背景:白血病抑制因子(LIF)是IL-6细胞因子家族的多功能成员,通过与细胞表面上由LIFR和gp130组成的异二聚体结合来激活下游信号传导途径。先前的研究表明,LIF在各种肿瘤组织中高表达(例如胰腺癌,乳腺癌,前列腺癌,和结肠直肠癌)并促进癌细胞增殖,迁移,入侵,和差异化。此外,LIF的过度表达与不良的临床病理特征相关。因此,我们假设LIF可能是一个有希望的癌症治疗靶点.在这项工作中,我们开发了针对LIF的拮抗剂抗体1G11,并在小鼠模型中研究了其抗肿瘤机制及其治疗效果。
结果:从幼稚的人scFv噬菌体文库中筛选了一系列靶向LIF的单链可变片段(scFv)。这些scFv以完全IgG形式重建并由哺乳动物瞬时表达系统产生。在抗体中,1G11对人表现出优异的结合活性,食蟹猴和小鼠LIF。功能分析表明1G11可以阻断LIF与LIFR的结合并抑制细胞内STAT3磷酸化信号。有趣的是,1G11不阻断LIF与gp130的结合,gp130是另一种参与与LIFR-起形成受体复合物的LIF受体。在体内,在结直肠癌的CT26和MC38模型中,腹膜内施用1G11抑制肿瘤生长。IHC分析显示肿瘤组织中p-STAT3和Ki67降低,而c-caspase3增加。此外,1G11治疗改善CD3+,肿瘤组织中CD4+和CD8+T细胞浸润。
结论:我们从幼稚的人scFv噬菌体文库中开发了靶向LIF/LIFR信号通路的拮抗剂抗体。拮抗剂抗LIF抗体通过特异性降低p-STAT3发挥抗肿瘤作用。进一步的研究表明,抗LIF抗体1G11增加了肿瘤组织中的免疫细胞浸润。
BACKGROUND: Leukemia inhibitory factor (
LIF) is a multifunctional member of the IL-6 cytokine family that activates downstream signaling pathways by binding to the heterodimer consisting of LIFR and gp130 on the cell surface. Previous research has shown that LIF is highly expressed in various tumor tissues (e.g. pancreatic cancer, breast cancer, prostate cancer, and colorectal cancer) and promotes cancer cell proliferation, migration, invasion, and differentiation. Moreover, the overexpression of
LIF correlates with poor clinicopathological characteristics. Therefore, we hypothesized that
LIF could be a promising target for the treatment of cancer. In this work, we developed the antagonist antibody 1G11 against LIF and investigated its anti-tumor mechanism and its therapeutic efficacy in mouse models.
RESULTS: A series of single-chain variable fragments (scFvs) targeting LIF were screened from a naive human scFv phage library. These scFvs were reconstructed in complete IgG form and produced by the mammalian transient expression system. Among the antibodies, 1G11 exhibited the excellent binding activity to human, cynomolgus monkey and mouse LIF. Functional analysis demonstrated 1G11 could block LIF binding to LIFR and inhibit the intracellular STAT3 phosphorylation signal. Interestingly, 1G11 did not block
LIF binding to gp130, another
LIF receptor that is involved in forming the receptor complex together with LIFR. In vivo, intraperitoneal administration of 1G11 inhibited tumor growth in CT26 and MC38 models of colorectal cancer. IHC analysis demonstrated that p-STAT3 and Ki67 were decreased in tumor tissue, while c-caspase 3 was increased. Furthermore, 1G11 treatment improves CD3+, CD4 + and CD8 + T cell infiltration in tumor tissue.
CONCLUSIONS: We developed antagonist antibodies targeting LIF/LIFR signaling pathway from a naive human scFv phage library. Antagonist anti-
LIF antibody exerts antitumor effects by specifically reducing p-STAT3. Further studies revealed that anti-LIF antibody 1G11 increased immune cell infiltration in tumor tissues.