Phytochemical Investigation of Egyptian Riverhemp: A Potential Source of Antileukemic Metabolites

Abdelgawad, Shimaa M.; Hetta, Mona H.; Ibrahim, Mohamed A.; Balachandran, Premalatha; Zhang, Jin; Wang, Mei; Eldehna, Wagdy M.; Fawzy, Ghada A.; El-Askary, Hesham I.; Ross, Samir A.

doi:https://doi.org/10.1155/2022/8766625

Journal of Chemistry

On this page

Abstract Introduction Materials and Methods Results and Discussion Conclusion Data Availability Conflicts of Interest Acknowledgments Supplementary Materials References Copyright Related Articles

Special Issue

The Development of Novel Biologically Active Small Molecules 2022

View this Special Issue

Research Article | Open Access

Volume 2022 | Article ID 8766625 | https://doi.org/10.1155/2022/8766625

Phytochemical Investigation of Egyptian Riverhemp: A Potential Source of Antileukemic Metabolites

Shimaa M. Abdelgawad,^1,2Mona H. Hetta,²Mohamed A. Ibrahim,¹Premalatha Balachandran,¹Jin Zhang,¹Mei Wang,³Wagdy M. Eldehna,^4,5Ghada A. Fawzy ,⁶Hesham I. El-Askary,⁶and Samir A. Ross^1,7

Academic Editor: Ajaya Kumar Singh

Received19 Apr 2022

Revised11 May 2022

Accepted24 Jun 2022

Published29 Aug 2022

Abstract

As part of our research group’s continuous efforts to find alternative treatments for cancer, the aqueous ethanol extract of Sesbania sesban L. Merr. (SS, Egyptian riverhemp) demonstrated an antileukemic activity against K562 cell line. Bioguided fractionation of SS leaves hydroethanolic extract resulted in the isolation of one new compound (33) named as hederatriol 3-O-β-D-glucuronic acid methyl ester as well as 34 known compounds. Seven compounds ((34), (22), (20), (24), (21), (19), and (35)) showed high antiproliferative effects (IC₅₀ = 22.3, 30.8, 31.3, 33.7, 36.6, 37.5, and 41.5 μM, respectively), while four compounds ((32), (5), (29), and (1)) showed milder activities (IC₅₀ = 56.4, 67.6, 83.3, and 112.3 μM, respectively). A mechanistic study was further carried out on a molecular genetics level against several transcription factors signaling pathways that are incorporated in the incidence of cancer. The results showed that compounds (22) and (21) demonstrated a specific inhibition of Wnt pathway (IC₅₀ = 3.8 and 4.6 μM, respectively), while compound (22) showed a specific inhibition of Smad pathway (IC₅₀ = 3.8 μM). Compound (34) strongly altered the signaling of Smad and E2F pathways (IC₅₀ = 5 μM). The bioactive metabolites were further investigated in silico by docking against several targets related to K562 cell line. The results showed that compounds (22) and (34) exhibited a strong binding affinity towards topoisomerase (docking score = −7.81 and −9.30 Kcal/Mole, respectively). Compounds (22) and (34) demonstrated a strong binding affinity towards EGFR-tyrosine kinase (docking score = −7.12 and −7.35 Kcal/Mole, respectively). Moreover, compound (34) showed a strong binding affinity towards Abl kinase (docking score = −7.05 Kcal/Mole).

1. Introduction

In our previous study, the antileukemic activities of 56 medicinal plants grown in Upper Egypt were checked, where Sesbania sesban (SS) leaves extract demonstrated a promising antileukemic activity against the chronic myeloid leukemia cell line (K562) [1]. Considering the limited studies on the phytoconstituents of SS leaves, we carried out this research with the aim of bioguided isolation of the antileukemic metabolites from SS leaves.

Chronic myeloid leukemia (CML) is the third most regular mortality-causing neoplasm and the fourth predominant cancer among the Egyptian population [2, 3]. CML has a trademark and explicit chromosomal irregularity known as Philadelphia (Ph) chromosome (Bcr-Abl gene) that is produced from a reciprocal translocation between the Abl gene on chromosome 9 and the Bcr gene on chromosome 22 in the pluripotent hematopoietic stem cells [4].

The value of natural products in the arsenal of leukemia therapies is clearly correlated with several agents such as L-asparaginase, Daunorubicin, Anthracyclines, Vinca alkaloids (Vincristine and Vinblastine) [5, 6], Homoharringtonine, Indirubin, Flavopiridol, Maytansinoids, Meisoindigo [6], and the Podophyllotoxin derivatives (Etoposide and Teniposide) [7, 8].

S. sesban (Egyptian riverhemp or Sesbania aegyptiaca Pers.) is a perennial legume tree belonging to family Fabaceae (Figure 1). It has several names like Aeschynomene aegyptiaca (Pers.) Steud., Aeschynomene sesban L., Sesbania confaloniana Chiov., Sesbania pubescens sensu auct., Sesbania punctata DC., and Emerus sesban (L.) Kuntze [9].

Traditionally, SS leaves were used as a poultice for inflammatory conditions such as boils and abscesses as well as rheumatic swellings, and the juice of the fresh leaves was used as anthelmintic [10–12].

The methanolic extract of SS leaves exhibited several biological activities such as anti-inflammatory [13], antioxidant [14], molluscicidal [15, 16], antidiabetic [17], potent spermicidal [18], antimicrobial [16], and anthelmintic activities [10–12]. Phytochemical analyses of the leaves showed the presence of steroids such as β-sitosterol and campesterol [13], triterpenoids such as betulinic and oleanolic acids [13, 19, 20], saponins of oleanolic acid [15, 18], and flavonoids [21].

The aim of this study is to isolate the antileukemic metabolites of SS leaves through extensive biological and phytochemical investigations.

2. Materials and Methods

2.1. General Experimental Procedures

High-resolution electrospray ionization mass spectrometry (HRESIMS) data were acquired using a Bruker BioApex-FTMS with electrospray ionization (ESI). 1D (¹HNMR, DEPTQ) and 2D (HSQC, HMBC, and TOCSY) NMR spectra were recorded on a Bruker 400 and 500 MHz spectrometer. Optical activity was measured using AA-65 series automatic polarimeter (Cambridgeshire, PE26 1NF, England). HPLC analysis was conducted using an Agilent 1100 HPLC system (Supplementary Materials). GC-MS analysis was performed with an Agilent 7890B gas chromatograph (Supplementary Materials). Sephadex LH-20 (Mitsubishi Kagaku, Tokyo, Japan) and silica gel (60–120 μM·mesh, Merck, Darmstadt, Germany), reversed phase silica (40–63 μM, Sorbent Technologies, 5955 Peachtree Corners East, Suite A, Norcross, GA 30071 USA), and Diaion® HP-20 (250 μm, Supelco, Bellefonte, PA 16823-00048, USA) were used for column chromatography (CC). SPE cartridges silica gel and C18 (Supelco Inc., Bellefonte, PA, USA) were used in the fractionation work. Fractions from CC were monitored by TLC using precoated aluminum sheets (silica 60 F254, 0.25 mm (Merck, Darmstadt, Germany)). The fractions were dissolved in the appropriate solvents and spots were applied manually using the capillary micropipette, and the plates were then dried and developed in different mobile phase systems such as n-hexane: ethyl acetate (8 : 2), n-hexane: acetone (8 : 2), and DCM: methanol (9 : 1). The spots on the developed plates were detected using UV light (254 and 366 nm) and by spraying with 2% p-anisaldehyde-H₂SO₄ reagent followed by heating for 5–10 min (105°C). Number of spots, colors, and retardation factors (Rf values) for each of the spots were determined and recorded [22].

2.2. Plant Material

The leaves of Sesbania sesban were purchased from the medicinal, aromatic, and poisonous plants experimental station, Faculty of Pharmacy, Cairo University, Egypt, in April 2018. The plant material was authenticated by Professor Dr. Abdelhalim Mohamed, Flora and Phytotaxonomy Research Department, Horticulture Research Institute (HRI), Agricultural Research Center, Giza, Egypt. A voucher specimen (FUPD-45) was kept at the Herbarium of Pharmacognosy Department, Faculty of Pharmacy, Fayoum University, Egypt.

2.3. Extraction and Bioguided Isolation

The shade-dried leaves (1 kg) were grounded and extracted with 75% ethanol at room temperature three times 24 h each. The total extract was concentrated to afford a crude residue (177 g). 83 g of the 75% ethanolic extract was suspended in water and exposed to successive fractionation using n-hexane and EtOAc to afford 9 g and 12 g, respectively. The remaining aqueous fraction was lyophilized to afford 61 g. All fractions were phytochemically investigated and tested against K562 cell line. The bioactive fractions were subjected to bioguided isolation techniques as described in the consequent text.

The n-hexane fraction (6 g) of SS leaves was saponified according to the published procedure [23] and the fatty acid methyl ester (FAME) and unsaponifiable matter (USM) were then subjected to GC/MS analysis. USM (78.8 mg) was chromatographed on silica gel column by gradient elution with n-hexane/EtOAc (2.5% gradient) to give 9 subfractions (A1–A9). Fr-A-6 (eluted by 7.5% EtOAc) was one spot on TLC and afforded compound 1 (11 mg). Fr-A-8 (eluted by 10% EtOAc) was one spot on TLC and afforded compound 2 (25 mg).

The EtOAc fraction (10 g) was chromatographed on a silica gel solid phase extraction (Si-SPE) column by gradient elution with n-hexane/EtOAc to give fractions (B1–B5). The subfraction Fr-B-2 (520 mg) was first subjected to silica gel SPE column chromatography and eluted with n-hexane/EtOAc gradient (2% gradient) to afford 5 subfractions. Subfraction Fr-B-2-2 was further purified using silica gel column and eluted with n-hexane/EtOAc (2.5% gradient) to afford compound (3) (5 mg) (eluted by 10% EtOAc). The subfraction Fr-B-2-4 was further chromatographed on silica gel column and eluted with n-hexane/acetone (2.5% gradient) to afford compounds (4) (1 mg), (5) (15.6 mg), (6) (1 mg), and (7) (2.5 mg) (eluted by 7.5%, 10%, 12.5%, and 15% acetone, respectively). The subfraction Fr-B-5 (4.5 g) was chromatographed on VLC silica gel column by gradient elution with DCM/MeOH (2.5% gradient) to afford 5 subfractions. Fr-B-5-4 (614.8 mg) (eluted by 20% MeOH) was subjected to a column of Sephadex LH-20 and eluted with methanol to give 8 subfractions. Fr-B-5-4-5 (164.2 mg) was chromatographed on a silica gel column and eluted with DCM/MeOH (2.5% gradient) to afford compound (8) (8 mg) (eluted by 25% MeOH). Fr-B-5-4-6 (46 mg) was subjected to a column chromatography on Sephadex LH-20 and eluted with MeOH to give compound (9) (2 mg). Fr-B-5-4-8 (34.3 mg) was subjected to column chromatography on a Sephadex LH-20 and eluted with MeOH to give compounds (10) (14.5 mg), (11) (11 mg), (12) (4 mg), and (13) (3 mg). Fr-B-5-5 (2.35 g) was subjected to a column of Sephadex LH-20 and eluted with methanol to afford 8 subfractions. Fr-B-5-5-4 (358 mg) was subjected to a column of Sephadex LH-20 and eluted with methanol to afford 5 subfractions. Fr-B-5-5-4-3 (27 mg) was purified by HPLC RP column and eluted with water/methanol gradient to afford compounds (14) (20 mg) and (15) (2.36 mg) at retention times (RT) = 9.8 and 10.3 minutes, respectively (Fig. S1, Supplementary Materials). Fr-B-5-5-4-5 (30 mg) was purified by HPLC RP column and eluted with water/methanol gradient to afford compounds (16) (15 mg) and (17) (3 mg) at RT = 8.0 and 8.29 minutes, respectively (Fig. S2, Supplementary Materials). Fr-B-5-5-6 (80 mg) was chromatographed on a silica gel column by gradient elution with DCM/MeOH (2.5% gradient) to afford compound (18) (6 mg) (eluted by 25% MeOH).

The aqueous fraction (60.0 g) was subjected to a column chromatography using HP-20 ion exchange resin and eluted with MeOH/water (25% gradient) to afford five fractions (C1–C5). Fr-C-5 (5 g) (eluted by 100% MeOH) was further chromatographed on a silica gel column by gradient elution with DCM/MeOH (2.5% gradient) to afford 9 subfractions. Fr-C-5-2 (511 mg) (eluted by 2.5% MeOH) was further chromatographed on a silica gel column using gradient elution with DCM/EtOAc (5% gradient) to afford compounds (19) (17.7 mg), (20) (33.3 mg), (21) (5 mg), (22) (8 mg), (23) (2 mg), (24) (52 mg), (25) (1.9 mg), and (26) (1.7 mg). Fr-C-5-4 (511 mg) was chromatographed on RP-18-silica gel column by gradient elution with MeOH/water (5% gradient) to give four subfractions. Fr-C-5-4-1 (7 mg) was purified by RP-18 preparative HPLC to give compounds (27) (20 mg) and (28) (6 mg) at retention times (RT) = 17.17 and 18.14 minutes, respectively (Fig. S3, Supplementary Materials). Fr-C-5-4-3 (198 mg) was chromatographed on a silica gel column using DCM/MeOH (5% gradient) to afford compounds (29) (45 mg), (30) (2 mg), (31) (2 mg), (23) (3 mg), (33) (2 mg), and (34) (22.5 mg). Fr-C-5-5 (672 mg) was chromatographed on reversed phase silica gel solid phase extraction (RP-SPE) column by gradient elution with MeOH/water (5% gradient) to give four subfractions. Fr-C-5-5-2 (400 mg) was further chromatographed on a silica gel column by gradient elution with DCM/MeOH (5% gradient) to afford compound (35) (64.8 mg).

Compounds (1)–(35) were identified using ¹HNMR, DEPT-Q, HSQC, HMBC, and HRESIMS spectroscopic techniques as well as comparing these data with the literature.

2.4. Hederatriol 3-O-β-D-Glucuronic Acid Methyl Ester (33)

White powder (MeOH), [α] ^25°_D = +34.5 (c = 0.15 in MeOH). ¹H and DEPT-Q data (Table 1); HRESIMS m/z 649.45082 [M+H] ⁺ (calcd. C₃₇H₆₁O₉ m/z 649.43156).

2.5. Cytotoxicity Assay

K562 cells from the American Type Culture Collection (ATCC) were plated in clear 384-well plates at an initial density of 2500 cells/well in 40 μL of growth medium (DMEM with 10% FBS and 1% Pen/step). Next day, the test agents were added in quadruplicates at the specified concentration and the treatment continued for 48 h and the cell viability was finally assessed using WST-8 assay Cell Counting Kit from Bimake, according to manufacturer’s instructions. The results were calculated by measuring the absorbance at 450 nm using SpectraMax M5 plate reader (Molecular Devices). Cell viability was calculated in comparison to DMSO as a negative control, as well as Taxol and Doxorubicin as positive controls [24]. The extract and fractions were screened primarily at concentrations of 50 and 75 μg/mL and the percentage inhibitions were calculated. Compounds (1)–(35) were tested at six concentrations (5, 10, 25, 50, 75, and 100 μg/mL), where compounds (1), (5), (19), (20), (21), (22), (24), (29), (32), (34), and (35) were shown to be bioactive and their IC₅₀ values were calculated.

2.6. Transfection and Luciferase Assays

Hela cells from ATCC were plated in white opaque 384-well plates at a density of 4300 cells/well in 30 μL of growth medium (DMEM with 10% FBS and 1% Pen/step). Next day, the medium was aspirated and replaced with DMEM containing 1% FBS. The cells were transfected with respective plasmids [25] using X-tremeGENE HP transfection reagent (Roche). After 24 h of transfection, the test agents were added in duplicates to the transfected cells, followed 30 min later by an inducing agent (IL-6 for Stat 3, TGF-β for Smad, m-Wnt3a for Wnt, and PMA for AP-1, NFk-B, E2F, Myc, ETS, Notch, and Hedgehog). No inducer was added for pTK, FOXO, miR-2, K-Ras, and AhR. After 4 h or 6 h of induction, the cells were lysed by the addition of One-Glo luciferase assay system (Promega, Madison, WI, USA). The light output was detected in a GloMax-Multi+ Detection System with Instinct Software (Promega, Madison, WI, USA). This luciferase assay determines if the test agent was able to inhibit the activation of cancer-related signaling pathways. In the case of FOXO and mi-R21, the enhanced luciferase activity by the test agent was assessed [26].

2.7. Molecular Docking

All docking simulations were conducted using MOE 2019 software (https://www.chemcomp.com). The receptors and the ligands were prepared using the standard structure optimization protocol of the software. The receptors were obtained from the protein data bank, PDB IDs: 3QX3, 3QRJ, 1M17, and 2SRC for topoisomerase, Abl kinase, EGFR-tyrosine kinase, and SRC kinase, respectively. Then they were energy-minimized under AMBER12:EHT force field. The active sites were set as where the cocrystalized ligand was bound. The docking was performed using a molecular structure of compounds isolated from SS leaves using the general protocol of MOE DOCKTITE Wizard. The validation of docking experiments was achieved through the redocking of the cocrystalized ligands into their corresponding active sites and then the root means square deviation (RMSD) was calculated. The docking results were visualized and analyzed by DS Visualizer available from BIOVIA Inc. and the affinity of binding (docking scores) was calculated in Kcal/Mole for each compound against the selected targets.

3. Results and Discussion

Through our ongoing efforts to discover antileukemic natural compounds from Egyptian plants, SS leaves showed a potential promising result against K562 cell line in vitro [1]. Accordingly, SS leaves were selected for further phytochemical and biological investigations with the aim of isolating the bioactive metabolites. The total extract and the successive fractions were screened against K562 cell line at a concentration of 20 μg/mL and the percentage inhibitions were calculated. The extracts of 75% ethanol, n-hexane, ethyl acetate, and the aqueous fractions showed mild antileukemic activities with percent inhibitions of 13%, 16%, 23%, and 9%, respectively, at a concentration of 20 μg/mL. All the fractions were phytochemically investigated, and their columns subfractions were screened for their antileukemic activities as described in the biological study results.

3.1. Phytochemical Study

3.1.1. Bioguided Phytochemical Investigation of the n-Hexane Fraction

The study resulted in the identification of 16 compounds in the saponifiable matter (Table S1, Supplementary Materials) with five majors identified as methyl palmitate, methyl elaidate, methyl linoleate, methyl linolenate, and ethyl linolenate at retention times = 33.1, 37.5, 37.8, 38.4, and 39.6 minutes, respectively. Moreover, 12 compounds were identified in the unsaponifiable matter (Table S2, Supplementary Materials) with five majors identified as phytol, stigmasta-5,22-dien-3-ol acetate (3, beta, 22 Z), stigmastan-3,5,22-trien, stigmasta-3,5-diene, and stigmasterol at retention times = 37.2, 47, 47.2, 52.6, and 66.1 minutes, respectively. Additionally, the unsaponifiable matter was purified and resulted in the isolation of two compounds identified as phytol (1) [27] and stigmasterol (2) [28] (Fig. S4, Supplementary Materials).

3.1.2. Bioguided Phytochemical Investigation of the EtOAc Fraction

The result was the isolation of 16 known compounds identified as liquidambaric lactone (3) [29], maslinic lactone (4) [30], oleanolic acid (5) [31], 11α,12α-epoxy-oleanolic lactone (6) [32], oleanderolide (7) [33], stigmasterol glucoside (8) [34], P-aminophenol (9) [35], quercetin-3-O-α-L-rhamnopyranoside (10) [36], quercetin-3-O-β-D-glucopyranoside (11) [36], quercetin-3-O-α-L-arabinoside (12) [37], quercetin-3-O-β-D-xyloside (13) [37], mauritianin (14) [38], isorhamnetin 3-O-(2,6-di-O-α-rhamnosyl)-β-D-galactopyranoside (15) [39], clitorin (16) [40], 3-[(O-6-deoxy-α-L-mannopyranosyl-(1⟶2)-O-[6-deoxy-α-L-mannopyranosyl-(1⟶6)]-β-D-glucopyranosyl) oxy]-5,7-dihydroxy-2-(3-hydroxy-4-methoxyphenyl)-4H-1-benzopyran-4-one (17) [41], and kaempferol-3-O-α-L-rhamnopyranosyl-(1⟶2)[α-L-rhamnopyranosyl-(1⟶6)]-(4-O-E-p-coumaroyl-β-D-galactopyranoside) (18) [42] (Fig. S4 and Table S3, Supplementary Materials).

3.1.3. Bioguided Phytochemical Investigation of the Aqueous Fraction

The result was the isolation of one new compound (33) (Hederatriol 3-O-β-D-glucuronic acid methyl ester), as well as 16 known compounds identified as betulinic acid (19) [31,43], ursolic acid (20) [44], 3β-O-(cis-p-coumaroyl)-2α-hydroxyurs-12-en-28-oic acid (21) [45], 3β-O-(trans-p-coumaroyl)-2α-hydroxyurs-12-en-28-oic acid (known as jacoumaric acid) (22) [45], obtuslin (23) [44], corosolic acid (24) [45], β-sitosterol glucoside (25) [46], hederagenin-3-O-β-D-glucuronopyranoside 6′-O-methyl ester (26) [47], kaempferol-3-O-rutinoside (27) [48], isorhamentin-3-O-rutinoside (28) [49], hederagenin-3-O-β-D-glucuronopyranoside (29) [47], 11-ketocorosolic acid (30) [50], hederagenin (31) [51], oleanolic acid 3-O-β-D-glucuronopyranoside 6′ methyl ester (32) [52], oleanolic acid 3-O-β-D-glucuronopyranoside (34) [18], and chikusetsusaponin II (35) [53] Fig. S4 and Table S3, (Supplementary Materials).

Compounds (2), (5), (19), and (34) were previously reported in SS leaves [18,19], while this is the first report of compounds (1), (3)-(4), (6)–(18), (20)–(33), and (35) from this species.

3.2. Structure Elucidation of Compound (33)

Compound (33) was isolated as a white powder (2 mg, soluble in MeOH), [α] ^25°_D = +34.5 (c = 0.15 in MeOH). Its molecular formula, C₃₇H₆₀O₉, was derived from the ¹D NMR data (Table 1) and the supportive HRESIMS ion at m/z 649.45082 [M+H] ⁺ (calcd. for C₃₇H₆₁O₉, 649.43156). The molecular formula revealed the hydrogen deficiency index to be 29.6 ppm. From the ¹H NMR spectrum, the seven distinct methyls [δ_H 3.67 (3H, s), 1.09 (3H, s), 0.87 (6H, s), 0.86 (3H, s), 0.78 (3H, S), 0.71 (3H, s), and 0.59 (3H, s)] were observed alongside with the signal assignable to one olefinic proton [δ_H 5.76 (1H, t)]. Two oxygenated methylenes were detected (δ_H 3.51 (2H, S), 3.07 (1H, d, J = 10.5 Hz), and 3.44 (1H, d, J = 11 Hz)). Five oxygenated methines (δ_H 3.67 (1H), 3.50 (IH), 3.28 (1H, t), 3.16 (1H, t), 2.98 (1H, t)) alongside the one anomeric proton (δ_H 4.33 (d, J = 8 Hz)) were detected. The DEPTQ-135 spectra of (33) showed 37 carbon signals (Table 1) of which 30 signals were assigned to the aglycone moiety (hederatriol) and 7 to a glucuronic acid methyl ester moiety. The anomeric carbon resonances (δ_C/_H 105.1/4.33) as well as four oxygenated methine signals at δ_C/_H 74.1/2.98, 75.9/3.16, 72.1/3.28, and 76.4/3.67 alongside the carbonyl ester (δ_C 170.1) and the attached methoxy signal (δ_C/_H 52.3/3.67) disclosed the presence of glucuronic acid 6′ methyl ester moiety. The anomeric configuration of the glucuronoid unit was determined to be based on the coupling constant (J_H1=8 Hz). There were 30 carbons left for the skeleton, indicating a triterpenoid core of (33). The skeletal carbons, particularly the six methyl carbons (δ_C 33.3, 26.0, 23.9, 17.4, 16.0, and 13.3), alongside the two oxygenated methylenes [δ_C 62.8 (C-24) and 70.2 (C-28)] implied an olean-type triterpenes alcohol derivative (hederatriol) scaffold for (33) [54]. Free alcoholic methylene at position C28 was confirmed by 2D HMBC & TOCSY correlations between the OH-proton at C28 (δ_H 3.5) and C18 in HMBC spectrum. Moreover, protons H16, H19, and H21 showed TOCSY correlations to H28. Free alcohol OH signal (δ_H 3.79) showed TOCSY correlation with H28 methylene. All of the previous data supported the presence of hederatriol. The HMBC cross-peaks enabled the substituent groups to be positioned. The glucuronoid moiety was connected to C-3 as suggested by the HMBC correlation of H-1′ with δ_C 80.4 (C-3) and H-3 to the anomeric carbon at δ_C105.1 (C-1′) of the glucuronoid moiety (Figure 2). The structure of compound (33) is considered to be new due to the 3-O-glycosylation of hederatriol with glucuronic acid methyl ester. Thus, compound (33) was elucidated as hederatriol 3-O-β-D-glucuronic acid methyl ester (NMR data, Figures S5–S11, Supplementary Materials).

3.3. Biological Study

3.3.1. Cytotoxicity against Leukemia K562 Cell Line

The n-hexane fraction was fractionated by saponification with KOH, where the yielded saponifiable and unsaponifiable matters were tested against K562 cells and showed inhibitions of IC₅₀ = 75.7 and 73.9 μg/mL, respectively. Accordingly, the phytoconstituents in both fractions were investigated using GC-MS analysis.

The EtOAc fraction (Fr-B) was fractionated using Si-SPE column and the five resulted subfractions were tested against K562 cells at concentrations of 50 and 75 μg/mL. The results showed that subfractions Fr-B-2 and Fr-B-5 were the bioactive fractions with percent inhibitions of 93.7% and 47.3%, respectively, at a concentration of 75 μg/mL (Figure S12, Supplementary Materials). The subfraction Fr-B-2 was chromatographed using silica gel SPE column and the resulting subfractions were tested against K562 cells. Subfractions Fr-B-2-2 and Fr-B-2-4 were the most cytotoxic with percent inhibitions of 45% and 47%, respectively, at a concentration of 75 μg/mL (Fig. S13, Supplementary Materials). The subfraction Fr-B-5 was chromatographed using VLC silica gel column and the resulting subfractions were tested against K562 cells. Subfractions Fr-B-5-4 and Fr-B-5-5 were the most cytotoxic with percent inhibitions of 28% and 85%, respectively, at a concentration of 75 μg/mL (Fig. S13, Supplementary Materials).

The aqueous fraction (Fr-C) was chromatographed using HP-20 ion exchange resin and the five resulting subfractions that were screened for their activity against K562 cells at concentrations of 50 and 75 μg/mL. The results showed that subfraction Fr-C-5 was the bioactive fraction with 94.4% percent of inhibition at concentrations of 50 and 75 μg/mL (Fig. S14, Supplementary Materials). Subfraction Fr-C-5 was chromatographed on silica gel column and the eight produced subfractions that were screened for their activity against K562 cells at concentrations of 50 and 75 μg/mL. The results showed that subfractions Fr-C-5-2 and Fr-C-5-4 were the most active ones with 65% and 63% percent of inhibitions, respectively, at a concentration of 75 μg/mL (Fig. S15, Supplementary Materials).

All the detected bioactive subfractions were subjected to several chromatographic techniques as mentioned in the Materials and Methods section with the aim of isolating and elucidating the structures of the bioactive constituents.

Compounds (1)–(35) were screened against K562 cell line at concentrations of 50 and 75 μg/mL and IC₅₀ values were calculated. Oleanolic acid 3-O-β-D-glucuronopyranoside (34) was identified as the most antiproliferative compound isolated from SS extract with IC₅₀ value of 22.3 μM compared to Taxol (∼2 μM) and Doxorubicin (∼4 μM). Other active triterpenes were betulinic acid (19), ursolic acid (20), 3β-O-(cis-p-coumaroyl)-2α-hydroxyurs-12-en-28-oic acid (21), jacoumaric acid (22), corosolic acid (24), and chikusetsusaponin II (35) with IC₅₀ values of 37.5, 31.3, 36.6, 30.8, 33.7, and 41.5 μM, respectively. Phytol (1), oleanolic acid (5), hederagenin 3-O-β-D-glucuronopyranoside (29), and oleanolic acid 3-O-β-D-glucuronopyranoside methyl ester (32) showed mild activity against K562 cell line with IC₅₀ values of 112.3, 67.6, 83.3, and 56.4 μM, respectively (Figure 3). Out of the 35 isolated compounds, the triterpene glycosides were shown to be the most active antileukemic compounds in this plant. Antileukemic activities of betulinic acid (19) [55], ursolic acid (20) [56], oleanolic acid (5) [57], and phytol (1) [58] were previously reported in literature but this is the first report of antileukemic activities of compounds (21), (22), (24), (29), (32), (34), and (35). Moreover, methyl linolenate, the major constituent of the saponifiable matter of the n-hexane fraction of SS leaves, was previously reported for its cytotoxicity against K562 cell line [59]. Other isolated compounds showed no cytotoxicity against K562 cell line.

3.3.2. Triterpenes Selectively Target Cancer Signaling Pathways

The most active isolated compounds (21), (22), (24), and (34) against K562 cell line were further evaluated for targeting the cancer signaling pathways. These compounds were tested for their influence on the battery of 15-plex transcription factor assay. The results (Table 2 and Figure 4) showed that compounds (22) and (21) showed specific inhibition of Wnt pathway (IC₅₀ = 3.8 and 4.6 μM, respectively) while compound (22) showed specific inhibition of Smad (IC₅₀ = 3.8 μM). Compound (34) can strongly alter the signaling pathways of Smad and E2F (IC₅₀ = 5 μM). From these results, we can conclude the correlation between the inhibition of Smad, E2F, and Wnt signaling pathways and the leukemia K562 cell line growth inhibition (Figure 5).

As cancer signaling plays a complex role in CML, the observed results from this study could be helpful in finding new therapeutic agents and molecular genetics-based treatment targeting these metabolic pathways. Despite the fortune available research on the molecular details of TGF-β signaling, little is known regarding how TGF-β and Smad influence CML [60]. One of these realized relationships is that the constitutively active tyrosine kinase produced by the specific Bcr-Abl fusion gene on the Philadelphia chromosome can enhance the resistance of malignant cells to TGF-β1-induced growth inhibition and apoptosis [61]. Previous studies reported ursolic acid and oleanolic acid as antagonists of TGF-β1 binding to its receptor in Balb/c 3T3 cells [62]. A recent study revealed that inhibition of E2F1 reduced CML cell proliferation, leading to p53-mediated apoptosis [63]. The BCR-ABL1 protein kinase-dependent pathway intervened by the upregulation of hsa-mir183, the downregulation of its direct target early growth response 1 (EGR1), and, as a consequence, upregulation of E2F1 [58]. Wnt signaling assumes a significant part in the malignancies of the hematopoietic system [64]. Several compounds that are natural in origin were reported in literature for targeting Wnt pathway as a mechanism for anticancer activity such as ursolic acid and corosolic acid that were reported as antagonists of the Wnt/β-catenin pathway [65–67]. Correlating the previous literature to the results, a mechanism of action of the active compounds could be supposed. The active compounds were mainly oleanolic acid and ursolic acid derivatives which exhibited their antileukemic activities by targeting Smad, E2F, and Wnt signaling pathways (Figure 5).

3.3.3. Molecular Docking

Multitarget therapies are crucial in the field of complex diseases such as cancers due to activation of compensatory mechanisms and consecutive cellular pathways [68]. The recent research showed a correlation between CML cell line K562 and several proteins which are considered as targets whose inhibition leads to antiproliferative effect in this cell line. Four target proteins, human topoisomerase II beta in complex with DNA and etoposide (PDB ID: 3QX3), epidermal growth factor receptor complexed with erlotinib (PDB ID: 1M17), human Abl-1 kinase (PDB ID: 3QRJ), and SRC Kinase (PDB ID: 2SRC), were reported as docking targets in the K562 cell line [69, 70].

Based on the previous biological results, compounds (22) and (34) were selected and screened against the selected targets and the affinity of binding was calculated (Table 3). Compounds (22) and (34) showed a strong binding affinity towards topoisomerase (docking score = −7.81 and −9.30 Kcal/Mole, respectively) compared to etoposide (docking score = −7.10 Kcal/Mole). Compounds (22) and (34) demonstrated a strong binding affinity towards EGFR-tyrosine kinase (docking score = −7.12 and −7.35 Kcal/Mole, respectively) compared to erlotinib (docking score = −6.99 Kcal/Mole). Moreover, compound (34) showed a strong binding affinity towards Abl kinase (docking score = −7.05 Kcal/Mole) compared to rebastinib (docking score = −7.86 Kcal/Mole) (Table 3 and Figure 6).

(a)

(b)

(c)

3.3.4. Structure Activity Relationship (SAR) Study

The structures of the isolated compounds could be classified as oleane triterpenoids, oleane lactones triterpenoids, ursane triterpenoids, lupane triterpenes, steroids, and flavonoid glycosides (Fig. S3 and Table S4, Supplementary Materials).

Among the different classes of isolated compounds, triterpenes were shown as the bioactive antileukemic molecules present in the SS leaves. The potency of the isolated triterpenoids prompted us to investigate the structure activity relationships underlying their antileukemic activities.

In this study, three types of triterpenoids aglycones were isolated and identified: lupane triterpenoids represented by betulinic acid, oleane type represented by oleanolic acid, and ursane type represented by ursolic acid. The results showed that the most active antileukemic triterpenoid is ursane type followed by lupane and the least active type is oleane triterpenoids (Figure 7).

From ursane type triterpenoids, six compounds (20)–(24) and (30) were isolated. Comparing their structures and antileukemic activities, two positions, C-2 and C-3, were believed to affect the activity. Hydroxylation at C-2 position slightly decreases the antileukemic activity. Also, the esterification at C-3 position by coumaric acid can increase or decrease the activity depending on the configuration of coumaric acid moiety. The trans coumaroyl derivative, jacoumaric acid (22), showed higher antileukemic activity than corosolic acid (24), while cis derivative, 3β-O-(cis-p-coumaroyl)-2α-hydroxyurs-12-en-28-oic acid (21), showed lower activity and these results support the idea of the higher activity of trans conformers than the cis one (Figure 8).

The SAR study showed that the activity of oleanolic acid can be greatly increased when esterified at C-3 position by glucuronic acid. Compound (34) showed strong antileukemic activity compared to compound (5) due to the presence of glucuronic acid moiety attached at C-3. In addition, the free OH group in glucuronic acid is crucial for the activity and, as the results show, when it is methylated as in compound (32) or glycosylated as in compound (35), the activity decreased. Meanwhile, for oleane triterpenes, the methylation at C-23 as in compounds (5), (32), and (34) increases the activity compared to compound (29) (Figure 9).

4. Conclusion

One new compound (33) and 34 known compounds were isolated from the leaves of Sesbania sesban encompassing 19 triterpenoids ((3)–(7), (19)–(24), (26), and (29)–(35)), three steroids ((2), (8), and (25)), 11 flavonoid glycosides ((10)–(18) and (27)-(28)), one fatty alcohol (1), and one phenolic compound (9). The antileukemic activity of the isolated compounds was evaluated against leukemia K562 cell line in vitro and 11 triterpenoids exhibited remarkable antileukemic activities against this cell line. Compounds (34) (oleanolic acid 3-O-β-D-glucuronopyranoside) and (22) (jacoumaric acid) exhibited the strongest activities with IC₅₀ values of 22.3 and 30.8 μM, respectively. The mechanism of the antileukemic activity of these compounds could be attributed to the attenuation of Smad, Wnt, and E2F signaling pathways. Compound (22) demonstrated a specific inhibition of Wnt and Smad signaling with an IC₅₀ value of 3.8 μM for both pathways. Meanwhile compound (34) showed an inhibition of E2F with an IC₅₀ value of 5 μM. Moreover, the binding patterns of the bioactive molecules against leukemia cell related targets were explored by docking them against topoisomerase, EGFR-tyrosine kinase, and Abl kinase. Compounds (22) and (34) exhibited a strong binding affinity towards topoisomerase (docking score = −7.81 and −9.30 Kcal/Mole, respectively). Furthermore, SAR study reveals that the presence of a carboxyl group either in the triterpene or in the sugar moiety was identified as contributing to the cytotoxic activity.

Data Availability

Data are available in the Supplementary Materials.

Conflicts of Interest

The authors declare no conflicts of interest.

Acknowledgments

This work was supported by the Egyptian Ministry of Higher Education Missions Sector (Grant no. JS-3770). Support from National Center for Natural Products Research (NCNPR), University of Mississippi, is gratefully acknowledged.

Supplementary Materials

Table S1: results of GC/MS analysis of fatty acid methyl ester (FAME) of the n-hexane fraction of SS leaves. Table S2: results of GC/MS analysis of unsaponifiable matter of the n-hexane fraction of SS leaves. Table S3: list of compounds isolated from Sesbania sesban L. leaves. Figure S1: HPLC (RP column) purification of compounds (14) and (15) using MeOH/water gradient. Figure S2: HPLC (RP column) purification of compounds (16) and (17) using MeOH/water gradient. Figure S3: HPLC (RP column) purification of compounds (27) and (28) using MeOH/water gradient. Figure S4: chemical structures of compounds isolated from SS leaves. Figure S5: ¹H NMR spectrum of compound (33). Figure S6: deptq135 spectrum of compound (33). Figure S7: dept spectrum of compound (33). Figure S8: HSQC spectrum of compound (33). Figure S9: TOCSY spectrum of compound (33). Figure S10: TOCSY spectrum of compound (33). Figure S11: ESI(-) MS spectrum of compound (33). Figure S12: bioguided screening of EtOAc (Fr-B) column subfractions against leukemia K562 cell line. Figure S13: bioguided screening of Fr-B-2 column subfractions and Fr-B-5 column subfractions against leukemia K562 cell line. Figure S14: bioguided screening of aqueous (Fr-C) column subfractions against leukemia K562 cell line. Figure S15: bioguided screening of Fr-C-5 column subfractions against leukemia K562 cell line. (Supplementary Materials)

References

M. H. H. Shimaa Mohammed Abdelgawad, G. Ahmed Fawzy, and H. I. El-Askary, “In vitro antileukemic activity of extracts of some medicinal plants from upper Egypt in human chronic leukemia K562 cell line,” Tropical Journal of Natural Products Research, vol. 5, no. 12, pp. 2115–2122, 2021.
View at: Google Scholar
M. A. Aboul-Soud, H. A. El-Shemy, K. M. Aboul-Enein, A. M. Mahmoud, A. M. Al-Abd, and D. A. Lightfoot, “Effects of plant-derived anti-leukemic drugs on individualized leukemic cell population profiles in Egyptian patients,” Oncology letters, vol. 11, no. 1, pp. 642–648, 2016.
View at: Google Scholar
A. S. Ibrahim, H. M. Khaled, N. N. Mikhail, H. Baraka, and H. Kamel, “Cancer incidence in Egypt: results of the national population-based cancer registry program,” Journal of cancer epidemiology, vol. 2014, Article ID 437971, 18 pages, 2014.
View at: Publisher Site | Google Scholar
S. Eskandari and R. Yazdanparast, “Bcl6 gene-silencing facilitates PMA-induced megakaryocyte differentiation in K562 cells,” Journal of Cell Communication and Signaling, vol. 11, no. 4, pp. 357–367, 2017.
View at: Publisher Site | Google Scholar
M. Karon, E. J. Freireich, E. Frei et al., “The role of vincristine in the treatment of childhood acute leukemia,” Clinical Pharmacology & Therapeutics, vol. 7, no. 3, pp. 332–339, 1966.
View at: Publisher Site | Google Scholar
D. M. Lucas, P. C. Still, L. Bueno Perez, M. R. Grever, and A. Douglas Kinghorn, “Potential of plant-derived natural products in the treatment of leukemia and lymphoma,” Current Drug Targets, vol. 11, no. 7, pp. 812–822, 2010.
View at: Publisher Site | Google Scholar
J. M. S. van Maanen, J. Retel, J. De Vries, and H. M. Pinedo, “Mechanism of action of antitumor drug etoposide: a review,” JNCI Journal of the National Cancer Institute, vol. 80, no. 19, pp. 1526–1533, 1988.
View at: Publisher Site | Google Scholar
K. Hande, “Etoposide: four decades of development of a topoisomerase II inhibitor,” European Journal of Cancer, vol. 34, no. 10, pp. 1514–1521, 1998.
View at: Publisher Site | Google Scholar
T. G. Heuzé, D. Bastianelli, and F. Lebas, “Sesban (Sesbania sesban),” 2015, https://www.feedipedia.org/node/253.
View at: Google Scholar
E. Debela, A. Tolera, L. O. Eik, and R. Salte, “Condensed tannins from Sesbania sesban and Desmodium intortum as a means of Haemonchus contortus control in goats,” Tropical Animal Health and Production, vol. 44, no. 8, 2012.
View at: Publisher Site | Google Scholar
E. G. Kamel, M. A. El-Emam, S. S. Mahmoud, F. M. Fouda, and F. E. Bayaumy, “Parasitological and biochemical parameters in Schistosoma mansoni-infected mice treated with methanol extract from the plants Chenopodium ambrosioides, Conyza dioscorides and Sesbania sesban,” Parasitology International, vol. 60, no. 4, pp. 388–392, 2011.
View at: Publisher Site | Google Scholar
A. D. Soren, R. P. Chen, and A. K. Yadav, “In vitro and in vivo anthelmintic study of Sesbania sesban var. bicolor, a traditionally used medicinal plant of Santhal tribe in Assam, India,” Journal of Parasitic Diseases, vol. 45, no. 1, pp. 1–9, 2021.
View at: Publisher Site | Google Scholar
A. U. Tatiya, P. R. Dande, R. E. Mutha, and S. J. Surana, “Effect of saponins from of Sesbania sesban L.(Merr) on acute and chronic inflammation in experimental induced animals,” Journal of Biological Sciences, vol. 13, no. 3, pp. 123–130, 2013.
View at: Publisher Site | Google Scholar
S. N. Fitriansyah, I. Fidrianny, and K. Ruslan, “Correlation of total phenolic, flavonoid and carotenoid content of Sesbania sesban (L. Merr) leaves extract with DPPH scavenging activities,” International Journal of Pharmacognosy and Phytochemical Research, vol. 9, no. 1, pp. 89–94, 2017.
View at: Google Scholar
A.-C. Dorsaz, M. Hostettmann, and K. Hostettmann, “Molluscicidal saponins from Sesbania sesban,” Planta Medica, vol. 54, no. 03, pp. 225–227, 1988.
View at: Publisher Site | Google Scholar
M. N. Walekhwa, T. K. Ogeto, M. K. Murithi, and Z. L. Malago, “Pharmacological effects of Sesbania sesban: a systematic review,” International Journal of Research in Medical Sciences, vol. 8, no. 3, p. 1185, 2020.
View at: Publisher Site | Google Scholar
R. B. Pandhare, B. Sangameswaran, P. B. Mohite, and S. G. Khanage, “Antidiabetic activity of aqueous leaves extract of Sesbania sesban (L) Merr. in streptozotocin induced diabetic rats,” Avicenna Journal of Medical Biotechnology, vol. 3, no. 1, pp. 37–43, 2011.
View at: Google Scholar
N. Das, P. Chandran, and S. Chakraborty, “Potent spermicidal effect of oleanolic acid 3-beta-D-glucuronide, an active principle isolated from the plant Sesbania sesban Merrill,” Contraception, vol. 83, no. 2, pp. 167–175, 2011.
View at: Publisher Site | Google Scholar
M. Farooq, I. Varshney, and M. S. Khan, “Saponins and sapogenins IV,” Journal of the American Pharmaceutical Association, vol. 48, no. 8, pp. 466–468, 1959.
View at: Publisher Site | Google Scholar
S. Manoharan and J. Kaur, “Anticancer, antiviral, antidiabetic, antifungal and phytochemical constituents of medicinal plants,” American Journal of PharmTech Research, vol. 3, pp. 149–169, 2013.
View at: Google Scholar
H. Dianhar, Y. M. Syah, D. Mujahidin, E. H. Hakim, and L. D. Juliawaty, “A flavone derivative from Sesbania sesban leaves and its cytotoxicity against murine leukemia P-388 cells,” AIP Conference Proceedings, vol. 1589, no. 1, pp. 411–413, 2014.
View at: Google Scholar
M. Namadina, S. Yahuza, H. Haruna et al., “Pharmacognostic and antioxidant activity of bryophyllum pinnatum leaves,” International Journal Of Science for Global Sustainability, vol. 6, no. 2, 2020.
View at: Google Scholar
I. Finar, Organic Chemistry, ELBS, Ghitorni, India, 6 edition, 1973.
M. Kageyama, K. Li, S. Sun et al., “Anti-tumor and anti-metastasis activities of honey bee larvae powder by suppressing the expression of EZH2,” Biomedicine & Pharmacotherapy, vol. 105, pp. 690–696, 2018.
View at: Publisher Site | Google Scholar
P. Balachandran, M. A. Ibrahim, J. Zhang, M. Wang, D. S. Pasco, and I. Muhammad, “Crosstalk of cancer signaling pathways by cyclic hexapeptides and anthraquinones from rubia cordifolia,” Molecules, vol. 26, no. 3, p. 735, 2021.
View at: Publisher Site | Google Scholar
M. A. Zaki, P. Balachandran, S. Khan et al., “Cytotoxicity and modulation of cancer-related signaling by (Z)-and (E)-3, 4, 3′, 5′-tetramethoxystilbene isolated from Eugenia rigida,” Journal of Natural Products, vol. 76, no. 4, pp. 679–684, 2013.
View at: Publisher Site | Google Scholar
D. Arigoni, W. Eisenreich, C. Latzel et al., “Dimethylallyl pyrophosphate is not the committed precursor of isopentenyl pyrophosphate during terpenoid biosynthesis from 1-deoxyxylulose in higher plants,” Proceedings of the National Academy of Sciences, vol. 96, no. 4, pp. 1309–1314, 1999.
View at: Publisher Site | Google Scholar
P. Jain, S. Bari, and S. Surana, “Isolation of stigmasterol and γ-sitosterol from petroleum ether extract of woody stem of Abelmoschus manihot,” Asian journal of biological sciences, vol. 2, no. 4, pp. 112–117, 2009.
View at: Publisher Site | Google Scholar
Y. Zhu, Y. J. Guan, Q. Z. Chen et al., “Pentacyclic Triterpenes from the resin of Liquidambar formosana have anti-angiogenic properties,” Phytochemistry, vol. 184, Article ID 112676, 2021.
View at: Publisher Site | Google Scholar
G. R. Mallavarapu and E. Muralikrishna, “Maslinic lactone from the heartwood of Terminalia alata,” Journal of Natural Products, vol. 46, no. 6, pp. 930-931, 1983.
View at: Publisher Site | Google Scholar
M. A. Hossain and Z. Ismail, “Isolation and characterization of triterpenes from the leaves of Orthosiphon stamineus,” Arabian Journal of Chemistry, vol. 6, no. 3, pp. 295–298, 2013.
View at: Publisher Site | Google Scholar
Q. Fu, L. Qiu, H.-M. Yuan, T. Yu, L. Zou, and D. and E. Paeonenoides, “Two new nortriterpenoids fromPaeonia lactifloraand their inhibitory activities on NO production,” Helvetica Chimica Acta, vol. 99, no. 1, pp. 46–49, 2016.
View at: Publisher Site | Google Scholar
L. Fu, S. Zhang, N. Li et al., “Three new triterpenes from nerium oleander and biological activity of the isolated compounds,” Journal of Natural Products, vol. 68, no. 2, pp. 198–206, 2005.
View at: Publisher Site | Google Scholar
S. K. Nigam, G. Misra, and C. R. Mitra, “Stigmasterol glucoside a constituent of Adenanthera pavonina seed and leaf,” Planta Medica, vol. 23, no. 2, pp. 145–148, 1973.
View at: Publisher Site | Google Scholar
R. Eyanagi, Y. Hisanari, and H. Shigematsu, “Studies of paracetamol/phenacetin toxicity: isolation and characterization of p-aminophenol-glutathione conjugate,” Xenobiotica, vol. 21, no. 6, pp. 793–803, Jun 1991.
View at: Publisher Site | Google Scholar
K. R. Markham and B. Ternai, “13C NMR of flavonoids—II,” Tetrahedron, vol. 32, no. 21, pp. 2607–2612, 1976.
View at: Publisher Site | Google Scholar
M. Kalegari, M. D. Miguel, JdF. G. Dias et al., “Phytochemical constituents and preliminary toxicity evaluation of leaves from Rourea induta Planch.(Connaraceae),” Brazilian Journal of Pharmaceutical Sciences, vol. 47, no. 3, pp. 635–642, 2011.
View at: Publisher Site | Google Scholar
I. Dini, G. Carlo Tenore, and A. Dini, “Phenolic constituents of Kancolla seeds,” Food Chemistry, vol. 84, no. 2, pp. 163–168, 2004/02/01/2004.
View at: Google Scholar
F. Ferreres, D. M. Pereira, P. Valentão, P. B. Andrade, R. M. Seabra, and M. Sottomayor, “New phenolic compounds and antioxidant potential of catharanthus roseus,” Journal of Agricultural and Food Chemistry, vol. 56, no. 21, pp. 9967–9974, 2008/11/12 2008.
View at: Publisher Site | Google Scholar
N. Morita, M. Arisawa, M. Nagase, H. Hsu, and Y. Chen, “Studies on the constituents of formosan leguminosae. I. The constituents in the leaves of Clitoria ternatea L,” Yakugaku zasshi: Journal of the Pharmaceutical Society of Japan, 1977.
View at: Google Scholar
R. S. Mohammed, A. H. Abou Zeid, E. A. El-Kashoury, A. A. Sleem, and D. A. Waly, “A new flavonol glycoside and biological activities of Adenanthera pavonina L. leaves,” Natural Product Research, vol. 28, no. 5, pp. 282–289, 2014.
View at: Publisher Site | Google Scholar
G. C. Kite, E. R. Rowe, G. P. Lewis, and N. C. Veitch, “Acylated flavonol tri- and tetraglycosides in the flavonoid metabolome of Cladrastis kentukea (Leguminosae),” Phytochemistry, vol. 72, no. 4-5, pp. 372–384, 2011/04/01/2011.
View at: Google Scholar
H. H. F. Koolen, E. R. Soares, F. M. A. da. Silva, A. Q. L. de. Souza, E. Rodrigues Filho, and A. D. L. de. Souza, “Triterpenes and flavonoids from the roots of Mauritia flexuosa,” Revista Brasileira de Farmacognosia, vol. 22, no. 1, pp. 189–192, 2012.
View at: Publisher Site | Google Scholar
W. Seebacher, N. Simic, R. Weis, R. Saf, and O. Kunert, “Complete assignments of 1H and 13C NMR resonances of oleanolic acid, 18α-oleanolic acid, ursolic acid and their 11-oxo derivatives,” Magnetic Resonance in Chemistry, vol. 41, no. 8, pp. 636–638, 2003.
View at: Publisher Site | Google Scholar
C.-K. Lee, “Ursane triterpenoids from leaves of Melaleuca leucadendron,” Phytochemistry, vol. 49, no. 4, pp. 1119–1122, 1998.
View at: Publisher Site | Google Scholar
A. Hamed, M. Ismail, M. M. El-Metwally et al., “Diverse polyketides and alkaloids from Penicillium sp. KHMM: structural elucidation, biological and molecular docking studies,” Zeitschrift fuer Naturforschung, C: Journal of Biosciences, vol. 74, no. 5-6, pp. 131–137, 2019.
View at: Publisher Site | Google Scholar
S. Srivastava and D. Jain, “Triterpenoid saponins from plants of Araliaceae,” Phytochemistry, vol. 28, no. 2, pp. 644–647, 1989.
View at: Publisher Site | Google Scholar
R. González-Laredo, M. Flores De La Hoya, M. Quintero-Ramos, and J. Karchesy, “Flavonoid and cyanogenic contents of chaya (spinach tree),” Plant Foods for Human Nutrition, vol. 58, no. 3, pp. 1–8, 2003.
View at: Publisher Site | Google Scholar
A. M. El-Hawiet, S. M. Toaima, A. M. Asaad, M. M. Radwan, and N. A. El-Sebakhy, “Chemical constituents from Astragalus annularis forssk. and A. Trimestris L., Fabaceae,” Revista Brasileira de Farmacognosia, vol. 20, no. 6, pp. 860–865, 2010.
View at: Publisher Site | Google Scholar
A. Ikuta, H. Tomiyasu, Y. Morita, and K. Yoshimura, “Ursane-and oleanane-type triterpenes from ternstroemia g ymnanthera callus tissues,” Journal of Natural Products, vol. 66, no. 8, pp. 1051–1054, 2003.
View at: Publisher Site | Google Scholar
B. S. Joshi, K. L. Singh, and R. Roy, “Complete assignments of 1H and 13C NMR spectra of the pentacyclic triterpene hederagenin from Nigella sativa Linn,” Magnetic Resonance in Chemistry, vol. 37, no. 4, pp. 295–298, 1999.
View at: Google Scholar
D. Zhang, Y. Fu, J. Yang et al., “Triterpenoids and their glycosides from Glinus oppositifolius with antifungal activities against Microsporum gypseum and Trichophyton rubrum,” Molecules, vol. 24, no. 12, p. 2206, 2019.
View at: Publisher Site | Google Scholar
T.-D. Lin and J. Shoji, “Constituents ofPanacis JaponiciRhizoma. VI the structure of chikusetsu-saponin II, IVC,” Journal of the Chinese Chemical Society, vol. 26, no. 1, pp. 29–35, 1979.
View at: Publisher Site | Google Scholar
K. Tori, S. Seo, A. Shimaoka, and Y. Tomita, “Carbon-13 NMR spectra of olean-12-enes. Full signal assignments including quaternary carbon signals assigned by use of indirect 13C, 1H spin couplings,” Tetrahedron Letters, vol. 15, no. 48, pp. 4227–4230, 1974.
View at: Publisher Site | Google Scholar
W. Qiuling, H. Jing, F. Jun, and H. Mei, “Antitumor effect of betulinic acid on human acute leukemia K562 cells in vitro,” Journal of Huazhong University of Science and Technology - Medical sciences, vol. 30, no. 4, pp. 453–457, 2010.
View at: Publisher Site | Google Scholar
B. Wu, X. Wang, Zf Chi et al., “Ursolic acid-induced apoptosis in K562 cells involving upregulation of PTEN gene expression and inactivation of the PI3K/Akt pathway,” Archives of Pharmacal Research, vol. 35, no. 3, pp. 543–548, 2012.
View at: Publisher Site | Google Scholar
S. Pan, J. Hu, T. Zheng, X. Liu, Y. Ju, and C. J. C. Xu, “Oleanolic acid derivatives induce apoptosis in human leukemia K562 cell involved in inhibition of both Akt1 translocation and pAkt1 expression,” Cytotechnology, vol. 67, no. 5, pp. 821–829, 2015.
View at: Publisher Site | Google Scholar
S. Anuchapreeda et al., “Antileukemic cell proliferation of active compounds from kaffir lime (citrus hystrix) leaves,” Molecules, vol. 25, no. 6, p. 1300, 2020.
View at: Publisher Site | Google Scholar
H. Ge, X. Kong, L. Shi, L. Hou, Z. Liu, and P. Li, “Gamma-linolenic acid induces apoptosis and lipid peroxidation in human chronic myelogenous leukemia K562 cells,” Cell Biology International, vol. 33, no. 3, pp. 402–410, 2009.
View at: Publisher Site | Google Scholar
G. M. Møller, V. Frost, J. V. Melo, and A. Chantry, “Upregulation of the TGFβ signalling pathway by Bcr‐Abl: implications for haemopoietic cell growth and chronic myeloid leukaemia,” FEBS Letters, vol. 581, no. 7, pp. 1329–1334, 2007.
View at: Publisher Site | Google Scholar
S. Mohan, A. B. Abdul, S. I. Abdelwahab et al., “Typhonium flagelliforme inhibits the proliferation of murine leukemia WEHI-3 cells in vitro and induces apoptosis in vivo,” Leukemia Research, vol. 34, no. 11, pp. 1483–1492, 2010.
View at: Publisher Site | Google Scholar
S. Murakami, H. Takashima, M. Sato-Watanabe et al., “Ursolic acid, an antagonist for transforming growth factor (TGF)-β1,” FEBS Letters, vol. 566, no. 1-3, pp. 55–59, 2004.
View at: Publisher Site | Google Scholar
F. Pellicano, L. Park, L. E. M. Hopcroft et al., “hsa-mir183/EGR1–mediated regulation of E2F1 is required for CML stem/progenitor cell survival,” Blood, vol. 131, no. 14, pp. 1532–1544, 2018.
View at: Publisher Site | Google Scholar
F. J. Staal, F. Famili, L. Garcia Perez, and K. Pike-Overzet, “Aberrant Wnt signaling in leukemia,” Cancers, vol. 8, no. 9, p. 78, 2016.
View at: Publisher Site | Google Scholar
D. Liu, L. Chen, H. Zhao, N. D. Vaziri, S.-C. Ma, and Y.-Y. Zhao, “Small molecules from natural products targeting the Wnt/β-catenin pathway as a therapeutic strategy,” Biomedicine & Pharmacotherapy, vol. 117, Article ID 108990, 2019.
View at: Google Scholar
M. Sheremet, S. Kapoor, P. Schröder, K. Kumar, S. Ziegler, and H. Waldmann, “Small molecules inspired by the natural product withanolides as potent inhibitors of wnt signaling,” ChemBioChem, vol. 18, no. 18, pp. 1797–1806, 2017.
View at: Publisher Site | Google Scholar
M. Ishibashi, “Screening for natural products that affect Wnt signaling activity,” Journal of Natural Medicines, vol. 73, no. 4, pp. 697–705, 2019.
View at: Publisher Site | Google Scholar
Z. i. Skok, N. Zidar, D. Kikelj, and J. Ilaš, “Dual inhibitors of human DNA topoisomerase II and other cancer-related targets,” Journal of Medicinal Chemistry, vol. 63, no. 3, pp. 884–904, 2019.
View at: Publisher Site | Google Scholar
A. R. James, B. Unnikrishnan, R. Priya et al., “Computational and mechanistic studies on the effect of galactoxyloglucan: imatinib nanoconjugate in imatinib resistant K562 cells,” Tumor Biology, vol. 39, no. 3, Article ID 101042831769594, 2017.
View at: Publisher Site | Google Scholar
N. H. Zamakshshari, G. C. L. Ee, I. S. Ismail, Z. Ibrahim, and S. H. Mah, “Cytotoxic xanthones isolated from Calophyllum depressinervosum and Calophyllum buxifolium with antioxidant and cytotoxic activities,” Food and Chemical Toxicology, vol. 133, Article ID 110800, 2019.
View at: Google Scholar

Copyright

Copyright © 2022 Shimaa M. Abdelgawad et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

PDF Download Citation

Download other formats

Order printed copies

Views

461

Downloads

394

Citations