iBET, Instituto de Biologia Experimental e Tecnológica, Oeiras, Portugal.
Instituto de Tecnologia Química e Biológica António Xavier, Oeiras, Portugal.
J Exp Clin Cancer Res. 2020 Aug 17;39(1):161. doi: 10.1186/s13046-020-01653-4.
Estrogen receptor α (ERα) signaling is a defining and driving event in most breast cancers; ERα is detected in malignant epithelial cells of 75% of all breast cancers (classified as ER-positive breast cancer) and, in these cases, ERα targeting is the main therapeutic strategy. However, the biological determinants of ERα heterogeneity and the mechanisms underlying therapeutic resistance are still elusive, hampered by the challenges in developing experimental models recapitulative of intra-tumoral heterogeneity and in which ERα signaling is sustained. Ex vivo cultures of human breast cancer tissue have been proposed to retain the original tissue architecture, epithelial and stromal cell components and ERα. However, loss of cellularity, viability and ERα expression are well-known culture-related phenomena.
BC samples were collected and brought to the laboratory. Then they were minced, enzymatically digested, entrapped in alginate and cultured for 1 month. The histological architecture, cellular composition and cell proliferation of tissue microstructures were assessed by immunohistochemistry. Cell viability was assessed by measurement of cell metabolic activity and histological evaluation. The presence of ERα was accessed by immunohistochemistry and RT-qPCR and its functionality evaluated by challenge with 17-β-estradiol and fulvestrant.
We describe a strategy based on entrapment of breast cancer tissue microstructures in alginate capsules and their long-term culture under agitation, successfully applied to tissue obtained from 63 breast cancer patients. After 1 month in culture, the architectural features of the encapsulated tissue microstructures were similar to the original patient tumors: epithelial, stromal and endothelial compartments were maintained, with an average of 97% of cell viability compared to day 0. In ERα-positive cases, fibers of collagen, the main extracellular matrix component in vivo, were preserved. ERα expression was at least partially retained at gene and protein levels and response to ERα stimulation and inhibition was observed at the level of downstream targets, demonstrating active ER signaling.
The proposed model system is a new methodology to study ex vivo breast cancer biology, in particular ERα signaling. It is suitable for interrogating the long-term effects of anti-endocrine drugs in a set-up that closely resembles the original tumor microenvironment, with potential application in pre- and co-clinical assays of ERα-positive breast cancer.
雌激素受体 α（ERα）信号是大多数乳腺癌的决定性和驱动事件；75%的所有乳腺癌（归类为 ER 阳性乳腺癌）的恶性上皮细胞中均检测到 ERα，在这些情况下，靶向 ERα 是主要的治疗策略。然而，ERα 异质性的生物学决定因素和治疗耐药的机制仍不清楚，这是因为在开发能够重现肿瘤内异质性且维持 ERα 信号的实验模型方面存在挑战。已经提出用人乳腺癌组织的体外培养来保留原始组织的结构、上皮和基质细胞成分以及 ERα。然而，细胞减少、活力丧失和 ERα 表达是众所周知的与培养相关的现象。
收集 BC 样本并带到实验室。然后将其切成小块，进行酶消化，包裹在藻酸盐中并培养 1 个月。通过免疫组织化学评估组织微结构的组织学结构、细胞组成和细胞增殖。通过测量细胞代谢活性和组织学评估来评估细胞活力。通过免疫组织化学和 RT-qPCR 评估 ERα 的存在，并通过用 17-β-雌二醇和氟维司群挑战来评估其功能。
我们描述了一种基于将乳腺癌组织微结构包裹在藻酸盐胶囊中并在搅拌下长期培养的策略，该策略成功应用于 63 名乳腺癌患者的组织。在培养 1 个月后，包裹的组织微结构的结构特征与原始患者肿瘤相似：上皮、基质和内皮隔室得以维持，与第 0 天相比，细胞活力平均为 97%。在 ERα 阳性病例中，纤维胶原，即体内主要的细胞外基质成分，得以保留。ERα 的表达在基因和蛋白水平上至少部分保留，并且在下游靶标水平上观察到对 ERα 刺激和抑制的反应，证明了活跃的 ER 信号。
所提出的模型系统是研究体外乳腺癌生物学，特别是 ERα 信号的新方法。它适用于在与原始肿瘤微环境非常相似的设置中研究抗内分泌药物的长期影响，具有 ERα 阳性乳腺癌的临床前和临床前检测的潜在应用。
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