Glycomic Signatures of Plasma IgG Improve Preoperative Prediction of the Invasiveness of Small Lung Nodules

Abstract

Preoperative assessment of tumor invasiveness is essential to avoid overtreatment for patients with small-sized ground-glass nodules (GGNs) of 10 mm or less in diameter. However, it is difficult to determine the pathological state by computed tomography (CT) examination alone. Aberrant glycans has emerged as a tool to identify novel potential disease biomarkers. In this study, we used a lectin microarray-based strategy to investigate whether glycosylation changes in plasma immunoglobulin G (IgG) provide additional information about the invasiveness of small GGNs before surgery. Two independent cohorts (discovery set, n = 92; test set, n = 210) of GGN patients were used. Five of 45 lectins (Sambucus nigra agglutinin, SNA; Datura stramonium agglutinin, DSA; Galanthus nivalis agglutinin, GNA; Euonymus europaeus lectin, EEL; and Vicia villosa agglutinin, VVA) were identified as independent factors associated with pathological invasiveness of small GGNs (p < 0.01). Receiver-operating characteristic (ROC) curve analysis indicated the combination of these five lectins could significantly improve the accuracy of CT in diagnosing invasive GGNs, with an area under the curve (AUC) of 0.792 (p < 0.001), a sensitivity of 74.6%, and specificity of 74.4%, which was superior to current clinical biomarkers. These results suggest that the multilectin assay based on plasma IgG glycosylation may be a useful in vitro complementary test to enhance preoperative determination of the invasiveness of GGNs and guide surgeons to select proper clinical management to avoid overtreatment.

Keywords: glycobiomarker; ground glass nodule (GGN); immunoglobulin G (IgG); lectin microarray; multilectin assay.

Figure 1

The flow diagram of the study population in this study. GGN, ground glass nodule; SCC, squamous cell carcinoma; d, diameter.

Figure 2

Differentially expressed IgG glycopatterns between noninvasive and invasive GGNs detected by lectin microarray. (A) The layout of the lectin microarray and representative array profiling of plasma IgG from noninvasive (benign, AAH, and AIS) and invasive (MIA and IA) GGN patients are shown. Differential signal patterns of eight lectins between the two groups are indicated in the open-box illustration in the array layout. Five lectins with signal > 1000 are marked with white frames in the representative profiling images. (B) The comparison of intensities of the five lectin signals (SNA, DSA, NPA, GNA, and WGA) between noninvasive (n = 90) and invasive (n = 120) GGNs less than 10 mm. * p < 0.05 and ** p < 0.01 from the Mann–Whitney U test. IgG, immunoglobin G; GGN, ground glass nodule; AAH, atypical adenomatous hyperplasia; AIS, adenocarcinoma in situ; MIA, minimally invasive adenocarcinoma; IA, invasive adenocarcinoma; SNA, Sambucus nigra agglutinin; DSA, Datura stramonium agglutinin; NPA, Narcissus pseudonarcissus agglutinin; GNA, Galanthus nivalis agglutinin; WGA, Weat germ agglutinin.

Figure 3

Validation of differential signals of SNA between noninvasive and invasive GGNs by lectin blot. (A) SNA-blot of plasma IgG were performed in 16 noninvasive (2 benign and 14 AIS) and 32 invasive (16 MIA and 16 IA) GGN patients. The intensity of IgG detected by Western blot was used as an internal control. (B) The expression of SNA signal was normalized to IgG. The ratio (SNA/IgG) between noninvasive and invasive GGNs showed a significant difference (p = 0.009). (C) Correlation analysis between the intensity of SNA signal from lectin blot and lectin microarray was performed (Spearman correlation coefficient = 0.412, p = 0.004). GGN, ground glass nodule; SNA, Sambucus nigra agglutinin.

Figure 4

Diagnostic accuracy for prediction of invasive GGNs in the test set. (A) Receiver-operating characteristic (ROC) curves for differentiating invasive GGNs from noninvasive GGNs by measuring the CT values, five tumor biomarkers CEA, Cyfra21-1, SCC-Ag, NSE, and CA125 (named “Clinical biomarkers”), five significantly differential lectin signals SNA, DSA, GNA, EEL, and VVA (named “Lectins”), and the combined models with CT values (named “CT + Clinical biomarkers” and “CT + Lectins”, respectively). The area under the ROC curve (AUC) of the indicators was compared by the Delong test (CT vs. CT + Lectins, p = 0.007). (B) Diagnostic performance of the CT value and the new indicators for the assessment of invasive GGNs. GGN, ground glass nodule; CEA, carcinoembryonic antigen; Cyfra21-1, cytokeratin 19 fragments; SCC-Ag, squamous cell carcinoma antigen; NSE, neuron-specific enolase; CA125, carbohydrate antigen 125; SNA, Sambucus nigra agglutinin; DSA, Datura stramonium agglutinin; GNA, Galanthus nivalis agglutinin; EEL, Euonymus europaeus lectin; VVA, Vicia villosa agglutinin.