Several studies were performed in an attempt to elucidate the precise molecular
reason behind the resistance development in T790M-mutated EGFR. Initially, the
loss of affinity observed for EGFRi gefitinib (1) and erlotinib (2) was associated with
steric constraints in the binding pocket due the substitution of the smaller threonine
by the bulkier methionine residue, hampering a proper interaction with the
ATP-binding site [65]. However, crystallographic data revealed no significant differences in the crystal structures of gefitinib (1) co-crystalized with EGFR wt and
EGFR L858R/T790M . These experimental results weakened the steric clash
hypothesis [66].
Subsequently, Yun and colleagues demonstrated that the lower potency of
gefitinib (1) and erlotinib (2) for EGFR T790M inhibition was actually a consequence
of an altered enzymatic kinetic. These authors clearly showed that ATP has a higher
affinity for EGFR wt and EGFR T790M when compared to the sensible EGFR L858R -
mutated form. Consequently, competitive reversible inhibitors, e.g., gefitinib (1) and
erlotinib (2), have shorter residence times and are easily displaced by ATP in the
clinical nonresponsive EGFR variants. On the other hand, as ATP has a lower
affinity for EGFR L858R , first-generation EGFRi result in a better clinical outcome
for patients harboring this EGFR mutation [60].
Moreover, these data indicate that NSCLC patients harboring EGFR T790M or
EGFR L858R/T790M are not expected to respond adequately to treatment with firstgeneration EGFRi drugs, highlighting the critical need for the discovery of novel
inhibitors active against these mutated forms [62, 67, 68].
2.4 Second-Generation EGFRi and the Strategy
to Circumvent Resistance Development Mediated by
EGFR T790M Mutation
Some years after the first reports of acquired resistance to gefitinib (1) mediated by
EGFR T790M mutation, Kwak and coworkers suggested the benefits of designing
irreversible EGFRi to overcome this issue, due to their ability to inhibit diverse
EGFR mutant forms, including EGFR T90M [69]. The proposed irreversible inhibition
would take place in two steps, comprising the formation of an initial reversible
ligand-receptor complex, through complementary molecular recognition and
induced fit of the ligand to the target protein, followed by a reaction between an
electrophilic group in the structure of the EGFRi and a nucleophilic amino acid to
generate a covalent bond. Usually, the nucleophilic amino acid contains a hydroxyl
or thiol side chain, while the electrophilic site consists of a reactive chemical scaffold
placed in a strategic position in the ligand structure after considering the chemical
reactivity of the nucleophile and the topology of the target binding site [70–72].
The longer residence time observed for covalent inhibitors would be enough to
inactivate the target protein until its de novo synthesis, allowing wider dosage
intervals. Moreover, permanent inhibition of EGFR mutant forms and slow
Case Study on Receptor Tyrosine Kinases EGFR, VEGFR, and PDGFR
167
reason behind the resistance development in T790M-mutated EGFR. Initially, the
loss of affinity observed for EGFRi gefitinib (1) and erlotinib (2) was associated with
steric constraints in the binding pocket due the substitution of the smaller threonine
by the bulkier methionine residue, hampering a proper interaction with the
ATP-binding site [65]. However, crystallographic data revealed no significant differences in the crystal structures of gefitinib (1) co-crystalized with EGFR wt and
EGFR L858R/T790M . These experimental results weakened the steric clash
hypothesis [66].
Subsequently, Yun and colleagues demonstrated that the lower potency of
gefitinib (1) and erlotinib (2) for EGFR T790M inhibition was actually a consequence
of an altered enzymatic kinetic. These authors clearly showed that ATP has a higher
affinity for EGFR wt and EGFR T790M when compared to the sensible EGFR L858R -
mutated form. Consequently, competitive reversible inhibitors, e.g., gefitinib (1) and
erlotinib (2), have shorter residence times and are easily displaced by ATP in the
clinical nonresponsive EGFR variants. On the other hand, as ATP has a lower
affinity for EGFR L858R , first-generation EGFRi result in a better clinical outcome
for patients harboring this EGFR mutation [60].
Moreover, these data indicate that NSCLC patients harboring EGFR T790M or
EGFR L858R/T790M are not expected to respond adequately to treatment with firstgeneration EGFRi drugs, highlighting the critical need for the discovery of novel
inhibitors active against these mutated forms [62, 67, 68].
2.4 Second-Generation EGFRi and the Strategy
to Circumvent Resistance Development Mediated by
EGFR T790M Mutation
Some years after the first reports of acquired resistance to gefitinib (1) mediated by
EGFR T790M mutation, Kwak and coworkers suggested the benefits of designing
irreversible EGFRi to overcome this issue, due to their ability to inhibit diverse
EGFR mutant forms, including EGFR T90M [69]. The proposed irreversible inhibition
would take place in two steps, comprising the formation of an initial reversible
ligand-receptor complex, through complementary molecular recognition and
induced fit of the ligand to the target protein, followed by a reaction between an
electrophilic group in the structure of the EGFRi and a nucleophilic amino acid to
generate a covalent bond. Usually, the nucleophilic amino acid contains a hydroxyl
or thiol side chain, while the electrophilic site consists of a reactive chemical scaffold
placed in a strategic position in the ligand structure after considering the chemical
reactivity of the nucleophile and the topology of the target binding site [70–72].
The longer residence time observed for covalent inhibitors would be enough to
inactivate the target protein until its de novo synthesis, allowing wider dosage
intervals. Moreover, permanent inhibition of EGFR mutant forms and slow
Case Study on Receptor Tyrosine Kinases EGFR, VEGFR, and PDGFR
167