Anti-metastatic potential of OR3 pigment from Streptomyces coelicolor JUACT03 on MDA-MB-231 breast cancer in vitro and in vivo

Abstract

Objective: To investigate the mechanism of anticancer activity of a pigment OR3 from Streptomyces coelicolor in in vitro and in vivo metastatic breast cancer models and to characterize the pigment.

Methods: The anticancer mechanism was analyzed in MDA-MB-231 cells using MTT, lactate dehydrogenase, caspase, DNA fragmentation, clonogenic, flow cytometry, Western blot, and scratch assays. The effects of OR3 on xenograft mouse models were evaluated by tumor volume measurement, hematological analysis, and histopathological observation. The characterization of OR3 was also performed using gas chromatograohy-mass spectrometry and nuclear magnetic resonance spectroscopy.

Results: OR3 exhibited potent cytotoxicity against MDA-MB-231 cells, with no observed effects on HEK-293 cells. Caspase-9 activation was detected in OR3-treated MDA-MB-231 cells. Flow cytometry showed a dose-dependent induction of apoptosis and cell cycle arrest at the sub-G₁ and S phases. Furthermore, OR3 completely inhibited MDA-MB-231 cell migration and demonstrated anti-proliferative effects by downregulating the protein expression of KPNA2, XPO1, RAB5B, and p38 MAPK. In in vivo studies, OR3 was non-toxic to mice, inhibited tumor xenograft growth, and maintained normal hematological parameters and tissue architecture. Nuclear magnetic resonance spectroscopy demonstrated the presence of a prodigiosin-like compound, while gas chromatography-mass spectrometry analysis identified additional compounds in OR3.

Conclusions: Our findings validate OR3 pigment as a promising compound for the treatment of metastatic breast cancer, warranting further studies.

Keywords

Breast cancer / MDA-MB-231 / Tumor growth inhibition / Apoptosis / OR3 pigment / Streptomyces coelicolor / Metastasis

Conflict of interest statement

The authors declare that they have no conflict of interest.

Acknowledgments

The authors are grateful to JAIN (Deemed-to-be University), Bengaluru for providing the infrastructural facilities to carry out the research. The author, Mr. Somasekhara D would like to acknowledge the financial support by way of fellowship provided by CSIR, Govt. of India.

Funding

The authors received no extramural funding for the study.

Data availability statement

The data supporting the findings of this study are available from the corresponding authors upon request.

Authors’ contributions

VKN and SD contributed to the conceptualization. SD designed the experiment, investigations, methodology and formal analysis. SD contributed to writing the initial draft. Both VKN and SD contributed to the final version of the manuscript, and approved it. VKN supervised the project.

Publisher’s note

The Publisher of the Journal remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

References

[1] Mehrotra R, Yadav K. Breast cancer in India: Present scenario and the challenges ahead. World J Clin Oncol 2022; 13(3): 209-218.

[2] Liang Y, Zhang D, Tse GM, Lin H, Kruyt FAE. Editorial: The molecular mechanisms of metastasis and therapeutic resistance in breast cancer. Front Oncol 2023; 13. doi: 10.3389/fonc.2023.1194858.

[3] Kavya D, Nadumane VK. A combination of semi-purified L-methioninase with tamoxifen citrate to ameliorate breast cancer in athymic nude mice. Mol Biol Rep 2023; 50: 2925-2932.

[4] Hoshyar R, Mohaghegh Z, Torabi N, Abolghasemi A. Antitumor activity of aqueous extract of Ziziphus jujube fruit in breast cancer: An in vitro and in vivo study. Asian Pac J Reprod 2015; 4(2): 116-122.

[5] Kroschinsky F, Stolzel F, von Bonin S, Beutel G, Kochanek M, Kiehl M, et al. New drugs, new toxicities: Severe side effects of modern targeted and immunotherapy of cancer and their management. Crit Care 2017; 21(1): 89.

[6] Newman DJ, Cragg GM. Natural products as sources of new drugs over the last 40 years. J Nat Prod 2020; 83(3): 770-803.

[7] Atanasov AG, Zotchev SB, Dirsch VM, Orhan IE, Banach M, Rollinger JM, et al. Natural products in drug discovery: Advances and opportunities. Nat Rev Drug Discov 2021; 20(3): 200-216.

[8] Tan LT, Ser HL, Yin WF, Chan KG, Lee LH, Goh BH. Investigation of antioxidative and anticancer potentials of Streptomyces sp. MUM256 isolated from Malaysia mangrove soil. Front Microbiol 2015; 6. doi: 10.3389/fmicb.2015.01316.

[9] Finger M, Sentek F, Hartmann L, Palacio-Barrera AM, Schlembach I, Rosenbaum MA, et al. Insights into Streptomyces coelicolor A3(2) growth and pigment formation with high-throughput online monitoring. Eng Life Sci 2022; 23(1). doi: 10.1002/elsc.202100151.

[10] Law JW, Law LN, Letchumanan V, Tan LT, Wong SH, Chan KG, et al. Anticancer drug discovery from microbial sources: The unique mangrove Streptomycetes. Molecules 2020; 25(22). doi: 10.3390/molecules25225365.

[11] D S, Dammalli M, Nadumane VK. Proteomic analysis of human breast cancer MCF-7 cells to identify cellular targets of the anticancer pigment OR3 from Streptomyces coelicolor JUACT03. Appl Biochem Biotechnol 2023; 195(1): 236-252.

[12] Prashanthi K, Suryan S, Varalakshmi KN. In vitro anticancer property of yellow pigment from Streptomyces griseoaurantiacus JUACT 01. Braz Arch Biol Technol 2015; 58(6): 869-876.

[13] Venkatachalam P, Nadumane VK. Overexpression of p53 and Bax mediating apoptosis in cancer cell lines induced by a bioactive compound from Bacillus endophyticus JUPR15. Process Biochem 2018; 73: 170-179.

[14] Pozarowski P, Darzynkiewicz Z. Analysis of cell cycle by flow cytometry. In: Schonthal AH (ed.) Checkpoint controls and cancer. Methods in molecular biology. USA, New Jersey: Humana Press; 2004, p. 281, 301-311.

[15] Buranrat B, Junking M. Piperine suppresses growth and migration of human breast cancer cells through attenuation of Rac1 expression. Asian Pac J Trop Biomed 2022; 12(1): 39-46.

[16] Bolla SR, Mohammed Al-Subaie A, Yousuf Al-Jindan R, Papayya Balakrishna J, Kanchi Ravi P, Veeraraghavan VP, et al. In vitro wound healing potency of methanolic leaf extract of Aristolochia saccata is possibly mediated by its stimulatory effect on collagen-1 expression. Heliyon 2019; 5(5). doi: 10.1016/j.heliyon.2019.e01648.

[17] Arivuselvam R, Dera AA, Parween Ali S, Alraey Y, Saif A, Hani U, et al. Isolation, identification, and antibacterial properties of prodigiosin, a bioactive product produced by a new Serratia marcescens JSSCPM1 strain: Exploring the biosynthetic gene clusters of serratia species for biological applications. Antibiotics 2023; 12(9): 1466. doi: 10.3390/antibiotics12091466.

[18] Faraag AH, El-Batal AI, El-Hendawy HH. Characterization of prodigiosin produced by Serratia marcescens strain isolated from irrigation water in Egypt. Nat Sci 2017; 15(5): 55-68.

[19] Salim N, Santhiagu A, Joji K. Purification, characterization, and anticancer evaluation of l-methioninase from Trichoderma harzianum. 3 Biotech 2020; 10: 501. doi: 10.1007/s13205-020-02494-w.

[20] Lopez J, Tait SW. Mitochondrial apoptosis: Killing cancer using the enemy within. Br J Cancer 2015; 112: 957-962.

[21] Ravikumar S, Fredimoses M, Gnanadesigan M. Anticancer property of sediment actinomycetes against MCF-7 and MDA-MB-231 cell lines. Asian Pac J Trop Biomed 2012; 2(2): 92-96.

[22] Aftab U, Zechel DL, Sajid I. Antitumor compounds from Streptomyces sp. KML-2, isolated from Khewra salt mines Pakistan. Biol Res 2015; 48: 58. doi: 10.1186/s40659-015-0046-3.

[23] Jeong SY, Shin H, Kim T, Lee HS, Park SK, Kim HM. Streptokordin, a new cytotoxic compound of the methylpyridine class from a marine-derived Streptomyces sp. KORDI-3238. J Antibiot 2006; 59: 234-240.

[24] Hughes CC, MacMillan JB, Gaudencio SP, Jensen PR, Fenical W. The ammosamides: Structures of cell cycle modulators from a marine-derived Streptomyces species. Angew Chem Int Ed Engl 2009; 48(4): 725-727.

[25] Poon IK, Hulett M D, Parish CR. Molecular mechanisms of late apoptotic/necrotic cell clearance. Cell Death Differ 2010; 17(3): 381-397.

[26] Elmallah MI, Micheau O. Marine drugs regulating apoptosis induced by tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Mar Drugs 2015; 13: 6884-6909.

[27] Dutta S, Mitra SK, Bir A, T RP, Ghosh A. Enhancing anti-cancer activity: Green synthesis and cytotoxicity evaluation of turmeric-gold nanocapsules on A549 lung cancer cells. Cureus 2023; 15(8). doi: 10.7759/cureus.43087.

[28] Narod SA, Sopik V. Is invasion a necessary step for metastases in breast cancer? Breast Cancer Res Treat 2018; 169: 9-23.

[29] Qu XY, Ren JW, Peng AH, Lin SQ, Lu DD, Du QQ, et al. Cytotoxic, anti-migration, and anti-invasion activities on breast cancer cells of angucycline glycosides isolated from a marine-derived Streptomyces sp. Mar Drugs 2019; 17. doi: 10.3390/md17050277.

[30] Vicente J, Stewart AK, van Wagoner RM, Elliott E, Bourdelais AJ, Wright JL. Monacyclinones, new angucyclinone metabolites isolated from Streptomyces sp. M7_15 associated with the Puerto Rican sponge Scopalina ruetzleri. Mar Drugs 2015; 13(8): 4682-4700.

[31] Alapati V, Noolvi MN, Manjula SN, Pallavi KJ, Patel HM, Tippeswamy BS, et al. In vivo anti-tumour activity of novel quinazoline derivatives. Eur Rev Med Pharmacol Sci 2012; 16(13): 1753-1764.