(Background). Canine mammary tumors (CMTs) have emerged as an important model for understanding pathophysiological aspects of human disease. Liquid biopsy (LB), which relies on blood-borne biomarkers and offers minimal invasiveness, holds promise for reflecting the disease status of patients. Small extracellular vesicles (SEVs) and their protein cargo have recently gained attention as potential tools for disease screening and monitoring. (Objectives). This study aimed to isolate SEVs from canine patients and analyze their proteomic profile to assess their diagnostic and prognostic potential. (Methods). Plasma samples were collected from female dogs grouped into CMT (malignant and benign), healthy controls, relapse, and remission groups. SEVs were isolated and characterized using ultracentrifugation (UC), nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM). Proteomic analysis of circulating SEVs was conducted using liquid chromatography–mass spectrometry (LC–MS). (Results). While no significant differences were observed in the concentration and size of exosomes among the studied groups, proteomic profiling revealed important variations. Mass spectrometry identified exclusive proteins that could serve as potential biomarkers for mammary cancer. These included Inter-alpha-trypsin inhibitor heavy chain (ITIH2 and ITI4), phosphopyruvate hydratase or alpha enolase (ENO1), eukaryotic translation elongation factor 2 (eEF2), actin (ACTB), transthyretin (TTR), beta-2-glycoprotein 1 (APOH) and gelsolin (GSN) found in female dogs with malignant tumors. Additionally, vitamin D-binding protein (VDBP), also known as group-specific component (GC), was identified as a protein present during remission. (Conclusions). The results underscore the potential of proteins found in SEVs as valuable biomarkers in CMTs. Despite the lack of differences in vesicle concentration and size between the groups, the analysis of protein content revealed promising markers with potential applications in CMT diagnosis and monitoring. These findings suggest a novel approach in the development of more precise and effective diagnostic tools for this challenging clinical condition.
（背景）犬乳腺肿瘤已成为理解人类疾病病理生理学方面的重要模型。液体活检依赖于血液中的生物标志物，具有微创性，有望反映患者的疾病状态。小细胞外囊泡及其携带的蛋白质作为疾病筛查和监测的潜在工具，近年来备受关注。（目的）本研究旨在从犬类患者体内分离小细胞外囊泡，并分析其蛋白质组学特征，以评估其诊断和预后潜力。（方法）采集雌性犬的血浆样本，分为乳腺肿瘤组（恶性和良性）、健康对照组、复发组和缓解组。采用超速离心法分离小细胞外囊泡，并通过纳米颗粒追踪分析和透射电子显微镜进行表征。使用液相色谱-质谱联用技术对循环小细胞外囊泡进行蛋白质组学分析。（结果）虽然各组间外泌体的浓度和大小未观察到显著差异，但蛋白质组学分析揭示了重要变化。质谱鉴定出可作为乳腺肿瘤潜在生物标志物的特异性蛋白质，包括在恶性肿瘤雌性犬中发现的α-胰蛋白酶抑制剂重链、磷酸丙酮酸水合酶或α-烯醇化酶、真核翻译延伸因子2、肌动蛋白、转甲状腺素蛋白、β2-糖蛋白1和凝溶胶蛋白。此外，维生素D结合蛋白（也称为群体特异性成分）被确定为缓解期存在的蛋白质。（结论）研究结果强调了小细胞外囊泡中蛋白质作为犬乳腺肿瘤重要生物标志物的潜力。尽管各组间囊泡浓度和大小无差异，但蛋白质含量分析揭示了在犬乳腺肿瘤诊断和监测中具有潜在应用前景的标志物。这些发现为开发针对这一临床挑战的更精准有效的诊断工具提供了新思路。
肿瘤(癌症)患者之家