Fig 2: ERO1L activity is essential for its growth-promoting role in PDAC. (A) Western blotting analysis of AsPC1 and BxPC3 cells after overexpressing either wide type ERO1L or its inactive mutant C394A. (B) Cell viability of ERO1L-WT or ERO1L-C394A overexpressing AsPC1 and BxPC3 cells (n = 3). (C) The long-term effect of ERO1L-WT or ERO1L-C394A overexpression in AsPC1 and BxPC3 cells was revealed by colony formation assay (n = 3). (D-E) AsPC1-vector, AsPC1-ERO1L-WT, and AsPC1-ERO1L-C394A cells were subcutaneously injected into the left and right hind limbs of 5 nude mice; gross xenografts (D) and tumor weights (E) were shown. *P < 0.05.

Fig 3: EMT contributes to ERO1L-mediated hepatic metastasis of PDAC. (a) Down reg (log2(HM/PT) < −0.25) and otherwise (log2(HM/PT) ≥ −0.25) were divided, based on the rate of ERO1L expression in hepatic metastasis and primary tumor. (b) The relatively lower ERO1L expression in hepatic metastasis leading to the lower activity of EMT. (c) GSEA suggesting lower ERO1L expression in hepatic metastasis would remarkably inhibit the activity of EMT pathway and whereas improve the activity of G2M CHECPIONT pathway positively correlated with pancreatic cancer metastasis. (d) Western blotting analysis of Capan-2 and MiaPaCa-2 cells after inhibiting ERO1L expression with shRNAs and EN460.

Fig 4: Pharmacological inhibition of ERO1L suppresses hepatic metastasis in vivo. a, b In vivo growth assay suggesting that ERO1L inhibitor, EN460 significantly decreased the number of liver nodules from Panc02 cell transplanted mice. Gross xenograft (a) and HE-staining images (b) were shown. (c) Kaplan-Meier survival curves showing the survival time of Panc02 PDAC cell transplanted mice treated with/without EN460 (log-rank test, P = 0.005). (d) Representative immunohistochemistry images showing EN460 suppressed liver nodules growth of PDAC in vivo. Scale bar: 50 μm.

Fig 5: Pan-cancer perspective of ERO1L gene expression profile and prognostic potential. (A) Comparison of ERO1L expression level in 9664 tumor tissue samples and 5539 normal cases. Data were derived from TCGA cohort and GTEx. (B) Comparison of ERO1L expression level in tumor tissue samples with different TNM stage. (C) The expression level of ERO1L in different tumor tissues and their normal counterparts. (D) Kaplan-Meier graphs showing significant association of ERO1L expression with patients' survival. (E) Prognostic analysis of ERO1L in different human cancers. The median expression value of ERO1L was used as a cutoff. HR: hazard ratio. ACC, Adrenocortical carcinoma; BLCA, Bladder urothelial carcinoma; BRCA, Breast invasive carcinoma; CESC, Cervical squamous cell carcinoma and endocervical adenocarcinoma; CHOL, Cholangio carcinoma; COAD, Colon adenocarcinoma; DLBC, Lymphoid neoplasm diffuse large B-cell lymphoma; ESCA, Esophageal carcinoma; GBM, Glioblastoma multiforme; HNSC, Head and neck squamous cell carcinoma; KICH, Kidney chromophobe; KIRC, Kidney renal clear cell carcinoma; KIRP, Kidney renal papillary cell carcinoma; LAML, Acute myeloid leukemia; LGG, Brain lower grade glioma; LIHC, Liver hepatocellular carcinoma; LUAD, Lung adenocarcinoma; LUSC, Lung squamous cell carcinoma; MESO, Mesothelioma; OV, Ovarian serous cystadenocarcinoma; PAAD, Pancreatic adenocarcinoma; PCPG, Pheochromocytoma and paraganglioma; PRAD, Prostate adenocarcinoma; READ, Rectum adenocarcinoma; SARC, Sarcoma; SKCM, Skin cutaneous melanoma; STAD, Stomach adenocarcinoma; TGCT, Testicular germ cell tumors; THCA, Thyroid carcinoma; THYM, Thymoma; UCEC, Uterine corpus endometrial carcinoma; UCS, Uterine carcinosarcoma; UVM, Uveal melanoma.