Fig 1: Malate dehydrogenase 2 as a diagnostic biomarker in distinguishing stage I patients with NSCLC from control subjects. A, Comparison by ELISA of the MDH2 concentration in urine between control subjects (n = 239) and patients with stage I NSCLC (n = 318) of the retrospective cohort P <.0001 by Mann‐Whitney U tests. B, ROC analysis of the detectable efficiency of MDH2 in the retrospective cohort in control subjects vs patients with stage I NSCLC (AUC = 0.7679, 95% CI:0.7291‐0.8066, P <.0001). C, Comparison of the MDH2 concentration in urine between control subjects (n = 493) and patients with NSCLC (n = 769) of the prospective cohort by ELISA P <.0001 by Mann‐Whitney U tests. D, ROC analysis of the detectable efficiency of MDH2 in the prospective cohort in control subjects vs patients with NSCLC (AUC = 0.6791, 95% CI: 0.6495‐0.7088, P <.0001). E, Comparison of the MDH2 concentration in urine between control subjects (n = 493) and patients with stage I NSCLC (n = 503) of the prospective cohort using ELISA P <.0001 by Mann‐Whitney U tests. F, ROC analysis of the detectable efficiency of MDH2 in the prospective cohort in control subjects vs stage I NSCLC patients (AUC = 0.7234, 95% CI: 0.6916‐0.7551, P <.0001). Scatter diagrams are represented as median with interquartile range and the vertical axis shows the square root (sqrt) of urinary MDH2 concentration. Black arrows show the cut‐off point
Fig 2: Malate dehydrogenase 2 expression is upregulated in lung cancer tissues. A, MDH2 RNA expression levels in paired normal lung tissue and cancer tissue (n = 19) were analyzed using qPCR. MDH2 expression is higher in lung cancer tissues compared with normal lung tissues. P =.0067 using Wilcoxon test. B, MDH2 protein levels in paired normal lung tissue and lung cancer tissue (n = 8) determined in triplicate using western blot analysis. The column graph indicates semi-quantitative analysis. P =.0078 using Wilcoxon test. C, Representative images of immunohistochemical staining of MDH2 in LUAD, LUSC and normal lung tissue. Magnification ×10 (top) and ×20 (bottom). LUAD, lung adenocarcinoma; LUSC, lung squamous carcinoma
Fig 3: Knockdown of MDH2 inhibits lung cancer cell proliferation. A, MDH2 expression levels in different lung cancer cell lines are significantly higher compared with that in the Bease2B cell (all P <.05). B, MDH2 expression levels in the A549, PC9, and Hop 62 cell lines are 6.3‐, 6.9‐, and 5.3‐fold lower after knockdown. All P <.0001 indicate Mann‐Whitney U tests. C, Cell proliferation assay of A549 (left), Hop62 (middle) and PC9 (right) after MDH2 knockdown. Red dots show the MDH2‐knockdown groups and blue dots show the control groups. Differences were calculated on the last day. All P ≤.0001 used paired t test. D, Colony‐forming analysis of A549, Hop62, and PC9 lines with MDH2 knockdown. Representative figures are shown on the left. Comparison of the clone counts between control and MDH2 knockdown cells. Data are presented as mean ± SEM. A549: P =.002, Hop62: P =.0014, PC9: P =.0014, using one‐way ANOVA. Con, control; Dox, doxycycline; KD, knockdown; OD, optical density
Fig 4: High urinary MDH2 levels were determined using DIA MS both in patients with NSCLC and lung cancer model mice. A, Schematic of discovery and assessment of lung cancer urinary biomarkers. B, Volcano plot of urine quantitative proteome results when comparing patients with lung cancer and healthy controls. Red dots show upregulated expression and green dots show downregulated expression in cancer. C, Heat map shows the top 20 most significantly upregulated urine proteins (fold change >2, P <.05) between patients with lung cancer (represented by T1-T4) and healthy controls (represented by N1-N4). D, Heat map shows the top 4 highly regulated urine proteins between EGFRL858R-driven and EGFR19DEL-driven lung cancer model mice and wild-type mice, which were consistent with those in human (fold change >2, P <.05). CTL, controls; WT, wild-type mice
Supplier Page from Abcam for Anti-MDH2 antibody [EPR14883(B)]