Iranian Journal of Basic Medical Sciences

Iranian Journal of Basic Medical Sciences

Traditional dairy-derived Lactobacillus pentosus enhances the anticancer activity of 5-fluorouracil against breast cancer cells

Articles in Press

Document Type : Original Article

Authors
Tahmineh Ghanei Yazdi 1
Nazila Arbab-Soleimani 1
Farzad Rahmani 2, 3
Majid Khazaei 3, 4
1 Department of Biology, Da.C, Islamic Azad University, Damghan, Iran
2 Kashmar School of Medical Sciences, Mashhad University of Medical Sciences, Mashhad, Iran
3 Metabolic Syndrome Research Center, Mashhad University of Medical Sciences, Mashhad, Iran
4 Department of Medical Physiology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
10.22038/ijbms.2026.96617.20807
Abstract
Objective(s): Breast cancer remains a significant global health challenge and a leading cause of cancer-related mortality. Probiotics, key regulators of microbial and immune homeostasis, have emerged as promising candidates for cancer prevention and therapy. This study investigated the anticancer effects of Lactobacillus-derived cell-free supernatants (CFS) in in vitro and in vivo breast cancer models.
Materials and Methods: Lactobacillus strains were characterized for growth, viability, and antimicrobial activity. Standardized Lactobacillus cultures were used to prepare CFS by centrifugation and membrane filtration. The effects of CFS, alone or in combination with 5-fluorouracil (5-FU; 5 mg/kg every other day), on 4T1 breast cancer cell proliferation, migration, apoptosis, and inflammation were evaluated using MTT, flow cytometry, qPCR, and ELISA. An in vivo 4T1 breast cancer mouse model was established to assess the effects of CFS on tumor growth and associated molecular and histopathological changes. Statistical analyses were performed using Student’s t-test or one-way ANOVA with Tukey’s post hoc test; P<0.05 was considered significant.
Results: Co-treatment with CFS and 5-FU inhibits cell proliferation and cell-cycle progression by modulating p53 and cyclin D1 expression (P<0.05). CFS decreases tumor growth and promotes apoptosis by regulating BCL-2 and Bax (P<0.05). Moreover, co-treatment enhanced the inhibitory effect of 5-FU on cancer cell migration by modulating E-cadherin, MMP2, and MMP9 expression (P<0.05).
Conclusion: Lactobacillus-derived CFS may enhance the antiproliferative and pro-apoptotic effects of 5-FU in breast cancer models. These findings provide a preclinical basis for further investigation into their therapeutic potential.
Keywords
5-fluorouracil
Apoptosis
Breast cancer
Cell-free supernatant
Inflammation
Lactic acid bacterium
Postbiotic
Subjects
1.    Lei H, Fu J, Gu W, Qiao H, Guo H, Chen Z, et al. Breast cancer: Molecular pathogenesis, targeted therapy, screening, and prevention. MedComm (2020) 2026;7:e70560.
2.    Rahmani F, Ferns GA, Talebian S, Nourbakhsh M, Avan A, Shahidsales S. Role of regulatory miRNAs of the PI3K/AKT signaling pathway in the pathogenesis of breast cancer. Gene 2020;737:144459.
3.    Plaza-Diaz J, Álvarez-Mercado AI. The interplay between microbiota and chemotherapy-derived metabolites in breast cancer. Metabolites 2023;13:703.
4.    Abdelqader EM, Mahmoud WS, Gebreel HM, Kamel MM, Abu-Elghait M. Correlation between gut microbiota dysbiosis, metabolic syndrome and breast cancer. Sci Rep 2025;15:6652.
5.    Chapadgaonkar SS, Bajpai SS, Godbole MS. Gut microbiome influences incidence and outcomes of breast cancer by regulating levels and activity of steroid hormones in women. Cancer Rep (Hoboken) 2023;6:e1847.
6.    Zhu Z, Yang Y, Pan L, Ma L, Fang L. Gut microbiota metabolic reprogramming drives endocrine and immune resistance in hormone-dependent cancers. Cancers (Basel) 2026;18:1218.
7.    Rezaei S, Ghorbani E, Al-Asady AM, Avan A, Soleimanpour S, Khazaei M, et al. Evaluating the therapeutic efficacy of Lactobacillus strains in the management of ulcerative colitis: an overview of recent advances. Curr Pharm Des 2025;31:413-421.
8.    Rezai S, Ghorbani E, Nazari SE, Rahmani F, Hassanian SM, Afshari A, et al. Investigation of Lactobacillus probiotics derived from traditional dairy products in eliciting anti-tumor responses in mouse colorectal cancer model. Iran J Med Sci 2025;50:247-259.
9.    Thu MS, Ondee T, Nopsopon T, Farzana IAK, Fothergill JL, Hirankarn N, et al. Effect of probiotics in breast cancer: A systematic review and meta-analysis. Biology (Basel) 2023;12:280.
10.    Oyedokun PA, Oyeleke BT, Akanji OO, Oyelaran AO, O KP, Akanbi GB, et al. The role of the microbiome in gynecological cancers: implications for diagnosis and treatment. Front Immunol 2026;17:1718883.
11.    Sun J, Chen F, Wu G. Potential effects of gut microbiota on host cancers: focus on immunity, DNA damage, cellular pathways, and anticancer therapy. ISME J 2023;17:1535-1551.
12.    Prazdnova EV, Mazanko MS, Chistyakov VA, Bogdanova AA, Refeld AG, Kharchenko EY, et al. Antimutagenic activity as a criterion of potential probiotic properties. Probiotics Antimicrob Proteins 2022;14:1094-1109.
13.    Shi Q, Wang J, Zhou M, Zheng R, Zhang X, Liu B. Gut Lactobacillus contribute to the progression of breast cancer by affecting the anti-tumor activities of immune cells in the TME of tumor-bearing mice. Int Immunopharmacol 2023;124:111039.
14.    Barchelouei NK, Yazdi MH, Haghighat S. Probiotic intervention alters immune gene expression and tumor characteristics in experimental breast cancer. Mol Biol Rep 2025;52:809.
15.    Ranjbar S, Seyednejad SA, Azimi H, Rezaeizadeh H, Rahimi R. Emerging roles of probiotics in prevention and treatment of breast cancer: A comprehensive review of their therapeutic potential. Nutr Cancer 2019;71:1-12.
16.    Nandhini G, Prasanth S, Selvi KS, Sundaresan S. Isolation and characterization of probiotic lactic acid bacteria isolated from fermented South Indian cereals. Int J Nutr Pharmacol Neurol Dis 2025;15:135-141.
17.    Marchwińska K, Gwiazdowska D. Isolation and probiotic potential of lactic acid bacteria from swine feces for feed additive composition. Arch Microbiol 2021;204:61.
18.    Biswas S, Rahman L, Rahman MT, Chowdhury S, Khatun F, Nahar A, et al. In-vitro evaluation of probiotic potential of gut microbes isolated from retail chicken. PLoS One 2026;21:e0340981.
19.    Mulaw G, Sisay Tessema T, Muleta D, Tesfaye A. In vitro evaluation of probiotic properties of lactic acid bacteria isolated from some traditionally fermented Ethiopian food products. Int J Microbiol 2019;2019:7179514.
20.    Hirad AH, Ahmad J, Alkhedhairy AA, Bahkali AH, Khan ST. Bacterial isolates exhibiting multidrug resistance, hemolytic activity, and high 16S rRNA gene similarity with well-known pathogens found in camel milk samples of Riyadh region. APMIS 2018;126:215-226.
21.    Behbahani BA, Barzegar H, Mehrnia MA, Sheikhjan MG. Probiotic characterization of Limosilactobacillus fermentum isolated from local yogurt: Interaction with pathogenic bacteria and Caco-2 enteric cell line. Nutr Food Sci Res 2023;10:37-45.
22.    Falqueto A, Rodrigues RdS, Souza LV, Carvalho AFd, Caggia C, Nero LA, et al. Bactericidal and antibiofilm activity of lactic acid bacteria-derived cell free extracts against dairy-associated spoilage and pathogenic bacteria. Front Microbiol 2026;17:1783760.
23.    Drumond MM, Tapia-Costa AP, Neumann E, Nunes AC, Barbosa JW, Kassuha DE, et al. Cell-free supernatant of probiotic bacteria exerted antibiofilm and antibacterial activities against Pseudomonas aeruginosa: A novel biotic therapy. Front Pharmacol 2023;14:1152588.
24.    Keeratikunakorn K, Kaewchomphunuch T, Kaeoket K, Ngamwongsatit N. Antimicrobial activity of cell free supernatants from probiotics inhibits against pathogenic bacteria isolated from fresh boar semen. Sci Rep 2023;13:5995.
25.    Clarridge JE 3rd. Impact of 16S rRNA gene sequence analysis for identification of bacteria on clinical microbiology and infectious diseases. Clin Microbiol Rev 2004;17:840-862.
26.    Hashemzehi M, Yavari N, Rahmani F, Asgharzadeh F, Soleimani A, Shakour N, et al. Inhibition of transforming growth factor-beta by tranilast reduces tumor growth and ameliorates fibrosis in colorectal cancer. EXCLI J 2021;20:601-613.
27.    Rahmani F, Hashemzehi M, Avan A, Barneh F, Asgharzadeh F, Moradi Marjaneh R, et al. Rigosertib elicits potent anti-tumor responses in colorectal cancer by inhibiting Ras signaling pathway. Cell Signal 2021;85:110069.
28.    Rezaei N, Al-Asady AM, Hashemzehi M, Binabaj MM, Rahmani F, Avan A, et al. Crocin potentiates anti-tumor properties of 5-FU by regulating cell proliferation and tumor necrosis in breast cancer. Lett Drug Des Discov 2024;21:3161-3168.
29.    Raheem A, Liang L, Zhang G, Cui S. Modulatory effects of probiotics during pathogenic infections with emphasis on immune regulation. Front Immunol 2021;12:616713.
30.    Mishra VH, Gupta P, Khade AM, Deshkar AT. Use of probiotics in managing gastrointestinal, metabolic, and immune-related diseases. J Datta Meghe Inst Med Sci Univ 2025;20:214-217.
31.    Dehghani N, Tafvizi F, Jafari P. Cell cycle arrest and anti-cancer potential of probiotic Lactobacillus rhamnosus against HT-29 cancer cells. Bioimpacts 2021;11:245-252.
32.    Rahmani F, Zandigohar M, Safavi P, Behzadi M, Ghorbani Z, Payazdan M, et al. The interplay between noncoding RNAs and p21 signaling in gastrointestinal cancer: from tumorigenesis to metastasis. Curr Pharm Des 2023;29:766-776.
33.    Maroof H, Hassan ZM, Mobarez AM, Mohamadabadi MA. Lactobacillus acidophilus could modulate the immune response against breast cancer in murine model. J Clin Immunol 2012;32:1353-1359.
34.    Aindelis G, Chlichlia K. Modulation of anti-tumour immune responses by probiotic bacteria. Vaccines (Basel) 2020;8:329.
35.    Fortingo N, Melnyk S, Sutton SH, Watsky MA, Bollag WB. Innate immune system activation, inflammation and corneal wound healing. Int J Mol Sci 2022;23:14933.
36.    Kapoor G, Prakash S, Jaiswal V, Singh AK. Chronic inflammation and cancer: key pathways and targeted therapies. Cancer Invest 2025;43:1-23.
37.    Zhao H, Wu L, Yan G, Chen Y, Zhou M, Wu Y, et al. Inflammation and tumor progression: signaling pathways and targeted intervention. Signal Transduct Target Ther 2021;6:263.
38.    Yue Y, Ye K, Lu J, Wang X, Zhang S, Liu L, et al. Probiotic strain Lactobacillus plantarum YYC-3 prevents colon cancer in mice by regulating the tumour microenvironment. Biomed Pharmacother 2020;127:110159.
39.    Kaktcham PM, Zambou NF, Tchouanguep FM, El-Soda M, Choudhary MI. Antimicrobial and safety properties of lactobacilli isolated from two Cameroonian traditional fermented foods. Sci Pharm 2012;80:189-203.
40.    Korhonen J, Van Hoek AHA, Saarela M, Huys G, Tosi L, Mayrhofer S, et al. Antimicrobial susceptibility of Lactobacillus rhamnosus. Benef Microbes 2010;1:75-80.
41.    Afshari A, Hashemi M, Tavassoli M, Eraghi V, Noori SMA. Probiotic bacteria from 10 different traditional Iranian cheeses: isolation, characterization, and investigation of probiotic potential. Food Sci Nutr 2022;10:2009-2020.
42.    Safavi P, Moghadam KB, Haghighi Z, Ferns GA, Rahmani F. Interplay between LncRNA/miRNA and TGF-β signaling in the tumorigenesis of gynecological cancer. Curr Pharm Des 2024;30:352-361.
43.    Peng D, Fu M, Wang M, Wei Y, Wei X. Targeting TGF-β signal transduction for fibrosis and cancer therapy. Mol Cancer 2022;21:104.
44.    Park JS, Choi JW, Jhun J, Kwon JY, Lee BI, Yang CW, et al. Lactobacillus acidophilus improves intestinal inflammation in an acute colitis mouse model by regulation of Th17 and Treg cell balance and fibrosis development. J Med Food 2018;21:215-224.
45.    Kim H, Nam BY, Park J, Song S, Kim WK, Lee K, et al. Lactobacillus acidophilus KBL409 reduces kidney fibrosis via immune modulatory effects in mice with chronic kidney disease. Mol Nutr Food Res 2022;66:e2101105.
46.    Won SM, Lee NY, Oh KK, Gupta H, Sharma SP, Kim KH, et al. Gut Lactobacillus and probiotics Lactobacillus lactis/rhamnosis ameliorate liver fibrosis in prevention and treatment. J Microbiol 2023;61:245-257.
47.    Rahmani F, Avan A, Hashemy SI, Hassanian SM. Role of Wnt/β-catenin signaling regulatory microRNAs in the pathogenesis of colorectal cancer. J Cell Physiol 2018;233:811-817.
48.    Rahmani F, Tabrizi AT, Hashemian P, Alijannejad S, Rahdar HA, Ferns GA, et al. Role of regulatory miRNAs of the Wnt/β-catenin signaling pathway in tumorigenesis of breast cancer. Gene 2020;754:144892.
49.    Rahmani F, Hashemian P, Tabrizi AT, Ghorbani Z, Ziaeemehr A, Alijannejad S, et al. Regulatory role of miRNAs on Wnt/β-catenin signaling in tumorigenesis of glioblastoma. Indian J Cancer 2023;60:295-302.
50.    Paolillo M, Schinelli S. Extracellular matrix alterations in metastatic processes. Int J Mol Sci 2019;20:4947.
51.    Eble JA, Niland S. The extracellular matrix in tumor progression and metastasis. Clin Exp Metastasis 2019;36:171-198.
Iranian Journal of Basic Medical Sciences

Articles in Press, Accepted Manuscript
Available Online from 11 October 2026