Targeting HER2 Exon 20 Insertion-Mutant Lung Adenocarcinoma with a Novel Tyrosine Kinase Inhibitor Mobocertinib

Han Han, Shuai Li, Ting Chen, Michael Fitzgerald, Shengwu Liu, Chengwei Peng, Kwan Ho Tang, Shougen Cao, Johara Chouitar, Jiansheng Wu, David Peng, Jiehui Deng, Zhendong Gao, Theresa E Baker, Fei Li, Hua Zhang, Yuanwang Pan, Hailin Ding, Hai Hu, Val Pyon, Cassandra Thakurdin, Eleni Papadopoulos, Sittinon Tang, Francois Gonzalvez, Haiquan Chen, Victor M Rivera, Rachael Brake, Sylvie Vincent, Kwok-Kin Wong, Han Han, Shuai Li, Ting Chen, Michael Fitzgerald, Shengwu Liu, Chengwei Peng, Kwan Ho Tang, Shougen Cao, Johara Chouitar, Jiansheng Wu, David Peng, Jiehui Deng, Zhendong Gao, Theresa E Baker, Fei Li, Hua Zhang, Yuanwang Pan, Hailin Ding, Hai Hu, Val Pyon, Cassandra Thakurdin, Eleni Papadopoulos, Sittinon Tang, Francois Gonzalvez, Haiquan Chen, Victor M Rivera, Rachael Brake, Sylvie Vincent, Kwok-Kin Wong

Abstract

No targeted treatments are currently approved for HER2 exon 20 insertion-mutant lung adenocarcinoma patients. Mobocertinib (TAK-788) is a potent irreversible tyrosine kinase inhibitor (TKI) designed to target human epidermal growth factor receptor 2 (HER2/ERBB2) exon 20 insertion mutations. However, the function of mobocertinib on HER2 exon 20 insertion-mutant lung cancer is still unclear. Here we conducted systematic characterization of preclinical models to understand the activity profile of mobocertinib against HER2 exon 20 insertions. In HER2 exon 20 insertion-mutant cell lines, the IC50 of mobocertinib was higher than poziotinib and comparable with or slightly lower than afatinib, neratinib, and pyrotinib. Mobocertinib had the lowest HER2 exon 20 insertion IC50/wild-type (WT) EGFR IC50 ratio, indicating that mobocertinib displayed the best selectivity profile in these models. Also, mobocertinib showed strong inhibitory activity in HER2 exon 20YVMA allograft and patient-derived xenograft models. In genetically engineered mouse models, HER2 exon 20G776>VC lung tumors exhibited a sustained complete response to mobocertinib, whereas HER2 exon 20YVMA tumors showed only partial and transient response. Combined treatment with a second antibody-drug conjugate (ADC) against HER2, ado-trastuzumab emtansine (T-DM1), synergized with mobocertinib in HER2 exon 20YVMA tumors. In addition to the tumor cell autonomous effect, sustained tumor growth control derived from M1 macrophage infiltration and CD4+ T-cell activation. These findings support the ongoing clinical development of mobocertinib (NCT02716116) and provide a rationale for future clinical evaluation of T-DM1 combinational therapy in HER2 exon 20YVMA insertion-mutant lung adenocarcinoma patients. SIGNIFICANCE: This study elucidates the potent inhibitory activity of mobocertinib against HER2 exon 20 insertion-mutant lung cancer and the synergic effect of combined mobocertinib and T-DM1, providing a strong rationale for clinical investigation.

©2021 The Authors; Published by the American Association for Cancer Research.

Figures

Graphical abstract
Graphical abstract
Figure 1.
Figure 1.
HER2 exon 20 insertion mutations are sensitive to mobocertinib. A, IC50 of mobocertinib, osimertinib, neratinib, poziotinib, afatinib, and pyrotinib on Ba/F3-HER2 exon 20 insertion–mutant cell lines. B, IC50 of mobocertinib, poziotinib, osimertinib, and afatinib on H1781 (HER2G776>VC) cell line. C, The ratio of Ba/F3-HER2 exon 20 insertion–mutant IC50 to EGFR WT IC50. D, Western blot of HER2 signaling (total HER2, pHER2, total AKT, pAKT, total ERK, pERK, and actin) of H1781 (HER2 Exon 20G776>VC) cell line treated with 0.01 μmol/L, 0.1 μmol/L, and 1 μmol/L mobocertinib for 6 hours. E, Western blot of HER2 signaling (total HER2, pHER2, total AKT, pAKT, total ERK, pERK, and actin) of Ba/F3-HER2 exon 20YVMA cell line treated with 0.1 μmol/L of mobocertinib for 6 hours. F, Quantification of colony formation assay on H1781 (HER2 exon 20G776>VC) cell line treated with DMSO control, 0.01 μmol/L, and 0.1 μmol/L of mobocertinib. G, Tumor volumes of Ba/F3-HER2 exon 20YVMA allograft treated with vehicle, mobocertinib 50 and 100 mpk. H, Tumor volume change of NSCLC HER2 exon 20YVMA PDX treated with vehicle, 15 and 30 mpk mobocertinib. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
Figure 2.
Figure 2.
HER2 exon 20G776>VC GEMM showed sustained response to mobocertinib. A, Working model of genetically engineered mice with HER2 exon 20G776>VC insertion mutation using the LoxP-STOP-LoxP system. B, Different time points of tumor volume quantification on MRI of HER2 exon 20G776>VC–mutant mice after induction. C, Representative image of hematoxylin and eosin (H&E) staining and immunochemistry of TTF1, p63, and SPC on HER2 exon 20G776>VC–mutant tumors. D, Dot plot of tumor volume changes on MRI from HER2 exon 20G776>VC insertion–mutant GEMMs treated with mobocertinib for 2 and 4 weeks. E, Tumor volume change for up to 16 weeks on MRI from HER2 exon 20G776>VC insertion–mutant GEMMs treated with mobocertinib. ****, P < 0.0001.
Figure 3.
Figure 3.
HER2 exon 20YVMA GEMM initially responded to mobocertinib but became acquired resistant upon continuous treatment. A, Tumor volume change on MRI from HER2 exon 20YVMA insertion–mutant GEMMs treated with mobocertinib. B, Representative immunochemistry image and quantification of HER2 protein on HER2 exon 20YVMA insertion–mutant GEMMs treated with vehicle or mobocertinib together with the acquired resistant tumors. C, Representative immunochemistry image and quantification of pHER2 protein on HER2 exon 20YVMA insertion–mutant GEMMs treated with vehicle or mobocertinib together with the acquired resistant tumors. D, Representative immunochemistry image and quantification of pERK protein on HER2 exon 20YVMA insertion–mutant GEMMs treated with vehicle or mobocertinib together with the acquired resistant tumors. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
Figure 4.
Figure 4.
Pathways enriched in HER2 exon 20YVMA acquired resistant tumors and mobocertinib increased HER2 expression at the cell membrane. A, Dot plot of enriched pathways (NOM P < 0.05 and FDR q < 0.25) in GSEA result from mobocertinib acquired resistant tumors versus response (treated for 3 days) tumor nodules. B, Enrichment plot of “G2M Checkpoint” gene set. C, Enrichment plot of “Mitotic Spindle” gene set. D, Enrichment plot of “mTOR1 Signaling” gene set. E, Mean fluorescence intensity of surface HER2 receptor on H1781 (HER2G776>VC) cell line treated with 0.01, 0.1, and 1 μmol/L mobocertinib using flow cytometry. F, Mean fluorescence intensity of surface HER2 receptor on the Ba/F3 HER2 exon 20YVMA cell line treated with 0.01, 0.1, and 1 μmol/L mobocertinib using flow cytometry. G, Relative mRNA change of HER2 on Ba/F3-HER2 exon 20YVMA cell line treated with 0.01, 0.1, and 1 μmol/L mobocertinib using real-time PCR. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
Figure 5.
Figure 5.
Mobocertinib and T-DM1 combination is the most effective among all three combinations. A, Dot plot of tumor volume change on MRI at the time point of weeks 2, 4, and 6 from HER2 exon 20YVMA insertion–mutant GEMMs treated with monotherapy (mobocertinib, T-DM1, alisertib, and sapanisertib) and combinations (mobocertinib combined with T-DM1, alisertib, and sapanisertib). B, Representative MRI image of HER2 exon 20YVMA insertion–mutant GEMMs treated with mobocertinib and T-DM1 combination. C, Survival curve of HER2 exon 20YVMA insertion–mutant GEMMs treated with vehicle, mobocertinib, T-DM1, and combination. D, Tumor volume change of acquired resistant HER2 exon 20YVMA insertion–mutant tumors treated with monotherapies (mobocertinib and T-DM1) and combinations (mobocertinib plus alisertib or T-DM1). E, Representative MRI image of acquired resistant HER2 exon 20YVMA insertion–mutant tumors treated with mobocertinib and T-DM1 combination therapy. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; NS, nonsignificant.
Figure 6.
Figure 6.
M1 macrophage infiltration and CD4+ T-cell activation contributed to the synergic effect of mobocertinib and T-DM1 combination. A, Dot plot of enriched pathways (NOM P < 0.05 and FDR q < 0.25) in GSEA resulting from mobocertinib and T-DM1–treated tumors versus response (treated for 3 days) tumor nodules. B, Heat map of 22 different immune cell populations from RNA-seq data of tumor nodules treated with vehicle, mobocertinib, and mobocertinib plus T-DM1 combination using CIBERSORT analysis. C–H, Comprehensive immuno-profiling of HER2 exon 20YVMA insertion-mutant GEMMs treated with vehicle, mobocertinib, T-DM1, and combination therapy using multiparameter flow cytometry. I, Multi-immunofluorescence of DAPI, TTF1, and F4/80 on mobocertinib and T-DM1–treated tumor. J, Dot plot of tumor volume change on MRI from HER2 exon 20YVMA insertion–mutant GEMMs treated with vehicle, Clodrosome, mobocertinib, mobocertinib plus T-DM1, and mobocertinib combined with T-DM1 plus Clodrosome. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; NS, nonsignificant.

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