Intra-Individual Variation in Disease-Specific IgG Fc Glycoform Ratios to Monitor the Disease Progression of Lung Cancer.

Aberrant protein glycosylation is an active pathological alteration related to the progression of cancers. The speed of progression varies among individuals, increasing the difficulties of prognosis assessment. Hence, evaluating variation in glycosylation using patients themselves as their own controls is a potential way to reduce the impact of individual differences on progression monitoring. Here, following a longitudinal follow-up study involving 125 lung cancer (LC) patients with progressive disease, we isolated disease-specific IgG from serum using polyacrylamide gel electrophoresis, obtained IgG glycoform ratios using mass spectrometry, and then set a fold-change cutoff of 1.5 to utilize the intra-individual variation in IgG glycosylation to monitor PD. We found that the serial monitoring of 15 types of glycoform ratios provided an effective way for monitoring LC progression. Over 1.5-fold changes in glycoform ratios relative to the first observed value were detected in 117 of 125 LC patients (93.6%). Our established method predicted LC progression 55.8 (IQR 31.1-90.1) weeks earlier than imaging examination did. In summary, intra-individual variation in IgG glycoform ratios is useful to monitor LC progression, expanding our knowledge about the relationship between IgG glycosylation and cancer prognosis. The raw data files are available via the ProteomeXchange Consortium with the identifier PXD037541.

Fe3O4@PANI: a magnetic polyaniline nanomaterial for highly efficient and handy enrichment of intact N-glycopeptides.

Glycosylation of proteins plays important roles in the occurrence and development of chronic diseases. In this study, we report an enrichment method of intact N-glycopeptides using a magnetic polyaniline nanomaterial (Fe3O4@PANI). Under the synergistic effect of hydrogen bonding and electrostatic adsorption, Fe3O4@PANI can rapidly and easily enrich N-glycopeptides derived from standard protein (bovine fetuin and transferrin) tryptic digests and serum haptoglobin tryptic digests. Finally we have detected 63 glycopeptides in the glycosylation sites of both N204 and N211 from the serum haptoglobin beta chain using MALDI FTICR MS. Compared with non-magnetic materials, Fe3O4@PANI can achieve complete separation from complex biological samples, meeting the requirement of the high purity of samples for mass spectrometric detection. Overall, Fe3O4@PANI exhibits great application potential in the highly efficient enrichment of intact N-glycopeptides due to its stability and convenient preparation.

Disease-Specific IgG Fc Glycosylation Ratios as Personalized Biomarkers to Differentiate Non-Small Cell Lung Cancer from Benign Lung Diseases.

PURPOSE: The authors aimed to separate Fc N-glycopeptides of disease-specific immunoglobulin G (DSIgG) as personalized biomarkers to distinguish non-small cell lung cancer (NSCLC) from benign lung diseases (BLDs). EXPERIMENTAL DESIGN: DSIgG from 509 BLDs patients and 477 NSCLC patients was isolated using native polyacrylamide gel electrophoresis and then the Fc glycosylation was determined using mass spectrometry. RESULTS: For the patients below 60 years of age, a combination of the glycopeptides ratios with one fucose residue difference of DSIgG1 and DSIgG2 can differentiate NSCLC from BLDs, with area under curve (AUC) values of >0.76, sensitivities of >87%, and specificities of >61%. For the patients above 60 years of age, a combination of the glycopeptides ratios with one monosaccharide residue of DSIgG2 can differentiate NSCLC from BLDs, with AUC values of >0.78, sensitivities of >91%, and specificities of >54%. For the same participants, the commonly used clinical biomarkers have AUC values of 0.5-0.621, sensitivities of 15.8-32.9%, and specificities of 75.7-90.5%. CONCLUSIONS: These findings indicate that these DSIgG Fc glycoforms are potential personalized biomarkers to differentiate NSCLC from BLDs.

Disease-specific haptoglobin-ß chain N-glycosylation as biomarker to differentiate non-small cell lung cancer from benign lung diseases.

Background: The association of pathological states with N-glycosylation of haptoglobin-ß has attracted increasing attention. Materials & Methods: In the present study, disease-specific haptoglobin-ß (DSHp-ß) was separated from serum immunoinflammation-related protein complexes (IIRPCs) of 600 participants including 300 patients with benign lung diseases (BLDs) and 300 patients with non-small cell lung cancer (NSCLC). The enriched glycopeptides of the tryptic digests of the DSHp-ß were analyzed using matrix assisted laser desorption/ionization-Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FTICR MS). Results: 20 of glycopeptides were detected for each sample. The statistical analysis has indicated that significant changes in the sialylation of DSHp-ß between BLDs and NSCLC patients were observed. The age- and sex-matched participants were randomly clarified into the training set and the validation set. Receiver operating characteristic (ROC) analysis has revealed that the level ratio of glycopeptides (G2G3/G2G3S4) at the sites of Asn207/211 has potential capability to distinguish BLDs from NSCLC, with the sensitivity of 74.4%, the specificity of 82.8%, and the area under curve (AUC) of 0.805. Conclusion: The glycosylation of DSHp-ß can distinguish NSCLC from BLDs with high diagnostic accuracy compared with current clinical available serum markers.

Monitoring changes of docosahexaenoic acid-containing lipids during the recovery process of traumatic brain injury in rat using mass spectrometry imaging.

Brain lipid homoeostasis is critical during recovery process after traumatic brain injury (TBI). In this study, we integrated liquid extraction and electrosonic spray ionization technology to develop an ionization device coupled with a Fourier transform ion cyclotron resonance mass spectrometer for imaging of docosahexaenoic acid (DHA)-containing lipids on rat brain tissues. The ion images of the brain tissue sections from the normal rats and the rats after TBI at acute phase (0 and 1 day) and chronic phase (3, 5, and 7 days) were obtained. The imaging results indicate that the levels of DHA and lyso-phosphatidylethanolamine (22:6) in the injury area of TBI rats increased significantly at the acute phase and subsequently decreased at the chronic phase. But the levels of DHA-containing phospholipids including phosphatidylethanolamine (PE)(P-18:0/22:6), PE(18:0/22:6), and phosphatidylserine (18:0/22:6) decreased at the acute phase and gradually increased at the chronic phase in the injury area accompanied by the morphogenesis and wound healing. These findings indicate that the DHA may participate in the recovery process of brain injury. This is the first report to in situ detect the changes in the levels of DHA and DHA-containing lipids in the TBI model.

Facile and Selective Enrichment of Intact Sialoglycopeptides Using Graphitic Carbon Nitride.

Combining powerful selectivity, high stability, convenient operation, mild condition, and eco-friendliness, a novel graphitic carbon nitride (g-C3N4)-based enrichment method of intact sialoglycopeptides (SGs) was developed. The intact SGs could be simply enriched and separated from protein tryptic digests by hydrogen bonding without damage of glycan structures due to the specific structure of g-C3N4. By optimizing the enrichment and elution conditions, 45 and 38 SGs were detected from the tryptic digests of bovine fetuin and transferrin, respectively. Under the synergistic effect of hydrogen bonding and electrostatic adsorption, the SGs could be enriched simply in less than 2 h with a detection limit of 50 fmol. The method is repeatable due to the high stability of g-C3N4 and the simple protocol of the method, indicating the potential application of g-C3N4 in efficient and selective enrichment of intact SGs.

Mass spectrometry imaging of small molecules in biological tissues using graphene oxide as a matrix.

With the development of matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI), molecular interrogation of tissue sections over a wide mass range has become feasible, but small molecule analysis is still far from being fully reached due to the limited sensitivity and matrix interference. Herein, graphene oxide (GO) is used as a MALDI matrix to image small molecules in tissues in negative ion mode. Finally, 212 of molecules including 190 of lipids and 22 of low molecular weight metabolites were detected and spatially visualized in mouse brain tissue sections without the interference of matrix ions/clusters, and the structures of 69 of the lipids were confirmed by using in situ tandem mass spectrometry. A further application of GO matrix could reveal distinct spatio-molecular signatures in viable and necrotic tumor regions derived from a mouse breast cancer tissue. In addition, GO as a MALDI matrix has exhibited a better performance in MSI of lipids relative to N-(1-naphthyl) ethylenediamine dihydrochloride and 9-aminoacridine.