Evaluation of in vitro and in vivo anticancer activities of potassium koetjapate: a solubility improved formulation of koetjapic acid against human colon cancer

Fatemeh Jafari , Maryam Keshavarzi, Amin MalikShah AbdulMajid, Fouad Saleih R. Al-Suede, Muhammad Asif, Mohamed B. Khadeer Ahamed, Md Shamsuddin Sultan Khan, Loiy Ahmed Elsir Hassan, Aman Shah Abdul Majid, Mohsen Naseri


Background and purpose: The previous work on koetjapic acid (KA) isolated from Sandoricum koetjape showed its efficacy towards colorectal cancer however KA has poor water solubility which poses the biggest hindrance to its efficacy. In the present paper, an attempt was made to study the anti-colon cancer efficacy of KA’s potassium salt i.e. potassium koetjapate (KKA) applying in vitro and in vivo methods.

Experimental approach: KKA was produced by a semi-synthetic method. A human apoptosis proteome profiler array was applied to determine the protein targets responsible for the stimulation of apoptosis. Three doses of KKA were studied in athymic nude mice models to examine the in vivo anti-tumorigenic ability of KKA.

Findings/Results: The results of this study demonstrated that KKA regulates the activities of various proteins. It downregulates the expression of several antiapoptotic proteins and negative regulators of apoptosis including HSP60, HSP90, Bcl-2, and IGF-1 in HCT 116 cells with consequent upregulation of TRAILR-1 and TRAILR-2, p27, CD40, caspase 3, and caspase 8 proteins. Additionally, KKA showed an in vitro antimetastatic effect against HCT 116 cells. These results are feasibly related to the down-regulation of Notch, Wnt, hypoxia, and MAPK/JNK and MAPK/ERK signalling pathways in HCT 116 cells besides the up-regulation of a transcription factor for cell cycle (pRb-E2F) pathways. In addition, KKA revealed potent inhibition of tumor growth.

Conclusion and implications: In sum, the findings indicate that KKA can be a promising candidate as a chemotherapeutic agent against colorectal cancer.



Apoptosis; Colon cancer; Hypoxia; MAPK signalling pathways; Potassium koetjapate; TRAILR-1&2.

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Bailly C. The health benefits of santol fruits and bioactive products isolated from Sandoricum koetjape merr. J Food Biochem. 2022;46(7): e14152.DOI: 10.1111/JFBC.14152.

Powell RG, Mikolajczak KL, Zilkowski BW, Mantus EK, Cherry D, Clardy J. Limonoid antifeedants from seed of Sandoricum koetjape. J Nat Prod. 1991;54(1):241-246.DOI: 10.1021/np50073a025.

Ismail IS, Ito H, Hatano T, Taniguchi S, Yoshida T. Modified limonoids from the leaves of Sandoricum koetjape. Phytochemistry. 2003;64(8):1345-1349.DOI: 10.1016/S0031-9422(03)00500-4.

Ismail IS, Ito H, Hatano T, Taniguchi S, Yoshida T. Two new analogues of trijugin-type limonoids from the leaves of Sandoricum koetjape. Chem Pharm Bull. 2004;52(9):1145-1147.DOI: 10.1248/cpb.52.1145.

Pancharoen O, Pipatanapatikarn A, Taylor WC, Bansiddhi J. Two new limonoids from the leaves of Sandoricum koetjape. Nat Prod Res. 2009;23(1):10-16.DOI: 10.1080/14786410601133426.

Kaneda N, Pezzuto JM, Kinghorn AD, Farnsworth NR, Santisuk T, Tuchinda P, et al. Plant anticancer agents, L. cytotoxic triterpenes from Sandoricum koetjape stems. J Nat Prod. 1992;55(5):654-659.DOI: 10.1021/np50083a016.

Tanaka T, Koyano T, Kowithayakorn T, Fujimoto H, Okuyama E, Hayashi M, et al. New Multiflorane-type triterpenoid acids from Sandoricum indicum. J Nat Prod. 2001;64(9):1243-1245.DOI: 10.1021/np010196w.

Nassar ZD, Aisha AFA, Idris N, Ahamed MBK , Ismail Z, Salah KMA, et al. Koetjapic acid, a natural triterpenoid, induces apoptosis in colon cancer cells. Oncol. 2012;27(3):727-733.DOI: 10.3892/OR.2011.1569.

Jafari SF, Ahamed MBK, Iqbal MA, Al Suede FSR, Khalid SH, Haque RA, et al. Increased aqueous solubility and proapoptotic activity of potassium koetjapate against human colorectal cancer cells. J Pharm Pharmacol. 2014;66(10):1394-1409.DOI: 10.1111/jphp.12272.

Jafari SF, Al-Suede FSR, Yehya AHS, Ahmad MBK, Shafaei A, Asif M, et al. Pharmacokinetics and antiangiogenic studies of potassium koetjapate in rats. Biomed. Pharmacother. 2020;130:110602,1-11.DOI: 10.1016/j.biopha.2020.110602.

Benassi-Zanqueta E, Marques CF, Valone LM, Pellegrini BL, Bauermeister A, Ferreira ICP , et al. Evaluation of anti-HSV-1 activity and toxicity of hydroethanolic extract of Tanacetum parthenium (L.) Sch.Bip. (Asteraceae). Int J Phytomedicine.2019;55:249-254.DOI: 10.1016/j.phymed.2018.06.040.

Liang CC, Park AY, Guan JL. In vitro scratch assay: a convenient and inexpensive method for analysis of cell migration in vitro. Nat Protoc. 2007;2(2): 329-333.DOI: 10.1038/nprot.2007.30.

Asif M, Shafaei A, Jafari SF, Mohamed SK, Ezzat MO, Majid ASA, et al. Isoledene from Mesua ferrea oleo-gum resin induces apoptosis in HCT 116 cells through ROS-mediated modulation of multiple proteins in the apoptotic pathways: A mechanistic study. Toxicol Lett. 2016;22:257,84-96.DOI: 10.1016/j.toxlet.2016.05.027.

Yehya AHS, Al-mansoub MA, Al-suede FSR, Sultan SBB, Abduraman MA, Azmi NA, et al. Anti-tumor activity of NuvastaticTM (C5OSEW5050ES) of Orthosiphon stamineus and rosmarinic acid in an athymic nude mice model of breast cancer. J Angiother. 2022;6(2):1-8.DOI: 10.25163/angiotherapy.624302.

Rajput A, Martin IDS, Rose R, Beko A, Levea C, Sharratt E, et al. Characterization of HCT116 human colon cancer cells in an orthotopic model. J Surg Res. 2008;147(2):276-281.DOI: 10.1016/j.jss.2007.04.021.

Al-Suede FSR, Ahamed MBK, Abdul Majid AS, Saghir SAM, Oon CE, Abdul Majid AMS. Immunomodulatory and antiangiogenic mechanisms of polymolecular botanical drug extract C5OSEW5050ESA OS derived from Orthosiphon stamineus. J Angiother.2021;5(1):1-9.DOI: 10.25163/angiotherapy.51211411913130321.

Teicher BA. Anticancer drug development guide: preclinical screening, clinical trials, and approval. Springer Science & Business Media; 2013. pp. 75.

Gan Y, Ai G, Wu J, Luo H, Chen L, Huang Q, et al. Patchouli oil ameliorates 5-fluorouracil-induced intestinal mucositis in rats via protecting intestinal barrier and regulating water transport. J Ethnopharmacol. 2020;250:112519,1-12.DOI: 10.1016/J.JEP.2019.112519.

Park JH, Kim JK. Pristimerin, a naturally occurring triterpenoid, attenuates tumorigenesis in experimental colitis-associated colon cancer. Phytomedicine. 2018;42:164-171.DOI: 10.1016/J.PHYMED.2018.03.033.

Dzubak P, Hajduch M, Vydra D, Hustova A, Kvasnica M, Biedermann D, et al. Pharmacological activities of natural triterpenoids and their therapeutic implications. Nat Prod Rep. 2006;23(3):394-411.DOI: 10.1039/b515312n.

Zorofchian Moghadamtousi S, Karimian H, Rouhollahi E, Paydar M, Fadaeinasab M, Abdul Kadir H. Annona muricata leaves induce G1 cell cycle arrest and apoptosis through mitochondria-mediated pathway in human HCT-116 and HT-29 colon cancer cells. J Ethnopharmacol. 2014;156:277-289.DOI: 10.1016/J.JEP.2014.08.011.

Lee K, Zhang H, Qian DZ, Rey S, Liu JO, Semenza GL. Acriflavine inhibits HIF-1 dimerization, tumor growth, and vascularization. Proc Natl Acad Sci. 2009;106(42):17910-17915.DOI: 10.1073/pnas.0909353106.

Ruan W, Wang Y, Ma Y, Xing X, Lin J, Cui J, et al. HSP60, a protein downregulated by IGFBP7 in colorectal carcinoma. J Exp Clin Cancer Res. 2010;29:41,1-9.DOI: 10.1186/1756-9966-29-41.

Takayama S, Reed JC, Homma S. Heat-shock proteins as regulators of apoptosis. Oncogene 2003 22:56. 2003;22(56):9041-9047.DOI: 10.1038/sj.onc.1207114.

Alimardan Z, Abbasi M, Khodarahmi GA, Kashfi K, Hasanzadeh F, Aghaei M. Identification of new small molecules as dual FoxM1 and Hsp70 inhibitors using computational methods. Res Pharm Sci. 2022;17(6):635-656.DOI: 10.4103/1735-5362.359431.

Sinicrope FA, Penington RC, Tang XM. Tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis is inhibited by Bcl-2 but restored by the small molecule Bcl-2 inhibitor, HA 14-1, in human colon cancer cells. Clin. Cancer Res. 2004;10(24):8284-8294. DOI: 10.1158/1078-0432.CCR-04-1289.

Wang Z, Banerjee S, Li Y, Rahman KMW, Zhang Y, Sarkar FH. Down-regulation of Notch-1 inhibits invasion by inactivation of nuclear factor-κB, vascular endothelial growth factor, and matrix metalloproteinase-9 in pancreatic cancer cells. Cancer Res. 2006;66(5): 2778-2784.DOI: 10.1158/0008-5472.CAN-05-4281.

Wei EK, Ma J, Pollak MN, Rifai N, Fuchs CS, Hankinso SE, et al. A prospective study of C-peptide, insulin-like growth factor-I, insulin-like growth factor binding protein-1, and the risk of colorectal cancer in women. Cancer Epidemiol Biomark Prev. 2005;14(4):850-855.DOI: 10.1158/1055-9965.EPI-04-0661.

Bach LA. Recent insights into the actions of IGFBP-6. J Cell Commun Signal. 2015;9(2):189-200. DOI: 10.1007/s12079-015-0288-4.

Tang Z, Gillatt D, Rowe E, Koupparis A, Holly JMP, Perks CM. IGFBP-2 acts as a tumour suppressor and plays a role in determining chemosensitivity in bladder cancer cells. Oncotarget. 2019;10(66): 7043-7057.DOI: 10.18632/oncotarget.27355.

Murillo G, Salti GI, Kosmeder JW, Pezzuto JM, Mehta RG. Deguelin inhibits the growth of colon cancer cells through the induction of apoptosis and cell cycle arrest. Eur J Cancer. 2002;38(18): 2446-2454.DOI: 10.1016/s0959-8049(02)00192-2.

Fre S, Pallavi SK, Huyghe M, Laé M, Janssen KP, Robine S, et al. Notch and Wnt signals cooperatively control cell proliferation and tumorigenesis in the intestine. Proc Natl Acad Sci. 2009;106(15): 6309-6314. DOI: 10.1073/pnas.0900427106.

Sikandar SS, Pate KT, Anderson S, Dizon D, Edwards RA, Waterman ML, et al. NOTCH signaling is required for formation and self-renewal of tumor-initiating cells and for repression of secretory cell differentiation in colon cancer. Cancer Res. 2010;70(4):1469-1478.DOI: 10.1158/0008-5472.CAN-09-2557.

Novellasdemunt L, Antas P, Li VS. Targeting Wnt signaling in colorectal cancer. A review in the theme: cell signaling: proteins, pathways and mechanisms. Am J Physiol Cell Physiol. 2015;309(8): C511-C521.DOI: 10.1152/ajpcell.00117.2015.

Schatoff EM, Leach BI, Dow LE. Wnt signaling and colorectal cancer. Curr Colorectal Cancer Rep. 2017;13(2):101-110.DOI: 10.1007/s11888-017-0354-9.

Roy SK, Srivastava RK, Shankar S. Inhibition of PI3K/AKT and MAPK/ERK pathways causes activation of FOXO transcription factor, leading to cell cycle arrest and apoptosis in pancreatic cancer. J Mol Signal. 2010;5:10,1-13. DOI: 10.1186/1750-2187-5-10.

Asif M, Yehya AHS, Sabbar Dahham SS, Mohamed SK, Shafaei A, Ezzat MO, et al. Establishment of in vitro and in vivo anti-colon cancer efficacy of essential oils containing oleo-gum resin extract of Mesua ferrea. Biomed Pharmacother. 2019;109:1620-1629.DOI: 10.1016/j.biopha.2018.10.127.

Ishitsuka H. Capecitabine: preclinical pharmacology studies. Invest New Drugs. 2000;18(4): 343-354.DOI: 10.1023/a:1006497231579.

Westman J, Grinstein S, Marques PE. Phagocytosis of necrotic debris at sites of injury and inflammation. Front Immunol. 2020;10:3030,1-18.DOI: 10.3389/fimmu.2019.03030.


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