A Preliminary Study Of Human Prostate Cancer CD44+ Cells Before And After ADT Using Metabolomics Profiling

Objective:To establish human prostate cancer cell models and to obtain sufficient number of tumor cells.To establish a highly successive method for human prostate cancer epithelial cells.This study well simulates the process which prostate cancer progresses from Androgen-dependent prostate cancer(ADPC)to Castration-resistant prostate cancer(CRPC).Primary cultures may eliminate many man-made interference factors and well represent the condition in vivo.We attempt to study human PCa CD44+ cells before and after ADT using metabolomics profiling.This may help us understand the mechanism of prostate cancer progression from ADPC to CRPC and also discover the new small-molecule biomarker for prostate cancer.This study is the very beginning of the great hypothesis.We tested the feasibility of the method and sum up future improvements from this pre-experiment experience.Methods:First,we cultured cells directly from human prostate cancer specimens.Second,two treatments were introduced to imitate the vivo environment when ADT was given or not given.For ADPC group,lOnM of DHT was given to maintain the hormone level as it is in human bodies while the ADT group’s hormone level was restricted by giving only lnM of DHT and casodex(an anti-androgen agent).Cells were then sorted by MACS using CD44 microbeads.In this way one sample was divided into 4 groups.Next,cellular metabolites was extracted and sent for metabolomics analysis.Results:A total of 106 metabolites have been detected using CE-MS,including 64 metabolites under cation mode and 42 under anion mode.24 metabolites including overlapping ones are statistically significant(Wilcoxon,p<0.05),which 15 under cation mode and 9 under anion mode.N-Methylaniline,acetylcholine,piperidine,isobutylamine,aminophenol(in forms of three isomers m/o/p-Aminophenol)valeric acid and benzoic acid are the 7 metabolites that have a greater amount in CD44+ cells before ADT than it is after ADT.Conclusion:The 7 differential chemicals we found may play an important role in CD44+ cells survival against androgen deprivation.However,key metabolites in glycolysis are not detected due to the limited CD44+ cells from primary culture,making it difficult to identify an import metabolic pathway through differential metabolites.Among the limited metabolites detected,none of them appears to be involved in a same pathway,making the data interpretation rather hard.Future improvements may be having larger sample sizes and optimizing the cell number for primary culture required for more effective metabolome detection.