NanoLuc Binary Technology as a methodological approach: an important new tool for studying the localization of androgen receptor and androgen receptor splice variant V7 homo and heterodimers.

BACKGROUND: The androgen/androgen receptor (AR)-signaling axis plays a central role in prostate cancer (PCa). Upon androgen-binding the AR dimerizes with another AR, and translocates into the nucleus where the AR-dimer activates/inactivates androgen-dependent genes. Consequently, treatments for PCa are commonly based on androgen deprivation therapy (ADT). The clinical benefits of ADT are only transitory and most tumors develop mechanisms allowing the AR to bypass its need for physiological levels of circulating androgens. Clinical failure of ADT is often characterized by the synthesis of a constitutively active AR splice variant, termed AR-V7. AR-V7 mRNA expression is considered as a resistance mechanism following ADT. AR-V7 no longer needs androgenic stimuli for nuclear entry and/or dimerization. METHODS: Our goal was to mechanistically decipher the interaction between full-length AR (AR-FL) and AR-V7 in AR-null HEK-293 cells using the NanoLuc Binary Technology under androgen stimulation and deprivation conditions. RESULTS: Our data point toward a hypothesis that AR-FL/AR-FL homodimers form in the cytoplasm, whereas AR-V7/AR-V7 homodimers localize in the nucleus. However, after androgen stimulation, all the AR-FL/AR-FL, AR-FL/AR-V7 and AR-V7/AR-V7 dimers were localized in the nucleus. CONCLUSIONS: We showed that AR-FL and AR-V7 form heterodimers that localize to the nucleus, whereas AR-V7/AR-V7 dimers were found to localize in the absence of androgens in the nucleus.

Combined miR-486 and GP88 (Progranulin) Serum Levels Are Suggested as Supportive Biomarkers for Therapy Decision in Elderly Prostate Cancer Patients.

Our study aimed to assess the applicability of miR-486 in combination with soluble GP88 protein as a diagnostic and/or predictive biomarker for prostate cancer (PCa) patients. miR-486 and GP88 levels in serum samples from 136 patients undergoing MRI-guided biopsy of the prostate were assessed by qRT−PCR and ELISA, respectively. Of these, 86 patients received a histologically confirmed diagnosis of PCa. Neither marker showed an association with the diagnosis of cancer. PCa patients were separated based on (i) treatment into patients with active surveillance or patients with any type of curative treatment and (ii) age into elderly (>68 years) patients and younger patients (≤68 years). In elderly patients (N = 41) with the intention of curative treatment at optimized cut-off values, significantly higher GP88 levels (p = 0.018) and lower miR-486 levels (p = 0.014) were observed. The total PSA level and ISUP biopsy grade were used in a baseline model for predicting definitive therapy. The baseline model exhibited an area under the curve (AUC) of 0.783 (p = 0.005). The addition of the serum biomarkers miR-486 and GP88 to the baseline model yielded an improved model with an AUC of 0.808 (p = 0.002). Altogether, combined miR-486 and GP88 serum levels are associated with and are therefore suggested as supportive biomarkers for therapy decisions, particularly in elderly PCa patients.

Analysis of CXCL9, PD1 and PD-L1 mRNA in Stage T1 Non-Muscle Invasive Bladder Cancer and Their Association with Prognosis.

Non-muscle invasive bladder cancer (NMIBC), which is characterized by a recurrence rate of approximately 30% and very long treatment times, remains a major unresolved problem for patients and the health care system. The immunological interplay between tumor cells and the immune environment is important for tumor development. Therefore, we analyzed the mRNA of three immune markers, CXCL9, PD1 and PD-L1, in NMIBC by qRT-PCR. The results were subsequently correlated with clinicopathological parameters and prognostic data. Altogether, as expected, higher age was an independent prognostic factor for overall survival (OS) and disease-specific survival (DSS), but not for recurrence-free survival (RFS). Lower CXCL9 mRNA was observed in multivariate Cox's regression analysis to be an independent prognostic parameter for reduced OS (relative risk; RR = 2.08; p = 0.049), DSS (RR = 4.49; p = 0.006) and RFS (RR = 2.69; p = 0.005). In addition, PD-L1 mRNA was an independent prognostic factor for DSS (RR = 5.02; p = 0.042) and RFS (RR = 2.07; p = 0.044). Moreover, in univariate Cox's regression analysis, the stratification of patients revealed that low CXCL9 or low PD1 mRNA was associated with reduced RFS in the younger patient group (≤71 years), but not in the older patient group (>71 years). In addition, low CXCL9 or low PD-L1 was associated with shorter RFS in patients with higher tumor cell proliferation and in patients without instillation therapy. In conclusion, the characterization of mRNA levels of immune markers differentiates NIMBC patients with respect to prognosis.