RT Journal Article SR Electronic T1 VHH212 nanobody targeting the hypoxia-inducible factor 1α suppresses angiogenesis and potentiates gemcitabine therapy in pancreatic cancer in vivo JF Cancer Biology and Medicine JO Cancer Biol Med FD China Anti-Cancer Association SP 772 OP 787 DO 10.20892/j.issn.2095-3941.2020.0568 VO 18 IS 3 A1 Guangbo Kang A1 Min Hu A1 He Ren A1 Jiewen Wang A1 Xin Cheng A1 Ruowei Li A1 Bo Yuan A1 Yasmine Balan A1 Zixuan Bai A1 He Huang YR 2021 UL http://www.cancerbiomed.org/content/18/3/772.abstract AB 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.