Analysis of the interactions of botanical extract combinations against the viability of prostate cancer cell lines

Abstract

Herbal medicines are often combinations of botanical extracts that are assumed to have additive or synergistic effects. The purpose of this investigation was to compare the effect of individual botanical extracts with combinations of extracts on prostate cell viability. We then modeled the interactions between botanical extracts in combination isobolographically. Scutellaria baicalensis, Rabdosia rubescens, Panax-pseudo ginseng, Dendranthema morifolium, Glycyrrhiza uralensis and Serenoa repens were collected, taxonomically identified and extracts prepared. Effects of the extracts on cell viability were quantitated in prostate cell lines using a luminescent ATP cell viability assay. Combinations of two botanical extracts of the four most active extracts were tested in the 22Rv1 cell line and their interactions assessed using isobolographic analysis. Each extract significantly inhibited the proliferation of prostate cell lines in a time- and dose-dependent manner except S. repens. The most active extracts, S. baicalensis, D. morifolium, G. uralensis and R. rubescens were tested as two-extract combinations. S. baicalensis and D. morifolium when combined were additive with a trend toward synergy, whereas D. morifolium and R. rubescens together were additive. The remaining two-extract combinations showed antagonism. The four extracts together were significantly more effective than the two-by-two combinations and the individual extracts alone. Combining the four herbal extracts significantly enhanced their activity in the cell lines tested compared with extracts alone. The less predictable nature of the two-way combinations suggests a need for careful characterization of the effects of each individual herb based on their intended use.

Figure 1

Herbal extracts inhibit proliferation in prostate cell lines. I. S. baicalensis II. G. uralensis III. D. morifolium IV. R. rubescens V. P. ginseng VI. S. repens. Cells were exposed to increasing concentrations of herbal extracts for 48 hours. Cell proliferation was determined via the CellTiter-Glo^® Luminescent Cell Viability Assay. Data are expressed as a percentage of vehicle treated controls, mean ± SE (n = 3). ^*P < 0.01 compared to vehicle controls.

Figure 2

Herbal combinations inhibit proliferation in 22Rv1 prostate cancer cell line. Cells were exposed to two by two and a four way mixture (Quad) of the four herbs tested at EC50 levels for the 22Rv1 cell line for 48 hours. Data are expressed as a percentage of vehicle treated controls, mean ± SE (n ≥ 3). Asterisks indicate significance from vehicle controls (single asterisk: P ≤ 0.001, double asterisk: P ≤ 0.05).

Figure 3

Isobologram demonstrating the interaction of two-by-two herbal combinations. The solid line represents the line of additivity (dose: µg/mL); the broken line represents proportions of the mixtures. Point A represents the calculated additive response where point B indicates the response achieved by testing