Borrelli Matthew J, Buensuceso Adrian, Ramos Valdes Yudith, Johnston Tiffany P A, Haagsma Jacob, Shepherd Trevor G
The Mary and John Knight Translational Ovarian Cancer Research Unit, Verspeeten Family Cancer Centre, London, ON, N6A 5W9, Canada.
Department of Anatomy and Cell Biology, Western University, London, ON, N6A 5C1, Canada.
Sci Rep. 2025 Jul 17;15(1):25913. doi: 10.1038/s41598-025-11584-9.
Epithelial ovarian cancer (EOC) metastasizes predominantly through multicellular aggregates known as spheroids, which disseminate within the peritoneal cavity and initiate secondary disease upon reattachment at distant sites. EOC spheroids resist detachment-induced cell death by upregulating stress responses including AMP-activated protein kinase (AMPK) signaling and AMPK-dependent macroautophagy (autophagy), highlighting these pathways as potential therapeutic targets. Previously, we used a pharmacological approach to putatively identify Ca/calmodulin-dependent protein kinase kinase 2 (CAMKKβ, encoded by CAMKK2) as the primary activator of AMPK in EOC spheroids. Herein we have generated CAMKK2 knockout EOC cell lines via CRISPR-Cas9 genome editing to confirm this function of CAMKKβ and explore the impacts of its loss using in vitro and in vivo models of metastatic EOC. CAMKK2 knockout spheroids exhibited decreased AMPK activation, autophagic flux, cell viability, and metastatic potential relative to parental spheroids, and intraperitoneal xenograft tumours lacking CAMKKβ grew slower than their CAMKKβ-intact counterparts. Effect magnitudes varied between cell line models, suggesting context-dependent roles for CAMKKβ in EOC and rationalizing further studies to characterize the underlying mechanisms. Altogether, our findings highlight CAMKKβ as an important contributor to metabolic reprogramming in EOC spheroids and as a potential therapeutic target in the setting of advanced disease.
上皮性卵巢癌（EOC）主要通过称为球体的多细胞聚集体发生转移，这些球体在腹腔内扩散，并在远处重新附着时引发继发性疾病。EOC球体通过上调包括AMP激活的蛋白激酶（AMPK）信号传导和AMPK依赖性巨自噬（自噬）在内的应激反应来抵抗脱离诱导的细胞死亡，突出了这些途径作为潜在治疗靶点的地位。此前，我们采用药理学方法推测性地确定钙/钙调蛋白依赖性蛋白激酶激酶2（CAMKKβ，由CAMKK2编码）是EOC球体中AMPK的主要激活剂。在此，我们通过CRISPR-Cas9基因组编辑产生了CAMKK2基因敲除EOC细胞系，以证实CAMKKβ的这一功能，并使用转移性EOC的体外和体内模型探索其缺失所产生的影响。相对于亲本球体，CAMKK2基因敲除球体表现出AMPK激活、自噬通量、细胞活力和转移潜力降低；缺乏CAMKKβ 的腹腔内异种移植肿瘤比具有完整CAMKKβ 的肿瘤生长得更慢。细胞系模型之间的效应大小有所不同,表明CAMKKβ在EOC中具有依赖于背景的作用，并为进一步研究以阐明潜在机制提供了理论依据。总之，我们的研究结果突出了CAMKKβ是EOC球体代谢重编程的重要贡献者，也是晚期疾病背景下的潜在治疗靶点。
