Integrated mRNA and miRNA Transcriptomic Analyses Reveals Divergent Mechanisms of Sunitinib Resistance in Clear Cell Renal Cell Carcinoma (ccRCC)
Abstract
:Simple Summary
Abstract
1. Introduction
2. Results
2.1. Generating In Vitro Models of Sunitinib Resistance
2.2. Non-Coding RNA and Gene Expression in Sunitinib-Resistant Cells
2.3. Interaction Network Analysis between Differentially Expressed Genes and miRNAs
2.4. Gene Ontology and Pathway Analyses
2.5. Silencing of PD-L1 in 786-O Sunitinib-Resistant Cells Results in an Increased Sensitivity to Sunitinib
3. Discussion
4. Materials and Methods
4.1. Generation of Sunitinib-Resistant ccRCC Cell Lines
4.2. Cell Proliferation Assay
4.3. RNA Extraction and Microarray Analysis
4.4. Interaction Network Analysis
4.5. Quantitative RT-PCR (qRT-PCR)
4.6. PDL-1 Silencing
4.7. Western Blotting
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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IC50 Value | ||||
---|---|---|---|---|
Cell Line | Biological Replicate | Parental | Clone c1 (p-Value) | Clone c2 (p-Value) |
786-O | A | 5.7 | 17.32 | 13.54 |
B | 11.39 | 23.3 | 20.07 | |
C | 6.7 | 20.4 | 12.8 | |
Average | 7.93 | 20.34 ** | 15.47 * | |
A498 | A | 10.48 | 11.92 | 14.36 |
B | 7.682 | 12.42 | 16.04 | |
C | 7.18 | 12.01 | 10.54 | |
Average | 8.44 | 12.12 * | 13.64 * | |
Caki-1 | A | 9.5 | 16.8 | 18 |
B | 9 | 14.6 | 17.3 | |
C | 12.2 | 16.2 | 14.7 | |
Average | 10.2 | 15.87 ** | 16.67 * |
Cell | miRNA/Gene | miRNA | Target Gene(s) |
---|---|---|---|
786-0 | ↑/↓ | hsa-miR-663a | CDKN1A |
↑/↓ | hsa-miR-572 | CDKN1A | |
↑/↓ | hsa-miR-638 | PTEN | |
↑/↓ | hsa-miR-612 | SP1 | |
↑/↓ | hsa-miR-212-3p | RFXAP | |
↓/↑ | hsa-miR-106a-5p | SIRPA, MMP2, HIF1A | |
↓/↑ | hsa-miR-30d-5p | KPNB1, BNIP3L, BECN1 | |
↓/↑ | hsa-miR-140-3p | ATP6AP2, FN1 | |
↓/↑ | hsa-miR-26b-5p | PTGS2, ST8SIA4 | |
↓/↑ | hsa-miR-17-5p | MMP2, SIRPA, GPR137B, EPAS1, VLDLR, HIF1A | |
↓/↑ | hsa-miR-200a-3p | WASF3,CD274 | |
↓/↑ | hsa-miR-200b-3p | LOX, FN1, CD274, WASF3, MSN, FERMT2, FSCN1, RAB23 | |
↓/↑ | hsa-miR-210-3p | HIF1A, BDNF, NCAM1, EHD2, TFRC | |
↓/↑ | hsa-miR-328-3p | PTPRJ, MMP16 | |
↓/↑ | hsa-miR-34a-5p | BECN1, VAMP2, FUT8, INHBB, NOTCH2, MAGEA2, MAGEA3, L1CAM, AXL, PAM, SYT1, CD274 | |
↓/↑ | hsa-miR-21-5p | BASP1, RECK, NFIB, SATB1, RHOB, PIAS3, DUSP10, LRP6 | |
↓/↑ | hsa-miR-146a-5p | L1CAM, NOTCH2, PTGS2, HOXD10, RAC1 | |
↓/↑ | hsa-miR-20a-5p | SIRPA, EPAS1, KIF26B, CRIM1, HIF1A, TSG101 | |
↓/↑ | hsa-miR-17-3p | ITGA5, ITGB3 | |
↓/↑ | hsa-miR-99a-5p | SMARCA5 | |
↓/↑ | hsa-miR-18a-5p | PIAS3, TNFAIP3, HIF1A, TBPL1 | |
↓/↑ | hsa-miR-25-5p | PRKCZ | |
A498 | ↑/↓ | hsa-miR-34c-5p | ITPR1, HNF4A |
↓/↑ | hsa-miR-145-5p | ITGB8, CTGFL, VPS51, EGFR | |
Caki-1 | ↑/↓ | hsa-miR-148b-3p | SLC2A1 |
↑/↓ | hsa-miR-192-5p | ITGB3, ITGAV, CAV1, WNK1 | |
↑/↓ | hsa-miR-29b-3p | DNMT3B | |
↓/↑ | hsa-miR-138-5p | CCND1 | |
↓/↑ | hsa-miR-193b-3p | CCND1, AKR1C2 | |
↓/↑ | hsa-miR-92a-5p | KLF2 | |
↓/↑ | hsa-miR-296-3p | ICAM1 | |
↓/↑ | hsa-miR-106b-5p | BMP2, RND3, CCND1 | |
↓/↑ | hsa-miR-106b-3p | BMP2 | |
↓/↑ | hsa-miR-130b-3p | IRF1 | |
↓/↑ | hsa-miR-708-5p | CCND1 | |
↓/↑ | hsa-miR-18a-5p | TNFAIP3, CTGF | |
↓/↑ | hsa-miR-17-5p | CCND1, BMP2, TCEAL1, RND3 | |
↓/↑ | hsa-miR-1180-3p | TCEAL1 | |
↓/↑ | hsa-miR-550a-5p | CPEB4 |
Cell Line | 786-O P | 786-O c1 | 786-O c2 | Caki-1 P | Caki-1 c1 | Caki-1 c2 | A498 P | A498 c1 | A498 c2 |
---|---|---|---|---|---|---|---|---|---|
PD-L1 | 100 | 161.3 | 131.3 | 100 | 16.9 | 35.8 | 100 | 1599.3 | 966.4 |
HIF1α | ND | ND | ND | ND | ND | ND | 100 | 18.3 | 14.8 |
Cyclin D1 | 100 | 119.9 | 77.5 | 100 | 56.5 | 34.9 | 100 | 122.0 | 338.8 |
HIF2α | 100 | 45.8 | 24.2 | ND | ND | ND | 100 | 128.5 | 60.8 |
Cell Line | P-scr | P-si | c1-scr | c1-P | c2-scr | c2-P |
---|---|---|---|---|---|---|
A498 | 100 | 0 | 100 | 15.8 | 100 | 4.5 |
786-O | 100 | 35.7 | 100 | 28.5 | 100 | 20.2 |
Cell Line | Replicate | P-SCR | P-siRNA | c1-SCR | c1-siRNA | c2-SCR | c2- siRNA |
---|---|---|---|---|---|---|---|
786-O | A | 5.9 | 4.2 | 11.0 | 6.5 | 10.6 | 6.6 |
B | 3.8 | 4.9 | 11.7 | 9.6 | 8.2 | 6.5 | |
C | 4.7 | 6.8 | 12.3 | 9.5 | 9.4 | 8.0 | |
Average | 4.8 | 5.3 | 11.7 | 8.5 * | 9.4 | 7.0 * | |
A498 | A | 5.5 | 6.31 | 6.7 | 7.7 | 9.6 | 13.3 |
B | 9.8 | 11.8 | 10.6 | 12.8 | 10.2 | 11.4 | |
C | 14.1 | 14.8 | 13 | 15.4 | 15.3 | 20.6 | |
Average | 9.8 | 11 | 10.1 | 12 | 11.7 | 15.1 | |
Caki-1 | A | 9.3 | 10.8 | 9 | 11.4 | 10 | 11.5 |
B | 6.4 | 6.7 | 9.5 | 16.1 | 11 | 11.9 | |
C | 9.7 | 9.2 | 11 | 11.4 | 12 | 12.4 | |
Average | 8.4 | 8.9 | 9.8 | 13 | 11 | 11.9 |
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Armesto, M.; Marquez, M.; Arestin, M.; Errarte, P.; Rubio, A.; Manterola, L.; López, J.I.; Lawrie, C.H. Integrated mRNA and miRNA Transcriptomic Analyses Reveals Divergent Mechanisms of Sunitinib Resistance in Clear Cell Renal Cell Carcinoma (ccRCC). Cancers 2021, 13, 4401. https://doi.org/10.3390/cancers13174401
Armesto M, Marquez M, Arestin M, Errarte P, Rubio A, Manterola L, López JI, Lawrie CH. Integrated mRNA and miRNA Transcriptomic Analyses Reveals Divergent Mechanisms of Sunitinib Resistance in Clear Cell Renal Cell Carcinoma (ccRCC). Cancers. 2021; 13(17):4401. https://doi.org/10.3390/cancers13174401
Chicago/Turabian StyleArmesto, María, Maitane Marquez, María Arestin, Peio Errarte, Ane Rubio, Lorea Manterola, Jose I. López, and Charles H. Lawrie. 2021. "Integrated mRNA and miRNA Transcriptomic Analyses Reveals Divergent Mechanisms of Sunitinib Resistance in Clear Cell Renal Cell Carcinoma (ccRCC)" Cancers 13, no. 17: 4401. https://doi.org/10.3390/cancers13174401