Nano-Fluidic Proteomic Assay and Desorption Electrospray Ionization Mass Spectrometry...
Transcript of Nano-Fluidic Proteomic Assay and Desorption Electrospray Ionization Mass Spectrometry...
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Nano-Fluidic Proteomic Assay and Desorption Electrospray Ionization
Mass Spectrometry Signatures Detect KRAS-altered Lipid Metabolism
in Lung Adenocarcinoma
Arvin M. Gouw, Ph.D.
TNT Fellow, Stanford University
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Glutamine metabolism
Fatty Acid Synthesis Pathway
Glucose metabolism
KRASKnown
Background
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Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 1: Does KRAS alter lipid metabolism?
?
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Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 2: Does altered lipid metabolism yield signaling and phospholipid signatures?
?Signaling
Membrane
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Glutamine metabolism
Fatty Acid Synthesis Pathway
Glucose metabolism
KRASKnown
Background
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Energy
Glucose
Normal Cells
Cutting off Cancer’s Fuel Supply
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Energy
Glutamine
Glucose
Lactate
Cancer Cells
Cutting off Cancer’s Fuel Supply
KRAS
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EnergyLipids
Signaling Membrane
Glutamine
Glucose
MYC
Lactate
CurrentResearch
Cancer Cells
Cutting off Cancer’s Fuel Supply
Gouw AM, Felsher DW Oncotarget 2015
KRAS
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Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 1: Does KRAS alter lipid metabolism?
?
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KRAS Lung Adenocarcinoma Model
• The KRAS oncogene can be conditionally activated in the
lungs of transgenic mice under the control of a Tet-On system.
• Removal of tetracycline/doxycycline induces KRAS
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KRAS upregulates fatty acid synthesis
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Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 2: Does altered lipid metabolism yield signaling and phospholipid signatures?
?Signaling
Membrane
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Capillary
Isoelectric point7.06.56.05.55.0
pERK2
pERK1
ppERK2ppERK1
ERK2
ERK1
Chem
ilum
inescence
(RLU
)
• Capillary-based microfluidic instrument (Nanopro1000)
• Isoelectric focusing for separation of native proteins
• Detects proteins in lysates of 6-25 cells per measurement, picogramrange of protein
• Uses antibody detection, luminescent readout
• Mean CV= 5% in clinical specimens
Nano-immunoassay analysis of clinical specimens
(NIA)
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Desorption Electro Spray Ionization Mass Spectrometry Imaging (DESI-MSI)
DESI analyzes and determines spatial localization and molecular
distribution of target molecules within a variety of samples.
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Fatty Acid Synthesis Pathway
Phospholipids
KRAS
Conclusion
Signaling
Membrane
NIA Signature
DESI-MSI Signature
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Acknowledgments
Felsher Lab, Zare lab, TNT Program