, the PDB structures
5JH6 vs. 5E7R).
Various warhead chemistries were evaluated by Yang et al. to covalently trap
GSK-3β [127]. Out of their series, one of the most promising inhibitors was
fluoroacetamide 72 (Fig. 25c; IC 50 ¼ 17 nM). Labeling of Cys199 was suggested
by experiments with a fluorescent competitor probe and by digestion/MS using an
acrylamide-based analog. In contrast, two derivatives equipped with
2-chloropyridine moieties as S N Ar-type warheads proved to be reversible inhibitors
albeit with similar biochemical potency. Another interesting study was conducted by
Wissner et al. who installed two independent warheads on a quinazoline scaffold to
generate dual irreversible inhibitors of EGFR and vascular endothelial growth factor
receptor 2 (VEGFR2, also termed KDR) as exemplified by compound 73
[128]. While the 4-(dimethylamino)crotonamide residue was directed toward the
aforementioned F2 cysteine in EGFR, the substituted benzoquinone moiety was
meant to target Cys1045 located at the D1 position of VEGFR2, presumably via a
radical-based reductive addition mechanism. Very recently, α-cyanoacrylamide 73
has been described as dual GSK-3β/CK-1δ inhibitor [129]. While being only
moderately potent, the compound covalently modified GSK-3β as shown by X-ray
crystallography (PDB ID: 6H0U) while no covalent interaction can be assumed for
CK-1δ, which is devoid of cysteine moieties in the active site.
2.8 Development of Inhibitors Targeting Remote Cysteines or
Inactive Conformations
Remote cysteines are located outside of the regular kinase domain. Such cysteines
may also be amenable to covalent targeting with ATP pocket ligands, provided that
the cysteine of interest is positioned in spatial proximity to ATP binding cleft. A first
76
M. Gehringer