ABSTRACT
[This corrects the article DOI: 10.1371/journal.pone.0116388.].
ABSTRACT
PURPOSE: Researchers are currently seeking relevant lung cancer biomarkers in order to make informed decisions regarding therapeutic selection for patients in so-called "precision medicine." However, there are challenges to obtaining adequate lung cancer tissue for molecular analyses. Furthermore, current molecular testing of tumors at the genomic or transcriptomic level are very indirect measures of biological response to a drug, particularly for small molecule inhibitors that target kinases. Kinase activity profiling is therefore theorized to be more reflective of in vivo biology than many current molecular analysis techniques. As a result, this study seeks to prove the feasibility of combining a novel minimally invasive biopsy technique that expands the number of lesions amenable for biopsy with subsequent ex vivo kinase activity analysis. METHODS: Eight patients with lung lesions of varying location and size were biopsied using the novel electromagnetic navigational bronchoscopy (ENB) technique. Basal kinase activity (kinomic) profiles and ex vivo interrogation of samples in combination with tyrosine kinase inhibitors erlotinib, crizotinib, and lapatinib were performed by PamStation 12 microarray analysis. RESULTS: Kinomic profiling qualitatively identified patient specific kinase activity profiles as well as patient and drug specific changes in kinase activity profiles following exposure to inhibitor. Thus, the study has verified the feasibility of ENB as a method for obtaining tissue in adequate quantities for kinomic analysis and has demonstrated the possible use of this tissue acquisition and analysis technique as a method for future study of lung cancer biomarkers. CONCLUSIONS: We demonstrate the feasibility of using ENB-derived biopsies to perform kinase activity assessment in lung cancer patients.
Subject(s)
Bronchoscopy/methods , Gene Expression Profiling/methods , Lung Neoplasms/pathology , Phosphotransferases/metabolism , Protein Kinase Inhibitors/pharmacology , Aged , Aged, 80 and over , Crizotinib , Electromagnetic Phenomena , Erlotinib Hydrochloride , Female , Gene Expression Profiling/instrumentation , Humans , Lapatinib , Lung Neoplasms/enzymology , Male , Middle Aged , Precision Medicine/methods , Pyrazoles/pharmacology , Pyridines/pharmacology , Quinazolines/pharmacologyABSTRACT
We report a novel, low-resource malaria diagnostic platform inspired by the coffee ring phenomenon, selective for Plasmodium falciparum histidine-rich protein-II (PfHRP-II), a biomarker indicative of the P. falciparum parasite strain. In this diagnostic design, a recombinant HRP-II (rcHRP-II) biomarker is sandwiched between 1 µm Ni(II)nitrilotriacetic acid (NTA) gold-plated polystyrene microspheres (AuPS) and Ni(II)NTA-functionalized glass. After rcHRP-II malaria biomarkers had reacted with Ni(II)NTA-functionalized particles, a 1 µL volume of the particle-protein conjugate solution is deposited onto a functionalized glass slide. Drop evaporation produces the radial flow characteristic of coffee ring formation, and particle-protein conjugates are transported toward the drop edge, where, in the presence of rcHRP-II, particles bind to the Ni(II)NTA-functionalized glass surface. After evaporation, a wash with deionized water removes nonspecifically bound materials while maintaining the integrity of the surface-coupled ring produced by the presence of the protein biomarker. The dynamic range of this design was found to span 3 orders of magnitude, and rings are visible with the naked eye at protein concentrations as low as 10 pM, 1 order of magnitude below the 100 pM PfHRP-II threshold recommended by the World Health Organization. Key enabling features of this design are the inert and robust gold nanoshell to reduce nonspecific interactions on the particle surface, inclusion of a water wash step after drop evaporation to reduce nonspecific binding to the glass, a large diameter particle to project a large two-dimensional viewable area after ring formation, and a low particle density to favor radial flow toward the drop edge and reduce vertical settling to the glass surface in the center of the drop. This robust, antibody-free assay offers a simple user interface and clinically relevant limits of biomarker detection, two critical features required for low-resource malaria detection.
