Gut Bacteria Fatty Acid Blocks Key Enzyme and Halts Endometrial Cancer Cell Growth in Lab Study

Sodium butyrate, a short-chain fatty acid produced when gut microbes ferment dietary fiber, suppresses the growth of endometrial cancer cells in laboratory experiments by strongly suppressing thymidylate synthase, a key enzyme in de novo thymidylate synthesis, and by broadly reprogramming pyrimidine metabolism. The findings, published in Genes & Nutrition, come from a study of two human endometrial cancer cell lines, HEC1A and AN3CA, and represent the first demonstration of thymidylate synthase suppression by sodium butyrate in endometrial cancer.

Endometrial cancer is the most common gynecologic malignancy and the sixth leading cause of cancer-related death among women worldwide. Its incidence has increased across multiple countries over time, particularly in countries undergoing rapid socioeconomic change, and is expected to more than double by 2030. Although overall survival has improved, mortality from the disease continues to rise, and there is limited evidence that current chemotherapy, hormonal therapy, and radiotherapy significantly improve survival in patients with high-risk, recurrent, or advanced endometrial cancer. These clinical limitations highlight the need for novel therapeutic targets.

The research team, led by Nayeon Kim, Sooyeon Park, and Changwon Yang, selected HEC1A and AN3CA as comparative models because the two lines differ in proliferation rates and tumor aggressiveness and have been widely used to assess changes in cell migration and invasiveness through epithelial-mesenchymal transition marker analyses. Sodium butyrate was applied at concentrations ranging from 0.5 to 10 millimolar, and the cells were evaluated with a battery of assays covering viability, aggregation, migration, apoptosis, mitochondrial membrane potential, cell cycle distribution, and gene and protein expression of pyrimidine metabolism enzymes. All experiments were performed in three independent replicates.

The growth-suppressive effects were clear and dose-dependent. Sodium butyrate reduced cell viability in both cell lines, with HEC1A cells showing significantly greater sensitivity than AN3CA cells. It also suppressed cell aggregation and cell migration in a dose-dependent manner in both lines, again with a greater reduction in migratory capacity in HEC1A cells. The authors note that the pH of the culture medium remained within the physiological range across all tested concentrations and time points and did not differ significantly from untreated controls, indicating that the effects were not attributable to physicochemical changes in the culture conditions. In addition, sodium butyrate did not reduce the viability of normal endometrial cells even at concentrations that markedly suppressed endometrial cancer cell growth.

Epithelial-mesenchymal transition, a process associated with cancer cell invasion, metastasis, and therapeutic resistance, was modulated in a cell line-specific manner. In HEC1A cells, sodium butyrate induced a dose-dependent increase in the epithelial marker E-cadherin while N-cadherin levels decreased, and vimentin remained relatively unchanged. AN3CA cells, which are known to lack E-cadherin expression, showed no detectable E-cadherin after treatment, but N-cadherin decreased in a dose-dependent manner while vimentin was largely unaffected. Because both lines showed dose-dependent suppression of migration and aggregation, the authors conclude that sodium butyrate inhibits motility regardless of baseline epithelial-mesenchymal transition status.

Apoptosis, however, diverged sharply between the two models. In HEC1A cells, the apoptotic cell population rose dose-dependently from 1.7 percent to 20.8 percent, and cleaved PARP, a well-established apoptosis marker, increased relative to full-length PARP. AN3CA cells showed only a modest, statistically insignificant increase in apoptotic cells, from 1.9 percent to 4.6 percent, and no detectable cleaved PARP at any concentration tested. Despite this resistance, mitochondrial membrane depolarization occurred in both lines: at 10 millimolar sodium butyrate, mitochondrial membrane potential fell to 30.4 percent of control in HEC1A cells and to 10.5 percent in AN3CA cells, comparable to the effect of the uncoupler FCCP, and the population of cells with depolarized mitochondria rose from 11.7 percent to 34.4 percent in HEC1A and from 3.4 percent to 44.1 percent in AN3CA. The authors suggest that AN3CA cells may harbor compensatory survival mechanisms that uncouple mitochondrial damage from apoptotic execution, potentially involving death receptor signaling, autophagy, or non-apoptotic pathways such as ferroptosis, noting recent work showing that sodium butyrate can induce ferroptosis in endometrial cancer via the RBM3/SLC7A11 axis.

Cell cycle analysis revealed distinct patterns that matched the apoptotic responses. HEC1A cells showed a marked dose-dependent increase in the subG1 population, consistent with extensive apoptosis, accompanied by decreases in G1, S, and G2/M phases. AN3CA cells showed a moderate subG1 increase with concurrent G1 accumulation and G2/M reduction, suggesting cell cycle arrest. At the transcript level, sodium butyrate downregulated the transcription factors c-Myc, FOXM1, and E2F1, the cyclins CCNB1, CCNA2, CCNE1, and CCND1, and the kinase CDK2 in HEC1A cells. In AN3CA cells, c-Myc, FOXM1, CCNB1, and CCNA2 were downregulated, whereas E2F1, CDK2, CCNE1, and CCND1 were paradoxically upregulated, a pattern the authors interpret as a compensatory transcriptional program supporting survival and cell cycle re-entry.

The most novel finding concerns thymidylate synthase, the enzyme that carries out the key step in de novo dTMP synthesis and the primary target of the chemotherapy drug 5-fluorouracil. Sodium butyrate potently suppressed thymidylate synthase at both mRNA and protein levels in both cell lines in a dose-dependent manner. Thymidine kinase 1, a salvage pathway enzyme, showed the opposite pattern and increased in both lines, while RRM2, DHFR, and DTYMK remained relatively unchanged at the protein level. Because thymidylate synthase inhibition disrupts S-phase progression, induces replication stress, and activates DNA damage checkpoints, the authors argue that this suppression is a prominent molecular event associated with sodium butyrate treatment in endometrial cancer cells and support further preclinical investigation of whether modulation of this pathway may influence fluoropyrimidine response in endometrial cancer models.

Beyond thymidylate synthase, the treatment broadly reprogrammed pyrimidine metabolism. At the mRNA level, the de novo synthesis enzymes CAD, DHODH, and UMPS were decreased in HEC1A, with UMPS showing particularly strong suppression, while in AN3CA, CAD and UMPS were decreased but DHODH was increased. DHFR and RRM2 were downregulated in HEC1A but increased in AN3CA, and MTHFR showed opposite patterns between the lines. In the salvage pathway, TK1 and TYMP were consistently upregulated in both cell lines, whereas UCK2 and UPP1 showed variable responses, and the degradation enzymes DPYS and UPB1 were decreased in both. The authors propose that because DHODH is a mitochondria-dependent step in de novo pyrimidine synthesis, sodium butyrate-induced mitochondrial depolarization may further exacerbate nucleotide depletion by limiting DHODH activity, mechanistically linking mitochondrial dysfunction to pyrimidine insufficiency. The concurrent salvage activation appeared insufficient to prevent viability loss, implying that co-targeting salvage enzymes could further intensify nucleotide stress and enhance efficacy.

The authors are careful about the limits of their work. The effective concentrations of sodium butyrate used in this in vitro study were in the millimolar range, and such concentrations are unlikely to be readily achievable systemically in humans under physiological or standard therapeutic conditions, so the findings should not be interpreted as direct evidence supporting sodium butyrate as a systemic therapeutic agent for endometrial cancer. Instead, the results are viewed as demonstrating the mechanistic potential of sodium butyrate as a metabolic modulator in endometrial cancer cells and as a rationale for further investigation in settings where local delivery or context-specific exposure may be more feasible. Additional non-malignant endometrial cell models will be required to define selectivity more clearly, and in vivo studies will be necessary to determine whether the observed metabolic and molecular effects are recapitulated in a more physiologically relevant tumor environment. The differential responses between HEC1A and AN3CA nevertheless suggest that cellular context shapes the downstream consequences of sodium butyrate exposure, supporting further mechanistic and preclinical studies that consider the molecular and metabolic heterogeneity of endometrial cancer.

Subject of Research: Effects of sodium butyrate on thymidylate synthase and pyrimidine metabolism in endometrial cancer cells

Article Title: Sodium butyrate suppresses endometrial cancer cell growth by inhibiting thymidylate synthase and reprogramming pyrimidine metabolism

Article References: Kim, N., Park, S., & Yang, C. (2026). Sodium butyrate suppresses endometrial cancer cell growth by inhibiting thymidylate synthase and reprogramming pyrimidine metabolism. Genes & Nutrition, 21(1), Article 13. https://doi.org/10.1186/s12263-026-00805-z

Image Credits: AI Generated

DOI: 10.1186/s12263-026-00805-z

Keywords: sodium butyrate, endometrial cancer, thymidylate synthase, pyrimidine metabolism, short-chain fatty acid, gut microbiota, apoptosis, cell cycle, mitochondrial membrane potential, epithelial-mesenchymal transition, in vitro study, HEC1A