Rapid, Multiplexed Phosphoprotein Profiling Using Silicon Photonic Sensor Arrays

Extracellular signaling is commonly mediated through post-translational protein modifications that propagate messages from membrane-bound receptors to ultimately regulate gene expression. Signaling cascades are ubiquitously intertwined, and a full understanding of function can only be gleaned by observing dynamics across multiple key signaling nodes. Importantly, targets within signaling cascades often represent opportunities for therapeutic development or can serve as diagnostic biomarkers. Protein phosphorylation is a particularly important post-translational modification that controls many essential cellular signaling pathways. Not surprisingly, aberrant phosphorylation is found in many human diseases, including cancer, and phosphoprotein-based biomarker signatures hold unrealized promise for disease monitoring. Moreover, phosphoprotein analysis has wide-ranging applications across fundamental chemical biology, as many drug discovery efforts seek to target nodes within kinase signaling pathways. For both fundamental and translational applications, the analysis of phosphoprotein biomarker targets is limited by a reliance on labor-intensive and/or technically challenging methods, particularly when considering the simultaneous monitoring of multiplexed panels of phosphoprotein biomarkers. We have developed a technology based upon arrays of silicon photonic microring resonator sensors that fills this void, facilitating the rapid and automated analysis of multiple phosphoprotein levels from both cell lines and primary human tumor samples requiring only minimal sample preparation.

[1]  Seamus J. Martin,et al.  Acid Sphingomyelinase–Deficient Human Lymphoblasts and Mice Are Defective in Radiation-Induced Apoptosis , 1996, Cell.

[2]  Ryan C Bailey,et al.  Rapid, multiparameter profiling of cellular secretion using silicon photonic microring resonator arrays. , 2011, Journal of the American Chemical Society.

[3]  A. Herr,et al.  Microchamber Western blotting using poly-L-lysine conjugated polyacrylamide gel for blotting of sodium dodecyl sulfate coated proteins. , 2013, Analytical chemistry.

[4]  Y. Yamada,et al.  Radiation necrosis following treatment of high grade glioma—a review of the literature and current understanding , 2012, Acta Neurochirurgica.

[5]  S. Kingsmore,et al.  Multiplexed protein profiling on microarrays by rolling-circle amplification , 2002, Nature Biotechnology.

[6]  S. Hoyer Glucose metabolism and insulin receptor signal transduction in Alzheimer disease. , 2004, European journal of pharmacology.

[7]  Ira Mellman,et al.  Antibody Therapeutics in Cancer , 2013, Science.

[8]  Amy E Herr,et al.  Microfluidic Western blotting , 2012, Proceedings of the National Academy of Sciences.

[9]  Muzammil Iqbal,et al.  Label-Free Biosensor Arrays Based on Silicon Ring Resonators and High-Speed Optical Scanning Instrumentation , 2010, IEEE Journal of Selected Topics in Quantum Electronics.

[10]  Tae-Min Kim,et al.  A developmental taxonomy of glioblastoma defined and maintained by MicroRNAs. , 2011, Cancer research.

[11]  Z. Fuks,et al.  Radiation-induced apoptosis of endothelial cells in the murine central nervous system: protection by fibroblast growth factor and sphingomyelinase deficiency. , 2000, Cancer research.

[12]  J. Sage,et al.  Cellular mechanisms of tumour suppression by the retinoblastoma gene , 2008, Nature Reviews Cancer.

[13]  I. Kramer,et al.  Prologue: Signal Transduction, Origins, and Ancestors , 2009 .

[14]  James R Heath,et al.  Microfluidics-based single-cell functional proteomics for fundamental and applied biomedical applications. , 2014, Annual review of analytical chemistry.

[15]  Adam L. Washburn,et al.  Quantitative, label-free detection of five protein biomarkers using multiplexed arrays of silicon photonic microring resonators. , 2010, Analytical chemistry.

[16]  E. Petricoin,et al.  Application of molecular technologies for phosphoproteomic analysis of clinical samples , 2014, Oncogene.

[17]  Johannes G. Reiter,et al.  The molecular evolution of acquired resistance to targeted EGFR blockade in colorectal cancers , 2012, Nature.

[18]  Ralph Weissleder,et al.  Cancer Cell Profiling by Barcoding Allows Multiplexed Protein Analysis in Fine-Needle Aspirates , 2014, Science Translational Medicine.

[19]  W. N. Burnette,et al.  "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. , 1981, Analytical biochemistry.

[20]  R. Kennedy,et al.  Western blotting using microchip electrophoresis interfaced to a protein capture membrane. , 2013, Analytical chemistry.

[21]  R. Kennedy,et al.  Western blotting using capillary electrophoresis. , 2011, Analytical chemistry.

[22]  A. J. van der Kogel,et al.  Mechanisms of radiation injury to the central nervous system: implications for neuroprotection. , 2004, Molecular interventions.

[23]  M. Rajer,et al.  Quantitative analysis of fine needle aspiration biopsy samples , 2005 .

[24]  Jingchun Zhu,et al.  Realizing the Promise of Reverse Phase Protein Arrays for Clinical, Translational, and Basic Research: A Workshop Report , 2014, Molecular & Cellular Proteomics.

[25]  L. Hood,et al.  Integrated barcode chips for rapid, multiplexed analysis of proteins in microliter quantities of blood , 2008, Nature Biotechnology.

[26]  Stephen L. Brown,et al.  Mechanisms of radiation-induced brain toxicity and implications for future clinical trials , 2008, Journal of Neuro-Oncology.

[27]  S. M. Rubin Deciphering the retinoblastoma protein phosphorylation code. , 2013, Trends in biochemical sciences.

[28]  L. Cantley,et al.  What a tangled web we weave: emerging resistance mechanisms to inhibition of the phosphoinositide 3-kinase pathway. , 2013, Cancer discovery.

[29]  I. Vivanco,et al.  Targeting molecular addictions in cancer , 2014, British Journal of Cancer.

[30]  Xingyu Jiang,et al.  Microfluidic Western blot. , 2010, Analytical chemistry.

[31]  G. Mills,et al.  Reverse phase protein array: validation of a novel proteomic technology and utility for analysis of primary leukemia specimens and hematopoietic stem cells , 2006, Molecular Cancer Therapeutics.

[32]  J. Heath,et al.  Chemical methods for the simultaneous quantitation of metabolites and proteins from single cells. , 2015, Journal of the American Chemical Society.

[33]  N. Gray,et al.  Targeting cancer with small molecule kinase inhibitors , 2009, Nature Reviews Cancer.

[34]  Gabriel A Kwong,et al.  DNA-encoded antibody libraries: a unified platform for multiplexed cell sorting and detection of genes and proteins. , 2007, Journal of the American Chemical Society.

[35]  Amy E. Herr,et al.  Single-cell western blotting , 2014, Nature Methods.

[36]  Amy E Herr,et al.  Single-Cell Western Blotting. , 2015, Methods in molecular biology.

[37]  I. Kramer,et al.  Phosphorylation and dephosphorylation: Protein kinases A and C , 2002 .

[38]  V. Quaranta,et al.  What Lies Beneath: Looking Beyond Tumor Genetics Shows the Complexity of Signaling Networks Underlying Drug Sensitivity , 2013, Science Signaling.

[39]  I. Bayazitov,et al.  Differentiated Horizontal Interneurons Clonally Expand to Form Metastatic Retinoblastoma in Mice , 2007, Cell.

[40]  R. Weinberg,et al.  The retinoblastoma protein and cell cycle control , 1995, Cell.

[41]  Abraham J. Qavi,et al.  Subpicogram per milliliter detection of interleukins using silicon photonic microring resonators and an enzymatic signal enhancement strategy. , 2013, Analytical chemistry.

[42]  J. Zierath,et al.  Characterization of signal transduction and glucose transport in skeletal muscle from type 2 diabetic patients. , 2000, Diabetes.

[43]  P. Steeg Metastasis suppressors alter the signal transduction of cancer cells , 2003, Nature Reviews Cancer.

[44]  J. Engelman,et al.  Bypass Mechanisms of Resistance to Receptor Tyrosine Kinase Inhibition in Lung Cancer , 2013, Science Signaling.