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Abstract

High-risk neuroblastoma (HR-NB) is an aggressive form of childhood cancer with a five-year survival rate of less than 50%, underscoring the need for more efficacious and less toxic treatments. The glycoprotein vitronectin (VN) has been linked to poor prognosis in patients with HR-NB, thus inhibitors of VN function represent a promising avenue for molecular mechanotherapy. This study investigated the binding affinity between the somatomedin B (SMB) domain of VN and natural compounds from the medicinal plant, Olax subscorpioidea, targeting the plasminogen activator inhibitor-1 (PAI-1). The therapeutic potential of α-amyrin (AMY), lupeol (LUP), and olax chalcone A (olax CHA) was tested in combination with an integrin antagonist of VN, cilengitide (CLG), using the SK-N-BE(2) HR-NB cell line as a model. Molecular docking studies indicated protein-ligand interactions for all compounds, with CLG showing the most favorable binding free energy, followed by LUP, AMY, and olax CHA. Molecular dynamics simulations indicated the SMB domain of VN binding site of PAI-1 initially exhibited flexibility, with alpha-carbon root mean square deviation (RMSD) stabilizing at 1.8-2.1 Å. All compounds reduced SK-N-BE(2) cell viability in a dose-dependent manner. CLG showed the strongest antiproliferative effect (IC₅₀ = 15.25 µM). Olax CHA had higher efficacy than AMY and LUP (IC₅₀ = 74.23 µM vs. 125.45 µM and 103.36 µM). Combining the compounds with CLG further decreased viability and IC₅₀ values. Synergy analysis showed only olax CHA plus CLG had a synergistic effect. This suggests olax CHA plus CLG as a promising therapeutic strategy against neuroblastoma cells.

References

Adelegan, A. A., Talabi, A. A., Dokunmu, T. M., & Iweala, E. E. (2024). Anticancer activity of ethyl acetate fraction and ethanol leaf extract of olax subscorpioidea against DMBA-Induced female rats. Tropical Journal of Natural Product Research, 8(1).

Alam, P., Al-Yousef, H. M., Siddiqui, N. A., Alhowiriny, T. A., Alqasoumi, S. I., Amina, M., Hassan, W. H. B., Abdelaziz, S., & Abdalla, R. H. (2018). Anticancer activity and concurrent analysis of ursolic acid, β-sitosterol and lupeol in three different Hibiscus species (aerial parts) by validated HPTLC method. Saudi Pharmaceutical Journal, 26(7), 1060-1067.

Alnajjar, R., Mostafa, A., Kandeil, A., & Al-Karmalawy, A. A. (2020). Molecular docking, molecular dynamics, and in vitro studies reveal the potential of angiotensin II receptor blockers to inhibit the COVID-19 main protease. Heliyon, 6(12).

Blanco-Luquin, I., Lázcoz, P., Celay, J., Castresana, J. S., & Encío, I. J. (2021). In vitro assessment of the role of p53 on chemotherapy treatments in neuroblastoma cell lines. Pharmaceuticals, 14(11), 1184.

Burgos-Panadero, R., El Moukhtari, S. H., Noguera, I., Rodríguez-Nogales, C., Martín-Vañó, S., Vicente-Munuera, P., Cañete, A., Navarro, S., Blanco-Prieto, M. J., & Noguera, R. (2021). Unraveling the extracellular matrix-tumor cell interactions to aid better targeted therapies for neuroblastoma. International Journal of Pharmaceutics, 608, 121058.

Burgos-Panadero, R., Noguera, I., Cañete, A., Navarro, S., & Noguera, R. (2019). Vitronectin as a molecular player of the tumor microenvironment in neuroblastoma. BMC Cancer, 19(1), 479. https://doi.org/10.1186/s12885-019-5693-2

Carroll, M. J., Mauldin, R. V., Gromova, A. V., Singleton, S. F., Collins, E. J., & Lee, A. L. (2012). Evidence for dynamics in proteins as a mechanism for ligand dissociation. Nature chemical biology, 8(3), 246-252.

Chaurasiya, N. D., Leon, F., Muhammad, I., & Tekwani, B. L. (2022). Natural products inhibitors of monoamine oxidases—Potential new drug leads for neuroprotection, neurological disorders, and neuroblastoma. Molecules, 27(13), 4297.

Chinot, O. L. (2014). Cilengitide in glioblastoma: when did it fail? The Lancet Oncology, 15(10), 1044-1045.

Chou, T.-C. (2010). Drug combination studies and their synergy quantification using the Chou-Talalay method. Cancer research, 70(2), 440-446.

Constantinescu, T., & Mihis, A. G. (2022). Two important anticancer mechanisms of natural and synthetic chalcones. International Journal of Molecular Sciences, 23(19), 11595.

Echavidre, W., Picco, V., Faraggi, M., & Montemagno, C. (2022). Integrin-αvβ3 as a Therapeutic Target in Glioblastoma: Back to the Future? Pharmaceutics, 14(5). https://doi.org/10.3390/pharmaceutics14051053

Fan, H., & Mark, A. E. (2003). Relative stability of protein structures determined by X‐ray crystallography or NMR spectroscopy: a molecular dynamics simulation study. Proteins: Structure, Function, and Bioinformatics: Structure, Function, and Bioinformatics, 53(1), 111-120.

Ferraris, G. M. S., Schulte, C., Buttiglione, V., De Lorenzi, V., Piontini, A., Galluzzi, M., Podestà, A., Madsen, C. D., & Sidenius, N. (2014). The interaction between uPAR and vitronectin triggers ligand‐independent adhesion signalling by integrins. The EMBO journal, 33(21), 2458-2472.

Forgacs, P., & Provost, J. (1981). Olaxoside, a saponin from Olax andronensis, Olax glabriflora and Olax psittacorum. Phytochemistry, 20(7), 1689-1691.

Gordon, A. T., Hosten, E. C., van Vuuren, S., & Ogunlaja, A. S. (2025). Copper (II)-photocatalyzed Hydrocarboxylation of Schiff bases with CO2: antimicrobial evaluation and in silico studies of Schiff bases and unnatural α-amino acids. Journal of Biomolecular Structure and Dynamics, 43(8), 4201-4214.

Granados-Aparici, S., Vieco-Martí, I., López-Carrasco, A., Navarro, S., & Noguera, R. (2024). Real-time morphometric analysis of targeted therapy for neuroblastoma cells in monolayer and 3D hydrogels using digital holographic microscopy. iScience, 27(11).

Idris, M. O., Yekeen, A. A., Alakanse, O. S., & Durojaye, O. A. (2021). Computer-aided screening for potential TMPRSS2 inhibitors: a combination of pharmacophore modeling, molecular docking and molecular dynamics simulation approaches. Journal of Biomolecular Structure and Dynamics, 39(15), 5638-5656.

Jemaà, M., Sime, W., Abassi, Y., Lasorsa, V. A., Bonne Køhler, J., Michaelis, M., Cinatl, J., Jr., Capasso, M., & Massoumi, R. (2020). Gene Expression Signature of Acquired Chemoresistance in Neuroblastoma Cells. International Journal of Molecular Sciences, 21(18). https://doi.org/10.3390/ijms21186811

Jensen, J. K., Durand, M. K., Skeldal, S., Dupont, D. M., Bødker, J. S., Wind, T., & Andreasen, P. A. (2004). Construction of a plasminogen activator inhibitor‐1 variant without measurable affinity to vitronectin but otherwise normal. FEBS letters, 556(1-3), 175-179.

Juvvuna, P. K., Mondal, T., Di Marco, M., Kosalai, S. T., Kanduri, M., & Kanduri, C. (2021). NBAT1/CASC15-003/USP36 control MYCN expression and its downstream pathway genes in neuroblastoma. Neuro-Oncology Advances, 3(1), vdab056.

Karplus, M., & McCammon, J. A. (2002). Molecular dynamics simulations of biomolecules. Nature Structural Biology, 9(9), 646-652. https://doi.org/10.1038/nsb0902-646

Khoddam, A., Vaughan, D., & Wilsbacher, L. (2025). Role of plasminogen activator inhibitor-1 (PAI-1) in age-related cardiovascular pathophysiology. The Journal of Cardiovascular Aging, 5(8).

Kumar, S., & Pandey, A. K. (2013). Chemistry and biological activities of flavonoids: an overview. The Scientific World Journal, 2013(1), 162750.

Leblond, P., Dewitte, A., Le Tinier, F., Bal-Mahieu, C., Baroncini, M., Sarrazin, T., Lartigau, E., Lansiaux, A., & Meignan, S. (2013). Cilengitide targets pediatric glioma and neuroblastoma cells through cell detachment and anoikis induction. Anti-Cancer Drugs, 24(8), 818-825.

Lee, J.-H., Lee, E.-J., Kwon, J.-S., Hwang, C.-J., & Kim, K.-N. (2014). Cytotoxicity comparison of the nanoparticles deposited on latex rubber bands between the original and stretched state. Journal of Nanomaterials, 2014(1), 567827.

Li, D., Park, Y., Hemati, H., & Liu, X. (2023). Cell aggregation activates small GTPase Rac1 and induces CD44 cleavage by maintaining lipid raft integrity. Journal of Biological Chemistry, 299(12).

Liu, K., Zhang, X., Xie, L., Deng, M., Chen, H., Song, J., Long, J., Li, X., & Luo, J. (2021). Lupeol and its derivatives as anticancer and anti-inflammatory agents: Molecular mechanisms and therapeutic efficacy. Pharmacological Research, 164, 105373.

Lönnroth, E.-C. (2005). Toxicity of medical glove materials: a pilot study. International Journal of Occupational Safety and Ergonomics, 11(2), 131-139.

Mas-Moruno, C., Rechenmacher, F., & Kessler, H. (2010). Cilengitide: the first anti-angiogenic small molecule drug candidate. Design, synthesis and clinical evaluation. Anti-Cancer Agents in Medicinal Chemistry (Formerly Current Medicinal Chemistry-Anti-Cancer Agents), 10(10), 753-768.

Matthay, K. K., Maris, J. M., Schleiermacher, G., Nakagawara, A., Mackall, C. L., Diller, L., & Weiss, W. A. (2016). Neuroblastoma. Nature Reviews Disease Primers, 2, 16078. https://doi.org/10.1038/nrdp.2016.78

Mishra, T., Arya, R. K., Meena, S., Joshi, P., Pal, M., Meena, B., Upreti, D., Rana, T., & Datta, D. (2016). Isolation, characterization and anticancer potential of cytotoxic triterpenes from Betula utilis bark. PloS one, 11(7), e0159430.

Muddagoni, N., Bathula, R., Dasari, M., & Potlapally, S. R. (2021). Homology modeling, virtual screening, prime-MMGBSA, AutoDock-identification of inhibitors of FGR protein. Biointerface Research in Applied Chemistry, 11(4), 11088-11103.

Narasaki, R., Xu, Z., Liang, Z., Fung, L., Donahue, D., Castellino, F., & Ploplis, V. (2012). The vitronectin‐binding domain of plasminogen activator inhibitor‐1 plays an important functional role in lipopolysaccharide‐induced lethality in mice. Journal of Thrombosis and Haemostasis, 10(12), 2618-2621.

Neto, S. F., Prada, A. L., Achod, L. D. R., Torquato, H. F. V., Lima, C. S., Paredes-Gamero, E. J., de Moraes, M. O. S., Lima, E. S., Sosa, E. H., & de Souza, T. P. (2021). α-amyrin-loaded nanocapsules produce selective cytotoxic activity in leukemic cells. Biomedicine & Pharmacotherapy, 139, 111656.

Nowakowska, Z. (2007). A review of anti-infective and anti-inflammatory chalcones. European Journal of Medicinal Chemistry, 42(2), 125-137.

Oladipupo, A. R., Alaribe, S. C., Ogunlaja, A. S., Beniddir, M. A., Gordon, A. T., Ogah, C. O., Okpuzor, J., & Coker, H. A. (2024). Structure-based molecular networking, molecular docking, dynamics simulation and pharmacokinetic studies of Olax subscorpioidea for identification of potential inhibitors against selected cancer targets. Journal of Biomolecular Structure and Dynamics, 42(3), 1110-1125.

Oladipupo, A. R., Alaribe, S. C. A., Ogunlaja, A. S., Beniddir, M. A., Ogah, C. O., Okpuzor, J., & Coker, H. A. (2023). Chemical and biological insights on Phaulopsis falcisepala: a source of bioactive compounds with multifunctional anticancer potentials. Chemistry Africa, 6(3), 1175-1189.

Oskouian, B., Lee, J. Y., Asgharzadeh, S., Khan, R., Zhang, M., Weisbrod, J. R., Choi, Y.-J., Puri, L., Aguilar, A. E., & Zhao, P. (2024). AF1q is a universal marker of neuroblastoma that sustains N-Myc expression and drives tumorigenesis. Oncogene, 43(16), 1203-1213.

Ouyang, Y., Li, J., Chen, X., Fu, X., Sun, S., & Wu, Q. (2021). Chalcone derivatives: role in anticancer therapy. Biomolecules, 11(6), 894.

Preissner, K. T. (1991). Structure and biological role of vitronectin. Annual Review of Cell and Developmental Biology, 7, 275-310. https://doi.org/10.1146/annurev.cb.07.110191.001423

Ross, R. A., & Spengler, B. A. (2007). Human neuroblastoma stem cells. Seminars in cancer biology

Sargsyan, K., Grauffel, C., & Lim, C. (2017). How molecular size impacts RMSD applications in molecular dynamics simulations. Journal of Chemical Theory and Computation, 13(4), 1518-1524.

Sernissi, L., Ricci, L., Scarpi, D., Bianchini, F., Arosio, D., Contini, A., & Occhiato, E. G. (2018). Stereodivergent synthesis of 5-aminopipecolic acids and application in the preparation of a cyclic RGD peptidomimetic as a nanomolar αVβ3 integrin ligand. Organic & Biomolecular Chemistry, 16(18), 3402-3414.

Shahdeo, D., Kesarwani, V., Suhag, D., Ahmed, J., Alshehri, S. M., & Gandhi, S. (2021). Self-assembled chitosan polymer intercalating peptide functionalized gold nanoparticles as nanoprobe for efficient imaging of urokinase plasminogen activator receptor in cancer diagnostics. Carbohydrate Polymers, 266, 118138.

Sillen, M., & Declerck, P. J. (2020). Targeting PAI-1 in cardiovascular disease: structural insights into PAI-1 functionality and inhibition. Frontiers in Cardiovascular Medicine, 7, 622473.

Sillen, M., Miyata, T., Vaughan, D. E., Strelkov, S. V., & Declerck, P. J. (2021). Structural insight into the two-step mechanism of PAI-1 inhibition by small molecule TM5484. International Journal of Molecular Sciences, 22(3), 1482.

Stupp, R., Hegi, M. E., Gorlia, T., Erridge, S. C., Perry, J., Hong, Y.-K., Aldape, K. D., Lhermitte, B., Pietsch, T., & Grujicic, D. (2014). Cilengitide combined with standard treatment for patients with newly diagnosed glioblastoma with methylated MGMT promoter (CENTRIC EORTC 26071-22072 study): a multicentre, randomised, open-label, phase 3 trial. The Lancet Oncology, 15(10), 1100-1108.

Teilum, K., Olsen, J. G., & Kragelund, B. B. (2009). Functional aspects of protein flexibility. Cellular and Molecular Life Sciences, 66(14), 2231-2247.

Tucci, M., Stucci, S., & Silvestris, F. (2014). Does cilengitide deserve another chance? The Lancet Oncology, 15(13), e584-e585.

Vora, J., Patel, S., Athar, M., Sinha, S., Chhabria, M. T., Jha, P. C., & Shrivastava, N. (2020). Pharmacophore modeling, molecular docking and molecular dynamics simulation for screening and identifying anti-dengue phytocompounds. Journal of Biomolecular Structure & Dynamics, 38(6), 1726–1740. https://doi.org/10.1080/07391102.2019.1615002

Wahab, R. M. A., Rozali, N. A. M., Senafi, S., Abidin, I. Z. Z., Ariffin, Z. Z., & Ariffin, S. H. Z. (2017). Impact of isolation method on doubling time and the quality of chondrocyte and osteoblast differentiated from murine dental pulp stem cells. PeerJ, 5, e3180.

Wang, L., Lin, X., & Sun, P. (2022). uPAR, beyond regulating physiological functions, has orchestrated roles in cancer (Review). International Journal of Oncology, 61(6). https://doi.org/10.3892/ijo.2022.5441

Wilson, P. G., Devkota, L., Payne, T., Crisp, L., Winter, A., & Wang, Z. (2012). Clonal populations of amniotic cells by dilution and direct plating: evidence for hidden diversity. Stem Cells International, 2012, 485950. https://doi.org/10.1155/2012/485950

Yasukawa, K., & Tabata, K. (2015). Promising natural products as anti-cancer agents against neuroblastoma. International Journal of Cancer Research and Prevention, 8, 267.

Yoneda, A. (2023). Role of surgery in neuroblastoma. Pediatric Surgery International, 39(1), 177. https://doi.org/10.1007/s00383-023-05459-1

Zafar, A., Wang, W., Liu, G., Wang, X., Xian, W., McKeon, F., Foster, J., Zhou, J., & Zhang, R. (2021). Molecular targeting therapies for neuroblastoma: Progress and challenges. Medicinal Research Reviews, 41(2), 961-1021. https://doi.org/10.1002/med.21750

Zhong, J., Yang, H.-C., Kon, V., Fogo, A. B., Lawrence, D. A., & Ma, J. (2014). Vitronectin-binding PAI-1 protects against the development of cardiac fibrosis through interaction with fibroblasts. Laboratory Investigation, 94(6), 633-644.

Zhou, A., Huntington, J. A., Pannu, N. S., Carrell, R. W., & Read, R. J. (2003). How vitronectin binds PAI-1 to modulate fibrinolysis and cell migration. Nature Structural & Molecular Biology, 10(7), 541-544.

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