Anticancer Potential of Sutherlandia frutescens and Xysmalobium undulatum in LS180 Colorectal Cancer Mini-Tumors.

Colorectal cancer remains to be one of the leading causes of death worldwide, with millions of patients diagnosed each year. Although chemotherapeutic drugs are routinely used to treat cancer, these treatments have severe side effects. As a result, the use of herbal medicines has gained increasing popularity as a treatment for cancer. In this study, two South African medicinal plants widely used to treat various diseases, Sutherlandia frutescens and Xysmalobium undulatum, were evaluated for potential activity against colorectal cancer. This potential activity for the treatment of colorectal cancer was assessed relative to the known chemotherapeutic drug, paclitaxel. The cytotoxic activity was considered in an advanced three-dimensional (3D) sodium alginate encapsulated LS180 colorectal cancer functional spheroid model, cultured in clinostat-based rotating bioreactors. The LS180 cell mini-tumors were treated for 96 h with two concentrations of each of the crude aqueous extracts or paclitaxel. S. frutescens extract markedly decreased the soluble protein content, while decreasing ATP and AK per protein content to below detectable limits after only 24 h exposure. X. undulatum extract also decreased the soluble protein content, cell viability, and glucose consumption. The results suggested that the two phytomedicines have potential to become a source of new treatments against colorectal cancer.

Cycloartanol and Sutherlandioside C peracetate from Sutherlandia frutescens and their immune potentiating effects.

A novel cycloartanol (1) and an acylated Sutherlandioside D (2) together with two known cycloartane derivatives, Sutherlandioside B (3) and Sutherlandioside A (4), were isolated from the aerial parts of Sutherlandia frutescens. The structures of these compounds were established by a combination of 1- and 2-D NMR techniques and further confirmed by high resolution ToF mass spectrometry (HRToFMS). Preliminary biological studies were also conducted to assess the activity of different plant extracts, fractions and compounds on cytokine expression. Compounds 1 and 2 prompted an increase in IL-6 expression while compound 4 showed a reduced IL-6 expression compared to the controls. Compound 1 is an effective suppressor of IL-10 expression. The plant compounds inhibited the expression of the two cytokines, IL-10 and TNFα. The results of the assays suggested that some components in the plant extract influence the immune system by suppressing the expression of IL-6, IL-10 and TNFα.

The mutagenic and antimutagenic activity of Sutherlandia frutescens extracts and marker compounds.

BACKGROUND: Sutherlandia frutescens (L.) R. Br is endemic to Southern Africa where it has been traditionally used for cancer and diabetes. In recent times it has been marketed for its reputed (but not proven) anticancer, antidiabetic and anti-HIV properties. Little is known about the mutagenic and antimutagenic potential of extracts and common marker compounds of Sutherlandia frutescens. Therefore this study aimed to investigate the putative efficacy and possible long-term adverse effects of using this herb. METHODS: Ethylacetate (EA) and 50% Methanol (MeOH) extracts were screened for mutagenic and antimutagenic activity using the Ames assay utilising TA97a, TA98, TA100 and TA102 in the presence and absence of metabolic activation. Four compounds, L-arginine, L-canavanine, GABA and D-pinitol known to occur in sutherlandia were also included. The total polyphenolic content of the both extracts was determined using the Folin-Ciocalteau method and FRAP and ABTS were used to determine the anti-oxidant potential of the extracts. RESULTS: The extracts and the standards did not show any cytotoxicity except in TA97a. The EA extract exhibited antimutagenicity against all the bacterial strains at all concentrations tested. The MeOH extract showed both pro-mutagenic and antimutagenic activities with 2-acetamidofluorene and aflatoxin B1 in the presence of metabolic activation of TA98 and TA100, respectively. All compounds, except L-canavanine exhibited antimutagenic activity against all strains. L-canavanine, on the other hand showed co-mutagenicity with 9-aminoacridine on TA97a, at all test concentrations. The extracts and pure compounds exhibited their antimutagenic activity in a dose response manner. L-arginine and GABA showed an some antimutagenic response. EA extract had three times the total phenolic content (12.56 µg GE / mg) observed in the MeOH extract. There was correlation between total phenolic content, antioxidant potential and antimutagenicity. CONCLUSION: Both extracts exhibited a protective effect, with the EA extract exhibiting greater potency. L-canavanine acted as a co-mutagen in a dose response manner without metabolic activation. It is suggested that the EA extract be priotized for future development work as it showed a better risk profile and activity.

Sutherlandia frutescens modulates adrenal hormone biosynthesis, acts as a selective glucocorticoid receptor agonist (SEGRA) and displays anti-mineralocorticoid properties.

ETHNOPHARMACOLOGICAL RELEVANCE: Sutherlandia frutescens is a traditional African medicinal plant used in the treatment of stress and anxiety, while also exhibiting anti-inflammatory properties. AIM OF STUDY: The study aimed at linking anti-stress and anti-inflammatory properties of S. frutescens to its influence on glucocorticoid biosynthesis and the inflammatory response via steroid receptor interaction. MATERIALS AND METHODS: The influence of S. frutescens extracts and sutherlandioside B (SUB),10 and 30µM, on key steroidogenic enzymes was assayed in COS-1 cells. Effects were also assayed on basal and stimulated hormone levels in the adrenal H295R cell model. Agonist activity for transactivation and transrepression of the extract and SUB with the glucocorticoid- (GR) and mineralocorticoid receptor (MR) was subsequently investigated. RESULTS: Inhibitory effects of the extract towards progesterone conversion by CYP17A1 and CYP21A2 were significant. SUB inhibited CYP17A1 and 3ß-HSD2, while not affecting CYP21A2. In H295R cells, SUB decreased cortisol and androgen precursors significantly. The extract decreased total steroid production (basal and stimulated) with cortisol and its precursor, deoxycortisol, together with mineralocorticoid metabolites significantly decreased under forskolin stimulated conditions. S. frutescens extracts and SUB repressed NF-κB-driven gene expression without activating GRE-driven gene expression and while neither activated MR mediated gene transcription, both antagonized the effects of aldosterone via the MR. CONCLUSION: Data provide evidence linking anti-stress, anti-inflammatory and anti-hypertensive properties of S. frutescens to inhibition of steroidogenic enzymes and modulation of adrenal hormone biosynthesis. Findings suggesting S. frutescens and SUB exhibit dissociated glucocorticoid characteristics underline potential therapeutic applications in the treatment of inflammation and hypertension.

Inhibition of Gli/hedgehog signaling in prostate cancer cells by "cancer bush" Sutherlandia frutescens extract.

Sutherlandia frutescens is a medicinal plant, traditionally used to treat various types of human diseases, including cancer. Previous studies of several botanicals link suppression of prostate cancer growth with inhibition of the Gli/hedgehog (Gli/Hh) signaling pathway. Here we hypothesized the anti-cancer effect of S. frutescens was linked to its inhibition of the Gli/Hh signaling in prostate cancer. We found a dose- and time-dependent growth inhibition in human prostate cancer cells, PC3 and LNCaP, and mouse prostate cancer cell, TRAMP-C2, treated with S. frutescens methanol extract (SLE). We also observed a dose-dependent inhibition of the Gli-reporter activity in Shh Light II and TRAMP-C2QGli cells treated with SLE. In addition, SLE can inhibit Gli/Hh signaling by blocking Gli1 and Ptched1 gene expression in the presence of a Gli/Hh signaling agonist (SAG). A diet supplemented with S. frutescens suppressed the formation of poorly differentiated carcinoma in prostates of TRAMP mice. Finally, we found Sutherlandioside D was the most potent compound in the crude extract that could suppress Gli-reporter in Shh Light II cells. Together, this suggests that the S. frutescens extract may exert anti-cancer effect by targeting Gli/Hh signaling, and Sutherlandioside D is one of the active compounds.

Unveiling the anti-inflammatory activity of Sutherlandia frutescens using murine macrophages.

Sutherlandia frutescens is a botanical widely used in southern Africa for treatment of inflammatory and other conditions. Previously, an ethanolic extract of S. frutescens (SFE) has been shown to inhibit the production of reactive oxygen species (ROS) and nitric oxide (NO) by murine neurons and a microglia cell line (BV-2 cells). In this study we sought to confirm the anti-inflammatory activities of SFE on a widely used murine macrophage cell line (i.e., RAW 264.7 cells) and primary mouse macrophages. Furthermore, experiments were conducted to investigate the anti-inflammatory activity of the flavonol and cycloartanol glycosides found in high quantities in S. frutescens. While the SFE exhibited anti-inflammatory activities upon murine macrophages similar to that reported with the microglia cell line, this effect does not appear to be mediated by sutherlandiosides or sutherlandins. In contrast, chlorophyll in our extracts appeared to be partly responsible for some of the activity observed in our macrophage-dependent screening assay.

Immuno-stimulatory activity of a polysaccharide-enriched fraction of Sutherlandia frutescens occurs by the toll-like receptor-4 signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Sutherlandia frutescens (L.) R. Br. is an indigenous plant of southern Africa that has been traditionally used for various cancers, infections, and inflammatory conditions. AIM OF THE STUDY: Our aim was to investigate the potential immuno-stimulatory activity of a polysaccharide-enriched fraction (SFPS) from a decoction of S. frutescens. MATERIALS AND METHODS: RAW 264.7 cells (a murine macrophage cell line) were used to determine the activities of SFPS on macrophage function. The production of reactive oxygen species (ROS), nitric oxide (NO), and inflammatory cytokines were evaluated in the cells treated with or without SFPS. CLI-095, a toll-like receptor (TLR) 4-specific inhibitor, was used to identify whether or not SFPS exerts its effects through TLR4. An antagonist of endotoxin, polymyxin B, was used to evaluate whether endotoxin present in SFPS contributed to its immune-stimulatory activity. RESULTS: SFPS exhibited potent immune-stimulatory activity by macrophages. The production of ROS, NO, and tumor necrosis factor (TNF-α) were increased upon exposure to SFPS in a dose-dependent manner. All of these activities were completely blocked by co-treatment with CLI-095, but only partially diminished by polymyxin B. CONCLUSION: We demonstrate for the first time potent immune-stimulatory activity in a decoction prepared from S. frutescens. We believe that this immune stimulatory activity is due, in part, to the action of polysaccharides present in the decoction that acts by way of TLR4 receptors and the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway. These findings provide a plausible mechanism through which we can understand some of the medicinal properties of S. frutescens.

Immunomodulating polysaccharides from Lessertia frutescens leaves: isolation, characterization and structure activity relationship.

ETHNOPHARMACOLOGICAL RELEVANCE: Sutherlandia frutescens (syn. Lessertia frutescens) is an indigenous plant in Southern Africa and has been extensively studied from the ethnobotanical point of view. Amongst the various traditional uses, several illnesses involving the immune system have been reported. Due to some of the therapeutic effects observed, in relation to the traditional uses reported by the "khoi san" and "nama" people on cancer related illnesses, the plant has been given the local name kankerbos (cancerbush). Recently the plant has also been used amongst HIV/AIDS patients to stimulate the immune system. MATERIALS AND METHODS: Leaves of Sutherlandia frutescens were extracted sequentially with ethanol, 50% ethanol/water, and water at 50 and 100°C. The polysaccharides were extracted with water and fractionated by ion exchange chromatography and gel filtration to obtain enriched polysaccharide fractions. The bioactivities of the fractions were tested in the complement assay. Some of the fractions were treated with the enzyme pectinase, and the fragments thus produced were separated by gel filtration and their activities tested. Monosaccharide compositions and linkage analyses were determined for the relevant fractions. RESULTS: The leaves of Sutherlandia frutescens contain polysaccharides of the pectin type. Fractions from both the water extracts of 50 and 100°C were bioactive. Fractions chosen for further studies showed that the fragment with the highest M(W) after the pectinase treatment had a substantially higher biological effect than the parent molecules. Based on a comparison of the different fractions it was concluded that galactose-rich regions were important for the bioactivity, these being of the AGII and AGI type, with the latter probably being more important than the former. Fragments rich in xylose also gave higher activity than those without it. CONCLUSIONS: Our theory that the polysaccharides present in the leaves of Sutherlandia frutescens could be of importance as immunomodulating agents was confirmed. It was also shown that certain types of polysaccharides had a higher effect in the complement system than others. Thus both the water extracts obtained at 50 and 100°C contain interesting biologically active polysaccharides.

Effects of Sutherlandia frutescens extracts on normal T-lymphocytes in vitro.

Sutherlandia frutescens (SF), a popular traditional medicinal plant found in various parts of southern Africa, is used for treatment or management of HIV/AIDS and other diseases including cancer. However, its toxicity profile has not been fully established. The aims of this study were to examine the effects of 70% ethanol (SFE) and deionised water (SFW) extracts on normal isolated human T cells. An experimental study on normal human lymphocytes treated with doses SF extract doses ranging from 0.25 to 2.5 mg/ml. Untreated, vehicle-treated (Ethanol) and camptothecin (CPT) treated normal T cells were used as controls. Induction of cell death, changes in intracellular ATP, caspase-3/-7 activity and nuclear changes were analysed using flow cytometry, luminometry and nuclear staining (Hoechst) respectively. The highest concentration (2.5 mg/ml) of SFE extract induced significant necrosis (95%), depletion of ATP (76%), and inhibition of caspase-3/-7 activity (11%) following a 24 hour incubation period (p< 0.001). The 2.5 mg/ml concentration of SFW showed the same trend but were less effective (necrosis- 26%, ATP- 91%, & caspase-3/-7- 15%). These effects showed a time-dependence over 48 hours of incubation, with high doses of SFE extracts eliminating viable cells by necrosis, depleting ATP levels and decreasing caspase-3/-7 activity (p< 0.001). The activity of SFE extract was independent of ethanol. The SFW extract dilutions were less toxic than the SFE extracts. Significant DNA fragmentation as demonstrated by Hoechst staining was also seen over 48-hour incubation for high doses of both types of SF extracts. These results showed that although high concentrations of SF extracts can be toxic to normal T cells in vitro, SFW fractions were relatively safe for use.

The immunomodulatory effects of Sutherlandia frutescens extracts in human normal peripheral blood mononuclear cells.

Sutherlandia frutescens (SF) is one of the medicinal plants used as an immune booster in the treatment of chronic ailments such as HIV/AIDS and cancer. Limited data suggest that its efficacy is based on its regulatory effect on cytokines, the critical components of the immune response. In this study, we investigated the in vitro immunomodulatory effects of SF extracts on normal human peripheral blood mononuclear cells (PBMCs). An ELISA-based assay was used to assess the levels of expression of 12 cytokines in treated cells. An adenosine triphosphate (ATP) assay was used to assess cell viability in relation to cytokine secretion. SF ethanol extracts induced changes in cytokine secretion relative to the dose of the extract. Generally cytokine expression and secretion was low in concentration because were not stimulated with any endotoxin. The high SFE dose (2.5 mg/ml) significantly (p<0.001) decreased some cytokines including TNF-α and IL 1ß. Low doses of this extract (0.5 mg/ml) did not change TNF-α and IL 1ß secretion from the baseline (untreated cells). Changes in cytokine secretion of SFE treated cells tracked changes in ATP levels (cell viability). The SFW extract-induced changes in cytokine secretion were independent of cell viability. TNF-α was decreased (p<0.001) by the high dose of SFW extract while IL 1ß and IFNγ were increased (p<0.01) by the same dose. High doses decreased cell viability which was reflected in cytokine secretion. It is evident, from these results, that SF extracts can modulate cytokine secretion in unstimulated normal PBMCs in vitro. Further studies in animal models are recommended to advance understanding of this immunomodulatory activity.