Preclinical Modeling of Osimertinib for NSCLC With EGFR Exon 20 Insertion Mutations

Yusoo Lee, Tae Min Kim, Dong Wan Kim, Soyeon Kim, Miso Kim, Bhumsuk Keam, Ja-Lok Ku, Dae Seog Heo

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Introduction: NSCLC with EGFR exon 20 insertion mutations is the third most common type of EGFR-mutant NSCLC and is resistant to EGFR tyrosine kinase inhibitors (TKIs). This study was conducted to evaluate the efficacies of first- to third-generation EGFR TKIs against NSCLC cells harboring EGFR exon 20 insertion mutations. Methods: We developed seven EGFR exon 20 insertion-mutant Ba/F3 models and one patient-derived NSCLC (SNU-3173) of subtypes A763insFQEA, V769insASV, D770insSVD, D770insNPG, P772insPR, H773insH, H773insNPH, and H773insAH. Cell viability assays, immunoblotting, and N-ethyl-N-nitrosourea mutagenesis screenings were performed. EGFR exon 20 insertion–mutant structures and couplings with osimertinib, a third-generation EGFR TKI, were modeled and compared. Results: EGFR exon 20 insertion–mutant NSCLC cells, excluding EGFR A763insFQEA, were resistant to first-generation EGFR TKIs (concentration that inhibits 50% [IC50], 1.1 ± 0.067 to 5.4 ± 0.115 μM). Mutants were sensitive to second-generation EGFR TKIs (IC50, 0.02 ± 0.0002 to 161.8 ± 18.7nM), except EGFR H773insH (IC50, 46.3 ± 8.0 to 352.5 ± 22.7nM). The IC50 ratios for mutant to wild-type cells were higher than those for third-generation EGFR TKIs. Third-generation EGFR TKI osimertinib was highly potent against EGFR exon 20 insertion–mutant cells (IC50, 14.7-62.7 nM), including EGFR H773insH, and spared wild-type EGFR cells. N-ethyl-N-nitrosourea mutagenesis screening of EGFR exon 20 insertion–mutant Ba/F3 cells showed various second sites for EGFR mutations, mostly at exons 20 and 21, including E762K, P794S, and G796D. In addition, osimertinib-resistant cells were established by stepwise exposure to osimertinib and harbored EGFR E762K mutation. Conclusions: Osimertinib is active against EGFR exon 20 insertion–mutant NSCLC and flexibly binds within drug-binding pockets in preclinical models.

Original languageEnglish
Pages (from-to)1556-1566
Number of pages11
JournalJournal of Thoracic Oncology
Volume14
Issue number9
DOIs
StatePublished - 1 Sep 2019

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Insertional Mutagenesis
Exons
Protein-Tyrosine Kinases
Inhibitory Concentration 50
Ethylnitrosourea
Mutagenesis
osimertinib
Mutation
Immunoblotting
Cell Survival

Keywords

  • EGFR exon 20 insertion
  • NSCLC
  • Osimertinib
  • Resistance

Cite this

Lee, Yusoo ; Kim, Tae Min ; Kim, Dong Wan ; Kim, Soyeon ; Kim, Miso ; Keam, Bhumsuk ; Ku, Ja-Lok ; Heo, Dae Seog. / Preclinical Modeling of Osimertinib for NSCLC With EGFR Exon 20 Insertion Mutations. In: Journal of Thoracic Oncology. 2019 ; Vol. 14, No. 9. pp. 1556-1566.
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title = "Preclinical Modeling of Osimertinib for NSCLC With EGFR Exon 20 Insertion Mutations",
abstract = "Introduction: NSCLC with EGFR exon 20 insertion mutations is the third most common type of EGFR-mutant NSCLC and is resistant to EGFR tyrosine kinase inhibitors (TKIs). This study was conducted to evaluate the efficacies of first- to third-generation EGFR TKIs against NSCLC cells harboring EGFR exon 20 insertion mutations. Methods: We developed seven EGFR exon 20 insertion-mutant Ba/F3 models and one patient-derived NSCLC (SNU-3173) of subtypes A763insFQEA, V769insASV, D770insSVD, D770insNPG, P772insPR, H773insH, H773insNPH, and H773insAH. Cell viability assays, immunoblotting, and N-ethyl-N-nitrosourea mutagenesis screenings were performed. EGFR exon 20 insertion–mutant structures and couplings with osimertinib, a third-generation EGFR TKI, were modeled and compared. Results: EGFR exon 20 insertion–mutant NSCLC cells, excluding EGFR A763insFQEA, were resistant to first-generation EGFR TKIs (concentration that inhibits 50{\%} [IC50], 1.1 ± 0.067 to 5.4 ± 0.115 μM). Mutants were sensitive to second-generation EGFR TKIs (IC50, 0.02 ± 0.0002 to 161.8 ± 18.7nM), except EGFR H773insH (IC50, 46.3 ± 8.0 to 352.5 ± 22.7nM). The IC50 ratios for mutant to wild-type cells were higher than those for third-generation EGFR TKIs. Third-generation EGFR TKI osimertinib was highly potent against EGFR exon 20 insertion–mutant cells (IC50, 14.7-62.7 nM), including EGFR H773insH, and spared wild-type EGFR cells. N-ethyl-N-nitrosourea mutagenesis screening of EGFR exon 20 insertion–mutant Ba/F3 cells showed various second sites for EGFR mutations, mostly at exons 20 and 21, including E762K, P794S, and G796D. In addition, osimertinib-resistant cells were established by stepwise exposure to osimertinib and harbored EGFR E762K mutation. Conclusions: Osimertinib is active against EGFR exon 20 insertion–mutant NSCLC and flexibly binds within drug-binding pockets in preclinical models.",
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Preclinical Modeling of Osimertinib for NSCLC With EGFR Exon 20 Insertion Mutations. / Lee, Yusoo; Kim, Tae Min; Kim, Dong Wan; Kim, Soyeon; Kim, Miso; Keam, Bhumsuk; Ku, Ja-Lok; Heo, Dae Seog.

In: Journal of Thoracic Oncology, Vol. 14, No. 9, 01.09.2019, p. 1556-1566.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Preclinical Modeling of Osimertinib for NSCLC With EGFR Exon 20 Insertion Mutations

AU - Lee, Yusoo

AU - Kim, Tae Min

AU - Kim, Dong Wan

AU - Kim, Soyeon

AU - Kim, Miso

AU - Keam, Bhumsuk

AU - Ku, Ja-Lok

AU - Heo, Dae Seog

PY - 2019/9/1

Y1 - 2019/9/1

N2 - Introduction: NSCLC with EGFR exon 20 insertion mutations is the third most common type of EGFR-mutant NSCLC and is resistant to EGFR tyrosine kinase inhibitors (TKIs). This study was conducted to evaluate the efficacies of first- to third-generation EGFR TKIs against NSCLC cells harboring EGFR exon 20 insertion mutations. Methods: We developed seven EGFR exon 20 insertion-mutant Ba/F3 models and one patient-derived NSCLC (SNU-3173) of subtypes A763insFQEA, V769insASV, D770insSVD, D770insNPG, P772insPR, H773insH, H773insNPH, and H773insAH. Cell viability assays, immunoblotting, and N-ethyl-N-nitrosourea mutagenesis screenings were performed. EGFR exon 20 insertion–mutant structures and couplings with osimertinib, a third-generation EGFR TKI, were modeled and compared. Results: EGFR exon 20 insertion–mutant NSCLC cells, excluding EGFR A763insFQEA, were resistant to first-generation EGFR TKIs (concentration that inhibits 50% [IC50], 1.1 ± 0.067 to 5.4 ± 0.115 μM). Mutants were sensitive to second-generation EGFR TKIs (IC50, 0.02 ± 0.0002 to 161.8 ± 18.7nM), except EGFR H773insH (IC50, 46.3 ± 8.0 to 352.5 ± 22.7nM). The IC50 ratios for mutant to wild-type cells were higher than those for third-generation EGFR TKIs. Third-generation EGFR TKI osimertinib was highly potent against EGFR exon 20 insertion–mutant cells (IC50, 14.7-62.7 nM), including EGFR H773insH, and spared wild-type EGFR cells. N-ethyl-N-nitrosourea mutagenesis screening of EGFR exon 20 insertion–mutant Ba/F3 cells showed various second sites for EGFR mutations, mostly at exons 20 and 21, including E762K, P794S, and G796D. In addition, osimertinib-resistant cells were established by stepwise exposure to osimertinib and harbored EGFR E762K mutation. Conclusions: Osimertinib is active against EGFR exon 20 insertion–mutant NSCLC and flexibly binds within drug-binding pockets in preclinical models.

AB - Introduction: NSCLC with EGFR exon 20 insertion mutations is the third most common type of EGFR-mutant NSCLC and is resistant to EGFR tyrosine kinase inhibitors (TKIs). This study was conducted to evaluate the efficacies of first- to third-generation EGFR TKIs against NSCLC cells harboring EGFR exon 20 insertion mutations. Methods: We developed seven EGFR exon 20 insertion-mutant Ba/F3 models and one patient-derived NSCLC (SNU-3173) of subtypes A763insFQEA, V769insASV, D770insSVD, D770insNPG, P772insPR, H773insH, H773insNPH, and H773insAH. Cell viability assays, immunoblotting, and N-ethyl-N-nitrosourea mutagenesis screenings were performed. EGFR exon 20 insertion–mutant structures and couplings with osimertinib, a third-generation EGFR TKI, were modeled and compared. Results: EGFR exon 20 insertion–mutant NSCLC cells, excluding EGFR A763insFQEA, were resistant to first-generation EGFR TKIs (concentration that inhibits 50% [IC50], 1.1 ± 0.067 to 5.4 ± 0.115 μM). Mutants were sensitive to second-generation EGFR TKIs (IC50, 0.02 ± 0.0002 to 161.8 ± 18.7nM), except EGFR H773insH (IC50, 46.3 ± 8.0 to 352.5 ± 22.7nM). The IC50 ratios for mutant to wild-type cells were higher than those for third-generation EGFR TKIs. Third-generation EGFR TKI osimertinib was highly potent against EGFR exon 20 insertion–mutant cells (IC50, 14.7-62.7 nM), including EGFR H773insH, and spared wild-type EGFR cells. N-ethyl-N-nitrosourea mutagenesis screening of EGFR exon 20 insertion–mutant Ba/F3 cells showed various second sites for EGFR mutations, mostly at exons 20 and 21, including E762K, P794S, and G796D. In addition, osimertinib-resistant cells were established by stepwise exposure to osimertinib and harbored EGFR E762K mutation. Conclusions: Osimertinib is active against EGFR exon 20 insertion–mutant NSCLC and flexibly binds within drug-binding pockets in preclinical models.

KW - EGFR exon 20 insertion

KW - NSCLC

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