In vitro Antiproliferative Effect of Earthworm Coelomic Fluid of Eudrilus Eugeniae, Eisenia Foetida, and Perionyx Excavatus on Squamous Cell Carcinoma-9 Cell Line: A Pilot Study

Introduction The earthworm coelomic fluid (ECF) has shown proven antiproliferative effect against breast, liver, gastrointestinal, and brain cancer, but it is least explored in oral cancer. The present in vitro study is an attempt to investigate the antiproliferative activity of ECF on oral cancer cell line squamous cell carcinoma (SCC)-9. Materials and Methods ECF was collected from the species Eudrilus eugeniae (EE), Eisenia foetida (EF), and Perionyx excavatus (PE) stored at -80°C. Percentage inhibition of ECF on squamous cell carcinoma-9 cells in vitro was recorded at 24 h. Protein estimation was done using Bradford protein assay validated by the biuret method. Cytotoxicity was tested at 2.5, 5, 10, 20, 40, and 80 μg/ml concentrations by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide assay in SCC-9 cells in vitro. GraphPad Prism 7.0 software was used to calculate the inhibitory concentration (IC50). Chi-square test was used to analyze the difference between samples. Results The test samples EE, EF, and PE inhibited the growth of SCC-9 cells significantly in a dose-dependent manner, and the IC50 values were found to be 4.6, 44.69, and 5.27 μg/ml, respectively. The antiproliferative effect was found to be variable among the three earthworm species with EE showing the most promising effect followed by PE and EF. Conclusion Establishing the antiproliferative effect of ECF on oral cancer cells could be an initial step toward drug development and future anticancer research. The preliminary investigation has shown that ECF has a promising antiproliferative effect on oral cancer cells in vitro. SUMMARY The present pilot study evaluated the in vitro antiproliferative effect of earthworm coelomic fluid (ECF) of Eudrilus eugeniae (EE), Eisenia foetida (EF), and Perionyx excavatus (PE) on squamous cell carcinoma-9 cell line. The ECF inhibitory activity was promising at inhibitory concentration values of 4.6, 44.69, and 5.27 μg/ml, respectively. Further studies pertaining to antiproliferative mechanism of EE, EF, and PE have been planned.Abbreviations Used: ECF: Earthworm coelomic fluid, EE: Eudrilus eugeniae, EF: Eisenia foetida, PE: Perionyx excavatus, MTT: 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, SCC: Squamous cell carcinoma, BSA: Bovine serum albumin, PBS: Phosphored buffered saline, ATCC: American Type Culture Collection.