Suppression of Melanoma Growth in a Murine Tumour Model Using Orthosiphon
stamineus Benth. Extract Loaded in Ethanolic Phospholipid Vesicles (Spherosome)

V.   Mansoureh   Nazari; Syed      Mahmood; Amin   Malik   Shah; Fouad Saleh   Resq   Al-Suede

Abstract

Background: Orthosiphon stamineus Benth (O.S) is a traditional south-east Asian herb. The extract of O.S is used in the formulation of ethanolic nanolipid vesicle system to have considerable potential for tumour therapeutics.

Methods: The research objective is to develop and characterise the anticancer and antiangiogenic effect of O.S extract in the form of nano-ethanolic spherosomes (ESP) using phospholipids in melanoma. Spherosomes formulation of O.S was developed using the thin-film re-hydration method and converted to gel using Acrypol 1%. The formulations were subjected to optimisation and physical-chemical characterisations like particle size, surface charge, DSC, FTIR, and TEM. Cytotoxicity of O.S and ESP was studied using an endothelial cell line (EA. hy926). Furthermore, anti-melanoma effect of O.S spherosome gel was studied in albino mice after topical administration.

Results: ESP-6 with the ratio of extract (O.S): cholesterol: phospholipid (1: 6: 0.5) showed the highest entrapment efficiency (80.56 ± 0.84%) using ultraviolet spectroscopy. In-vivo permeation/penetration studies revealed deeper absorption of ESP-6 compared to a hydroethanolic gel of O.S. In-vitro and in vivo anti-melanoma studies demonstrated the significant tumour-suppressing effect of ESP-6 on murine melanoma. Percentage inhibition of tumour growth by O.S and ESP-6 at 3000 mg/kg showed to be 63.98 ± 7.86% and 87.76 ± 7.90%, respectively.

Conclusion: Spherosome vesicles were developed with a smooth surface. The results demonstrated that O.S extract showed no toxicity when tested on the endothelial cell line. O.S loaded in spherosomes has the potential to lower the growth of melanoma in mice. The spherosomes loaded with O.S do not promote tumour growth or act as antiangiogenetic in melanoma.

Keywords: Orthosiphon stamineus, spherosomes, angiogenesis, melanoma, anti-tumour, spectroscopy.

Graphical Abstract

[1] 
Kim, S.; Paik, J. In Contradictory between morphology and phylogenetic trees of Orthosiphon spp. (Lamiaceae) from indonesia IOP conference Series. Earth Environ. Sci.,  2020, 457, m 012030.
[2] 
Xu, W.H.; Wang, H.T.; Sun, Y.; Xue, Z.C.; Liang, M.L.; Su, W.K. Antihyperuricemic and nephroprotective effects of extracts from Orthosi-phon stamineus in hyperuricemic mice. J. Pharm. Pharmacol.,  2020, 72(4), 551-560.
[http://dx.doi.org/10.1111/jphp.13222] [PMID:  31910301] 
[3] 
Gimbun, J.; Pang, S.F.; Yusoff, M.M. Orthosiphon stamineus (javatea).Nonvitamin and Nonmineral Nutritional Supplements; Elsevier, 2019, p. 327-333.
[4] 
AHMED. O. H.; KHAMEES, A. H.; NOMAN, S. K. A review on antiangiogenic activity medicinal plants available in Iraq. Int. J. Pharmac. Res.,  2020, 12(4)
[5] 
Nazari, M.; Nazari, M.; Arabani, S. Anti-inflammation prediction of orthosiphon stamineus extract against covid19 (in silico. study). Int. J. Eng. Sci.,  2021, 8(1), 14-18.
[http://dx.doi.org/10.15282/ijets.8.1.2021.1002] 
[6] 
Mohammed, H.A.; Khan, R.A.; Abdel-Hafez, A.A.; Abdel-Aziz, M.; Ahmed, E.; Enany, S.; Mahgoub, S.; Al-Rugaie, O.; Alsharidah, M.; Aly, M.S.A.; Mehany, A.B.M.; Hegazy, M.M. Phytochemical profiling, in vitro and in silico anti-microbial and anti-cancer activity evaluations and Staph GyraseB and h-TOP-IIβ receptor-docking studies of major constituents of Zygophyllum coccineum L. Aqueous-ethanolic extract and its subsequent fractions: An approach to validate traditional phytomedicinal knowledge. Molecules,  2021, 26(3), 577.
[http://dx.doi.org/10.3390/molecules26030577] [PMID:  33499325] 
[7] 
Mohamed, E.A.H.; Lim, C.P.; Ebrika, O.S.; Asmawi, M.Z.; Sadikun, A.; Yam, M.F. Toxicity evaluation of a standardised 50% ethanol extract of Orthosiphon stamineus. J. Ethnopharmacol.,  2011, 133(2), 358-363.
[http://dx.doi.org/10.1016/j.jep.2010.10.008] [PMID:  20937371] 
[8] 
Fallah, A.; Sadeghinia, A.; Kahroba, H.; Samadi, A.; Heidari, H.R.; Bradaran, B.; Zeinali, S.; Molavi, O. Therapeutic targeting of angiogenesis molecular pathways in angiogenesis-dependent diseases. Biomed. Pharmacother.,  2019, 110, 775-785.
[http://dx.doi.org/10.1016/j.biopha.2018.12.022] [PMID:  30554116] 
[9] 
Maya-Cano, D.A.; Arango-Varela, S.; Santa-Gonzalez, G.A. Phenolic compounds of blueberries (Vaccinium spp) as a protective strategy against skin cell damage induced by ROS: A review of antioxidant potential and antiproliferative capacity. Heliyon,  2021, 7(2), e06297.
[http://dx.doi.org/10.1016/j.heliyon.2021.e06297] [PMID:  33665449] 
[10] 
Al-Dulaimi, D.W.; Shah Abdul Majid, A.; Baharetha, M.H.; Ahamed, M.B.K.; Faisal, S.F.; Al Zarzour, R.H.; Ein Oon, C.; Abdul Majid, A.M.S.; Ahmed Hassan, L.E. Anticlastogenic, antimutagenic, and cytoprotective properties of Orthosiphon stamineus ethanolic leaves extract. Drug Chem. Toxicol.,  2022, 45(2), 641-650.
[11] 
Bennardo, L.; Bennardo, F.; Giudice, A.; Passante, M.; Dastoli, S.; Morrone, P.; Provenzano, E.; Patruno, C.; Nisticò, S.P. Local chemothera-py as an adjuvant treatment in unresectable squamous cell carcinoma: What do we know so far? Curr. Oncol.,  2021, 28(4), 2317-2325.
[http://dx.doi.org/10.3390/curroncol28040213] [PMID:  34201867] 
[12] 
Opatha, S.A.T.; Titapiwatanakun, V.; Chutoprapat, R. Transfersomes: A promising nanoencapsulation technique for transdermal drug deliv-ery. Pharmaceutics,  2020, 12(9), 855.
[http://dx.doi.org/10.3390/pharmaceutics12090855] [PMID:  32916782] 
[13] 
Sguizzato, M.; Ferrara, F.; Hallan, S.S.; Baldisserotto, A.; Drechsler, M.; Malatesta, M.; Costanzo, M.; Cortesi, R.; Puglia, C.; Valacchi, G.; Esposito, E. Ethosomes and transethosomes for mangiferin transdermal delivery. Antioxidants,  2021, 10(5), 768.
[http://dx.doi.org/10.3390/antiox10050768] [PMID:  34066018] 
[14] 
Anita, C.; Munira, M.; Mural, Q.; Shaily, L. Topical nanocarriers for management of Rheumatoid Arthritis: A review. Biomed. Pharmacother.,  2021, 141, 111880.
[http://dx.doi.org/10.1016/j.biopha.2021.111880] [PMID:  34328101] 
[15] 
Sudhakar, K.; Fuloria, S.; Subramaniyan, V.; Sathasivam, K.V.; Azad, A.K.; Swain, S.S.; Sekar, M.; Karupiah, S.; Porwal, O.; Sahoo, A.; Meenakshi, D.U.; Sharma, V.K.; Jain, S.; Charyulu, R.N.; Fuloria, N.K. Ultraflexible liposome nanocargo as a dermal and transdermal drug delivery system. Nanomaterials (Basel),  2021, 11(10), 2557.
[http://dx.doi.org/10.3390/nano11102557] [PMID:  34685005] 
[16] 
Majeed, I.; Raza, S.A.; Akhtar, N.; Siddiqui, F.A.; Iqbal, B. Formulation and in-vitro characterization of Capsaicin loaded ethosomes. Pak. J. Pharm. Sci.,  2019, 32(6), 2849-2857.
[PMID: 32024624] 
[17] 
Abd El-Alim, S.H.; Kassem, A.A.; Basha, M.; Salama, A. Comparative study of liposomes, ethosomes and transfersomes as carriers for enhancing the transdermal delivery of diflunisal: In vitro and in vivo evaluation. Int. J. Pharm.,  2019, 563, 293-303.
[http://dx.doi.org/10.1016/j.ijpharm.2019.04.001] [PMID:  30951860] 
[18] 
de Macedo, L.M.; Santos, É.M.D.; Militão, L.; Tundisi, L.L.; Ataide, J.A.; Souto, E.B.; Mazzola, P.G. Rosemary (Rosmarinus officinalis L., syn Salvia rosmarinus Spenn.) and its topical applications: A review. Plants,  2020, 9(5), 651.
[http://dx.doi.org/10.3390/plants9050651] [PMID:  32455585] 
[19] 
Kaushik, N.; Oh, H.; Lim, Y.; Kumar Kaushik, N.; Nguyen, L.N.; Choi, E.H.; Kim, J.H. Screening of Hibiscus and Cinnamomum plants and identification of major phytometabolites in potential plant extracts responsible for apoptosis induction in skin melanoma and lung adenocar-cinoma cells. Front. Bioeng. Biotechnol.,  2021, 9, 779393.
[http://dx.doi.org/10.3389/fbioe.2021.779393] [PMID:  34957073] 
[20] 
Furtado, R.A.; Ozelin, S.D.; Ferreira, N.H.; Miura, B.A.; Almeida, Junior S.; Magalhães, G.M.; Nassar, E.J.; Miranda, M.A.; Bastos, J.K.; Tavares, D.C. Antitumor activity of solamargine in mouse melanoma model: Relevance to clinical safety. J. Toxicol. Environ. Health A,  2022, 85(4), 131-142.
[http://dx.doi.org/10.1080/15287394.2021.1984348] [PMID:  34612163] 
[21] 
Safety assessment of P2Et extract in healthy voluntary subjects in colombia. 2019.
[22] 
Kirsch, A. Successful treatment of metastatic malignant melanoma with Viscum album extract (Iscador M). J. Altern. Complement. Med.,  2007, 13(4), 443-445.
[http://dx.doi.org/10.1089/acm.2007.6175] [PMID:  17532738] 
[23] 
Cho, H.; Shen, Q.; Zhang, L. H.; Okumura, M.; Kawakami, A.; Ambrose, J.; Sigoillot, F.; Miller, H. R.; Gleim, S.; Cobos-Correa, A. CYP27A1-dependent anti-melanoma activity of limonoid natural products targets mitochondrial metabolism. Cell Chem. Biol.,  2021, 28(10), 1407-1419. e6.
[http://dx.doi.org/10.1016/j.chembiol.2021.03.004] 
[24] 
Bouhlel Chatti, I.; Ben Toumia, I.; Krichen, Y.; Maatouk, M.; Chekir Ghedira, L.; Krifa, M. Assessment of rhamnus alaternus leaves extract: Phytochemical characterization and antimelanoma activity. J. Med. Food, 2021.
[http://dx.doi.org/10.1089/jmf.2020.0170] [PMID:  34152839] 
[25] 
Mirzavi, F.; Barati, M.; Soleimani, A.; Vakili-Ghartavol, R.; Jaafari, M.R.; Soukhtanloo, M. A review on liposome-based therapeutic ap-proaches against malignant melanoma. Int. J. Pharm.,  2021, 599, 120413.
[http://dx.doi.org/10.1016/j.ijpharm.2021.120413] [PMID:  33667562] 
[26] 
Barenholz, Y. Doxil®-the first FDA-approved nano-drug: Lessons learned. J. Control. Release,  2012, 160(2), 117-134.
[http://dx.doi.org/10.1016/j.jconrel.2012.03.020] [PMID:  22484195] 
[27] 
Zhong, H.; Chan, G.; Hu, Y.; Hu, H.; Ouyang, D. A comprehensive map of FDA-approved pharmaceutical products. Pharmaceutics,  2018, 10(4), 263.
[http://dx.doi.org/10.3390/pharmaceutics10040263] [PMID:  30563197] 
[28] 
Paliwal, R.; Kumar, P.; Chaurasiya, A.; Kenwat, R.; Katke, S.; Paliwal, S.R. Development of nanomedicines and nano-similars: Recent ad-vances in regulatory landscape. Curr. Pharm. Des.,  2022, 28(2), 165-177.
[http://dx.doi.org/10.2174/1381612827666211115170001] [PMID:  34781869] 
[29] 
Ventola, C.L. Progress in nanomedicine: Approved and investigational nanodrugs. P&T,  2017, 42(12), 742-755.
[PMID: 29234213] 
[30] 
Weng, J.; Liu, Y.; Ding, J. XU, Y. Study on the Preparation and Preliminary Stability of Three Liposomes with Different Solvent Medium. China Pharmacist,  2018, 824-827.
[31] 
Mahmood, S.; Kiong, K.C.; Tham, C.S.; Chien, T.C.; Hilles, A.R.; Venugopal, J.R. PEGylated lipid polymeric nanoparticle-encapsulated acy-clovir for in vitro controlled release and ex vivo gut sac permeation. AAPS PharmSciTech,  2020, 21(7), 285.
[http://dx.doi.org/10.1208/s12249-020-01810-0] [PMID:  33057878] 
[32] 
He, Y.; Luo, L.; Liang, S.; Long, M.; Xu, H. Influence of probe-sonication process on drug entrapment efficiency of liposomes loaded with a hydrophobic drug. Intern. J. Polym. Mat. Polym. Biomat.,  2019, 68(4), 193-197.
[http://dx.doi.org/10.1080/00914037.2018.1434651] 
[33] 
Yehya, A.H.; Asif, M.; Majid, A.M.A.; Oon, C.E. Complementary effects of Orthosiphon stamineus standardized ethanolic extract and ros-marinic acid in combination with gemcitabine on pancreatic cancer. Biomed. J.,  2021, 44(6), 694-708.
[PMID: 35166208] 
[34] 
Zhang, W.; Li, X.; Hua, Y.; Li, Z.; Chen, B.; Liu, A.; Lu, W.; Zhao, X.; Diao, Y.; Chen, D. Antioxidant product analysis of Hulu Tea (Tadeha-gi triquetrum). New J. Chem.,  2021, 45(43), 20257-20265.
[http://dx.doi.org/10.1039/D1NJ02639A] 
[35] 
Abel, S.D.A.; Baird, S.K. Honey is cytotoxic towards prostate cancer cells but interacts with the MTT reagent: Considerations for the choice of cell via bility assay. Food Chem.,  2018, 241, 70-78.
[http://dx.doi.org/10.1016/j.foodchem.2017.08.083] [PMID:  28958561] 
[36] 
Mahmood, S.; Chatterjee, B.; Mandal, U.K. Pharmacokinetic evaluation of the synergistic effect of raloxifene loaded transfersomes for trans-dermal delivery. J. Drug Deliv. Sci. Technol.,  2021, 63, 102545.
[http://dx.doi.org/10.1016/j.jddst.2021.102545] 
[37] 
Liu, Y.; Lunter, D.J. Systematic investigation of the effect of non-ionic emulsifiers on skin by confocal raman spectroscopy-A comprehen-sive lipid analysis. Pharmaceutics,  2020, 12(3), 223.
[http://dx.doi.org/10.3390/pharmaceutics12030223] [PMID:  32131544] 
[38] 
Natsheh, H.; Vettorato, E.; Touitou, E. Ethosomes for dermal administration of natural active molecules. Curr. Pharm. Des.,  2019, 25(21), 2338-2348.
[http://dx.doi.org/10.2174/1381612825666190716095826] [PMID:  31333087] 
[39] 
Uscanga-Palomeque, A.C.; Zapata-Benavides, P.; Saavedra-Alonso, S.; Zamora-Ávila, D.E.; Franco-Molina, M.A.; Arellano-Rodríguez, M.; Manilla-Muñoz, E.; Martínez-Torres, A.C.; Trejo-Ávila, L.M.; Rodríguez-Padilla, C. Inhibitory effect of Cuphea aequipetala extracts on mu-rine B16F10 melanoma in vitro and in vivo. BioMed research int.,  2019, 2019, 8560527.
[http://dx.doi.org/10.1155/2019/8560527] 
[40] 
Muhammad, H.; Omar, M. H.; Isa, M. L.; Thani, N.; Rasid, E. N. I.; Awang, N. Male reproductive toxicity studies of Orthosiphon stamineus aqueous extract in Sprague Dawley rats. J. Med. Plants,  2018, 6(5), 07-14.
[41] 
Arora, D.; Nanda, S. Quality by design driven development of resveratrol loaded ethosomal hydrogel for improved dermatological benefits via enhanced skin permeation and retention. Int. J. Pharm.,  2019, 567, 118448.
[http://dx.doi.org/10.1016/j.ijpharm.2019.118448] [PMID:  31226472] 
[42] 
Tripathi, P.K.; Gorain, B.; Choudhury, H.; Srivastava, A.; Kesharwani, P. Dendrimer entrapped microsponge gel of dithranol for effective topical treatment. Heliyon,  2019, 5(3), e01343.
[http://dx.doi.org/10.1016/j.heliyon.2019.e01343] [PMID:  30957038] 
[43] 
Mahmood, S.; Mandal, U.K.; Chatterjee, B. Transdermal delivery of raloxifene HCl via ethosomal system: Formulation, advanced characteri-zations and pharmacokinetic evaluation. Int. J. Pharm.,  2018, 542(1-2), 36-46.
[http://dx.doi.org/10.1016/j.ijpharm.2018.02.044] [PMID:  29501737] 
[44] 
Kamada, M.; Pierlot, C.; Molinier, V.; Aubry, J-M.; Aramaki, K. Rheological properties of wormlike micellar gels formed by novel bio-based isosorbide surfactants. Colloids Surf. A Physicochem. Eng. Asp.,  2018, 536, 82-87.
[http://dx.doi.org/10.1016/j.colsurfa.2017.07.037] 
[45] 
Yehya, A.H.S.; Asif, M.; Kaur, G.; Hassan, L.E.A.; Al-Suede, F.S.R.; Abdul Majid, A.M.S.; Oon, C.E. Toxicological studies of Orthosiphon stamineus (Misai Kucing) standardized ethanol extract in combination with gemcitabine in athymic nude mice model. J. Adv. Res.,  2018, 15, 59-68.
[http://dx.doi.org/10.1016/j.jare.2018.05.006] [PMID:  30581613] 
[46] 
Jiang, X.; Chen, Z.; Shen, G.; Jiang, Y.; Wu, L.; Li, X.; Wang, G.; Yin, T. Psychotropic agent thioridazine elicits potent in vitro and in vivo anti-melanoma effects. Biomed. Pharmacother.,  2018, 97, 833-837.
[http://dx.doi.org/10.1016/j.biopha.2017.11.012] [PMID:  29136758] 
[47] 
Nainwal, N.; Jawla, S.; Singh, R.; Saharan, V.A. Transdermal applications of ethosomes - a detailed review. J. Liposome Res.,  2019, 29(2), 103-113.
[http://dx.doi.org/10.1080/08982104.2018.1517160] [PMID:  30156120] 

Cite As

Current Drug Metabolism

Suppression of Melanoma Growth in a Murine Tumour Model Using Orthosiphon stamineus Benth. Extract Loaded in Ethanolic Phospholipid Vesicles (Spherosome)

Abstract

Graphical Abstract