Chatterjee Essha, Sharma Ram, Dey Biswajit, Singh Hoshiyar, Naik Aliva, Khan Anjesh, Basak Santanu, Bansode Ankush, Sachdeva Ritika, Sharma Anamika, Kumar Rahul, Naik Pradeep, Singh Pankaj Kumar, Nepali Kunal, Guru Santosh Kumar
Department of Biological Sciences, National Institute of Pharmaceutical Education and Research, Hyderabad, India.
Department of Biotechnology and Bioinformatics, Sambalpur University, Sambalpur, Odisha, India.
Front Pharmacol. 2025 Jun 5;16:1544666. doi: 10.3389/fphar.2025.1544666. eCollection 2025.
Lung cancer remains the leading cause of cancer-related deaths, necessitating novel therapeutic strategies. In this study, we developed RM-3-22, a TAZQ-based hydroxamic acid derivative with histone deacetylase (HDAC) inhibitory properties. We evaluated its anticancer activity in non-small cell lung cancer (NSCLC), using A549 adenocarcinoma cells as the primary model.
The anticancer efficiency of RM-3-22 was assessed in 2D and 3D cell culture models. Cell survivalism was analysed by MTT assay. Different microscopical staining methods, including acridine orange and DAPI, were employed to evaluate autophagy, nuclear changes, and apoptosis. Cell cycle progression, mitochondrial membrane potential, and apoptosis-necrosis profiles were assessed using flow cytometry. Protein and gene expression related to the RM-3-22 induced pathway were evaluated via immunofluorescence (IF), Western blotting, and RT-PCR. Functional gene analysis was performed using siRNA-mediated knockdown. Different in silico studies were also conducted to check the clinical relevance and expression pattern of the RM-3-22-induced gene. Additionally, the in vivo efficiency of the molecule was evaluated using the NOD/SCID xenograft model.
RM-3-22 potentially suppressed cell viability and decreased the tumor spheroid size of A549s in vitro. It induced autophagy via downregulation of PI3K/Akt/mTOR signalling pathway. Besides, flow cytometry confirmed increased apoptotic cell population and decreased mitochondrial membrane potential due to the exposure of RM-3-22. RM-3-22 also promoted G2/M arrest. Signalling cascade confirmed that autophagy regulates RM-3-22-mediated apoptosis and cell cycle arrest. Additionally, RM-3-22 upregulated FTH1, a tumor suppressor, reinforcing its anticancer potential. Notably, RM-3-22 exhibited lower toxicity to normal cells, underscoring its selectivity. In vivo, RM-3-22 markedly reduced tumor growth in the xenograft mouse model.
RM-3-22 demonstrates potent anticancer activity through different mechanisms, including inhibition of the PI3K/Akt/mTOR pathway and activation of autophagy, apoptosis, and cell cycle arrest. Further, in vivo validation also supports that RM-3-22 represents a promising therapeutic candidate against lung cancer.
肺癌仍然是癌症相关死亡的主要原因，因此需要新的治疗策略。在本研究中，我们开发了RM-3-22，一种基于TAZQ的异羟肟酸衍生物，具有组蛋白脱乙酰酶（HDAC）抑制特性。我们以A549腺癌细胞作为主要模型，评估了其在非小细胞肺癌（NSCLC）中的抗癌活性。
在二维和三维细胞培养模型中评估RM-3-22的抗癌效率。通过MTT法分析细胞存活率。采用包括吖啶橙和DAPI在内的不同显微镜染色方法评估自噬、细胞核变化和细胞凋亡。使用流式细胞术评估细胞周期进程、线粒体膜电位和凋亡-坏死情况。通过免疫荧光（IF）、蛋白质印迹和逆转录-聚合酶链反应（RT-PCR）评估与RM-3-22诱导途径相关的蛋白质和基因表达。使用小干扰RNA（siRNA）介导的敲低进行功能基因分析。还进行了不同的计算机模拟研究，以检查RM-3-22诱导基因的临床相关性和表达模式。此外，使用NOD/SCID异种移植模型评估该分子的体内效率。
RM-3-22在体外可能抑制A549细胞的活力并减小其肿瘤球大小。它通过下调PI3K/Akt/mTOR信号通路诱导自噬。此外，流式细胞术证实，由于RM-3-22的作用，凋亡细胞群体增加，线粒体膜电位降低。RM-3-22还促进G2/M期阻滞。信号级联反应证实自噬调节RM-3-22介导的细胞凋亡和细胞周期阻滞。此外，RM-3-22上调了肿瘤抑制因子FTH1，增强了其抗癌潜力。值得注意的是，RM-3-22对正常细胞的毒性较低，突出了其选择性。在体内，RM-3-22显著降低了异种移植小鼠模型中的肿瘤生长。
RM-3-22通过不同机制表现出强大的抗癌活性，包括抑制PI3K/Akt/mTOR途径以及激活自噬、细胞凋亡和细胞周期阻滞。此外，体内验证也支持RM-3-22是一种有前景的肺癌治疗候选药物。
