Yan Gao1, Ning Wu1,2, Shuai Wang1, Xue Yang1, Xin Wang2, Bo Xu3,4. 1. Department of Biochemistry and Molecular Biology, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Tianjin, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin, 300060, China. 2. The First Department of Breast Cancer, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Tianjin, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin, 300060, China. 3. Department of Biochemistry and Molecular Biology, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Tianjin, Tianjin's Clinical Research Center for Cancer, Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin, 300060, China. xubo@tmu.edu.cn. 4. Center for Intelligent Oncology, Chongqing University Cancer Hospital, Chongqing University School of Medicine, Chongqing, 400030, China. xubo@tmu.edu.cn.
Abstract
PURPOSE: HER2-positive breast cancer patients benefit from HER2-targeted therapies, among which the most commonly used is trastuzumab. However, acquired resistance typically happens within one year. The cellular heterogeneity of it is less clear. METHODS: Here we generated trastuzumab-resistant cells in two HER2-positive breast cancer cell lines, SK-BR-3 and BT-474. Cells at different time points during the resistance induction were examined by exome sequencing to study changes of genomic alterations over time. Single cell-targeted sequencing was also used to identify resistance-associated concurrent mutations. RESULTS: We found a rapid increase of copy number variation (CNV) regions and gradual accumulation of single nucleotide variations (SNVs). On the pathway level, non-synonymous SNVs for SK-BR-3 cells were enriched in the MAPK signaling pathway, while for BT-474 cells they were enriched in mTOR and PI3K-Akt signaling pathways. However, all of the three signaling pathways were in the downstream of the HER2 kinase. Putative trastuzumab-resistance-associated SNVs included AIFM1 P548L and ERBB2 M833R in SK-BR-3 cells, and ADAMTS19 V451L, OR5M9 D230N, COL9A1 R627T, and ITGA7 H911Q in BT-474 cells. Single-cell-targeted sequencing identified several concurrent mutations. By validation, we found that concurrent mutations (AIFM1 P548L and IL1RAPL2 S546C in SK-BR-3 cells, MFSD11 L242I and ANAPC4 E16K in BT-474 cells) led to a decrease of trastuzumab sensitivity. CONCLUSION: Taken together, our study revealed a common pathway level trastuzumab-resistance mechanism for HER2-positive breast cancer cells. In addition, our identification of concurrent SNVs associated with trastuzumab-resistance may be indicative of potential targets for the treatment of trastuzumab-resistant breast cancer patients.
PURPOSE:HER2-positive breast cancerpatients benefit from HER2-targeted therapies, among which the most commonly used is trastuzumab. However, acquired resistance typically happens within one year. The cellular heterogeneity of it is less clear. METHODS: Here we generated trastuzumab-resistant cells in two HER2-positive breast cancer cell lines, SK-BR-3 and BT-474. Cells at different time points during the resistance induction were examined by exome sequencing to study changes of genomic alterations over time. Single cell-targeted sequencing was also used to identify resistance-associated concurrent mutations. RESULTS: We found a rapid increase of copy number variation (CNV) regions and gradual accumulation of single nucleotide variations (SNVs). On the pathway level, non-synonymous SNVs for SK-BR-3 cells were enriched in the MAPK signaling pathway, while for BT-474 cells they were enriched in mTOR and PI3K-Akt signaling pathways. However, all of the three signaling pathways were in the downstream of the HER2 kinase. Putative trastuzumab-resistance-associated SNVs included AIFM1P548L and ERBB2M833R in SK-BR-3 cells, and ADAMTS19V451L, OR5M9 D230N, COL9A1R627T, and ITGA7H911Q in BT-474 cells. Single-cell-targeted sequencing identified several concurrent mutations. By validation, we found that concurrent mutations (AIFM1P548L and IL1RAPL2S546C in SK-BR-3 cells, MFSD11L242I and ANAPC4E16K in BT-474 cells) led to a decrease of trastuzumab sensitivity. CONCLUSION: Taken together, our study revealed a common pathway level trastuzumab-resistance mechanism for HER2-positive breast cancer cells. In addition, our identification of concurrent SNVs associated with trastuzumab-resistance may be indicative of potential targets for the treatment of trastuzumab-resistant breast cancerpatients.
Entities:
Keywords:
Concurrent mutations; HER2-positive breast cancer; Single cell sequencing; Trastuzumab-resistance
Authors: Edith A Perez; Edward H Romond; Vera J Suman; Jong-Hyeon Jeong; George Sledge; Charles E Geyer; Silvana Martino; Priya Rastogi; Julie Gralow; Sandra M Swain; Eric P Winer; Gerardo Colon-Otero; Nancy E Davidson; Eleftherios Mamounas; Jo Anne Zujewski; Norman Wolmark Journal: J Clin Oncol Date: 2014-10-20 Impact factor: 44.544
Authors: Sandra M Swain; José Baselga; Sung-Bae Kim; Jungsil Ro; Vladimir Semiglazov; Mario Campone; Eva Ciruelos; Jean-Marc Ferrero; Andreas Schneeweiss; Sarah Heeson; Emma Clark; Graham Ross; Mark C Benyunes; Javier Cortés Journal: N Engl J Med Date: 2015-02-19 Impact factor: 91.245
Authors: Sandra M Swain; Sung-Bae Kim; Javier Cortés; Jungsil Ro; Vladimir Semiglazov; Mario Campone; Eva Ciruelos; Jean-Marc Ferrero; Andreas Schneeweiss; Adam Knott; Emma Clark; Graham Ross; Mark C Benyunes; José Baselga Journal: Lancet Oncol Date: 2013-04-18 Impact factor: 41.316
Authors: Antonio C Wolff; M Elizabeth H Hammond; David G Hicks; Mitch Dowsett; Lisa M McShane; Kimberly H Allison; Donald C Allred; John M S Bartlett; Michael Bilous; Patrick Fitzgibbons; Wedad Hanna; Robert B Jenkins; Pamela B Mangu; Soonmyung Paik; Edith A Perez; Michael F Press; Patricia A Spears; Gail H Vance; Giuseppe Viale; Daniel F Hayes Journal: J Clin Oncol Date: 2013-10-07 Impact factor: 44.544