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ARC (NSC 188491) has identical activity to Sangivamycin (NSC 65346) including inhibition of both P-TEFb and PKC.

Stockwin LH, Yu SX, Stotler H, Hollingshead MG, Newton DL - BMC Cancer (2009)

Bottom Line: In this study, the mechanism of action of ARC was further investigated by comparing in vitro and in vivo activity with other anti-neoplastic purines.Structure-based homology searches were used to identify those compounds with similarity to ARC.Results demonstrated that sangivamycin, an extensively characterized pro-apoptotic nucleoside isolated from Streptomyces, had identical activity to ARC in terms of 1) cytotoxicity assays, 2) ability to induce a G2/M block, 3) inhibitory effects on RNA/DNA/protein synthesis, 4) transcriptomic response to treatment, 5) inhibition of protein kinase C, 6) inhibition of positive transcription elongation factor b (P-TEFb), 7) inhibition of VEGF secretion, and 8) activity within hollow fiber assays.

View Article: PubMed Central - HTML - PubMed

Affiliation: Developmental Therapeutics Program, SAIC-Frederick Inc, NCI- Frederick, Frederick, MD 21702, USA. Stockwin@ncifcrf.gov

ABSTRACT

Background: The nucleoside analog, ARC (NSC 188491) is a recently characterized transcriptional inhibitor that selectively kills cancer cells and has the ability to perturb angiogenesis in vitro. In this study, the mechanism of action of ARC was further investigated by comparing in vitro and in vivo activity with other anti-neoplastic purines.

Methods: Structure-based homology searches were used to identify those compounds with similarity to ARC. Comparator compounds were then evaluated alongside ARC in the context of viability, cell cycle and apoptosis assays to establish any similarities. Following this, biological overlap was explored in detail using gene-expression analysis and kinase inhibition assays.

Results: Results demonstrated that sangivamycin, an extensively characterized pro-apoptotic nucleoside isolated from Streptomyces, had identical activity to ARC in terms of 1) cytotoxicity assays, 2) ability to induce a G2/M block, 3) inhibitory effects on RNA/DNA/protein synthesis, 4) transcriptomic response to treatment, 5) inhibition of protein kinase C, 6) inhibition of positive transcription elongation factor b (P-TEFb), 7) inhibition of VEGF secretion, and 8) activity within hollow fiber assays. Extending ARC activity to PKC inhibition provides a molecular basis for ARC cancer selectivity and anti-angiogenic effects. Furthermore, functional overlap between ARC and sangivamycin suggests that development of ARC may benefit from a retrospective of previous sangivamycin clinical trials. However, ARC was found to be inactive in several xenograft models, likely a consequence of rapid serum clearance.

Conclusion: Overall, these data expand on the biological properties of ARC but suggest additional studies are required before it can be considered a clinical trials candidate.

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Related in: MedlinePlus

ARC, sangivamycin and toyocamycin inhibit VEGF secretion. MCF7 cells were grown in 6 well dishes to 50% confluence, washed twice with PBS and 2 mL fresh RPMI-1640 media added. Media was then supplemented with the appropriate drug to a final concentration of 8 μM. After 24 h supernatant was removed and any cellular debris depleted by centrifugation. Adherent cells were trypsinized and cell numbers determined. An ELISA-based method was used to measure supernatant concentrations of VEGF. To avoid changes in cell number negatively influencing levels of VEGF secretion, results are expressed as picogram VEGF/mL per cell.
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Figure 6: ARC, sangivamycin and toyocamycin inhibit VEGF secretion. MCF7 cells were grown in 6 well dishes to 50% confluence, washed twice with PBS and 2 mL fresh RPMI-1640 media added. Media was then supplemented with the appropriate drug to a final concentration of 8 μM. After 24 h supernatant was removed and any cellular debris depleted by centrifugation. Adherent cells were trypsinized and cell numbers determined. An ELISA-based method was used to measure supernatant concentrations of VEGF. To avoid changes in cell number negatively influencing levels of VEGF secretion, results are expressed as picogram VEGF/mL per cell.

Mentions: ARC has been shown to possess strong antiangiogenic activity in vitro [1]. To compare any inherent antiangiogenic potential between drugs, an ELISA-based assay was used to quantify secretion of vascular endothelial growth factor (VEGF) from cell supernatants (Fig. 6). Values of secreted VEGF were calculated as a function of cell number to correct for the effects of cell death (pg/mL VEGF/cell). Results demonstrated a 5fold decrease in VEGF secretion for cells treated with ARC, sangivamycin or toyocamycin after 24 h. Fludarabine treatment did not appear to influence secretion whilst thioguanine decreased VEGF levels by one third. This result confirms that ARC, sangivamycin and toyocamycin all significantly inhibit secretion of this important angiogenic factor.


ARC (NSC 188491) has identical activity to Sangivamycin (NSC 65346) including inhibition of both P-TEFb and PKC.

Stockwin LH, Yu SX, Stotler H, Hollingshead MG, Newton DL - BMC Cancer (2009)

ARC, sangivamycin and toyocamycin inhibit VEGF secretion. MCF7 cells were grown in 6 well dishes to 50% confluence, washed twice with PBS and 2 mL fresh RPMI-1640 media added. Media was then supplemented with the appropriate drug to a final concentration of 8 μM. After 24 h supernatant was removed and any cellular debris depleted by centrifugation. Adherent cells were trypsinized and cell numbers determined. An ELISA-based method was used to measure supernatant concentrations of VEGF. To avoid changes in cell number negatively influencing levels of VEGF secretion, results are expressed as picogram VEGF/mL per cell.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651907&req=5

Figure 6: ARC, sangivamycin and toyocamycin inhibit VEGF secretion. MCF7 cells were grown in 6 well dishes to 50% confluence, washed twice with PBS and 2 mL fresh RPMI-1640 media added. Media was then supplemented with the appropriate drug to a final concentration of 8 μM. After 24 h supernatant was removed and any cellular debris depleted by centrifugation. Adherent cells were trypsinized and cell numbers determined. An ELISA-based method was used to measure supernatant concentrations of VEGF. To avoid changes in cell number negatively influencing levels of VEGF secretion, results are expressed as picogram VEGF/mL per cell.
Mentions: ARC has been shown to possess strong antiangiogenic activity in vitro [1]. To compare any inherent antiangiogenic potential between drugs, an ELISA-based assay was used to quantify secretion of vascular endothelial growth factor (VEGF) from cell supernatants (Fig. 6). Values of secreted VEGF were calculated as a function of cell number to correct for the effects of cell death (pg/mL VEGF/cell). Results demonstrated a 5fold decrease in VEGF secretion for cells treated with ARC, sangivamycin or toyocamycin after 24 h. Fludarabine treatment did not appear to influence secretion whilst thioguanine decreased VEGF levels by one third. This result confirms that ARC, sangivamycin and toyocamycin all significantly inhibit secretion of this important angiogenic factor.

Bottom Line: In this study, the mechanism of action of ARC was further investigated by comparing in vitro and in vivo activity with other anti-neoplastic purines.Structure-based homology searches were used to identify those compounds with similarity to ARC.Results demonstrated that sangivamycin, an extensively characterized pro-apoptotic nucleoside isolated from Streptomyces, had identical activity to ARC in terms of 1) cytotoxicity assays, 2) ability to induce a G2/M block, 3) inhibitory effects on RNA/DNA/protein synthesis, 4) transcriptomic response to treatment, 5) inhibition of protein kinase C, 6) inhibition of positive transcription elongation factor b (P-TEFb), 7) inhibition of VEGF secretion, and 8) activity within hollow fiber assays.

View Article: PubMed Central - HTML - PubMed

Affiliation: Developmental Therapeutics Program, SAIC-Frederick Inc, NCI- Frederick, Frederick, MD 21702, USA. Stockwin@ncifcrf.gov

ABSTRACT

Background: The nucleoside analog, ARC (NSC 188491) is a recently characterized transcriptional inhibitor that selectively kills cancer cells and has the ability to perturb angiogenesis in vitro. In this study, the mechanism of action of ARC was further investigated by comparing in vitro and in vivo activity with other anti-neoplastic purines.

Methods: Structure-based homology searches were used to identify those compounds with similarity to ARC. Comparator compounds were then evaluated alongside ARC in the context of viability, cell cycle and apoptosis assays to establish any similarities. Following this, biological overlap was explored in detail using gene-expression analysis and kinase inhibition assays.

Results: Results demonstrated that sangivamycin, an extensively characterized pro-apoptotic nucleoside isolated from Streptomyces, had identical activity to ARC in terms of 1) cytotoxicity assays, 2) ability to induce a G2/M block, 3) inhibitory effects on RNA/DNA/protein synthesis, 4) transcriptomic response to treatment, 5) inhibition of protein kinase C, 6) inhibition of positive transcription elongation factor b (P-TEFb), 7) inhibition of VEGF secretion, and 8) activity within hollow fiber assays. Extending ARC activity to PKC inhibition provides a molecular basis for ARC cancer selectivity and anti-angiogenic effects. Furthermore, functional overlap between ARC and sangivamycin suggests that development of ARC may benefit from a retrospective of previous sangivamycin clinical trials. However, ARC was found to be inactive in several xenograft models, likely a consequence of rapid serum clearance.

Conclusion: Overall, these data expand on the biological properties of ARC but suggest additional studies are required before it can be considered a clinical trials candidate.

Show MeSH
Related in: MedlinePlus