Environmental Pollutants and AHR pathway in Uterine Leiomyoma

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Uterine leiomyomas (LM, fibroids) disrupt uterine function and cause recurrent pregnancy loss, excessive uterine bleeding, and anemia in 15-30% of reproductive-age women. No long-term medical treatment is available. Understanding how a LM develops is essential for identifying new non-surgical treatments. MED12 mutations (mut-MED12) occur in 70% of all LM and drive LM growth in a steroid hormone-dependent manner. Pre- and peri-menopausal women are widely exposed to endocrine disrupting chemicals (EDCs), e.g., phthalates, which are found in many consumer products and associated with a number of reproductive diseases. How a mut- MED12 influences LM growth and whether this process is enhanced by exposure to EDCs remain unknown. We recently found that tryptophan (Trp) 2,3-dioxygenase (TDO2) gene expression is strikingly upregulated in mut- but not wild-type (wt)-MED12 LM. TDO2 catalyzes a critical step in Trp breakdown to kynurenine (Kyn), an endogenous ligand for the aryl hydrocarbon receptor (AHR) whose activation stimulates the expression of pro- growth genes to promote cell survival and proliferation in various tissues. Kyn levels are markedly higher in LM vs myometrium, particularly in mut-MED12 LM. Trp and Kyn treatments of LM cells activated AHR and increased cell survival; blocking the Trp-Kyn-AHR pathway by siRNA knockdown or inhibitors of TDO2 or AHR abolished these effects, with mutant LM cells showing higher sensitivity to the treatments. Epidemiological studies have shown positive associations between LM and exposure to di(2-ethylhexyl) phthalate (DEHP). In vitro, mono(2- ethyl-5-hydroxyhexyl) phthalate (MEHHP), a major metabolite of DEHP, stimulated the expression of Trp transporters (LAT1 and LAT2), increased Trp uptake and Kyn production, activated AHR, and promoted LM cell survival. In addition, progesterone receptor is crucial for the expression of AHR and its nuclear translocator ARNT. Thus, the Trp-Kyn-AHR pathway appears to be a hub at which mut-MED12, steroid hormone action, and EDC effects converge enabling LM growth. Our overall hypothesis is that increased Trp uptake and metabolism, Kyn production, and AHR pathway activation, as a result of mut-MED12 or high exposure to the environmental pollutant DEHP, promote cell survival and proliferation and lead to LM growth. Using a xenograft mouse model and genome-wide studies, we will test our hypothesis in the following Aims: (1) Define the functional role of the Trp-Kyn-AHR pathway in LM tumorigenesis. Hypothesis: elevated expression of the TDO2 enzyme in mut- MED12 LM causes Kyn overproduction that activates AHR and promotes proliferation and survival of smooth muscle cells and tumor growth. (2) Determine whether DEHP stimulates LM growth via activation of the Trp- Kyn-AHR pathway. Hypothesis: exposure to DEHP and its metabolite MEHHP upregulates the expression of Trp transporters to increase its uptake, resulting in increased Kyn production and AHR activation leading to LM cell survival and tumor growth. The study will link, for the first time, abnormal amino acid metabolism and LM growth, opening a new avenue for translational research and the development of novel therapeutics for LM.