SOX4 Mediates BRAF Inhibitor Resistance in Melanoma through Regulation of IGF-1R Signaling: In Vitro Study
Objective::SOX4, a transcription factor, has been found to contribute to tumorigenesis in several cancers. This study was performed to determine whether SOX4 mediates BRAF inhibitor resistance in melanoma.Methods::Melanoma cell lines with acquired resistance to BRAF inhibitor (SK-MEL-5R, SK-MEL-28R, and A375R) were generated by adding escalating concentrations of PLX4032 into parental SK-MEL-5, SK-MEL-28, and A375 cells for >6 months. The expression of SOX4 and insulin-like growth factor 1 receptor (IGF-1R) was measured by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. The downstream signaling of IGF-1R was detected by Western blotting. SOX4 and IGF-1R overexpression or knockdown was conducted by lentivirus transfection. Cell viability and apoptosis were demonstrated by MTT and flow cytometry, respectively. The binding ability of SOX4 to IGF-1R promoter was determined by chromatin immunoprecipitation quantitative PCR assay.Results::SOX4 was upregulated in BRAF inhibitor-resistant melanoma cells as compared with parental cells (SK-MEL-5 group, 1.02
vs. 6.33; SK-MEL-28 group, 1.03
vs. 3.22; A375 group, 1.00
vs. 1.86;
t=°7.069, 29.26, and 5.291, respectively; all
P < 0.01), and PLX4032 treatment could not alter the expression of SOX4 in resistant cells. SOX4 overexpression attenuated the response of parental cells to PLX4032 (for cell viability, SK-MEL-5 group: 77.76%
vs. 104.28%,
F= 91.50; SK-MEL-28 group: 60.59%
vs. 93.13%,
F= 171.8; A375 group: 62.50%
vs. 80.87%,
F= 47.15. For apoptosis rates, SK-MEL-5 group: 34.90%
vs. 14.31%,
F= 4.781; SK-MEL-28 group, 40.8%
vs. 29.4%,
F= 13.32,
P= 0.063; A375 group: 40.20%
vs. 17.09%,
F= 11.39; all
P < 0.05, otherwise indicated). While SOX4 knockdown enhanced the response of resistant cells to PLX4032 (for cell viability, SK-MEL-5R group: 93.75%
vs. 69.53%,
F= 94.45, SK-MEL-28R group: 95.60%
vs. 66.79%,
F= 30.41, A375R group: 95.51%
vs. 59.98%,
F= 111.6; for apoptosis rates, SK-MEL-5R group: 16.2%
vs. 44.4%,
F= 25.67, SK-MEL-28R group: 26.59%
vs. 44.20%,
F= 158.0, A375R group: 5.98%
vs. 31.51 %,
F= 14.35, and all
P < 0.01). Chromatin immunoprecipitation quantitative PCR assay demonstrated that SOX4 binded to the promoter of IGF-1R (1.04
vs. 1.94 [-1044 to -920 bp] and 0.110
vs. 0.139 [GAPDH],
F= 534.5,
P < 0.01). In addition, SOX4 overexpression increased IGF-1R and its downstream phosphorylated ERK, phosphorylated AKT, and phosphorylated STAT3 expression, while SOX4 knockdown exerted the opposite effects. Moreover, IGF-1R knockdown overcame SOX4 overexpression-induced PLX4032 resistance (cell viability: 35.85%
vs. 52.79%
vs. 37.84% [A375 group, negative control group
vs. SOX4 overexpressing group
vs. SOX4 overexpressing + sh-IGF-1R group]; apoptosis rates: 25.30%
vs. 9.56%
vs. 22.26 [A375 group, negative control group
vs. SOX4 overexpressing group
vs. SOX4 overexpressing+ sh-IGF-1R group];
F= 13.01 and 41.87, respectively; all
P < 0.01), while IGF-1R overexpression abrogated SOX4 knockdown-induced response enhancement to PLX4032 for comparison of negative control group, sh-SOX4 group and sh-SOX4 + IGF-1R overexpressing group (cell viability: 96.62%
vs. 86.86%
vs. 97.26% (A375R), 98.15%
vs. 81.63%
vs. 98.49% [SK-MEL-5R]; apoptosis rates: 13.81%
vs. 32.00%
vs. 12.16 [A375R], 29.70%
vs. 41.40%
vs. 26.10% [SK-MEL-5R];
F= 13.56, 12.86, 38.81, and 39.85, respectively; all
P < 0.01).
Conclusion::SOX4 mediates BRAF inhibitor resistance in melanoma through regulation of IGF-1R signaling. SOX4 might serve as a potential target for the treatment of BRAF inhibitor-resistant melanoma.
SOX4、melanoma、BRAF inhibitor、drug resistance、IGF-1R
03
This work was supported by grants from the National Natural Science Foundation of ChinaNo. 81673917 and No. 81973582;Guangdong Basic and Applied Basic Research FoundationNo. 2019A1515110833;Fundamental Research Funds of Shandong UniversityNo. 2019GN043;This work was supported by grants from the National Natural Science Foundation of ChinaNo. 81673917 and No. 81973582;Guangdong Basic and Applied Basic Research FoundationNo. 2019A1515110833;Fundamental Research Funds of Shandong UniversityNo. 2019GN043
2023-05-30（万方平台首次上网日期，不代表论文的发表时间）
共10页
156-165
