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Fulltext In vivo toxicity and antitumor activity of essential oils extract from agarwood (Aquilaria crassna)

Dahham SS, Hassan LE, Ahamed MB, Majid AS, Majid AM, Zulkepli NN

BMC Complement Altern Med, 2016 Jul 22;16:236.
PMID: 27450078 DOI: 10.1186/s12906-016-1210-1

Aquilaria crassna has been used in traditional Asian medicine to treat vomiting, rheumatism, asthma, and cough. Furthermore, earlier studies from our laboratory have revealed that the essential oil extract from agarwood inhibited colorectal carcinoma cells. Despite of the wide range of ethno-pharmacological uses of agarwood, its toxicity has not been previously evaluated through systematic toxicological studies. Therefore, the potential safety of essential oil extract and its in vivo anti-tumor activity had been investigated.
Fulltext Correlation of antiangiogenic, antioxidant and cytotoxic activities of some Sudanese medicinal plants with phenolic and flavonoid contents

Hassan LE, Ahamed MB, Majid AS, Baharetha HM, Muslim NS, Nassar ZD, et al.

BMC Complement Altern Med, 2014 Oct 20;14:406.
PMID: 25331269 DOI: 10.1186/1472-6882-14-406

BACKGROUND: Consumption of medicinal plants to overcome diseases is traditionally belongs to the characteristics of most cultures on this earth. Sudan has been a host and cradle to various ancient civilizations and developed a vast knowledge on traditional medicinal plants. The present study was undertaken to evaluate the antioxidant, antiangiogenic and cytotoxic activities of six Sudanese medicinal plants which have been traditionally used to treat neoplasia. Further the biological activities were correlated with phytochemical contents of the plant extracts.
METHODS: Different parts of the plants were subjected to sequential extraction method. Cytotoxicity of the extracts was determined by dimethylthiazol-2-yl)- 2,5diphenyl tetrazolium bromide (MTT) assay on 2 human cancer (colon and breast) and normal (endothelial and colon fibroblast) cells. Anti-angiogenic potential was tested using ex vivo rat aortic ring assay. DPPH (1,1-diphenyl-2-picrylhydrazyl) assay was conducted to screen the antioxidant capabilities of the extracts. Finally, total phenolic and flavonoid contents were estimated in the extracts using colorimetric assays.
RESULTS: The results indicated that out of 6 plants tested, 4 plants (Nicotiana glauca, Tephrosia apollinea, Combretum hartmannianum and Tamarix nilotica) exhibited remarkable anti-angiogenic activity by inhibiting the sprouting of microvessels more than 60%. However, the most potent antiangiogenic effect was recorded by ethanol extract of T. apollinea (94.62%). In addition, the plants exhibited significant antiproliferative effects against human breast (MCF-7) and colon (HCT 116) cancer cells while being non-cytotoxic to the tested normal cells. The IC50 values determined for C. hartmannianum, N. gluaca and T. apollinea against MCF-7 cells were 8.48, 10.78 and 29.36 μg/ml, respectively. Whereas, the IC50 values estimated for N. gluaca, T. apollinea and C. hartmannianum against HCT 116 cells were 5.4, 20.2 and 27.2 μg/ml, respectively. These results were more or less equal to the standard reference drugs, tamoxifen (IC50 = 6.67 μg/ml) and 5-fluorouracil (IC50 = 3.9 μg/ml) tested against MCF-7 and HCT 116, respectively. Extracts of C. hartmannianum bark and N. glauca leaves demonstrated potent antioxidant effect with IC50s range from 9.4-22.4 and 13.4-30 μg/ml, respectively. Extracts of N. glauca leaves and T apollinea aerial parts demonstrated high amount of flavonoids range from 57.6-88.1 and 10.7-78 mg quercetin equivalent/g, respectively.
CONCLUSIONS: These results are in good agreement with the ethnobotanical uses of the plants (N. glauca, T. apollinea, C. hartmannianum and T. nilotica) to cure the oxidative stress and paraneoplastic symptoms caused by the cancer. These findings endorse further investigations on these plants to determine the active principles and their mode of action.
Fulltext Anti-Uterine Fibroid Effect of Standardized Labisia Pumila Var. Alata Extracts In Vitro and in Human Uterine Fibroid Cancer Xenograft Model

Zakaria N, Mohd KS, Ahmed Saeed MA, Ahmed Hassan LE, Shafaei A, Al-Suede FSR, et al.

Asian Pac J Cancer Prev, 2020 Apr 01;21(4):943-951.
PMID: 32334454 DOI: 10.31557/APJCP.2020.21.4.943

BACKGROUND: Uterine fibroids are a common type of solid tumor presenting in women of reproductive age. There are very few alternative treatment available from conventional treatment involving surgeries. Labisia pumila var. alata or locally known as 'Kacip Fatimah' was widely used as traditional medicine in Malaysia. This plant has been used to maintain a healthy female reproductive system. The present study aimed to evaluate anti fibroid potential of L. pumila extracts through in vitro apoptosis activity against uterine leiomyoma cells (SK-UT-1) and in uterine leiomyoma xenograft model. Evaluation of bioactive markers content were also carried out.
METHODS: Apoptotic induction of the extracts was determined by morphological examination of AO/PI dual staining assay by flourescent microscopy and flow cytometry analysis on Annexin V-FITC/PI stained cells. In vivo study was done in immune-compromised mouse xenograft model. HPLC analysis was employed to quantify marker compounds.
RESULTS: Morphological analysis showed L. pumila induced apoptosis in a dose dependent manner against SK-UT-1 cells. In vivo study indicated that L. pumila significantly suppressed the growth of uterine fibroid tumor. All tested extracts contain bioactive marker of gallic acid and cafeic acid.
CONCLUSION: This work provide significant data of the potential of L. pumila in management of uterine fibroids.
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Evaluation of in vitro and in vivo anti-inflammatory effects of (-)-pseudosemiglabrin, a major phytoconstituent isolated from Tephrosia apollinea (Delile) DC

Hassan LE, Dahham SS, Fadul SM, Umar MI, Majid AS, Khaw KY, et al.

J Ethnopharmacol, 2016 Aug 20.
PMID: 27553975 DOI: 10.1016/j.jep.2016.08.023

Tephrosia apollinea (Delile) DC (Leguminosae) has been used in folk medicine in Arabian countries to treat inflammatory disorders. The plant has been described to treat swelling, bone fracture, bronchitis, cough, earache and wounds.
Chemotherapeutic potentials of the stem bark of Balanite aegyptiaca (L.) Delile: an antiangiogenic, antitumor and antioxidant agent

Hassan LE, Dahham SS, Saghir SA, Mohammed AM, Eltayeb NM, Majid AM, et al.

BMC Complement Altern Med, 2016 Oct 19;16(1):396.
PMID: 27760539

Balanite aegyptiaca (L.) Delile, is a plant with extensive medicinal properties. Its stem bark is traditionally known for its spasmolytic and antiepileptic properties and used to treat yellow fever, jaundice and syphilis. Angiogenesis (sprouting of new blood vessels) is crucial for tumor growth and metastasis. The goal of this study is investigate the antiangiogenic, cytotoxicity and antioxidant activity as well as antitumor in vivo properties of B. aegyptiaca stem bark extracts.
Fulltext Ethyl-p-methoxycinnamate isolated from Kaempferia galanga inhibits inflammation by suppressing interleukin-1, tumor necrosis factor-α, and angiogenesis by blocking endothelial functions

Umar MI, Asmawi MZ, Sadikun A, Majid AM, Al-Suede FS, Hassan LE, et al.

Clinics (Sao Paulo), 2014 Feb;69(2):134-44.
PMID: 24519205 DOI: 10.6061/clinics/2014(02)10

The present study aimed to investigate the mechanisms underlying the anti-inflammatory and anti-angiogenic effects of ethyl-p-methoxycinnamate isolated from Kaempferia galanga.
Anti-inflammatory activities of cucurbitacin E isolated from Citrullus lanatus var. citroides: role of reactive nitrogen species and cyclooxygenase enzyme inhibition

Abdelwahab SI, Hassan LE, Sirat HM, Yagi SM, Koko WS, Mohan S, et al.

Fitoterapia, 2011 Dec;82(8):1190-7.
PMID: 21871542 DOI: 10.1016/j.fitote.2011.08.002

The in vivo and in vitro mechanistic anti-inflammatory actions of cucurbitacin E (CE) (Citrullus lanatus var. citroides) were examined. The results showed that LPS/INF-γ increased NO production in RAW264.7 macrophages, whereas L-NAME and CE curtailed it. CE did not reveal any cytotoxicity on RAW264.7 and WRL-68 cells. CE inhibited both COX enzymes with more selectivity toward COX-2. Intraperitoneal injection of CE significantly suppressed carrageenan-induced rat's paw edema. ORAC and FRAP assays showed that CE is not a potent ROS scavenger. It could be concluded that CE is potentially useful in treating inflammation through the inhibition of COX and RNS but not ROS.
Scopoletin, an active principle of tree tobacco (Nicotiana glauca) inhibits human tumor vascularization in xenograft models and modulates ERK1, VEGF-A, and FGF-2 in computer model

Tabana YM, Hassan LE, Ahamed MB, Dahham SS, Iqbal MA, Saeed MA, et al.

Microvasc Res, 2016 09;107:17-33.
PMID: 27133199 DOI: 10.1016/j.mvr.2016.04.009

We recently reported the antineovascularization effect of scopoletin on rat aorta and identified its potential anti-angiogenic activity. Scopoletin could be useful as a systemic chemotherapeutic agent against angiogenesis-dependent malignancies if its antitumorigenic activity is investigated and scientifically proven using a suitable human tumor xenograft model. In the present study, bioassay-guided (anti-angiogenesis) phytochemical investigation was conducted on Nicotiana glauca extract which led to the isolation of scopoletin. Further, anti-angiogenic activity of scopoletin was characterized using ex vivo, in vivo and in silico angiogenesis models. Finally, the antitumorigenic efficacy of scopoletin was studied in human colorectal tumor xenograft model using athymic nude mice. For the first time, an in vivo anticancer activity of scopoletin was reported and characterized using xenograft models. Scopoletin caused significant suppression of sprouting of microvessels in rat aortic explants with IC50 (median inhibitory concentration) 0.06μM. Scopoletin (100 and 200mg/kg) strongly inhibited (59.72 and 89.4%, respectively) vascularization in matrigel plugs implanted in nude mice. In the tumor xenograft model, scopoletin showed remarkable inhibition on tumor growth (34.2 and 94.7% at 100 and 200mg/kg, respectively). Tumor histology revealed drastic reduction of the extent of vascularization. Further, immunostaining of CD31 and NG2 receptors in the histological sections confirmed the antivascular effect of scopoletin in tumor vasculature. In computer modeling, scopoletin showed strong ligand affinity and binding energies toward the following angiogenic factors: protein kinase (ERK1), vascular endothelial growth factor A (VEGF-A), and fibroblast growth factor 2 (FGF-2). These results suggest that the antitumor activity of scopoletin may be due to its strong anti-angiogenic effect, which may be mediated by its effective inhibition of ERK1, VEGF-A, and FGF-2.
Anticlastogenic, antimutagenic, and cytoprotective properties of Orthosiphon stamineus ethanolic leaves extract

Al-Dulaimi DW, Shah Abdul Majid A, M Baharetha H, Ahamed MBK, Faisal SF, Al Zarzour RH, et al.

Drug Chem Toxicol, 2020 Apr 22.
PMID: 32321321 DOI: 10.1080/01480545.2020.1749652

Orthosiphon stamineus (O.S) is widely consumed for its medidcinal value including anti-inflammatory, anti-infective, and diuretic properties. The present study evaluates the cytoprotective, anti-mutagenic, and anticlastogenic efficacies of standardized extract of Orthosiphon stamineus. Normal liver cell line (WRL68) exposed to hydrogen peroxide and serum-deprived media as insults to evaluate cytoprotective and glutathione activation activities of (Et. O. s). Salmonella typhimurium TA98 and TA100 exposed to different concentrations of (Et. O. s). The influence of Et. O. s on mitotic, replicative indices as well as chromosomal aberration (CA) and sister chromatid exchange (SCE) induced in human peripheral blood lymphocytes by mitomycin C (MMC). The Et. O.s proved to be a potent scavenger for hydrogen peroxide and other free radicals in serum-depraved media, which showed to stimulate glutathione production in liver cells line. Moreover, it did not induce mutations in S. typhimurium subspecies TA98 and TA100. The standardized extract exhibited powerful antimutagenic activities as verified against both 2-nitrofluorene and sodium azide in S. typhimurium TA98 and TA100 cells, respectively. Cytogenetic tests showed high concentrations of Et. O. s to reduce the values of mitotic and replicative indices without any accompanying side effects, such as chromosomal abnormalities or SCE. To ameliorate MMC effects, pretreatment with the extract proofed to be efficient protocol. These data suggests that O. stamineus extract could be useful as cytoprotective, antimutagenic, and anticlastogenic efficacies, which owes to its potent chemoprevention, antioxidant, and glutathione activation properties.
Fulltext β-Caryophyllene Induces Apoptosis and Inhibits Angiogenesis in Colorectal Cancer Models

Dahham SS, Tabana Y, Asif M, Ahmed M, Babu D, Hassan LE, et al.

Int J Mol Sci, 2021 Sep 29;22(19).
PMID: 34638895 DOI: 10.3390/ijms221910550

Beta-Caryophyllene (BCP), a naturally occurring sesquiterpene abundantly found in cloves, hops, and cannabis, is the active candidate of a relatively new group of vascular-inhibiting compounds that aim to block existing tumor blood vessels. Previously, we have reported the anti-cancer properties of BCP by utilizing a series of in-vitro anti-tumor-related assays using human colorectal carcinoma cells. The present study aimed to investigate the effects of BCP on in-vitro, ex-vivo, and in-vivo models of anti-angiogenic assays and evaluate its anti-cancer activity in xenograft tumor (both ectopic and orthotopic) mice models of human colorectal cancer. Computational structural analysis and an apoptosis antibody array were also performed to understand the molecular players underlying this effect. BCP exhibited strong anti-angiogenic activity by blocking the migration of endothelial cells, tube-like network formation, suppression of vascular endothelial growth factor (VEGF) secretion from human umbilical vein endothelial cells and sprouting of rat aorta microvessels. BCP has a probable binding at Site#0 on the surface of VEGFR2. Moreover, BCP significantly deformed the vascularization architecture compared to the negative control in a chick embryo chorioallantoic membrane assay. BCP showed a remarkable reduction in tumor size and fluorescence molecular tomography signal intensity in all the mice treated with BCP, in a dose-dependent relationship, in ectopic and orthotopic tumor xenograft models, respectively. The histological analysis of the tumor from BCP-treated mice revealed a clear reduction of the density of vascularization. In addition, BCP induced apoptosis through downregulation of HSP60, HTRA, survivin, and XIAP, along with the upregulation of p21 expressions. These results suggest that BCP acts at multiple stages of angiogenesis and could be used as a promising therapeutic candidate to halt the growth of colorectal tumor cells.
Fulltext Crystal structure elucidation and anticancer studies of (-)-pseudosemiglabrin: a flavanone isolated from the aerial parts of Tephrosia apollinea

Ahmed Hassan LE, Khadeer Ahamed MB, Abdul Majid AS, Iqbal MA, Al Suede FS, Haque RA, et al.

PLoS One, 2014;9(6):e90806.
PMID: 24608571 DOI: 10.1371/journal.pone.0090806

Tephrosia apollinea is a perennial shrublet widely distributed in Africa and is known to have medicinal properties. The current study describes the bio-assay (cytotoxicity) guided isolation of (-)-pseudosemiglabrin from the aerial parts of T. apollinea. The structural and stereochemical features have been described using spectral and x-ray crystallographic techniques. The cytotoxicity of isolated compound was evaluated against nine cancer cell lines. In addition, human fibroblast was used as a model cell line for normal cells. The results showed that (-)-pseudosemiglabrin exhibited dose-dependent antiproliferative effect on most of the tested cancer cell lines. Selectively, the compound showed significant inhibitory effect on the proliferation of leukemia, prostate and breast cancer cell lines. Further studies revealed that, the compound exhibited proapoptotic phenomenon of cytotoxicity. Interestingly, the compound did not display toxicity against the normal human fibroblast. It can be concluded that (-)-pseudosemiglabrin is worthy for further investigation as a potential chemotherapeutic agent.
Fulltext Optimization of Cat's Whiskers Tea (Orthosiphon stamineus) Using Supercritical Carbon Dioxide and Selective Chemotherapeutic Potential against Prostate Cancer Cells

Al-Suede FS, Khadeer Ahamed MB, Abdul Majid AS, Baharetha HM, Hassan LE, Kadir MO, et al.

Evid Based Complement Alternat Med, 2014;2014:396016.
PMID: 25276215 DOI: 10.1155/2014/396016

Cat's whiskers (Orthosiphon stamineus) leaves extracts were prepared using supercritical CO2 (SC-CO2) with full factorial design to determine the optimum extraction parameters. Nine extracts were obtained by varying pressure, temperature, and time. The extracts were analysed using FTIR, UV-Vis, and GC-MS. Cytotoxicity of the extracts was evaluated on human (colorectal, breast, and prostate) cancer and normal fibroblast cells. Moderate pressure (31.1 MPa) and temperature (60°C) were recorded as optimum extraction conditions with high yield (1.74%) of the extract (B2) at 60 min extraction time. The optimized extract (B2) displayed selective cytotoxicity against prostate cancer (PC3) cells (IC50 28 µg/mL) and significant antioxidant activity (IC50 42.8 µg/mL). Elevated levels of caspases 3/7 and 9 in B2-treated PC3 cells suggest the induction of apoptosis through nuclear and mitochondrial pathways. Hoechst and rhodamine assays confirmed the nuclear condensation and disruption of mitochondrial membrane potential in the cells. B2 also demonstrated inhibitory effects on motility and colonies of PC3 cells at its subcytotoxic concentrations. It is noteworthy that B2 displayed negligible toxicity against the normal cells. Chemometric analysis revealed high content of essential oils, hydrocarbon, fatty acids, esters, and aromatic sesquiterpenes in B2. This study highlights the therapeutic potentials of SC-CO2 extract of cat's whiskers in targeting prostate carcinoma.
Fulltext Cucurbitacin L 2-O-β-Glucoside Demonstrates Apoptogenesis in Colon Adenocarcinoma Cells (HT-29): Involvement of Reactive Oxygen and Nitrogen Species Regulation

Abdelwahab SI, Hassan LE, Abdul Majid AM, Yagi SM, Mohan S, Elhassan Taha MM, et al.

Evid Based Complement Alternat Med, 2012;2012:490136.
PMID: 22685485 DOI: 10.1155/2012/490136

Emerging evidence suggests that reactive oxygen (ROS) and nitrogen (RNS) species can contribute to diverse signalling pathways of inflammatory and tumour cells. Cucurbitacins are a group of highly oxygenated triterpenes. Many plants used in folk medicine to treat cancer have been found to contain cucurbitacins displaying potentially important anti-inflammatory actions. The current study was designed to investigate the anti-ROS and -RNS effects of cucurbitacin L 2-O-β-glucoside (CLG) and the role of these signaling factors in the apoptogenic effects of CLG on human colon cancer cells (HT-29). This natural cucurbitacin was isolated purely from Citrullus lanatus var. citroides (Cucurbitaceae). The results revealed that CLG was cytotoxic to HT-29. CLG increased significantly (P < 0.05) RNA and protein levels of caspase-3 in HT-29 cells when verified using a colorimetric assay and realtime qPCR, respectively. The results showed that lipopolysaccharide/interferon-gamma (LPS/INF-γ) increased nitrous oxide (NO) production inR AW264.7macrophages, whereas N(G)-nitro-L-argininemethyl ester (L-NAME) and CLG curtailed it. This compound did not reveal any cytotoxicity on RAW264.7 macrophages and human normal liver cells (WRL-68) when tested using the MTT assay. Findings of ferric reducing antioxidant power (FRAP) and oxygen radical absorption capacity (ORAC) assays demonstrate the antioxidant properties of CLG. The apoptogenic property of CLG on HT-29 cells is thus related to inhibition of reactive nitrogen and oxygen reactive species and the triggering of caspase-3-regulated apoptosis.

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