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Flavopiridol synergizes with sorafenib to induce cytotoxicity and potentiate antitumorigenic activity in EGFR/HER-2 and mutant RAS/RAF breast cancer model systems.

ABSTRACT: Oncogenic receptor tyrosine kinase (RTK) signaling through the Ras-Raf-Mek-Erk (Ras-MAPK) pathway is implicated in a wide array of carcinomas, including those of the breast. The cyclin-dependent kinases (CDKs) are implicated in regulating proliferative and survival signaling downstream of this pathway. Here, we show that CDK inhibitors exhibit an order of magnitude greater cytotoxic potency than a suite of inhibitors targeting RTK and Ras-MAPK signaling in cell lines representative of clinically recognized breast cancer (BC) subtypes. Drug combination studies show that the pan-CDK inhibitor, flavopiridol (FPD), synergistically potentiated cytotoxicity induced by the Raf inhibitor, sorafenib (SFN). This synergy was most pronounced at sub-EC50 SFN concentrations in MDA-MB-231 (KRAS-G13D and BRAF-G464V mutations), MDA-MB-468 [epidermal growth factor receptor (EGFR) overexpression], and SKBR3 [ErbB2/EGFR2 (HER-2) overexpression] cells but not in hormone-dependent MCF-7 and T47D cells. Potentiation of SFN cytotoxicity by FPD correlated with enhanced apoptosis, suppression of retinoblastoma (Rb) signaling, and reduced Mcl-1 expression. SFN and FPD were also tested in an MDA-MB-231 mammary fat pad engraftment model of tumorigenesis. Mice treated with both drugs exhibited reduced primary tumor growth rates and metastatic tumor load in the lungs compared to treatment with either drug alone, and this correlated with greater reductions in Rb signaling and Mcl-1 expression in resected tumors. These findings support the development of CDK and Raf co-targeting strategies in EGFR/HER-2-overexpressing or RAS/RAF mutant BCs.

SUBMITTER: Nagaria TS

PROVIDER: S-EPMC3730045 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

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Flavopiridol synergizes with sorafenib to induce cytotoxicity and potentiate antitumorigenic activity in EGFR/HER-2 and mutant RAS/RAF breast cancer model systems.

Nagaria Teddy S TS Williams Julia L JL Leduc Charles C Squire Jeremy A JA Greer Peter A PA Sangrar Waheed W

Neoplasia (New York, N.Y.) 20130801 8

Oncogenic receptor tyrosine kinase (RTK) signaling through the Ras-Raf-Mek-Erk (Ras-MAPK) pathway is implicated in a wide array of carcinomas, including those of the breast. The cyclin-dependent kinases (CDKs) are implicated in regulating proliferative and survival signaling downstream of this pathway. Here, we show that CDK inhibitors exhibit an order of magnitude greater cytotoxic potency than a suite of inhibitors targeting RTK and Ras-MAPK signaling in cell lines representative of clinically ...[more]

PMID: 23908594

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Project description:Background Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) are standard treatment for advanced non-small cell lung cancer (NSCLC). However, the emergence of EGFR-TKIs resistance poses a big challenge to the treatment. Although several resistant mutations have been identified, our understanding of the mechanisms underlying acquired EGFR-TKIs resistance remains incomplete. This study aimed to identify novel mutations and mechanisms that could contribute to acquired EGFR-TKIs resistance in EGFR mutated NSCLC cells. Methods Erlotinib resistant cells (HCC827/ER cells) were generated from the EGFR mutated NSCLC cell line HCC827, and whole-exome sequencing was performed to identify gene mutations in HCC827/ER cells. The Spred-3 expression was determined using quantitative real-time PCR (qPCR) and Western blotting assays, and the p-p44/42, p44/42, p-Akt and Akt expression was determined using Western blotting. The half maximal inhibitory concentration (IC50 value) was measured using the MTS assay, and cell migration was detected with a Transwell migration assay. Results Whole-exome sequencing identified deletion mutation c.120delG at exon 1 of the Spred-3 gene, resulting in a p.E40fs change in amino acid, in HCC827/ER cells. The Spred-3 expression was much reduced in HCC827/ER cells as compared to the HCC827 cells at both mRNA and protein levels. Knocking out Spred-3 in HCC827 cells using CRISPR/Cas9 increased erlotinib resistance and cell migration, while overexpressing Spred-3 in HCC827/ER cells using a cDNA construct reduced erlotinib resistance and cell migration. We also showed the Ras/Raf/MAPK pathway was activated in HCC827/ER cells, and inhibiting ERK1/2 in HCC827/Spred-3-sgRNA cells resulted in reduced erlotinib resistance and cell migration. Conclusions The results of this study indicate that a loss-of-function mutation in Spred-3 resulted in activation of the Ras/Raf/MAPK pathway that confers resistance to EGFR-TKIs in NSCLC cells harboring an EGFR mutation.

Dataset Information

Flavopiridol synergizes with sorafenib to induce cytotoxicity and potentiate antitumorigenic activity in EGFR/HER-2 and mutant RAS/RAF breast cancer model systems.

Publications

Flavopiridol synergizes with sorafenib to induce cytotoxicity and potentiate antitumorigenic activity in EGFR/HER-2 and mutant RAS/RAF breast cancer model systems.

Similar Datasets

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