1. Organic Chemistry I (Chem 210) Final Presentation By Daniel I. Egbe Instructor: Professor Wan Jin Jahng

2. Potential Problems Recall in my last presentation that I was talking about the use of triterpenoid saponin in the in the fight against cancer. But the fact is, synthesizing this form of saponins is very expensive and so, producing it from the lab might not be feasible

3. Alternative Approach Using a cheaper and more effective form of saponins in the fight against cancer in the body, called steroid saponins, obtained from the rhizome of Paris polyphylla var. yunnanens.

4. Experimental Design I will use about 48 T739 inbred mice, both male and female (24 male and 24 female) of weight 18.0 ± 2.0 g. Use 4 groups for the experiment:  one control group (0.1ml/10g distilled water)  cyclophosphamide (CPH)- treated group (20mg/kg)  compound 1 (α-L-arabinofuranosyl (1→4) ) treated group (100mg/kg) and  Diosgenin treated group (200mg/kg).

5. Model System- In Vivo Keep the mice under pathogen-free conditions, and feed them with chow (food) and sterile water. Inject Lung LA795 adenocarcinoma cells under the skin, into the armpits of the mice. After injecting, the tumors will grow to a volume of 0.5*1.0 cm 3. Then 8 of the mice will be sacrificed by cutting them into pieces of 0.5- 1.0mm 3 and then dip them into 0.9% NaCl at 4 0 C to ensure that only tumor cells are cultivated.

6.  Inject the cultivated tumor cells into the remaining 40 mice (2*10 6 /ml, 0.2ml for each mouse)  5 days after, the 40 T739 mice cells bearing LA795 lung adenocarcinoma are divided into 4 groups of 10 each.  one control group (0.1ml/10g distilled water)  cyclophosphamide (CPH)- treated group (20mg/kg)  compound 1 (α-L-arabinofuranosyl (1→4) ) treated group (100mg/kg) and  Diosgenin treated group (200mg/kg).

7.  The experiment will run for 14 days.  Then, the body weight of each of the mouse is taken at the first and last days of the experiment.  On the last day, the mice will be sacrifice and autopsies are performed on the mice.  Major organs like lungs, liver, kidneys are then dissected and their weights are measured.

8. The tumor inhibitory rate of each of the compounds is calculated using the formula:. Tumor inhibitory rate (%)= 100% * (mean tumor weight in control exp-mean tumor weight in treated mice)/ mean tumor weight in control exp

9. Results Group Volume (mg/kg/d) Tumor Volume (mm 3 ) Tumor Weight (g) Inhibitory Growth Rate (%) Control - 5462.5 ± 105.5 6.3 ± 0.6 Cyclophosphamide 20 2912.7 ± 252.9 2.2 ± 0.4 70.09 Compound 1 100 4234.0 ± 486.4 4.4 ± 0.5 29.44 Diosgenin 200 4005.1 ± 762.4 4.2 ± 0.5 33.94

10. Expected Results in Humans. Since the immune system of humans are stronger and more resistant than those of mice, I expect that if compounds are administered in humans with cancer cells in the same dosage as those used in the mice, it will induce about 55% response.

11. Feasibility A lot of studies have been carried out at preclinical level to examine the tumor- suppressive properties of saponins. In a study by Gutterman et al, the purification and isolation of steroid saponins and two biologically active derivatives, referred to as avicins D and G from Acacia victoriae, an Australian desert tree, were reported. It was noticed that the compounds that were isolated greatly inhibited the growth of many tumor cell lines. They prompted cell cycle arrest of human breast cancer cell line and also induced apoptosis of T cell leukemia and breast cancer cell lines.

12. Conclusion The best and most active form of steroid saponins to use in the fight against cancers is cyclophosphamide (CPH) Also, the administration of the CPH in humans should be done using a greater dosage than that used for mice for it to have great effect on inhibiting the growth of tumor cells.

13. References Zhao, Y., Kang, LP., Liu, YX., Liang, YG., Tan, DW., Yu, ZY., Cong, YW., & Ma, BP. (2009, March). Steroidal saponins from the rhizome of Paris Polyphylla and their cytotoxic activities. Planta Med, 75(4), p. 356-63. Yan, L.L., Zhang, Y.J., Gao, W.Y., Man, S.L., & Wang, Y. (2009). In vitro and in vivo anticancer activity of steroid saponins of paris polyphylla var. yunnanensis. Experimental Oncology, 31(1), p. 27-32. Wu, X., Wang, L., Wang, H., Dai, Y., Ye, WC., & Li, YL. (2012, September). Steroidal saponins from paris polyphylla var. yunnanensis. Phytochemistry, 81, 133- 143.