Mycotoxins Show Estrogenic and Cytotoxic Effects in Breast Cancer Cell Lines

Fungal toxins that contaminate staple foods around the world can act on human breast cancer cells in sharply different ways depending on the toxin and the hormone receptor status of the cell line, according to a systematic review published in Frontiers in Public Health. The review, which followed PRISMA 2020 guidelines and covered literature up to December 2025, pooled evidence from 41 in vitro studies spanning 20 mycotoxin classes across 11 human breast cell lines. Its central finding is a receptor-dependent dichotomy: in estrogen receptor-positive models, the mycoestrogen zearalenone and its metabolite alpha-zearalenol behaved as potent estrogen agonists that drove proliferation, while triple-negative and ER-negative lines resisted hormone-driven growth but proved highly vulnerable to cytotoxic toxins such as aflatoxin B1, gliotoxin, and verrucarin A.

The quantitative signals are striking. Alpha-zearalenol showed the highest relative proliferative potency reported in the review, with an RPP of 7, normalized against 17β-estradiol as a reference baseline of 1.0. Parent zearalenone, meanwhile, increased viable MCF-7 cell numbers by 29.2 percent at 2 nanomolar, 154.5 percent at 32 nanomolar, and 190.4 percent at 96 nanomolar, while shifting cells from G1 into S phase, with the S-phase fraction rising roughly from 20 percent to 40 percent. On the cytotoxic side, aflatoxin B1 collapsed MCF-7 viability from 91.7 percent at 1.2 micrograms per milliliter to 31.1 percent at 9.6 micrograms per milliliter, with apoptosis rates reaching up to 46.4 percent. Gliotoxin suppressed proliferation across six breast cancer cell lines with IC50 values ranging from 38 to 985 nanomolar, and verrucarin A induced dose-dependent apoptosis with reported IC50 values of 0.29 and 0.41 micromolar.

These numbers come with important caveats. The review’s authors assessed risk of bias with an adapted QUIN tool for in vitro studies and found that none of the 41 included studies had a low risk of bias: 95.1 percent, or 39 studies, showed medium risk, and the remaining 4.9 percent, or two studies, showed high risk. The most common deficiencies were missing operator and outcome assessor blinding, absent randomization, and no formal sample size calculations. The authors explicitly caution that the findings should be interpreted with methodological caution, particularly when evaluating marginal cytotoxic thresholds. The evidence base is also entirely in vitro, so the review establishes biological plausibility in cell models rather than demonstrating any causal relationship between dietary mycotoxin exposure and human breast cancer.

The review’s scope reflects the scale of the underlying exposure problem. Mycotoxins are toxic secondary metabolites produced by filamentous fungi, principally from the genera Fusarium, Aspergillus, and Penicillium, which contaminate agricultural commodities under warm and humid conditions. The Food and Agriculture Organization has estimated that roughly 25 percent of the world’s food crops carry detectable mycotoxin contamination, a figure the review notes likely underestimates the total dietary burden when low-level contamination is included. Principal classes include zearalenone from Fusarium species, aflatoxins from Aspergillus species, ochratoxin A from Aspergillus and Penicillium species, and fumonisins from Fusarium species. The primary route of human exposure is consumption of contaminated foods, and cereal- and nut-heavy diets common in many developing regions are a major contributor.

The estrogenic arm of the review centers on zearalenone because its structure closely resembles 17β-estradiol, allowing it to bind estrogen receptors and trigger the same transcriptional programs as the body’s own hormones. In ER-positive luminal models such as MCF-7 and T47D, which were the most commonly used lines in the included studies, zearalenone enhanced expression of both ERα and ERβ, promoted G1 to S phase cell-cycle progression, and activated downstream signaling through the Ras/ERK cascade, cyclin D1, c-Myc, and upregulation of the anti-apoptotic protein Bcl-2. The authors also report that zearalenone increased the S-phase population by roughly 50 percent at 24 hours in some studies, with an eight-fold rise in cyclin B1-associated kinase activity, and that several studies demonstrated synergistic activity with estradiol, which the review interprets as tumor-promoting potential in hormone-sensitive tissue.

Among the zearalenone metabolites, alpha-zearalenol consistently showed the strongest proliferative potency, with a proliferative effect of 2.6 relative to untreated controls and the RPP of 7 relative to 17β-estradiol. The review attributes this superior potency to stereospecific reduction of the C-6 keto group, which produces a steric orientation more compatible with the receptor’s binding conformation. In contrast, hydrolyzed metabolites HZEN and DHZEN showed negligible estrogenicity, with potency roughly 50 to 10,000 times lower than intact zearalenone, because loss of the intact lactone ring abolishes ERα binding capacity. Zearalenone-14-glucoside, a masked mycotoxin, caused no direct toxicity up to 1 millimolar but was enzymatically converted by MCF-7 cells into active zearalenone, alpha-zearalenol, and beta-zearalenol over two to six hours, unmasking estrogenic potential through ER signaling.

The cytotoxic arm of the review covers a structurally diverse set of toxins with distinct mechanisms. Aflatoxin B1 requires bioactivation by liver cytochrome P450 enzymes to form DNA adducts, and the review describes its ability to target and mutate the TP53 tumor suppressor gene, alongside intracellular reactive oxygen species accumulation. Brefeldin A showed exceptionally high potency in triple-negative MDA-MB-231 suspension cultures, with an EC50 of approximately 0.016 micrograms per milliliter, acting through ER-stress activation, PARP cleavage, and preferential induction of anoikis, a form of detachment-induced apoptosis. Ochratoxin A produced cytotoxicity with a monolayer IC50 of roughly 3 micromolar at 24 hours via ROS-mediated oxidative stress and intrinsic mitochondrial apoptosis. Wortmannin, a PI3K/Akt and NF-κB pathway inhibitor, increased apoptosis from 1.75 percent at 1 nanomolar to 25.3 percent at 50 nanomolar, and in BT-20 cells it suppressed oncogenic microRNAs including miR-21 while upregulating tumor-suppressive species such as miR-19a/b and miR-22, alongside downregulation of the mesenchymal marker Vimentin and upregulation of the metastasis suppressor NME1.

Several toxins showed selective toxicity toward malignant cells over non-tumorigenic controls. Patulin, with an IC50 of roughly 25 micromolar in MCF-7 cells, reduced cancer cell viability to 12 percent at 100 micromolar while normal cell viability remained at 36 percent. Cytochalasin B showed an IC50 of approximately 12.5 micromolar in ZR-75-1 cells with 98.2 percent cell death, yet no toxicity was observed in normal MCF-10A cells. Under aflatoxin B1 exposure, non-tumorigenic MCF-10A cells showed apoptosis rates of 40.8 percent versus 46.4 percent in malignant MCF-7 cells, a contrast the review cites as evidence of selective tumor cell vulnerability. Alternariol and altertoxins took a different route, activating the aryl hydrocarbon receptor and inducing CYP1 enzyme activity to roughly 190 percent of controls, which shifts endogenous estrogen metabolism toward genotoxic catecholestrogens, an indirect pathway through which non-estrogenic mycotoxins may promote estrogenic carcinogenesis.

The review also highlights how experimental architecture shapes apparent toxicity. Three-dimensional spheroid cultures were markedly more resistant to mycotoxins such as sterigmatocystin and ochratoxin A than conventional two-dimensional monolayers, and the authors argue that 2D systems, lacking the spatial architecture and stromal interactions of the native tumor microenvironment, likely overestimate cellular sensitivity. Because of substantial methodological heterogeneity across the included studies, spanning exposure concentrations from picomolar to micromolar levels and durations from minutes to 12 days, quantitative meta-analysis was precluded, and the authors conducted a structured narrative synthesis stratified first by hormone receptor status and then by mycotoxin class. They further note that the near-exclusive examination of individual mycotoxins rather than realistic co-exposure mixtures, and a paucity of epigenetic endpoint studies, remain key limitations, although one included study showed that combined low-dose zearalenone and aflatoxin B1 promoted greater cell cycle progression in MCF-7 cells than either compound alone.

The authors frame their conclusions carefully. They state that the current evidence does not establish direct causality for human breast cancer but robustly demonstrates biological plausibility, and they note that the epidemiological literature directly linking mycotoxin exposure to human breast cancer remains sparse and conflicting, with only a small number of case-control studies examining zearalenone biomarkers and inconsistent results likely due to variations in biomarker selection, assay sensitivity, and dietary assessment methods. Future research priorities they identify include enhanced regulatory surveillance of zearalenone in food supplies, prospective epidemiological cohort studies with validated biomarkers, three-dimensional organoid and co-exposure experimental paradigms, and epigenetic endpoint characterization in breast epithelial models. Given the predominantly medium risk of bias across the evaluated in vitro literature, the review’s quantitative thresholds should be read as mechanistic signals from cell culture experiments rather than as established risks to human health, and the authors emphasize that mycotoxins cannot be treated as a single pharmacological entity but instead constitute a structurally and mechanistically varied group of fungal metabolites requiring further scientific and public health attention.

Subject of Research: Systematic review of estrogenic, proliferative, and cytotoxic effects of mycotoxins in human breast cancer cell line models

Article Title: Estrogenic, cytotoxic, proliferative effects of mycotoxins in human breast cancer cell line models: a systematic review

Article References: Sasidharan, S., Naina, P., Jyoti, D., Arora, T., Burman, A., Bansal, S., Gowardhan, S., Mishra, A., & Chandra, N. (2026). Estrogenic, cytotoxic, proliferative effects of mycotoxins in human breast cancer cell line models: a systematic review. Frontiers in Public Health, 14, 1950615. https://doi.org/10.3389/fpubh.2026.1950615

Image Credits: AI Generated

DOI: 10.3389/fpubh.2026.1950615

Keywords: mycotoxins, zearalenone, alpha-zearalenol, breast cancer, estrogen receptor, systematic review, aflatoxin B1, gliotoxin, verrucarin A, cell lines, in vitro, xenoestrogens