| Purpose: To screen the synthesized flavanone derivatives and evaluate the antiproliferative activity of candidate MSFTZ in vitro and in vivo and elucidate underlying molecular mechanisms providing with the basis for its chemical modification as well as clinical use.Methods:Synthesized 22 flavanone derivatives were screened using MTT assay before anti-cancer spectrum and activity of candidate compound (MSFTZ) were determined. The in vivo anti-tumor activity of MSFTZ was measured by mice bearing Sarcoma-180 (S180) and nude mice bearing human hepatocellular carcinoma SMMC-7721. Apoptosis was detected on both HL-60 and SMMC-7721 cells by flow cytometric analysis, DNA electrophoresis and AO/EB staining. Apoptosis-related protein level was determined on HL-60 cells by western blotting.Results:1. The results of in vitro screening of 22 flavanone derivatives indicatedthat No. 5 (MSFTZ) and 9 compound exhibited significant antiproliferative effects (IC50 < 20 pg/ml) with broad anti-cancer spectrum, more potent than their parent compound flavanone.2. Acute promyelocytic leukemia (NB4), Acute lymphoblastic leukemia (Molt-4), Chronic myelocytic leukemia (K562), Chronic myelocytic leukemia (76.1-fold resistance to vincristine)(K562R), Acute promyelocytic leukemia (HL-60), Acute promyelocytic leukemia (26.9-fold resistance to daunorubicin) (HL-60R), breast cancer (MCF-7), non-small cell lung cancer (A549), hepatocellular carcinoma (SMMC-7721 and Bel-7402), esophageal cancer (Eca-109), prostate carcinoma (PC-3) cell lines were treated with indicated concentrations of MSFTZ for 72 hours. The result showed that MSFTZ exhibited high antiproliferative activity in all tested cells, with IC50 values ranged from 0.58-10.58 μg/ml.3. The result of Sarcoma-180 (S180) tumor bearing mice tests approved that MSFTZ exhibited high experimental therapeutic activity on the tumor model. The inhibition rate was 48.2% (P < 0.001) at the dose of MSFTZ 80 mg/kg for 7-days consecutive administration. In SMMC-7721 hepatocellular carcinoma bearing nude mice models, the tumor weight of 80 mg/kg and 40 mg/kg MSFTZ groups were significantly decreased (P <0.01-0.05). The inhibition rate were 44.7% and 34.5%, respectively.4. Flow cytometric analysis confirmed that MSFTZ induced apoptosis in both HL-60 and SMMC-7721 cells in a dose dependent manner. MSFTZ showed little influence on cell cycle. DNA ladder, a hallmark of apoptosis, was observed on an agarose gel in MSFTZ-treated HL-60 cells. Human hepatocellular carcinoma SMMC-7721 cells were stained with AO/EB and themorphological changes demonstrated that MSFTZ could induce apoptosis.5. The results of western blot demonstrated that MSFTZ activated MAPK, cleaved procaspase-3 and procaspase-9, degraded PARP and eventually induced apoptosis in HL-60 cells.Conclusion:1. MSFTZ exerted high antiproliferative effects in all tested cancer cell lines detected, including 6 leukemia cell lines and 6 solid tumor cell lines. Interestingly, the cytotoxicity of MSFTZ does not show huge difference on drug-resistant cell lines and their parental cell lines compared with vincristine and daunorubicin.2. MSFTZ showed high experimental therapeutic activity on both murine bearing mice tumor and human tumor xenograft athymic mice.3. MSFTZ exerted antiproliferative property via induction of apoptosis of HL-60 leukemia cell lines and SMMC-7721 liver cancer cell lines.4. MSFTZ activated MAPK cell signal pathways, cleaved pro-caspase-9 and pro-caspase-3, downregulated XIAP and BcI-Xl, and degraded poly (ADP-ribose) polymerase (PARP).In summary, MSFFZ exerted the anti-cancer activity in vitro and in vivo via activation of MAPK cell signal pathways and caspase cascade.
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