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Objective: We aimed to develop a novel anti-HIF-1α intrabody to decrease gemcitabine resistance in pancreatic cancer patients. Methods: Surface plasmon resonance and glutathione S-transferase pull-down assays were conducted to identify the binding affinity and specificity of anti-HIF-1α VHH212 [a single-domain antibody (nanobody)]. Molecular dynamics simulation was used to determine the protein-protein interactions between hypoxia-inducible factor-1α (HIF-1α) and VHH212. The real-time polymerase chain reaction (PCR) and Western blot analyses were performed to identify the expressions of HIF-1α and VEGF-A in pancreatic ductal adenocarcinoma cell lines. The efficiency of the VHH212 nanobody in inhibiting the HIF-1 signaling pathway was measured using a dual-luciferase reporter assay. Finally, a PANC-1 xenograft model was developed to evaluate the anti-tumor efficiency of combined treatment. Immunohistochemistry analysis was conducted to detect the expressions of HIF-1α and VEGF-A in tumor tissues. Results: VHH212 was stably expressed in tumor cells with low cytotoxicity, high affinity, specific subcellular localization, and neutralization of HIF-1α in the cytoplasm or nucleus. The binding affinity between VHH212 and the HIF-1α PAS-B domain was 42.7 nM. Intrabody competitive inhibition of the HIF-1α heterodimer with an aryl hydrocarbon receptor nuclear translocator was used to inhibit the HIF-1/VEGF pathway in vitro. Compared with single agent gemcitabine, co-treatment with gemcitabine and a VHH212-encoding adenovirus significantly suppressed tumor growth in the xenograft model with 80.44% tumor inhibition. Conclusions: We developed an anti-HIF-1α nanobody and showed the function of VHH212 in a preclinical murine model of PANC-1 pancreatic cancer. The combination of VHH212 and gemcitabine significantly inhibited tumor development. These results suggested that combined use of anti-HIF-1α nanobodies with first-line treatment may in the future be an effective treatment for pancreatic cancer.

Objective: Cystic lesions of the pancreas have been increasingly recognized. Some lesions exhibit benign behavior, while others have unequivocal malignant potential. Thus, accurate identification of malignancy in patients diagnosed with pancreatic cystic neoplasms (PCNs) remains a major challenge. The aim of this study was to define a combined criterion to better predict malignant lesions in patients with PCNs.Methods: We retrospectively analyzed 165 patients who underwent resection of PCNs from October 2011 to May 2017. The relationship among malignancy and serum carbohydrate antigen 19-9 (CA19-9), preoperative neutrophil-to-lymphocyte ratio (NLR), and the presence of enhanced solid component on imaging was analyzed.Results: NLR before surgery in patients with malignant PCNs (2.81±2.14) was significantly higher than that in patients diagnosed with pancreatic neuroendocrine tumor (1.90±0.69, P=0.013) or healthy volunteers (1.40±0.48; P1.976 and presence of enhanced solid component were independent predictors of PCN malignancy. A combined criterion meeting any two or more of the three elements including CA19-9 ≥39 U/mL, NLR >1.976, and presence of enhanced solid component on computed tomography imaging is an indicator with a high positive predictive value of 80.5% and a high negative predictive value of 87.9%, and thus, represents a highly accurate test (86.1%).Conclusions: The new combined criterion is an effective predictor of tumor malignancy in patients with PCNs.