Bioactivity Study of Russelia equisetiformis Ethanol Extract as an MCF-7 Breast Cancer Cells Inhibitor with In vitro and In silico Approaches

Rizq Rachmad Ramiizah, Dini Attala Hefa Insyira, Najwa Salsabila Hakim, Muhammad Rafly Aditya Firmansyah, Tukiran Tukiran

Abstract

Russelia equisetiformis has been used in traditional medicine, but its molecular mechanisms against breast cancer remain unexplored. This study aimed to investigate the cytotoxic potential of R. equisetiformis ethanol extract and elucidate its molecular interactions through a combined in vitro and in silico approach. Cytotoxicity evaluation using the Resazurin assay demonstrated the extract’s activity against MCF-7 breast cancer cells with an IC50 value of 352.60 ± 0.23 ????g/mL. To understand the underlying mechanism, the identified bioactive compounds that passed the druglikeness test were analyzed using molecular docking against the Progesterone Receptor (PDB ID: 2W8Y) and Estrogen Receptor-alpha (PDB ID: 3ERT). Docking analysis revealed that these compounds form binding interactions with target receptors, providing insight into their potential mechanism of action, albeit with lower affinity compared to standard drugs (tamoxifen and anastrozole). Importantly, ADMET prediction using ProTox 3.0 highlighted apigenin, kaempferol, isokaempferide, and kaempferol 3-O-rhamnoside as the most promising primary candidates, demonstrating inactivity against hepatotoxicity, carcinogenicity, and cytotoxicity. These findings provide initial scientific evidence for the potential anticancer properties of R. equisetiformis compounds, indicating that all four compounds are safe and viable bioactive candidates for further development as non-toxic therapeutic agents.

References

Ahmed, E., S. Desoukey, M. Fouad, and M. Kamel (2016). A Pharmacognostical Study of Russelia equisetiformis Sch. & Cham. International Journal of Pharmacognosy and Phytochemical Research, 8; 174–192

Aldahasi, R. M., S. S. Alghamdi, N. Albulushi, H. Alhuraibi, R. S. Suliman, A. Alsubait, A. S. Alahmari, and A. E. Mohammed (2025). Metabolite Detection via QTOF-LCMS and Computational Analysis on Russelia equisetiformis Exploring Green Nanotechnology and Antifungal Against Phytopathogens. Chemical and Biological Technologies in Agriculture, 12(1); 142

Alsaraf, K. M., M. H. Mohammad, A. M. Al-Shammari, and I. S. Abbas (2019). Selective Cytotoxic Effect of Plantago lanceolata L. Against Breast Cancer Cells. Journal of the Egyptian National Cancer Institute, 31(1); 10

Amrulloh, L., N. Harmastuti, A. Prasetiyo, and R. Herowati (2023). Analysis of Molecular Docking and Dynamics Simulation of Mahogany (Swietenia macrophylla King) Compounds Against the PLpro Enzyme SARS-CoV-2. Jurnal Farmasi dan Ilmu Kefarmasian Indonesia, 10(3); 347–359

Arwansyah, A., L. Ambarsari, and T. I. Sumaryada (2014). Simulasi Docking Senyawa Kurkumin dan Analognya sebagai Inhibitor Reseptor Androgen pada Kanker Prostat. Current Biochemistry, 1(1); 11–19

Awe, E. O., J. M. Makinde, O. A. Olajide, and O. K.Wakeel (2004). Evaluation of the Anti-Inflammatory and Analgesic Properties of the Extract of Russelia equisetiformis (Schlecht & Cham) Scrophulariaceae. Inflammopharmacology, 12(4); 399–405

Bajalovic, N., Y. Z. Or, A. R. E.Woo, S. H. Lee, and V. C. L. Lin (2022). High Levels of Progesterone Receptor B in MCF-7 Cells Enable Radical Anti-Tumoral and Anti-Estrogenic Effects of Progestin. Biomedicines, 10(8); 1860

Banerjee, P., O. Ulker, I. Ozkan, and O. C. Ulker (2025). The Investigation of the Toxicity of Organophosphorus Flame Retardants (OPFRs) by Using the In Silico Toxicity Prediction Platform ProTox-3.0. Toxicology Mechanisms and Methods, 35(1); 32–42

Barthwal, R. and R. Mahar (2024). Exploring the Significance, Extraction, and Characterization of Plant-Derived Secondary Metabolites in Complex Mixtures. Metabolites, 14(2); 119

Barzegarparay, F., H. Najafzadehvarzi, R. Pourbagher, H. Parsian, S. M. Ghoreishi, and S. Mortazavi-Derazkola (2024). Green Synthesis of Novel Selenium Nanoparticles Using Crataegus Monogyna Extract (SeNPs@CM) and Investigation of Its Toxicity, Antioxidant Capacity, and Anticancer Activity Against MCF-7 as a Breast Cancer Cell Line. Biomass Conversion and Biorefinery, 14(20); 25369–25378

Basu, A. and S. Krishnamurthy (2010). Cellular Responses to Cisplatin-Induced DNA Damage. Journal of Nucleic Acids, 2010(1); 201367

Bray, F., M. Laversanne, H. Sung, J. Ferlay, R. L. Siegel, I. Soerjomataram, and A. Jemal (2024). Global Cancer Statistics 2022: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA: A Cancer Journal for Clinicians, 74(3); 229–263

Columba-Palomares, M. C., R. M. Montiel-Ruiz, L. C. Sánchez, D. Palafox-Gante, and V. Rodríguez-López (2021). Anti-Inflammatory, Antioxidant, and Antinociceptive Activities of Russelia equisetiformis (L.) Wettst. Avicenna Journal of Phytomedicine, 11(2); 101–108

Costa-Lotufo, L. V., P. C. Jimenez, D. V.Wilke, L. K. A. M. Leal, G. M. A. Cunha, E. R. Silveira, K. M. Canuto, G. S. B. Viana, M. E. A. Moraes, and M. O. d. Moraes (2003). Antiproliferative Effects of Several Compounds Isolated from Amburana cearensis AC Smith. Zeitschrift für Naturforschung C, 58(9–10); 675–680

Damayanti, S., K. Khonsa, and T. Amelia (2021). Antiviral Activity and Toxicity Prediction of Compounds Contained in Figs (Ficus carica L.) by in Silico Method. Indonesian Journal of Pharmaceutical Science and Technology, 8(1); 21–33

Diantini, A., A. Subarnas, K. Lestari, E. L. I. Halimah, Y. Susilawati, S. Supriyatna, E. Julaeha, T. H. Achmad, E. W. Suradji, and C. Yamazaki (2012). Kaempferol-3-O-Rhamnoside Isolated from the Leaves of Schima Wallichii Korth. Inhibits MCF-7 Breast Cancer Cell Proliferation Through Activation of the Caspase Cascade Pathway. Oncology Letters, 3(5); 1069–1072

Elango, A., I. Kannan, R. Ravichandar, and P. Kumaravelu (2024). Molecular Docking Analysis of Imeglimin and Its DerivativesWith Estrogen Receptor-Alpha. Bioinformation, 20(7); 711

Elya, B., R. Rosmalena, A. M. Fajrin, A. Tedjo, N. A. Ramadanti, N. N. Azizah, and N. B. M. Hashim (2025). Ethanol Extract from Ziziphus nummularia Stem Inhibits MCF-7 Breast Cancer Cell Proliferation Through TP53 Regulating Kinase (TP53RK)-Mediated p53 Activation: In Silico and Gene Expression Investigations. Narra J, 5(1); e1382

Faria, S. S., D. Giannarelli, V. C. Cordeiro de Lima, S. L. Anwar, C. Casadei, U. De Giorgi, G. Madonna, P. A. Ascierto, R. V. Mendoza López, and R. Chammas (2024). Development of a Prognostic Model for Early Breast Cancer Integrating Neutrophil to Lymphocyte Ratio and Clinical-Pathological Characteristics. The Oncologist, 29(4); e447–e454

Fiorelia, N. E., A. D.Wibowo, N. L. Lae, and O. Krisbianto (2022). Types of High-Performance Liquid Chromatography (HPLC) Columns: A Review. Food Tech, 5(1); 1–16

Gómez-Estrada, H., F. Díaz-Castillo, L. Franco-Ospina, J. Mercado-Camargo, J. Guzmán-Ledezma, J. D. Medina, and R. Gaitán-Ibarra (2011). FolkMedicine in the Northern Coast of Colombia: An Overview. Journal of Ethnobiology and Ethnomedicine, 7; 1–11

Hammami, A., A. H. Saputro, W. N. Auli, S. R. Hasanah, N. Fadila, A. P. Sunugroho, D. Y. Aristania, and P. Amanda (2025). Studi Penambatan Molekul Senyawa Mangiferin, Apigenin, Kukurbitasin E, dan Kuersetin sebagai Agen Antikanker Payudara terhadap Protein Target Reseptor Estrogen Alfa (ER-????). Jurnal Sains dan Kesehatan, 6(3); 1–10 (in Indonesia)

Harris, G. H. (2002). Chapter 10 - The Use of CCC in the Pharmaceutical Industry. In A. B. Berthod, editor, Countercurrent Chromatography, volume 38. pages 301–330

Haryoto, H. (2019). Cytotoxic Activities of Ethanol Extract, Nonpolar, Semipolar, and Polar Fractions of Dioscorea esculenta L., on MCF7 Cancer Cells. Journal of Nutraceuticals and Herbal Medicine, 2(1); 12–19

Hurria, A. Alfian, and R. Mahmudah (2026). Phytochemical Properties, In Silico and In Vitro Analysis of the Antibacterial Activity of Cinnamomum sulavesianum Bark from Sulawesi. Science and Technology Indonesia, 11(1); 121–136

Hussein, U. A.-R., B. Abd, H. A. Mahamda, A. F. Al-Rubaye, U. S. Altimari, and A. A. Al-Alwany (2025). Molecular Docking and Toxicity Profiling of Quercetin and Silibinin: A Computational Approach Against Key Hepatitis C Virus Proteins. Journal of Experimental Zoology India, 28(2); 1311–1318

Jayaram, B., T. Singh, G. Mukherjee, A. Mathur, S. Shekhar, and V. Shekhar (2012). Sanjeevini: A Freely Accessible Web-Server for Target-Directed Lead Molecule Discovery. BMC Bioinformatics, 13(Suppl 17); S7

Khudzaifi, M., S. A. Kalsum, and A. Z.Nisak (2024). Molecular Docking Senyawa Flavonoid Daun Sirsak (Annona muricata L.) Terhadap Reseptor Estrogen Alfa (RE-????) Sebagai Kandidat Obat Antikanker Payudara. Sains Medisina, 3(1); 1–9 (in Indonesia)

Kolawole, O. T. and S. O. Kolawole (2010). Effects of Russelia equisetiformis Methanol and Aqueous Extracts on Hepatic Function Indices. Biol Med, 2(3); 38–41

Konieczna, A. and M. Rosińska (2025). Early Predictive Markers and Histopathological Response to Neoadjuvant Endocrine Therapy in Postmenopausal Patients with HR+/HER2- Early Breast Cancer. Cancers, 17(14); 2319

Kumi, J., A. Aning, J. Ampofo, S. Hamidu, and E. N. Y. Nyarko (2026). In Vitro Cytotoxicity Assessment of Leaves of Tieghemella heckelii on Breast (MDA-MB-468), Liver (HepG2), and Prostate (PC3) Cancer Cell Lines. BioMed Research International, 2026(1); 2002140

Kunc, M., M. Popęda,W. Biernat, and E. Senkus (2021). Lost but Not Least–Novel Insights into Progesterone Receptor Loss in Estrogen Receptor-Positive Breast Cancer. Cancers, 13(19); 4755

Kurniawidjaja, L. M., F. Lestari, M. Tejamaya, and D. H. Ramdhan (2021). Konsep Dasar Toksikologi Industri. FKM UI (in Indonesia)

Latief, S., F. I. Syahruddin, I. Royani, and R. P. Juhamran (2023). Hubungan Tingkat Pengetahuan dan Sikap terhadap Deteksi Dini Kanker Payudara pada Pegawai RS Ibnu Sina. Fakumi Medical Journal: Jurnal Mahasiswa Kedokteran, 3(9); 685–694 (in Indonesia)

Lele, M. N. E., N. I. Ahmad, and Y. Bare (2022). Molecular Interaction Analysis of Homodihydrocapsaicin as a COX-2 Inhibitor. Spizaetus: Jurnal Biologi dan Pendidikan Biologi, 3(2); 36–42

Lipinski, C. A., F. Lombardo, B.W. Dominy, and P. J. Feeney (2012). Experimental and Computational Approaches to Estimate Solubility and Permeability in Drug Discovery and Development Settings. Advanced Drug Delivery Reviews, 64; 4–17

Lu, H.-F., Y.-J. Chie, M.-S. Yang, C.-S. Lee, J.-J. Fu, J.-S. Yang, T.-W. Tan, S.-H.Wu, Y.-S. Ma, and S.-W. Ip (2010). Apigenin Induces Caspase-Dependent Apoptosis in Human Lung Cancer A549 Cells Through Bax- and Bcl-2-Triggered Mitochondrial Pathway. International Journal of Oncology, 36(6); 1477–1484

Mala, N., Y. F. Yun, and L. S. Aisyah (2024). Aktivitas Antioksidan dan Antikanker dari Ekstrak Etanol, n-Heksana, Etil Asetat, dan Air Etanol Kalanchoe marmorata (Crassulaceae). Jurnal Kartika Kimia, 7(2); 121–128 (in Indonesia)

Manasathien, J. and K. Indrapichate (2017). Apoptosis ofMCF-7 Cancer Cells Induced by Pomegranate (Punica granatum L.) Peel Extract. Suranaree Journal of Science & Technology, 24(1)

Nasyanka, A. L., D. Ratnasari, J. Na’imah, and S. N. Asiyah (2023). Studi In Silico: Toksisitas Senyawa Potensial sebagai Antikanker Turunan N-((4-Fluorofenil) Karbamatiol) Benzamida. Journal of Pharmaceutical Care Anwar Medika (J-PhAM), 5(2); 1–16 (in Indonesia)

Nugraha, G. I., F. Amalia, F. Imadudda’wah, E. F. Ariyanto, M. Ghozali, and S. N. Fatimah (2024). A Combination of Diethylpropion with Dietary Intervention Results in Body Weight and Fat Loss with Preserved Muscle Mass in Obese Patients. Medicine, 103(39); e39908

Nugroho, E. D., R. Ardiansyah, N. Kurniawan,Widodo, D. A. Rahayu, and A. M. Sururi (2023). An In Silico Study on the Chemical Compounds from Macrophiothrix longipedia as Antiviral Compounds Against COVID-19. AACL Bioflux, 16(4); 2380–2390

Nursanti, O., A. Aziz, and G. Hadisoebroto (2022). Prediksi Toksisitas dan Farmakokinetika untuk Mendapatkan Kandidat Obat Analgesik. (in Indonesia)

Ojurongbe, O., J. A. Ojo, D. I. Adefokun, O. O. Abiodun, G. Odewale, and E. O. Awe (2015). In Vivo Antimalarial Activities of Russelia equisetiformis in Plasmodium berghei-Infected Mice. Indian Journal of Pharmaceutical Sciences, 77(4); 504

Oladeinde, F.O. (2005). Phytochemical Screening of Cnidoscolus aconitifolius and Russelia equisetiformis Used for the Treatment of Diabetes and Leukemia in Southwestern Nigeria. In African Healing Wisdom: From Tradition to Current Application and Research, International Conference.Washington DC, USA, pages 6–9

Patel, A., M. Bhatt, A. Soni, and P. Sharma (2021). Identification of Steroidal Saponins from Tribulus terrestris and Their In Silico Docking Studies. Journal of Cellular Biochemistry, 122(11); 1665–1685

Pebriani, S. A., W. S. Bhagawan, M. E. Prastiyanto, R. M. Rukmana, and R. Pratiwi (2026). Anticancer Efficacy of a Combination of Curcuma longa L. Rhizome and Annona muricata L. Leaf Extracts Against T47D Cells. Science and Technology Indonesia, 11(2)

Pinheiro, A. A., C. Y.-A. Lee, A. G. P. d. Santos, É. R. Pereira, M. S. d. Rosário, D. L. Ribeiro, J. M. Serpeloni, and C. Q. d. Rocha (2024). Chemical Characterization and Evaluation of the Antitumoral Activity of Annona tomentosa RE Fr. on Breast (MCF-7) Three-Dimensional (3D) Tumor Spheroids. Journal of the Brazilian Chemical Society, 35(8); e–20240037

Puranik, N. V., P. Srivastava, G. Bhatt, D. J. S. John Mary, A. M. Limaye, and J. Sivaraman (2019). Determination and Analysis of Agonist and Antagonist Potential of Naturally Occurring Flavonoids for Estrogen Receptor (ER????) by Various Parameters and Molecular Modelling Approach. Scientific Reports, 9(1); 7450

Puspitasari, Y. E., M. A. Alfikri, R. Sitanggang, J. E. Tambunan, and H. Hardoko (2023). In Silico Analysis of Phenolic Compounds from Ceriops decandra Griff. Leaves and Molecular Interaction as Anti-Diabetes. Science and Technology Indonesia, 8(4); 542–553

Qamar, S. U. R. (2021). Nanocomposites: Potential Therapeutic Agents for the Diagnosis and Treatment of Infectious Diseases and Cancer. Colloid and Interface Science Communications, 43; 100463

Rambitan, S. R., A. Manampiring, B. J. Kepel, F. Budiarso, and W. Bodhi (2021). Molecular Docking Senyawa Vitexin, Ursolic Acid, dan Flavonol dalam Tumbuhan Binahong (Andromeda cordifolia (Ten.) Steenis) yang Berpotensi sebagai Penghambat Pertumbuhan COVID-19. EBiomedik, 9(2); 201–207 (in Indonesia)

Ramiizah, R. R., A. M. Sururi, R. A. Sutriyansyah, D. A. Ramadhani, A. H. Tsana, H. Kristyandari, Tukiran, D. A. Rahayu, and N. B. Mamat (2025). LC-MS Identification of Bioactive Compounds from the Methanol Extract of Veitchia merrillii Seeds as Potential Antiviral Agents Targeting HIV-1 Protease, Integrase, and Reverse Transcriptase: An In Silico Study. Tropical Journal of Natural Product Research, 9(11); 5360–5370

Riaz, M., N. Rasool, I. H. Bukhari, M. Shahid, A. F. Zahoor, M. A. Gilani, and M. Zubair (2012). Antioxidant, Antimicrobial, and Cytotoxicity Studies of Russelia equisetiformis. African Journal of Microbiology Research, 6(27); 5700–5707

Riaz, M., N. Rasool, M. Iqbal, A. Tawab, F. E-Habib, A. Khan, and M. Farman (2017). Liquid Chromatography-Electrospray Ionization-Tandem Mass Spectrometry (LCESI-MS/MS) Analysis of Russelia equisetiformis Extract. Bulgarian Chemical Communications, 49; 354–359

Rock, K. D., H. M. Starnes, and S. M. Belcher (2024). Reproductive System, Female. In Encyclopedia of Toxicology (Fourth Edition), volume 8. pages 167–202

Romero-Cerecero, O., H. Reyes-Morales, L. Aguilar-Santamaría, M. Huerta-Reyes, and J. Tortoriello-Garcia (2009). Use of Medicinal Plants Among Patients with Diabetes Mellitus Type 2 in Morelos, Mexico. Boletín Latinoamericano y del Caribe de Plantas Medicinales y Aromáticas, 8(5); 380–388

Sabu, A., J. Selvaraj, R. Gayatridevi, and E. Dilipan (2023). In Vitro Anticancer Activity of Rauvolfia tetraphylla Extract on MCF-7 Breast Cancer Cell Lines. Bioinformation, 19(1); 43–47

Saibaba, S. V., M. S. Kumar, and P. S. Pandiyan (2016). Mini Review on LC/MS Techniques. World Journal of Pharmacy and Pharmaceutical Science, 5(4); 2381–2395

Sanjai, C., S. L. Gaonkar, and S. S. Hakkimane (2024). Harnessing Nature’s Toolbox: Naturally Derived Bioactive Compounds in Nanotechnology-Enhanced Formulations. ACS Omega, 9(43); 43302–43318

Setiyanto, A. E. R., M. D. Taufiqurrahman, T. Turhadi, and D. Siswanto (2025). LC-MS-Based Metabolite Profiling of Aqueous Garlic (Allium sativum L.) Bulb Extract. Berkala Penelitian Hayati, 31(2); 86–95

Sururi, A. M., D. K. Maharani, and F. A.Wati (2023). Potensi Senyawa Eugenol dari Cengkeh (Syzygium aromaticum) sebagai Inhibitor Protease HIV-1 (PR). Unesa Journal of Chemistry, 12(1); 26–30

Susanti, N., A. Mustika, J. Khotib, R. Muti, and M. Rochmanti (2023). Phytochemical, Metabolite Compound, and Antioxidant Activity of Clinacanthus nutans Leaf Extract from Indonesia. Science and Technology Indonesia, 8(1); 38–44

Tukiran, T. and M. Raihan (2025). Chemical Profiling Ethyl Acetate Extract of Basilicum Polystachyon Leaves and Exploration of Anticancer SIRT1 Inhibitors Using an In Silico Approach. Indonesian Journal of Chemistry, 25(1); 256–265

Widiastuti, N. L. G. K. (2019). Pendidikan Sains Terintegrasi Keterkaitan Konsep Ikatan Kimia dengan Berbagai Bidang Ilmu. Widya Accarya, 10(2); (in Indonesia)

Widiyana, A. P., T.Widiandani, and S. Siswodihardjo (2024). 5-O-Acetylpinostrobin Derivatives Inhibit Estrogen Alpha and Progesterone Receptors Through a Molecular Docking Approach. Pharmacy Education, 24(3); 244–250

Yang, Y., R. Zheng, P. Zhang, J.Wen, Y. Luo, Z. Zhao, L. You, and C.-T. Ho (2022). Combination Effects of Polyphenols Present in Sugarcane on Proliferation in MCF-7 Human Breast Cancer Cells. Sugar Tech, 24(3); 832–840

Yi, X., J. Zuo, C. Tan, S. Xian, C. Luo, S. Chen, L. Yu, and Y. Luo (2016). Kaempferol, a Flavonoid Compound from Gynura medica, Induced Apoptosis and Growth Inhibition in MCF-7 Breast Cancer Cells. African Journal of Traditional, Complementary and Alternative Medicines, 13(4); 210–215

Zhang, M.-Q. and B. Wilkinson (2007). Drug Discovery Beyond the ‘Rule-of-Five’. Current Opinion in Biotechnology, 18(6); 478–488

Authors

Rizq Rachmad Ramiizah
Dini Attala Hefa Insyira
Najwa Salsabila Hakim
Muhammad Rafly Aditya Firmansyah
Tukiran Tukiran
tukiran@unesa.ac.id (Primary Contact)
Ramiizah, R. R., Insyira, D. A. H. ., Hakim, N. S., Firmansyah, M. R. A. ., & Tukiran, T. (2026). Bioactivity Study of Russelia equisetiformis Ethanol Extract as an MCF-7 Breast Cancer Cells Inhibitor with In vitro and In silico Approaches. Science and Technology Indonesia, 11(3), 927–939. https://doi.org/10.26554/sti.2026.11.3.927-939

Article Details