曲妥珠单抗是唯一用于治疗Her2阳性进展期胃癌的一线单抗药物，其耐药已严重影响靶向治疗效果且分子机制尚未阐明。课题组前期研究成功筛选胃癌曲妥珠单抗耐药候选基因6磷酸果糖激酶-2/果糖双磷酸-2同工酶3（PFKFB3），初步证实抑制PFKFB3可恢复肿瘤血管正常化并增强治疗敏感性。预实验发现，耐药后PFKFB3发生核异位，且其核异位可促进CXCL8分泌，而这可能与Smad3转录激活相关。据此，我们提出假设：胃癌细胞PFKFB3核异位发挥激酶功能磷酸化Smad3，增强其转录活性调控CXCL8的转录及分泌，后者可通过激活MMPs降解血管基膜、破坏肿瘤屏障影响肿瘤血管正常化，从而诱导曲妥珠单抗耐药。项目立足肿瘤微环境血管正常化，结合分子生物学方法，体内外全方位阐明PFKFB3核异位介导胃癌曲妥珠单抗耐药的分子机制，为确立PFKFB3-CXCL8轴作为临床克服曲妥珠单抗耐药新靶点提供充分的科学依据。

Trastuzumab is the only first-line monoclonal antibody used in the treatment of advanced Her2-positive gastric cancer, but the resistance seriously affects the therapeutic effect and the molecular mechanism remains unknown. Previous transcriptional sequencing found that 6-Phosphofructo-2- Kinase/Fructose-2,6-Biphosphatase 3 (PFKFB3) increased in Her2 positive gastric cancer PDX model after trastuzumab resistance, and inhibition of PFKFB3 could restore tumor vascular normalization and enhance therapeutic sensitivity. In the preliminary experiments, we found that PFKFB3 nuclear translocation occurred after resistance, and its translocation could promote the secretion of downstream CXCL8, which may have related to Smad3 transcriptional activation. Therefore, we hypothesis that the nuclear translocation of PFKFB3 plays a kinase role in promoting the phosphorylation of Smad3 and activating its transcriptional activity, which then binds to CXCL8 promoter region to promote its transcription and extracellular secretion. Tumor-derived CXCL8 can affect tumor vascular normalization and induce drug resistance by degrading the vascular basement membrane and destroying the tumor barrier function via activating MMPs. Based on the vascular normalization in the tumor microenvironment and combined with molecular biology methods, the project comprehensively elucidated the molecular mechanism of PFKFB3 promoting trastuzumab resistance in gastric cancer in vivo and in vitro, providing sufficient scientific basis for establishing the PFKFB3-CXCL8 axis as a new target for overcoming clinical trastuzumab resistance.