Two new bracelet cyclotides from roots of Pombalia calceolaria with potential anticancer activity have been characterized in this work. The cyclotides Poca A and B (1 and 2) and the previously known CyO4 (3) were de novo sequenced by MALDI-TOF/TOF mass spectrometry (MS). The MS2 spectra were examined and the amino acid sequences were determined. The purified peptides were tested for their cytotoxicity and effects on cell migration of MDA-MB-231, a triple-negative breast cancer cell line. The isolated cyclotides reduced the number of cancer cells by more than 80% at 20 μM, and the concentration-related cytotoxic responses were observed with IC50 values of 1.8, 2.7, and 9.8 μM for Poca A (1), Poca B (2), and CyO4 (3), respectively. Additionally, the inhibition of cell migration (wound-healing assay) exhibited that CyO4 (3) presents an interesting activity profile, in being able to inhibit cell migration (50%) at a subtoxic concentration (2 μM). The distribution of these cyclotides in the roots was analyzed by MALDI imaging, demonstrating that all three compounds are present in the phloem and cortical parenchyma regions.

中文翻译：

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从 Pombalia calceolaria 中分离的环肽抑制乳腺癌细胞迁移。

来自 Pombalia calceolaria 根的两种具有潜在抗癌活性的新手镯环肽已在这项工作中得到表征。通过 MALDI-TOF/TOF 质谱 (MS) 对环肽 Poca A 和 B（1 和 2）以及先前已知的 CyO4（3）进行了从头测序。检查MS2谱并确定氨基酸序列。测试纯化的肽的细胞毒性和对三阴性乳腺癌细胞系 MDA-MB-231 的细胞迁移的影响。分离的环肽在 20 μM 时使癌细胞数量减少了 80% 以上，并且观察到浓度相关的细胞毒性反应，Poca A (1)、Poca B (2) 的 IC50 值为 1.8、2.7 和 9.8 μM , 和 CyO4 (3)，分别。此外，细胞迁移的抑制（伤口愈合试验）表明 CyO4 (3) 呈现出有趣的活性特征，能够在亚毒性浓度 (2 μM) 下抑制细胞迁移 (50%)。通过 MALDI 成像分析了这些环化物在根中的分布，表明所有三种化合物都存在于韧皮部和皮质薄壁组织区域。