Ramzan Rehana, Bukhari Shazia Anwer, Rasul Azhar
Department of Biochemistry, Government College University, Faisalabad, Pakistan.
Department of Zoology, Baba Guru Nanak University, Nankana Sahib, Pakistan.
PLoS One. 2025 Sep 8;20(9):e0331735. doi: 10.1371/journal.pone.0331735. eCollection 2025.
Secreted frizzled-related protein 4 (sFRP4) plays a fundamental role in the regulation of Wnt signalling, which is crucial for cellular proliferation and differentiation. The sFRP4 has garnered significant interest as a therapeutic target for metabolic diseases and cancer due to its mechanism of action. Although existing sFRP4 modulators show limited specificity and notable off-target effects, our study explores the potential of known bioactive compounds as more selective and less toxic alternatives. This study is based on the analysis of expression profiles, which demonstrated that the sFRP4 gene exhibits aberrant expression in multiple cancers, including breast cancer. The protein's primary involvement in cancer signaling pathways was determined through pathway enrichment analysis. The study employed molecular docking analyses and MD simulations to identify breast cancer-fighting small molecules with docking energies of less than -6 kcal/mol, targeting the sFRP4 binding hotspot using 100 natural or synthetic small molecules. Out of 100 screened compounds, Silibinin and Isotretinoin were selected based on docking results and further validated in vitro. In vitro investigations were carried out using the colorimetric MTT assay to assess cell viability and cytotoxicity based on metabolic activity. The potential of Silibinin and isotretinoin to upregulate the tumour suppressor sFRP4 was further examined using ELISA and real-time quantitative PCR. Our study identified potential compounds for high-potential drug candidates against sFRP4, demonstrating their effectiveness in cancer cell death and upregulating sFRP4 expression through improved drug design methods and experimental studies. In conclusion, our in-silico findings could facilitate the discovery of potential therapeutic agents against breast cancer. Silibinin and Isotretinoin impede cancer cell development in vitro; nonetheless, this study demonstrated that they directly upregulate sFRP4 and induce apoptosis in breast cancer cells.
分泌型卷曲相关蛋白4（sFRP4）在Wnt信号通路的调节中发挥着重要作用，而Wnt信号通路对细胞增殖和分化至关重要。由于其作用机制，sFRP4作为代谢性疾病和癌症的治疗靶点已引起了广泛关注。尽管现有的sFRP4调节剂显示出有限的特异性和明显的脱靶效应，但我们的研究探索了已知生物活性化合物作为更具选择性和更低毒性替代品的潜力。本研究基于表达谱分析，结果表明sFRP4基因在包括乳腺癌在内的多种癌症中存在异常表达。通过通路富集分析确定了该蛋白在癌症信号通路中的主要作用。该研究采用分子对接分析和分子动力学模拟，使用100种天然或合成小分子靶向sFRP4结合热点，以识别对接能量小于-6 kcal/mol的抗乳腺癌小分子。在筛选出的100种化合物中，根据对接结果选择了水飞蓟宾和异维A酸，并在体外进一步验证。体外研究采用比色MTT法，根据代谢活性评估细胞活力和细胞毒性。使用酶联免疫吸附测定（ELISA）和实时定量PCR进一步检测水飞蓟宾和异维A酸上调肿瘤抑制因子sFRP4的潜力。我们的研究确定了针对sFRP4的高潜力药物候选物的潜在化合物，通过改进的药物设计方法和实验研究证明了它们在癌细胞死亡和上调sFRP4表达方面的有效性。总之，我们的计算机模拟研究结果有助于发现潜在的抗乳腺癌治疗药物。水飞蓟宾和异维A酸在体外可阻碍癌细胞的发展；尽管如此，本研究表明它们可直接上调sFRP4并诱导乳腺癌细胞凋亡。
