PURPOSE: The first-generation ALK tyrosine kinase inhibitor (TKI) crizotinib is 
a standard therapy for patients with ALK-rearranged non-small cell lung cancer 
(NSCLC). Several next-generation ALK-TKIs have entered the clinic and have shown 
promising activity in crizotinib-resistant patients. As patients still relapse 
even on these next-generation ALK-TKIs, we examined mechanisms of resistance to 
the next-generation ALK-TKI alectinib and potential strategies to overcome this 
resistance.
EXPERIMENTAL DESIGN: We established a cell line model of alectinib resistance, 
and analyzed a resistant tumor specimen from a patient who had relapsed on 
alectinib. We developed Ba/F3 models harboring alectinib-resistant ALK mutations 
and evaluated the potency of other next-generation ALK-TKIs in these models. We 
tested the antitumor activity of the next-generation ALK-TKI ceritinib in the 
patient with acquired resistance to alectinib. To elucidate structure-activity 
relationships of ALK mutations, we performed computational thermodynamic 
simulation with MP-CAFEE.
RESULTS: We identified a novel V1180L gatekeeper mutation from the cell line 
model and a second novel I1171T mutation from the patient who developed 
resistance to alectinib. Both ALK mutations conferred resistance to alectinib as 
well as to crizotinib, but were sensitive to ceritinib and other next-generation 
ALK-TKIs. Treatment of the patient with ceritinib led to a marked response. 
Thermodynamics simulation suggests that both mutations lead to distinct 
structural alterations that decrease the binding affinity with alectinib.
CONCLUSIONS: We have identified two novel ALK mutations arising after alectinib 
exposure that are sensitive to other next-generation ALK-TKIs. The ability of 
ceritinib to overcome alectinib-resistance mutations suggests a potential role 
for sequential therapy with multiple next-generation ALK-TKIs.