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The antitumor ether lipid 1-Q-octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3) inhibits the association between Ras and Raf-1.
Anticancer Res. 2003 May-Jun; 23(3B):2291-5.AR

Abstract

BACKGROUND

Previous studies have shown that the antitumor ether lipid, 1-O-Octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3), inhibits the activation of the MAPK pathway in EGF- and serum-stimulated MCF-7 cells. The activation of the MAPK pathway subsequent to growth factor stimulation requires the recruitment of Raf-1 from the cytosol to the membrane. ET-18-OCH3 decreased the level of membrane-associated Raf-1 relative to untreated control cells. Since ET-18-OCH3 did not inhibit the activities of the kinases in the cascade, the reduced Raf-1 levels appeared to be the cause for the reduction in the magnitude and duration of MAPK activity. In this study we have investigated whether the reduced Raf-1 levels arise from a perturbation of the interaction of Raf-1 with activated Ras, which is the event that mediates the membrane recruitment.

MATERIALS AND METHODS

The interaction of Raf-1 with Ras was examined by investigating the association of cytosolic Raf-1, from ET-18-OCH3-treated and untreated cells with purified GST-Ras-GTP-gamma-S bound to agarose beads. The level of associating Raf was determined by Western blot analysis. The effect of naturally occurring phospholipids on the Raf-1-Ras interaction was also examined to assess the specificity of the results.

RESULTS

In cells preincubated with ET-18-OCH3, the interaction of GST-Ras-GTP-gamma-S with cytosolic Raf was reduced. The addition of ET-18-OCH3 to the cytosolic fraction isolated from untreated cells also reduced the binding of Raf to activated GST-Ras-GTP-gamma-S. Cytosolic Raf-1 from cells incubated with natural lysophospholipids was similar to that of controls.

CONCLUSION

Our findings suggest that ET-18-OCH3 associates specifically with Raf-1 in the cytosol and interferes in the interaction of Raf-1 with activated Ras, thereby reducing the levels that are translocated to the membrane for activation. Thus Raf-1 appears to be a molecular target of ET-18-OCH3.

Authors+Show Affiliations

Department of Biochemistry and Medical Genetics, University of Manitaoba, 770 Bannatyne Avenue, Winnipeg, Manitoba, Canada R3E 0W3.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

12894505

Citation

Samadder, Pranati, et al. "The Antitumor Ether Lipid 1-Q-octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3) Inhibits the Association Between Ras and Raf-1." Anticancer Research, vol. 23, no. 3B, 2003, pp. 2291-5.
Samadder P, Richards C, Bittman R, et al. The antitumor ether lipid 1-Q-octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3) inhibits the association between Ras and Raf-1. Anticancer Res. 2003;23(3B):2291-5.
Samadder, P., Richards, C., Bittman, R., Bhullar, R. P., & Arthur, G. (2003). The antitumor ether lipid 1-Q-octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3) inhibits the association between Ras and Raf-1. Anticancer Research, 23(3B), 2291-5.
Samadder P, et al. The Antitumor Ether Lipid 1-Q-octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3) Inhibits the Association Between Ras and Raf-1. Anticancer Res. 2003 May-Jun;23(3B):2291-5. PubMed PMID: 12894505.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - The antitumor ether lipid 1-Q-octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3) inhibits the association between Ras and Raf-1. AU - Samadder,Pranati, AU - Richards,Christina, AU - Bittman,Robert, AU - Bhullar,Rajinder Pal, AU - Arthur,Gilbert, PY - 2003/8/5/pubmed PY - 2003/8/29/medline PY - 2003/8/5/entrez SP - 2291 EP - 5 JF - Anticancer research JO - Anticancer Res VL - 23 IS - 3B N2 - BACKGROUND: Previous studies have shown that the antitumor ether lipid, 1-O-Octadecyl-2-O-methyl-rac-glycerophosphocholine (ET-18-OCH3), inhibits the activation of the MAPK pathway in EGF- and serum-stimulated MCF-7 cells. The activation of the MAPK pathway subsequent to growth factor stimulation requires the recruitment of Raf-1 from the cytosol to the membrane. ET-18-OCH3 decreased the level of membrane-associated Raf-1 relative to untreated control cells. Since ET-18-OCH3 did not inhibit the activities of the kinases in the cascade, the reduced Raf-1 levels appeared to be the cause for the reduction in the magnitude and duration of MAPK activity. In this study we have investigated whether the reduced Raf-1 levels arise from a perturbation of the interaction of Raf-1 with activated Ras, which is the event that mediates the membrane recruitment. MATERIALS AND METHODS: The interaction of Raf-1 with Ras was examined by investigating the association of cytosolic Raf-1, from ET-18-OCH3-treated and untreated cells with purified GST-Ras-GTP-gamma-S bound to agarose beads. The level of associating Raf was determined by Western blot analysis. The effect of naturally occurring phospholipids on the Raf-1-Ras interaction was also examined to assess the specificity of the results. RESULTS: In cells preincubated with ET-18-OCH3, the interaction of GST-Ras-GTP-gamma-S with cytosolic Raf was reduced. The addition of ET-18-OCH3 to the cytosolic fraction isolated from untreated cells also reduced the binding of Raf to activated GST-Ras-GTP-gamma-S. Cytosolic Raf-1 from cells incubated with natural lysophospholipids was similar to that of controls. CONCLUSION: Our findings suggest that ET-18-OCH3 associates specifically with Raf-1 in the cytosol and interferes in the interaction of Raf-1 with activated Ras, thereby reducing the levels that are translocated to the membrane for activation. Thus Raf-1 appears to be a molecular target of ET-18-OCH3. SN - 0250-7005 UR - https://www.unboundmedicine.com/medline/citation/12894505/The_antitumor_ether_lipid_1_Q_octadecyl_2_O_methyl_rac_glycerophosphocholine__ET_18_OCH3__inhibits_the_association_between_Ras_and_Raf_1_ L2 - https://antibodies.cancer.gov/detail/CPTC-RAF1-1 DB - PRIME DP - Unbound Medicine ER -