Materials and methods: The antiproliferative activity of koenimbin was examined using MTT, and the apoptotic detection was carried out by acridine orange/propidium iodide (AO/PI) double-staining and multiparametric high-content screening (HCS) assays. Caspase bioluminescence assay, reverse transcription polymerase chain reaction (RT-PCR), and immunoblotting were conducted to confirm the expression of apoptotic-associated proteins. Cell cycle analysis was investigated using flow cytometry. Involvement of nuclear factor-kappa B (NF-κB) was analyzed using HCS assay. Aldefluor™ and prostasphere formation examinations were used to evaluate the impact of koenimbin on PC-3 CSCs in vitro.
Results: Koenimbin remarkably inhibited cell proliferation in a dose-dependent manner. Koenimbin induced nuclear condensation, formation of apoptotic bodies, and G0/G1 phase arrest of PC-3 cells. Koenimbin triggered the activation of caspase-3/7 and caspase-9 and the release of cytochrome c, decreased anti-apoptotic Bcl-2 and HSP70 proteins, increased pro-apoptotic Bax proteins, and inhibited NF-κB translocation from the cytoplasm to the nucleus, leading to the activation of the intrinsic apoptotic pathway. Koenimbin significantly (P<0.05) reduced the aldehyde dehydrogenase-positive cell population of PC-3 CSCs and the size and number of PC-3 CSCs in primary, secondary, and tertiary prostaspheres in vitro.
Conclusion: Koenimbin has chemotherapeutic potential that may be employed for future treatment through decreasing the recurrence of cancer, resulting in the improvement of cancer management strategies and patient survival.
METHODS: By utilizing a panel of breast cancer cells and mammospheres culture as cell-based screening platforms, we performed high-throughput chemical library screens to identify agents that are effective against breast CSCs and non-CSCs. The hit molecules were paired with conventional chemotherapy to evaluate the combinatorial treatment effects on breast CSCs and non-CSCs.
RESULTS: We identified a total of 193 inhibitors that effectively targeting both breast CSCs and non-CSCs. We observed that histone deacetylase inhibitors (HDACi) synergized conventional chemotherapeutic agents (i.e., doxorubicin and cisplatin) in targeting breast CSCs and non-CSCs simultaneously. Further analyses revealed that quisinostat, a potent inhibitor for class I and II HDACs, potentiated doxorubicin-induced cytotoxicity in both breast CSCs and non-CSCs derived from the basal-like (MDA-MB-468 and HCC38), mesenchymal-like (MDA-MB-231), and luminal-like breast cancer (MCF-7). It was also observed that the basal-like breast CSCs and non-CSCs were more sensitive to the co-treatment of quisinostat with doxorubicin compared to that of the luminal-like breast cancer subtype.
CONCLUSION: In conclusion, this study demonstrates the potential of HDACi as therapeutic options, either as monotherapy or in combination with chemotherapeutics against refractory breast cancer.