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
Glutamine metabolism
Fatty Acid Synthesis Pathway
Glucose metabolism
KRASKnown
Background
Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 1: Does KRAS alter lipid metabolism?
?
Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 2: Does altered lipid metabolism yield signaling and phospholipid signatures?
?Signaling
Membrane
Glutamine metabolism
Fatty Acid Synthesis Pathway
Glucose metabolism
KRASKnown
Background
Energy
Glucose
Normal Cells
Cutting off Cancer’s Fuel Supply
Energy
Glutamine
Glucose
Lactate
Cancer Cells
Cutting off Cancer’s Fuel Supply
KRAS
EnergyLipids
Signaling Membrane
Glutamine
Glucose
MYC
Lactate
CurrentResearch
Cancer Cells
Cutting off Cancer’s Fuel Supply
Gouw AM, Felsher DW Oncotarget 2015
KRAS
Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 1: Does KRAS alter lipid metabolism?
?
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
KRAS upregulates fatty acid synthesis
Glutamine metabolism
Fatty Acid Synthesis Pathway
Phospholipids
Glucose metabolism
KRASKnown
Part 2: Does altered lipid metabolism yield signaling and phospholipid signatures?
?Signaling
Membrane
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)
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.
Fatty Acid Synthesis Pathway
Phospholipids
KRAS
Conclusion
Signaling
Membrane
NIA Signature
DESI-MSI Signature
Acknowledgments
Felsher Lab, Zare lab, TNT Program