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KRAS oncogene repression in colon cancer cell lines by G-quadruplex binding indolo[3,2- c ]quinolines

View Article: PubMed Central - PubMed

ABSTRACT

KRAS is one of the most frequently mutated oncogenes in human cancer, yet remaining undruggable. To explore a new therapeutic strategy, a library of 5-methyl-indolo[3,2-c]quinoline derivatives (IQc) with a range of alkyldiamine side chains was designed to target DNA and RNA G-quadruplexes (G4) in the promoter and 5′-UTR mRNA of the KRAS gene. Biophysical experiments showed that di-substituted IQc compounds are potent and selective KRAS G4 stabilizers. They preferentially inhibit the proliferation of KRAS mutant cancer cell lines (0.22 < IC50 < 4.80 μM), down-regulate KRAS promoter activity in a luciferase reporter assay, and reduce both KRAS mRNA and p21KRAS steady-state levels in mutant KRAS colon cancer cell lines. Additionally, IQcs induce cancer cell death by apoptosis, explained in part by their capacity to repress KRAS expression. Overall, the results suggest that targeting mutant KRAS at the gene level with G4 binding small molecules is a promising anticancer strategy.

No MeSH data available.


Related in: MedlinePlus

KRAS mRNA and protein steady-state expression after exposure of HCT116 and SW620 cells lines to equitoxic (IC50) concentrations of compounds TMPyP4, 2d, 3d and 3e for 72 h.(a) KRAS mRNA steady-state expression was evaluated by Taqman Real-time RT-PCR using specific Taqman Assays for KRAS and β-Actin for normalization. KRAS mRNA steady-state expression levels were calculated by the ΔΔCt method, using DMSO (vehicle control) for calibration; and (b) KRAS protein steady-state expression evaluated by immunoblot relative to control (DMSO vehicle) Results are expressed as mean ± SEM of at least three independent experiments; *p < 0.05 and §p < 0.01 from DMSO (vehicle control).
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f6: KRAS mRNA and protein steady-state expression after exposure of HCT116 and SW620 cells lines to equitoxic (IC50) concentrations of compounds TMPyP4, 2d, 3d and 3e for 72 h.(a) KRAS mRNA steady-state expression was evaluated by Taqman Real-time RT-PCR using specific Taqman Assays for KRAS and β-Actin for normalization. KRAS mRNA steady-state expression levels were calculated by the ΔΔCt method, using DMSO (vehicle control) for calibration; and (b) KRAS protein steady-state expression evaluated by immunoblot relative to control (DMSO vehicle) Results are expressed as mean ± SEM of at least three independent experiments; *p < 0.05 and §p < 0.01 from DMSO (vehicle control).

Mentions: The effects of compounds on KRAS transcription in colon cancer cells (HCT116 and SW620) were evaluated by quantifying KRAS mRNA steady-state levels using Taqman real-time RT-PCR. The results (Figure 6a) show that IQcs significantly reduced KRAS transcription in colon cancer cells. The extent of the effect of 3e on KRAS mRNA steady-state levels depends on cell phenotype, whereas the effects of 2d and 3d do not, since these two IQcs were able to reduce KRAS mRNA steady-state levels by 90% (2d) and 25% (3d) in both cell lines. TMPyP4 at 12 μM showed no ability to repress KRAS transcription in HCT116 cell line, despite the promoter activity reduction observed in the dual-reporter assay. It must be noted that the effect of TMPyP4 on SW620 cells could not be compared as this cell line showed reduced sensitivity to this compound (IC50 > 20 μM).


KRAS oncogene repression in colon cancer cell lines by G-quadruplex binding indolo[3,2- c ]quinolines
KRAS mRNA and protein steady-state expression after exposure of HCT116 and SW620 cells lines to equitoxic (IC50) concentrations of compounds TMPyP4, 2d, 3d and 3e for 72 h.(a) KRAS mRNA steady-state expression was evaluated by Taqman Real-time RT-PCR using specific Taqman Assays for KRAS and β-Actin for normalization. KRAS mRNA steady-state expression levels were calculated by the ΔΔCt method, using DMSO (vehicle control) for calibration; and (b) KRAS protein steady-state expression evaluated by immunoblot relative to control (DMSO vehicle) Results are expressed as mean ± SEM of at least three independent experiments; *p < 0.05 and §p < 0.01 from DMSO (vehicle control).
© Copyright Policy - open-access
Related In: Results  -  Collection

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f6: KRAS mRNA and protein steady-state expression after exposure of HCT116 and SW620 cells lines to equitoxic (IC50) concentrations of compounds TMPyP4, 2d, 3d and 3e for 72 h.(a) KRAS mRNA steady-state expression was evaluated by Taqman Real-time RT-PCR using specific Taqman Assays for KRAS and β-Actin for normalization. KRAS mRNA steady-state expression levels were calculated by the ΔΔCt method, using DMSO (vehicle control) for calibration; and (b) KRAS protein steady-state expression evaluated by immunoblot relative to control (DMSO vehicle) Results are expressed as mean ± SEM of at least three independent experiments; *p < 0.05 and §p < 0.01 from DMSO (vehicle control).
Mentions: The effects of compounds on KRAS transcription in colon cancer cells (HCT116 and SW620) were evaluated by quantifying KRAS mRNA steady-state levels using Taqman real-time RT-PCR. The results (Figure 6a) show that IQcs significantly reduced KRAS transcription in colon cancer cells. The extent of the effect of 3e on KRAS mRNA steady-state levels depends on cell phenotype, whereas the effects of 2d and 3d do not, since these two IQcs were able to reduce KRAS mRNA steady-state levels by 90% (2d) and 25% (3d) in both cell lines. TMPyP4 at 12 μM showed no ability to repress KRAS transcription in HCT116 cell line, despite the promoter activity reduction observed in the dual-reporter assay. It must be noted that the effect of TMPyP4 on SW620 cells could not be compared as this cell line showed reduced sensitivity to this compound (IC50 > 20 μM).

View Article: PubMed Central - PubMed

ABSTRACT

KRAS is one of the most frequently mutated oncogenes in human cancer, yet remaining undruggable. To explore a new therapeutic strategy, a library of 5-methyl-indolo[3,2-c]quinoline derivatives (IQc) with a range of alkyldiamine side chains was designed to target DNA and RNA G-quadruplexes (G4) in the promoter and 5&prime;-UTR mRNA of the KRAS gene. Biophysical experiments showed that di-substituted IQc compounds are potent and selective KRAS G4 stabilizers. They preferentially inhibit the proliferation of KRAS mutant cancer cell lines (0.22 &lt; IC50 &lt; 4.80 &mu;M), down-regulate KRAS promoter activity in a luciferase reporter assay, and reduce both KRAS mRNA and p21KRAS steady-state levels in mutant KRAS colon cancer cell lines. Additionally, IQcs induce cancer cell death by apoptosis, explained in part by their capacity to repress KRAS expression. Overall, the results suggest that targeting mutant KRAS at the gene level with G4 binding small molecules is a promising anticancer strategy.

No MeSH data available.


Related in: MedlinePlus