MSACL 

37. Discovery of Mammalian Cardenolide Hormone (MCH-518) using FT-ICR-MS and Tandem Mass Spectrometry
Poster: Mon 2:00-3:00PM
Mohammad Al-Ghoul
University of Louisville
Mohammad Al-Ghoul, Department of Pathology and Laboratory Medicine, Environmental Health Sciences Training Program, University of Louisville School of Medicine, Louisville, KY.
Edward P. Womack Jr, Department of Pathology and Laboratory Medicine, University of Louisville School of Medicine, Louisville, KY.
William W. Tucker, Department of Pathology and Laboratory Medicine, University of Louisville School of Medicine, Louisville, KY.
Bogdan Bogdanov, Department of Chemistry, Center for Regulatory and Environmental Analytical Metabolomics, Louisville, KY
Richard Higashi, Department of Chemistry, Center for Regulatory and Environmental Analytical Metabolomics, Louisville, KY
Roland Valdes, Jr. Department of Pathology and Laboratory Medicine,University of Louisville School of Medicine, Louisville, KY
Hypothesis: Recently discovered endogenous compound (MCH-518) from human serum is cytotoxic to breast cancer cells and not to normal breast cells.

Background: We have isolated a compound with physical properties similar to those of endogenous DLIF (digoxin-like immunoreactive factor). The compound was extracted from human serum and characterized by FT-ICR-MS and tandem mass spectrometry with a reproducible peak of high abundance at m/z of 518.3221. Several studies have suggested the beneficial effect of the plant-derived cardenolide digoxin in treating women with breast cancer. Although results using digoxin remain controversial, we posit that an endogenous mammalian counterpart, such as MCH-518, would have increased specificity in targeting cancer cells over normal cells. In this study we provide evidence that MCH-518 has the elemental composition of C28H47O7 and that the activity of this compound is more specific than digoxin at inducing apoptosis in breast cancer cells in culture.

Methodology: We isolated the MCH-518 compound from human serum using solid phase extraction followed by HPLC purification. The HPLC peak of interest was further analyzed by FT-ICR-MS and tandem mass spectrometry. For biological activity we used breast cancer cell lines MCF 7 and MDA-MB-231 and epithelial cells MCF 10A as a control. Cells were incubated with MCH-518 and separately with digoxin. Cell viability was measured by MTT assay. Apoptosis was confirmed by flow cytometry at the IC50 concentrations.

Results: The purified HPLC peak showed a peak with m/z = 518.3221 using FT-ICR-MS and LC-MS/MS. Diogoxin is cytotoxic to breast cancer cells at an IC50 concentration of 90 ± 20 nM and to normal cells at IC50 concentration of 210 ± 20 nM. DLIF is cytotoxic to breast cancer cells at 0.2 ± 0.1 nM and to normal cells at 1.0 ± 0.1 nM. However, we note that the concentration of MCH 518 was measured using an immunoassay based on digoxin equivalence.

Conclusion: We conclude that MCH-518 is much more toxic to breast cancer cells and is less toxic to epithelial cells than is digoxin. These findings suggest the possibility that subjects with appropriate blood concentration of MCH-518 may be inherently protected from developing breast cancers compared to those who do not produce enough MCH-518.
Email: [email protected]