Anti-Carcinogenic Properties of Chinese Herbs. Ryan Kelly Faculty Advisor: Ted Meigs

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1 Anti-Carcinogenic Properties of Chinese Herbs Ryan Kelly Faculty Advisor: Ted Meigs

2 Project Overview: The effects of several Chinese herbal extracts on cancer cell proliferation were quantified through analysis of protein content in cellular growth assays.

3 Typhonium giganteum Herbs Family: Araceae Common name: Typhonium rhizome Mandarin name: Bai Fu Zi English translation(literal): white appendage Traditional uses: expels wind and cold dampness, alleviates pain Clinical research: anti-tubercular tubercular effect

4 Ligustrum lucidum Herbs Family: Oleaceae Alternate name: Japanese privet Mandarin name: Nu Zhen Zi English translation (literal): female chastity seed Traditional Uses: nourishes and tonifies kidneys and liver Clinical research: diuretic

5 Oldenlandia diffusa Herbs Family: Rubiaceae Alternate name: Hedyotis diffusa Mandarin name: Bai Hua She She Cao English translation (literal): white patterned snake s tongue herb Traditional Uses: clears heat, reduces abscesses Clinical research: antimicrobial

6 Scutellaria barbata Family: Labiatae Alternate name: barbed skullcap Mandarin name: Huang Qin English translation (literal): none Traditional uses: cools the blood Clinical research: reduces diastolic blood pressure Herbs

7 Euphorbia pekinensis Herbs Family: Euphorbacea Alternate name: Peking spurge root Mandarin name: Da Ji English translation (literal): big lance from the capital Traditional uses: expels water, reduces swelling Clinical research: effects on gastrointestinal tract

8 Step 1 Development of BSA Standard Curve BSA Standard Curve y= x Protein Concentration (mg/ml)

9 Propagation and maintenance of cancer cell lines Step 2 K435 SW480 Human Breast Carcinoma Cells Human Colon Adenocarcinoma Cells

10 Step 3 Development of a Standard Growth Assay K435 and SW K435, Untreated SW480, Untreated

11 Step 4 Preparation of water-soluble extracts from herbs (taken from Dr. H Lau s procedure, Loma Linda University) - dried herbs were ground to a fine powder using an automatic coffee grinder - 10 g of herbal matter was decocted in 100 ml of sterilized water in a 56 C water bath for 1 hour -this solution was then centrifuged at 1200 x g for 15 minutes, and the supernatant was recentrifuged under the same conditions -the final supernatant was removed and passed through a.45µm cellulose acetate filter for sterilization

12 Step 5 Application of an herbal extract in the cell proliferation assay Experimental Design: - Cells were seeded in triplicate in 6 well plates - One well was dictated as control and received 0.5 ml sterilized water -Remaining 2 wells received varying concentrations of extract of 0.5 ml each

13 Results

14 Typhonium giganteum Little to no effect on either cell line K435, Treated with Typhonium giganteum Stock Concentration 1:100 Dilution SW480, Treated with Typhonium giganteum Stock Concentration 1:100 Dilution

15 Ligustrum lucidum Retarding of cell proliferation In both cancer cell lines K435, Treated with Ligustrum lucidium Stock Concentration 1:100 Dilution SW480, Treated with Ligustrum lucidium Stock Concentration 1:100 Dilution

16 Oldenlandia diffusa Varied response in each cancer cell line!! K435, Treated with Oldenlandia diffusa Stock Concentration 1:100 Dilution SW480, Treated with Oldenlandia diffusa Stock Concentration 1:100 Dilution

17 Scutellaria barbata Varied response in each cancer cell line K435, Treated with Scutellaria Barbata Stock Concentration 1:10 Dilution SW480, Treated with Scutellaria Barbata Stock Concentration 1:10 Concentration

18 K435, treated with Scutellaria barbata 1:10 Stock SW480 treated with Scutellaria barbata 1:10 Stock

19 Euphorbia pekinensis Varied response in each cancer cell line K435, Treated with Euphorbia pekinensis :4 Dilution 1:100 Dilution SW480, Treated with Euphorbia pekinensis :4 Dilution 1:100 Dilution

20 K435, treated with Euphorbia pekinensis 1:100 Stock SW480, treated with Euphorbia pekinensis 1:100 Stock

21 Application of Scutellaria barbata and Euphorbia pekinensis to a non-cancerous cell line, HEK293 (embryonic kidney cells) HEK293, Treated with Scutellaria barbata Stock Concentration 1:100 Dilution HEK293, Treated with Euphorbia pekinesis Stock Concentration 1:100 Dilution

22 Summary -Not all extracts had effects on cancer cell lines (TG), indicating that the extraction procedure was not generating nonspecifically toxic substances -Some extracts affected both cell lines in a similar fashion (LL), illustrating positive effects of the extract on cancer cell proliferation -Some extracts had different effects on each cancer cell line (OD), suggesting that the extract affected different signaling pathways specific to the cell type -Some of the extracts that elicited responses in the cancer cell lines, responded at different times throughout the growth assays - At least one herb showed effects on cancer cell proliferation without similar effects in a noncancerous cell line

23 Future Directions -More complete time course and concentration studies, and application of statistical analyses -Biochemically fractionate extracts into individual compounds and identify, test individual compound s effects on cancer cell proliferation, once a positive response is identified, explore the signaling pathway that is affected -Determine the effects these herbs exert on gene expression in the individual cell lines. Through techniques such as microarray analysis it could be determined if these extracts activate or deactivate specific genes

24 Acknowledgements Dr. Ted Meigs, Research Advisor The UNCA Undergraduate Research Program Herb provider: Joe Hollis at Mountain Gardens, Burnsville, NC Advisory Committee (Dr. David Clarke, Dr. Betsy Wilson)