Additional file 1 of DNA replication stress and mitotic catastrophe mediate sotorasib addiction in KRASG12C-mutant cancer
Creators: name: Chiou, Li-Wen
name: Chan, Chien-Hui
name: Jhuang, Yu-Ling
name: Yang, Ching-Yao
name: Jeng, Yung-Ming
Publisher: name: figshare
Year: 2023
Subjects: subject: Biophysics
subject: Biochemistry
subject: Cell Biology
subject: Molecular Biology
subject: Pharmacology
subject: Biotechnology
classificationCode: 39999
subject: Chemical Sciences not elsewhere classified
subject: Cancer
subject: Hematology
classificationCode: 110309
subject: Infectious Diseases
subject: FOS: Health sciences

description: Additional file 1: Table S1. Antibodies used for Western blotting. Table S2. Antibodies used for immunofluorescence staining. Table S3. STR profiling of MIA PaCa-2 and MIA-SR. Table S4. STR profiling of NCI-H23 and H23-SR. Table S5. STR profiling of LU65 and 65-SR. Table S6. Non-silent genetic variants present in sotorasib-resistant cells but not present in parental cells identified by whole exome sequencing. Figure S1. Relative growth rates of MIA-PaCa-2, NCI-H23, and LU65 cells and their sotorasib-resistantsublines MIA-SR, H23-SR, and 65-SR cultured in the different concentrations of sotorasib for 3 days. Data are presented in terms of the fold-change values relative to the growth rates of cells cultured without sotorasib. ****P &lt; 0.0001. Figure S2. Relative growth rates of the 65-SR cells cultured with or without 5 μM sotorasib. Data were normalized to the cell numbers noted after the 10-day-long treatment. Figure S3. A Quantification of the total ROS levels in the MIA-SR, H23-SR, and 65-SR cells cultured with or without 5 μM sotorasib for 3 days; the levels were measured through CellROX staining and flow cytometry. B The MIA-SR and H23-SR cells were cultured with or without 5 μM sotorasib in the presence of 2 mM NAC. The cells were counted on Day 11. Data are presented in terms of the mean ± standard deviation values of three cell culture replicates. ***P &lt; 0.001, and ****P &lt; 0.0001. Figure S4. Phase-contrast microscopy images of the MIA-SR and H23-SR cells cultured with or without 5 μM sotorasib for 8 days. Figure S5. Time-lapse microscopy images of the H23-SR cells cultured with or without 5 μM sotorasib for the indicated days. The yellow arrows denote mitotic cells. Figure S6. Effects of encorafenib on levels of p-ERK, ERK, p-RPA32, p-CHEK1, and p21 in the H23-SR cells levels with or without 5 μM sotorasib; protein levels were measured through Western blotting. Figure S7. Representative images of immunofluorescence staining and quantification of nucleoplasmic bridges and micronuclei in the H23-SR cells cultured with or without 5 μM sotorasib, 100 nM encorafenib, or both. The yellow arrows denote the locations of micronucleior nucleoplasmic bridges. *P &lt; 0.05, ***P &lt; 0.001, and ****P &lt; 0.0001. Figure S8. Results of annexin V/PI flow cytometric assay on the H23-SR cells cultured with or without 5 μM sotorasib, 100 nM encorafenib, or both. Figure S9. Effects of sotorasib withdrawal on the levels of p-MEK1/2 and p-ERK in the 65-SR cells; protein levels were measured through Western blotting. Figure S10. Effects of encorafenib on tumor sizes and body weights of experimental mice. The 65-SR cells were subcutaneously implanted into the flanks of 6-week-old female NOD/SCID mice. Sotorasib was orally administered at 10 mg/kg/day to all mice one day before injection and throughout the experiment. After the mean tumor volume reached at least 100 mm3, the mice were treated with encorafenib or vesicle control treated for 14 days. Tumor sizes and body weights of the mice were measured at the indicated time point.