Evaluation of Novel Spiro-pyrrolopyridazine Derivatives as Anticancer Compounds: In Vitro Selective Cytotoxicity, Induction of Apoptosis, EGFR Inhibitory Activity, and Molecular Docking Analysis.

Cancer, characterized by uncontrolled cell proliferation, remains a global health challenge. Despite advancements in cancer treatment, drug resistance and adverse effects on normal cells remain challenging. The epidermal growth factor receptor (EGFR), a transmembrane tyrosine kinase protein, is crucial in controlling cell proliferation and is implicated in various cancers. Here, the cytotoxic and apoptotic potential of 21 newly synthesized spiro-pyrrolopyridazine (SPP) derivatives was investigated on breast (MCF-7), lung (H69AR), and prostate (PC-3) cancer cells. XTT assay was used for cytotoxicity assessment. Flow cytometry and western blot (WB) analyses were conducted for apoptosis detection. Additionally, the EGFR inhibitory potential of these derivatives was evaluated via a homogeneous time-resolved fluorescence (HTRF) assay, and WB and molecular docking studies were conducted to analyze the binding affinities of SPP10 with EGFR. SPPs, especially SPP10, exhibit significant cytotoxicity across MCF-7, H69AR, and PC-3 cancer cells with IC50 values of 2.31 ± 0.3, 3.16 ± 0.8, and 4.2 ± 0.2 µM, respectively. Notably, SPP10 demonstrates selective cytotoxicity against cancer cells with a low impact on nontumorigenic cells (IC50 value: 26.8 ± 0.4 µM). Flow cytometric analysis demonstrated the potent induction of apoptotic cell death by SPP10 in all of the tested cancer cells. Western blot analysis revealed the involvement of key apoptotic proteins, with SPP10 notably inhibiting antiapoptotic Bcl-2 while inducing pro-apoptotic Bax and cytochrome c. SPP10 exhibited significant EGFR kinase inhibitory activity, surpassing the efficacy of the reference drug erlotinib. Molecular docking studies support these findings, revealing strong binding affinities of SPP10 with both wild-type and mutated EGFR. The study underscores the significance of heterocyclic compounds, particularly spiro-class heterocyclic molecules, in advancing cancer research. Overall, SPP10 emerges as a promising candidate for further investigations in cancer treatment, combining potent cytotoxicity, apoptotic induction, and targeted EGFR inhibition.

Design and synthesis of benzimidazole derivatives as apoptosis-inducing agents by targeting Bcl-2 protein.

Bcl-2, an anti-apoptotic protein, is a well-known and appealing cancer therapy target. Novel series of benzimidazole derivatives were synthesized and tested for their activity as Bcl-2 inhibitors on T98G glioblastoma, PC3 prostate, MCF-7 breast, and H69AR lung cancer cells. MTT assay was used to evaluate the cytotoxic effect. PI Annexin V Apoptosis Detection Kit was used to detect apoptosis. Expression levels of the Bcl-2 protein were examined by the Western blot analysis and qRT-PCR. All synthesized benzimidazole derivatives exhibited a cytotoxic effect on cancer cells with IC50 values in the range of 25.2-88.2 µg/mL. Among all derivatives, compounds C1 and D1 demonstrated a higher cytotoxic effect on cancer cells with IC50 values < 50 µg/mL, while a lower cytotoxic effect against human embryonic kidney cells with IC50 values of > 100 µg/mL. C1 and D1 caused a significant increase in the percentage of apoptotic cells in all types of cancer cell cells and both Bcl-2 mRNA and protein levels were significantly reduced. These results suggest that the novel benzimidazole derivatives may be candidates for apoptosis-inducing agents in cancer treatment by targeting anti-Bcl-2 proteins in cancer cells.

Synthesis of silver nanoparticles using Alpinia officinarum rhizome extract induces apoptosis through down-regulating Bcl-2 in human cancer cells.

In this study, silver nanoparticles were synthesized using Alpinia officinarum rhizome extract via an eco-friendly green synthesis method. The silver nanoparticles (AO-AgNPs) were characterized by UV-Vis spectrometry, scanning electron microscopy, energy-dispersive X-ray spectroscopy, and dynamic light scattering. Further, the cytotoxic and apoptotic effects of AO-AgNPs were investigated in human cancer cells with different tissue origins via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and flow cytometric analyses, respectively. The expression levels of anti-apoptotic Bcl-2 protein were evaluated via a real-time polymerase chain reaction. The synthesized AO-AgNPs induced a significant cytotoxic effect in all tested cancer cells but not in normal cells. AO-AgNPs induced the percentage of apoptotic cells and reduced the levels of anti-apoptotic Bcl-2 mRNA levels in cancer cells. These results demonstrate the potential application of AO-AgNPs in cancer treatment.

Liquidambar orientalis Mill. gum extract induces autophagy via PI3K/Akt/mTOR signaling pathway in prostate cancer cells.

Liquidambar orientalis Mill (LOM), is an endemic species having a local distribution in the southwestern coastal district of Turkey. Styrax liquidus gum (SLG), is a gum-like resinous which exudates in response to injury of the trunk of LOM. The aim of the study was to investigate the cytotoxic effects and the molecular mechanisms of the ethanolic SLG extract in human prostate cancer cells. GC-MS analysis was performed to identify the volatile compound composition. Cytotoxicity was determined by XTT analysis. Apoptosis and necrosis were evaluated via ELISA assay. Autophagic cell death was detected via monodansylcadaverine (MDC) staining and by measuring the levels of LC3I and LC3II. The protein levels of p-PI3K, p-Akt and p-mTOR were evaluated by western blot analysis. In the present study, it is shown that the SLG extract containing a considerable amount of ravidomycin derivate induced autophagic cell death in prostate cancer cells via inhibiting the PI3K/Akt/mTOR pathway.