The Impact Of Testicular Nuclear Receptor4on Chemo-sensitivity Of Docetaxel In Prostate Cancer

Background:Castration resistant prostate cancer (CRPC) is a top problem in clinical treatment and basic research. Docetaxel (DTX) is the recommended option of chemotherapeutic agent for CRPC. By interacting with β-tubulin, docetaxel breaks the normal process of mitosis, causing tumor cells to arrest in G2/M phase and eventually leads to cell apoptosis. Unfortunately, due to acquired or hereditary drug resistance, the chemo-sensitivity of docetaxel in these patients will gradually decrease.Coded by NR2C2gene, testicular nuclear receptor4(TR4) is a member of the nuclear receptor superfamily. As an important transcription factor, TR4can interact with several nuclear receptors. Via its various down-stream target genes, TR4plays an important role in intracellular ROS, oxidative stress resistance and DNA damage reparation. In vivo studies found that TR4gene knockout mice (TR4-/-) display impaired oxidative stress defense and DNA repair function. These functions are closely related with cancers. Our pre-experiments also showed that after being treated with docetaxel, PC3cell showed a higher TR4expression level. Which indicated that TR4might be related to prostate cancer, however, few studies have mentioned the relationship between TR4and prostate cancer.Objective:This study aimed to clarify the relationship between TR4and chemo-sensitivity of castration resistant prostate cancer at cell as well as clinical level and find out its possible mechanism.Methods:1. Using lentivirus infection, we were able to establish PCa cell lines with different TR4expression level. IC50assay was carried out using MTT experiment.2. Cell viability assay was carried out using MTT experiment to assess the long term effect of docetaxel on TR4-modified PC3cells.3. Use Annexin V-FITC/PI double staining to evaluate the influence of TR4on cell apoptosis.4. To confirm the results of cell experiments, we collected30biopsy samples from CRPC patients who were treated with docetaxel-based therapy to perform immunohistochemistry.Results:1. We successfully established2cell lines:TR4knockdown cell line (PC3scr/siTR4) and TR4overexpress cell line (PC3vector/TR4). MTT assay showed Knocking-down TR4in PC3cells led to a lower IC50dose while overexpression of TR4in PC3led to a higher IC50dose.2. MTT assay showed that knocking-down TR4in PC3cells led to poorer cell viability and overexpression of TR4in PC3led to better cell viability.3. Flow cytometer results revealed that under the influence of docetaxel, knocking-down TR4in PC3cells led to more cell apoptosis whereas overexpression of TR4in PC3led to less cell apoptosis.4. IHC results showed TR4expression inversely correlates with the outcome of chemotherapy in CRPC patients. Patients who failed to achieve PSA response (<50%PSA reduction from baseline) after docetaxel-based chemotherapy had a comparatively higher TR4expression than those who achieved PSA response (≥50%PSA reduction from baseline, last for8weeks).Conclusion:TR4enhances chemo-resistance of docetaxel in CRPC. It may serve as a biomarker to determine the prognosis of docetaxel-based therapy and becoming a potential therapy target to combine with docetaxel to better suppress CRPC.