Anti-cancer Activities Of The Storage Protein Of The Sweet Potatoes And Its Underlying Mechanisms

As the world's largest sweet potato producer, the major post-harvest processing of the sweetpotatoes in China is extracting of the starches. As a result, a large volume of waste water rich inproteins is being produced and directly emitted to the rivers, causing severe environmentalpollutions. Previous reports have shown that the storage protein in the roots, sporamin, has apotent anticancer effect against a variety of cancer cell lines. Therefore, in the current study, thefeasibility of a novel strategy of recovering the sporamins from the sweet potato starch slurry wasinvestigated and then the anticancer effect and its underlying molecular mechanisms of actionwere examined and explored in three malignant cancer cell lines. Finally, a preliminary clinicaltrial was carried out to examine the safety and effectiveness of orally consuming the sporamins.The study shows that the "natural fermentation+foam separation" process is a feasiblestrategy for the recovery of sweet potato protein from the processing wastewater. After starchextraction the slurry was naturally fermented at room temperature for1,4,7,10, and13days. Atthese days a sample was taken and examined for the feasibility of recovering the sporamins from itby a glass foam fractionation column with air as carrier gas (superficial air velocity90cm/min).Results: with the elongation of the fermentation time, both the pH and the protein concentration ofthe slurry gradually reduced, which were positively correlated with each other (R=0.828, P=0.042). After7days of fermentation, the pH decreased from6.57+0.15to3.40+0(P <0.05). Thepeak recovery and enrichment ratio of the protein were obtained at this time, which were87.9+6.07%and1.55+0.11, respectively. From a cost-benefit perspective, fermentation for4-7days isrecommended. SDS-PAGE showed that the recovered proteins were sporamins., but sporamin Adegraded more rapidly than sporamin B. Trypsin inhibitor (TI) activity staining showed that therecovered proteins still have the TI activity. Conclusion: the strategy of the "natural fermentation+foam separation" is feasible for the recovery of proteins from the sweet potato starch processingwastewater and has many advantages like low cost, simple process, and easy popularization etc..MTT assay and the crystal violet staining assay showed that sporamin potently suppressedthe proliferation of human hepatoma HepG2cells, human breast cancer BCAP-37cells, andhuman colon cancer HT-29cells, with a half inhibitory concentration (IC50) of4.44μM,5.12μM,and4.82μM, respectively. Wound healing and the de-adhesion assay showed that sporaminsignificantly inhibited the migration and adhesion capacity of the cancer cells, which werestimulated by a tumor promoter phorbol ester acid1-fourteen-13-acetate (PMA). Sporamin at1μMsignificantly reduced the migration of the three cancer cells (P <0.05). Gelatin zymography andplasminogen-casein zymography tests showed that the sporamin inhibited the activity of thematrix metalloproteinase MMP-2, MMP-9and the urokinase type plasminogen activator (uPA) inthe conditioned medium of HT-29cells. Western blot analysis showed a significant inhibition ofthe expression of uPA, MMP-2and MMP-9elicited by PMA. The phosphorylation of proteinkinase B (Akt) was also suppressed by sporamin. The secretion of uPA in the conditioned medium was also reduced. Reverse transcription-polymerase chain reaction (RT-PCR) showed that, at themRNA level, the expression of uPA was not significantly altered, but the expression of MMP-7,MMP-9, MMP-13, and vascular endothelial growth factor (VEGF) were significantly reduced.Hoechst33258nuclear staining showed that sporamin induced cell apoptosis. Together, theseresults indicated that sporamin may exert its effect by inhibition of the intracellular PI3K/Aktsignal transduction pathway and then the downstream uPA and MMPs systems and regulate theuPA level at the posttranscriptional level. Meanwhile, sporamin may have an inhibitory effect ontumor angiogenesis. Through these multiple effects, sporamin may play an inhibitory role in tumorcell proliferation, invasion, and angiogenesis.Finally, the sporamin was made into a granule dose for a randomized, double-blind, placebocontrolled clinical trial. In this trial, sporamin was consumed orally to treat patients with stage IIIcolon cancer who were accepting5-fluorouracil (5-FU) therapy at the same time. As of this report,10patients had finished a60-day trial with5of them taking the sporamin at12g/d and5takingonly a placebo. No adverse effects were observed by the investigator or reported by the patients.Self appraisal and mental status were improved by taking the protein. Detection of the serumtumor biomarkers revealed that consumption of the protein decreased the level of ferritin,carcinoembryonic antigen, and carbohydrate antigen242in female subjects. Because at presentstage the sample size is still small, it is obvious that with the increment of the sample size it willbe possible to obtain more significant evidence on the safety and effectiveness of orally taking theprotein for the treatment of colon cancer.