ABSTRACT
Downregulation of miR-221-3p expression in prostate cancer (PCa) predicted overall and cancer-specific survival of high-risk PCa patients. Apart from PCa, miR-221-3p expression levels predicted a response to tyrosine kinase inhibitors (TKI) in clear cell renal cell carcinoma (ccRCC) patients. Since this role of miR-221-3p was explained with a specific targeting of VEGFR2, we examined whether miR-221-3p regulated VEGFR2 in PCa. First, we confirmed VEGFR2/KDR as a target gene of miR-221-3p in PCa cells by applying Luciferase reporter assays and Western blotting experiments. Although VEGFR2 was mainly downregulated in the PCa cohort of the TCGA (The Cancer Genome Atlas) database, VEGFR2 was upregulated in our high-risk PCa cohort (n = 142) and predicted clinical progression. In vitro miR-221-3p acted as an escape mechanism from TKI in PC3 cells, as displayed by proliferation and apoptosis assays. Moreover, we confirmed that Sunitinib induced an interferon-related gene signature in PC3 cells by analyzing external microarray data and by demonstrating a significant upregulation of miR-221-3p/miR-222-3p after Sunitinib exposure. Our findings bear a clinical perspective for high-risk PCa patients with low miR-221-3p levels since this could predict a favorable TKI response. Apart from this therapeutic niche, we identified a partially oncogenic function of miR-221-3p as an escape mechanism from VEGFR2 inhibition.
ABSTRACT
The aim was to investigate the induction and repair of radiation-induced DNA double-strand breaks (DSBs) as a function of the absorbed dose to the blood of patients undergoing PET/CT examinations with [68Ga]Ga-PSMA. Blood samples were collected from 15 patients before and at four time points after [68Ga]Ga-PSMA administration, both before and after the PET/CT scan. Absorbed doses to the blood were calculated. In addition, blood samples with/without contrast agent from five volunteers were irradiated ex vivo by CT while measuring the absorbed dose. Leukocytes were isolated, fixed, and stained for co-localizing γ-H2AX+53BP1 DSB foci that were enumerated manually. In vivo, a significant increase in γ-H2AX+53BP1 foci compared to baseline was observed at all time points after administration, although the absorbed dose to the blood by 68Ga was below 4 mGy. Ex vivo, the increase in radiation-induced foci depended on the absorbed dose and the presence of contrast agent, which could have caused a dose enhancement. The CT-dose contribution for the patients was estimated at about 12 mGy using the ex vivo calibration. The additional number of DSB foci induced by CT, however, was comparable to the one induced by 68Ga. The significantly increased foci numbers after [68Ga]Ga-PSMA administration may suggest a possible low-dose hypersensitivity.
ABSTRACT
RATIONALE: High-precision stable isotope measurements in gas-source isotope ratio mass spectrometry are generally carried out by repeated comparison of the composition of an unknown sample with that of a working gas (WG) through a dual-inlet (DI). Due to the established DI protocols, however, most of the sample gas is wasted rather than measured, which is a major problem when sample size is limited. Here we propose a new methodology allowing the measurement of a much larger portion of the available sample. METHODS: We tested a new measurement protocol, the long-integration dual-inlet (LIDI) method, which consists of a single measurement of the sample for 200 to 600 seconds followed by a single measurement of the WG. The isotope ratios of the sample are calculated by comparison of the beam ratios of the WG and sample at equivalent intensities of the major ion beam. RESULTS: Three isotopically very different CO2 samples were analyzed. The LIDI measurements of large samples (50 to 100 µmol of CO2) measured at quasi-constant beam sizes, and of small samples (1.5 to 2 µmol of CO2) measured in micro-volume mode, generated results that are indistinguishable from the standard DI measurements for carbon, oxygen and clumped isotope compositions. The external precision of Δ47 using the LIDI protocol (~±0.007) is similar to that of the state of the art DI measurements. CONCLUSIONS: For traditional and clumped isotope measurements of CO2, the LIDI protocol allows the measurement of a much larger portion of the sample gas rather than only ~20% of it. In addition, the sample can be measured at higher signal intensity and for longer time, allowing the measurement of smaller samples while preserving precision. We suggest that other gases commonly used for stable isotope measurements with gas-source mass spectrometry would also benefit from this new protocol.