PI3K/AKT途径的失调在高级别浆液性卵巢癌（HGSOC）中常见，HGSOC中肿瘤抑制因子PTEN的丧失与预后差相关。PTEN丧失对HGSOC的细胞机制尚不清楚。我们利用HGSOC球状体的时间序列成像和机器学习方法来分类PTEN丧失的表型。PTEN缺陷诱导了富含PI（3,4,5）P3和依赖于其的细胞质膜突出进入细胞外基质（ECM），从而导致一种集体入侵表型。我们发现小的GTP酶ARF6在PTEN丧失时对HGSOC细胞至关重要。通过功能蛋白质组学CRISPR筛选ARF6相互作用物，我们确定ARF GTP酶激活蛋白（GAP）AGAP1和ECM受体β1整合素（ITGB1）是调控HGSOC中PTEN丧失相关入侵的关键ARF6相互作用物。ARF6通过控制内吞的活性β1整合素的回收以维持对ECM的入侵活性来促进入侵。CYTH2-ARF6-AGAP1复合物在HGSOC患者中的表达与预后呈负相关，可用于鉴定预后良好和差的患者群体。ARF6可能是PTEN缺失的HGSOC中的治疗靶点。© 2023 The Authors. Published under the terms of the CC BY 4.0 license.

Dysregulation of the PI3K/AKT pathway is a common occurrence in high-grade serous ovarian carcinoma (HGSOC), with the loss of the tumour suppressor PTEN in HGSOC being associated with poor prognosis. The cellular mechanisms of how PTEN loss contributes to HGSOC are largely unknown. We here utilise time-lapse imaging of HGSOC spheroids coupled to a machine learning approach to classify the phenotype of PTEN loss. PTEN deficiency induces PI(3,4,5)P3 -rich and -dependent membrane protrusions into the extracellular matrix (ECM), resulting in a collective invasion phenotype. We identify the small GTPase ARF6 as a crucial vulnerability of HGSOC cells upon PTEN loss. Through a functional proteomic CRISPR screen of ARF6 interactors, we identify the ARF GTPase-activating protein (GAP) AGAP1 and the ECM receptor β1-integrin (ITGB1) as key ARF6 interactors in HGSOC regulating PTEN loss-associated invasion. ARF6 functions to promote invasion by controlling the recycling of internalised, active β1-integrin to maintain invasive activity into the ECM. The expression of the CYTH2-ARF6-AGAP1 complex in HGSOC patients is inversely associated with outcome, allowing the identification of patient groups with improved versus poor outcome. ARF6 may represent a therapeutic vulnerability in PTEN-depleted HGSOC.© 2023 The Authors. Published under the terms of the CC BY 4.0 license.