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Blood proteome profiling using aptamer-based technology for rejection biomarker discovery in transplantation.

ABSTRACT: Face transplantation is a promising solution for patients with devastating facial injuries who lack other satisfactory treatment options. At the same time, this type of transplantation is accompanied with high risks of acute transplant rejection. The limitations of traditional skin biopsy and the need to frequently monitor the condition of face transplant call for less invasive biomarkers to better diagnose and treat acute rejection. Discovery of peripheral serum proteins accurately reflecting the transplant status would represent a reasonable solution to meet this demand. However, to date, there is no clinical data available to address the feasibility of this approach. In this study, we used the next generation aptamer-based SOMAscan proteomics platform to profile 1305 proteins of peripheral blood serum in twenty-four samples taken from 6 patients during no-rejection, nonsevere rejection, and severe rejection episodes. Also, we provide a detailed description of biosample processing and all steps to generate and analyze the SOMAscan dataset with hope it will assist in performing biomarker discovery in other transplantation centers using this platform.

SUBMITTER: Shubin AV

PROVIDER: S-EPMC6901551 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Blood proteome profiling using aptamer-based technology for rejection biomarker discovery in transplantation.

Shubin Andrey V AV Kollar Branislav B Dillon Simon T ST Pomahac Bohdan B Libermann Towia A TA Riella Leonardo V LV

Scientific data 20191209 1

Face transplantation is a promising solution for patients with devastating facial injuries who lack other satisfactory treatment options. At the same time, this type of transplantation is accompanied with high risks of acute transplant rejection. The limitations of traditional skin biopsy and the need to frequently monitor the condition of face transplant call for less invasive biomarkers to better diagnose and treat acute rejection. Discovery of peripheral serum proteins accurately reflecting t ...[more]

PMID: 31819064

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Project description:BackgroundThe interrogation of proteomes ("proteomics") in a highly multiplexed and efficient manner remains a coveted and challenging goal in biology and medicine.Methodology/principal findingsWe present a new aptamer-based proteomic technology for biomarker discovery capable of simultaneously measuring thousands of proteins from small sample volumes (15 µL of serum or plasma). Our current assay measures 813 proteins with low limits of detection (1 pM median), 7 logs of overall dynamic range (~100 fM-1 µM), and 5% median coefficient of variation. This technology is enabled by a new generation of aptamers that contain chemically modified nucleotides, which greatly expand the physicochemical diversity of the large randomized nucleic acid libraries from which the aptamers are selected. Proteins in complex matrices such as plasma are measured with a process that transforms a signature of protein concentrations into a corresponding signature of DNA aptamer concentrations, which is quantified on a DNA microarray. Our assay takes advantage of the dual nature of aptamers as both folded protein-binding entities with defined shapes and unique nucleotide sequences recognizable by specific hybridization probes. To demonstrate the utility of our proteomics biomarker discovery technology, we applied it to a clinical study of chronic kidney disease (CKD). We identified two well known CKD biomarkers as well as an additional 58 potential CKD biomarkers. These results demonstrate the potential utility of our technology to rapidly discover unique protein signatures characteristic of various disease states.Conclusions/significanceWe describe a versatile and powerful tool that allows large-scale comparison of proteome profiles among discrete populations. This unbiased and highly multiplexed search engine will enable the discovery of novel biomarkers in a manner that is unencumbered by our incomplete knowledge of biology, thereby helping to advance the next generation of evidence-based medicine.

Project description:BackgroundBlood-based protein measurement is a routine practice for detecting biomarkers in human disease. Comprehensive profiling of blood/plasma/serum proteome is a challenge due to an extremely large dynamic range, as exemplified by a small subset of highly abundant proteins. Antibody-based depletion of these abundant proteins alleviates the problem but introduces experimental variations. We aimed to establish a method for direct profiling of undepleted human serum and apply the method toward biomarker discovery for Alzheimer's disease (AD), as AD is the most common form of dementia without available blood-based biomarkers in clinic.MethodsWe present an ultra-deep analysis of undepleted human serum proteome by combining the latest 11-plex tandem-mass-tag (TMT) labeling, exhaustive two-dimensional liquid chromatography (LC/LC) fractionation (the 1st LC: 3 h for 180 fractions, and the 2nd LC: 3 h gradient per fraction), coupled with high resolution tandem mass spectrometry (MS/MS). AD (n = 6) and control (n = 5) sera were analyzed in this pilot study. In addition, we implemented a multiplexed targeted LC-MS3 method (TOMAHAQ) for the validation of selected target proteins.ResultsThe TMT-LC/LC-MS/MS platform is capable of analyzing 4826 protein components (4368 genes), covering at least 6 orders of magnitude in dynamic range, representing one of the deepest serum proteome analysis. We defined intra- and inter- group variability in the AD and control groups. Statistical analysis revealed differentially expressed proteins in AD (26 decreased and 4 increased). Notably, these altered proteins are enriched in the known pathways of mitochondria, fatty acid beta oxidation, and AGE/RAGE. Finally, we set up a TOMAHAQ method to confirm the decrease of PCK2 and AK2 in our AD samples.ConclusionsOur results show an ultra-deep serum discovery study by TMT-LC/LC-MS/MS, and a validation experiment by TOMAHAQ targeted LC-MS3. The MS-based discovery and validation methods are of general use for biomarker discovery from complex biofluids (e.g. serum proteome). This pilot study also identified deregulated proteins, in particular proteins associated with mitochondrial function in the AD serum samples. These proteins may serve as novel AD candidate biomarkers.

Project description:Lung cancer is the leading cause of cancer deaths worldwide. New diagnostics are needed to detect early stage lung cancer because it may be cured with surgery. However, most cases are diagnosed too late for curative surgery. Here we present a comprehensive clinical biomarker study of lung cancer and the first large-scale clinical application of a new aptamer-based proteomic technology to discover blood protein biomarkers in disease.We conducted a multi-center case-control study in archived serum samples from 1,326 subjects from four independent studies of non-small cell lung cancer (NSCLC) in long-term tobacco-exposed populations. Sera were collected and processed under uniform protocols. Case sera were collected from 291 patients within 8 weeks of the first biopsy-proven lung cancer and prior to tumor removal by surgery. Control sera were collected from 1,035 asymptomatic study participants with ? 10 pack-years of cigarette smoking. We measured 813 proteins in each sample with a new aptamer-based proteomic technology, identified 44 candidate biomarkers, and developed a 12-protein panel (cadherin-1, CD30 ligand, endostatin, HSP90?, LRIG3, MIP-4, pleiotrophin, PRKCI, RGM-C, SCF-sR, sL-selectin, and YES) that discriminates NSCLC from controls with 91% sensitivity and 84% specificity in cross-validated training and 89% sensitivity and 83% specificity in a separate verification set, with similar performance for early and late stage NSCLC.This study is a significant advance in clinical proteomics in an area of high unmet clinical need. Our analysis exceeds the breadth and dynamic range of proteome interrogated of previously published clinical studies of broad serum proteome profiling platforms including mass spectrometry, antibody arrays, and autoantibody arrays. The sensitivity and specificity of our 12-biomarker panel improves upon published protein and gene expression panels. Separate verification of classifier performance provides evidence against over-fitting and is encouraging for the next development phase, independent validation. This careful study provides a solid foundation to develop tests sorely needed to identify early stage lung cancer.

Dataset Information

Blood proteome profiling using aptamer-based technology for rejection biomarker discovery in transplantation.

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Blood proteome profiling using aptamer-based technology for rejection biomarker discovery in transplantation.

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