Department of Cancer Chemoprevention, Chittaranjan National Cancer Institute, 37, S. P. Mukherjee Road, Kolkata-700026, West Bengal, India.
Department of Pathology, N.R.S. Medical College, Kolkata-700014, West Bengal, India.
Food Funct. 2021 Feb 15;12(3):1063-1078. doi: 10.1039/d0fo02105a.
According to population-based studies, lung cancer has become one of the leading causes of death globally in males and is also rising in females at an alarming rate. The aim of this study was to exploit the inherent properties of eugenol to restrict the growth of cancer cells in a tobacco-related human carcinogen NDEA-induced lung carcinogenesis model in vivo as a chemopreventive agent. More precisely, by utilizing its abundance in nature, eugenol (a component of clove) was utilized to establish the molecular mechanism of chemoprevention in the NDEA-induced mouse lung carcinogenesis model in a substantial cost-effective manner and was validated in the A549 human lung cancer cell line. Our study especially targeted the tiny, drug-resistant, and most virulent subpopulation of cancer cells called CSCs by targeting their regulator molecule β-catenin. The non-toxic dosage of eugenol was shown to enhance apoptosis, simultaneously suppressing cell proliferation in the lung tissue of carcinogen-treated mice without affecting the normal mice. Combining cellular apoptosis and proliferation, eugenol showed an exceptional chemopreventive potential in this lung carcinogenesis model. Importantly, eugenol strongly restricted the lung carcinoma in the mild dysplastic stage as a chemopreventive agent. The molecular analysis remarkably depicted the restriction of β-catenin nuclear transportation. The minimized total β-catenin pool and induced N-terminal Ser37 phosphorylation form after eugenol treatment resulted in its cytoplasmic degradation. Consequently, CSC markers such as CD44, Oct4, EpCAM, and Notcht1, whose expression is dependent on β-catenin decreased significantly, as proven by IHC, ICC, and WB analysis both in vivo and in vitro. The in vitro secondary sphere formation assay also proved the remarkably repressed CSC population, and hence the virulence. In another way, eugenol was proven to significantly enhance the degradation of β-catenin when treated with the CK1α inhibitor D4476 in vitro by Western blot. CK1α in the Wnt/β-catenin pathway plays a crucial role for tagging with the N-terminal Ser45 phosphorylation of β-catenin, which ultimately opens a position for the decisive phosphorylation by GSK3β at the Ser37 residue to take place. Thus, the conclusive extermination of CSCs achieved that was associated with recurrence due to treatment failure. That can help to achieve a longer and better quality of life in a natural, economical way.
根据基于人群的研究，肺癌已成为男性全球范围内主要死亡原因之一，在女性中的发病率也以惊人的速度上升。本研究旨在利用丁香酚的固有特性，在体内烟草相关的人类致癌剂 NDEA 诱导的肺癌发生模型中作为化学预防剂来限制癌细胞的生长。更确切地说，利用其在自然界中的丰富度，以丁香酚（丁香的一种成分）为研究对象，以具有成本效益的方式在 NDEA 诱导的小鼠肺癌发生模型中建立化学预防的分子机制，并在 A549 人肺癌细胞系中进行验证。我们的研究特别针对被称为 CSCs 的微小、耐药且最具侵袭性的癌细胞亚群，通过靶向其调节分子β-catenin。非毒性剂量的丁香酚显示出增强凋亡的作用，同时抑制致癌物处理小鼠肺组织中的细胞增殖，而不影响正常小鼠。将细胞凋亡和增殖相结合，丁香酚在该肺癌发生模型中表现出卓越的化学预防潜力。重要的是，丁香酚作为化学预防剂强烈限制了轻度发育不良阶段的肺癌。分子分析显著描述了β-catenin 核转运的限制。丁香酚处理后总β-catenin 池减少，N 端 Ser37 磷酸化形成，导致其细胞质降解。因此，CSC 标志物（如 CD44、Oct4、EpCAM 和 Notcht1）的表达显著降低，免疫组化、免疫细胞化学和 WB 分析均证明了这一点，无论是在体内还是体外。体外二级球体形成实验也证明了 CSC 群体受到显著抑制，从而降低了其侵袭性。另一方面，通过 Western blot 证明，丁香酚与 CK1α 抑制剂 D4476 体外联合处理时，β-catenin 的降解显著增强。Wnt/β-catenin 通路中的 CK1α 在β-catenin 的 N 端 Ser45 磷酸化标记中起着至关重要的作用，最终为 GSK3β 在 Ser37 残基上进行决定性磷酸化开辟了位置。因此，实现了与治疗失败相关的 CSCs 的最终根除，这与复发有关。这可以帮助以自然、经济的方式实现更长、更好的生活质量。
