Aquaporin (AQP) channels in endometrial cancer (EC) cells are of interest as pharmacological targets to reduce tumor progression. A panel of compounds, including AQP1 ion channel inhibitors (AqB011 and 5-(phenoxymethyl) furan-2-carbaldehyde, PMFC), were used to test the hypothesis that inhibition of key AQPs can limit the invasiveness of low- and high-grade EC cells. We evaluated the effects on transwell migration in EC cell lines (Ishikawa, MFE-280) and primary EC cells established from surgical tissues (n= 8). Quantitative PCR uncovered classes ofAQPsnot previously reported in EC that are differentially regulated by hormonal signaling. With estradiol, Ishikawa showed increasedAQPs 5,11,12, and decreasedAQPs 0and4; MFE-280 showed increasedAQPs 0,1,3,4,8, and decreasedAQP11. Protein expression was confirmed by Western blot and immunocytochemistry. AQPs 1, 4, and 11 were colocalized with plasma membrane marker; AQP8 was intracellular in Ishikawa and not detectable in MFE-280. AQP1 ion channel inhibitors (AqB011; PMFC) reduced invasiveness of EC cell lines in transwell chamber and spheroid dispersal assays. In Ishikawa cells, transwell invasiveness was reduced ~41% by 80 µM AqB011 and ~55% by 0.5 mM 5-PMFC. In MFE-280, 5-PMFC inhibited invasion by ~77%. In contrast, proposed inhibitors of AQP water pores (acetazolamide, ginsenoside, KeenMind, TGN-020, IMD-0354) were not effective. Treatments of cultured primary EC cells with AqB011 or PMFC significantly reduced the invasiveness of both low- and high-grade primary EC cells in transwell chambers. We confirmed the tumors expressed moderate to high levels of AQP1 detected by immunohistochemistry, whereas expression levels of AQP4, AQP8, and AQP11 were substantially lower. The anti-invasive potency of AqB011 treatment for EC tumor tissues showed a positive linear correlation with AQP1 expression levels. In summary, AQP1 ion channels are important for motility in both low- and high-grade EC subtypes. Inhibition of AQP1 is a promising strategy to inhibit EC invasiveness and improve patient outcomes.
水通道蛋白（AQP）在子宫内膜癌（EC）细胞中作为减少肿瘤进展的药理学靶点备受关注。本研究采用一组化合物，包括AQP1离子通道抑制剂（AqB011和5-(苯氧甲基)呋喃-2-甲醛，PMFC），验证抑制关键AQP可限制低级别与高级别EC细胞侵袭能力的假说。我们评估了这些化合物对EC细胞系（Ishikawa、MFE-280）及手术组织来源的原代EC细胞（n=8）在Transwell迁移实验中的影响。定量PCR揭示了EC中既往未报道的AQP亚型，其表达受激素信号差异调控：雌二醇处理后，Ishikawa细胞中AQP5、11、12表达上调，AQP0、4下调；MFE-280细胞中AQP0、1、3、4、8上调，AQP11下调。蛋白表达经Western blot和免疫细胞化学验证，AQP1、4、11与质膜标志物共定位；AQP8在Ishikawa细胞内表达且未在MFE-280中检出。AQP1离子通道抑制剂（AqB011；PMFC）在Transwell小室和球体分散实验中均能降低EC细胞系的侵袭能力：80 µM AqB011使Ishikawa细胞侵袭降低约41%，0.5 mM 5-PMFC降低约55%；5-PMFC使MFE-280细胞侵袭抑制率达77%。而针对AQP水孔结构的抑制剂（乙酰唑胺、人参皂苷、KeenMind、TGN-020、IMD-0354）均未显效。AqB011或PMFC处理培养的原代EC细胞后，低级别与高级别原代EC细胞在Transwell中的侵袭能力均显著降低。免疫组化证实肿瘤组织表达中高水平AQP1，而AQP4、8、11表达显著较低。AqB011对EC肿瘤组织的抗侵袭效力与AQP1表达水平呈正线性相关。综上，AQP1离子通道对低级别与高级别EC亚型的运动能力至关重要，抑制AQP1是抑制EC侵袭性、改善患者预后的潜在策略。
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