In vivo 5-ethynyluridine (EU) labelling detects reduced transcription in Purkinje cell degeneration mouse mutants, but can itself induce neurodegeneration

Fluorescent staining of newly transcribed RNA via metabolic labelling with 5-ethynyluridine (EU) and click chemistry enables visualisation of changes in transcription, such as in conditions of cellular stress. Here, we tested whether EU labelling can be used to examine transcription in vivo in mouse models of nervous system disorders. We show that injection of EU directly into the cerebellum results in reproducible labelling of newly transcribed RNA in cerebellar neurons and glia, with cell type-specific differences in relative labelling intensities, such as Purkinje cells exhibiting the highest levels. We also observed EU-labelling accumulating into cytoplasmic inclusions, indicating that EU, like other modified uridines, may introduce non-physiological properties in labelled RNAs. Additionally, we found that EU induces Purkinje cell degeneration nine days after EU injection, suggesting that EU incorporation not only results in abnormal RNA transcripts, but also eventually becomes neurotoxic in highly transcriptionally-active neurons. However, short post-injection intervals of EU labelling in both a Purkinje cell-specific DNA repair-deficient mouse model and a mouse model of spinocerebellar ataxia 1 revealed reduced transcription in Purkinje cells compared to controls. We combined EU labelling with immunohistology to correlate altered EU staining with pathological markers, such as genotoxic signalling factors. These data indicate that the EU-labelling method provided here can be used to identify changes in transcription in vivo in nervous system disease models

The prognostic value of two different histopathological scoring systems for adrenocortical carcinomas.

AIMS: To compare two different multiparameter histopathological scoring indices and determine their prognostic value in patients presenting with adrenocortical carcinoma (ACC). METHODS AND RESULTS: Seventy-nine adrenal cortical tumours were divided into adenomas (n = 17), non-metastatic carcinomas (n = 24) and carcinomas with metastatic disease and/or local recurrence during follow-up (n = 19) or at time of presentation (n = 19). All cases were scored according to the Weiss revisited index (WRI) and the Van Slooten index (VSI). Both scoring indices yielded a significantly different score (P < 0.005) between adenomas and carcinomas. Non-metastasized carcinomas had a lower score with both indices compared with carcinomas with metastases at the time of presentation (VSI, P = 0.017; WRI, P = 0.019). The VSI also distinguished ACC that had metastasized at any time from those that had not (P = 0.015). Cancer-specific survival in patients with metastasized ACC correlated with the scores for both indices (VSI, P = 0.0078; WRI, P = 0.0025). Time from diagnosis of ACC to development of metastatic disease was correlated with the WRI (P = 0.036, r = -0.350). CONCLUSIONS: The VSI and the WRI have equal validity in the correct categorization of ACC and adenomas. Furthermore, both indices show a correlation with survival for metastasizing ACC

Purkinje-cell-specific DNA repair-deficient mice reveal that dietary restriction protects neurons by cell-intrinsic preservation of genomic health

Dietary restriction (DR) is a universal anti-aging intervention, which reduces age-related nervous system pathologies and neurological decline. The degree to which the neuroprotective effect of DR operates by attenuating cell intrinsic degradative processes rather than influencing non-cell autonomous factors such as glial and vascular health or systemic inflammatory status is incompletely understood. Following up on our finding that DR has a remarkably large beneficial effect on nervous system pathology in whole-body DNA repair-deficient progeroid mice, we show here that DR also exerts strong neuroprotection in mouse models in which a single neuronal cell type, i.e., cerebellar Purkinje cells, experience genotoxic stress and consequent premature aging-like dysfunction. Purkinje cell specific hypomorphic and knock-out ERCC1 mice on DR retained 40 and 25% more neurons, respectively, with equal protection against P53 activation, and alike results from whole-body ERCC1-deficient mice. Our findings show that DR strongly reduces Purkinje cell death in our Purkinje cell-specific accelerated aging mouse model, indicating that DR protects Purkinje cells from intrinsic DNA-damage-driven neurodegeneration

Improved polyvinylpyrrolidone microneedle arrays with non-stoichiometric cyclodextrin

Dissolving polymer microneedles have attracted much attention for their biocompatibility, fast dissolution, and high drug loading. Among them, polyvinylpyrrolidone (PVP) is widely used, but its high water absorption and poor mechanical properties constrain its broad applications. Herein we show that adding cyclodextrin (CD) to form PVP–CD inclusion complexes can alleviate these problems. The water absorption of PVP was reduced by 36–40% at different RHs as the PVP–CD inclusion complexes formed. Attractively, the water absorption at 10 and 20 days remained almost the same for the complexes while it could dramatically increase for the pure PVP samples, particularly in high humidity environments, indicating a possibly longer storage time for the complexes. It was also found that the Young's modulus and hardness of the PVP–CD could be greatly improved, especially for low molecular weight PVP. Furthermore, the glass transition temperature (Tg) of the PVP–CD increased by up to 39 °C. With the improved properties, the fabricated PVP–CD microneedles possessed much sharper needle tips and the patch had less cracks than those made from pure PVP. Pig skin application results suggested that the PVP–CD microneedle arrays were able to reliably pierce the stratum corneum of the skin while it was not achievable for the PVP microneedles with the same geometry. We anticipate that these PVP–CD complex microneedles are more suitable for vaccine and drug delivery because of their superior properties

A Phase i Study of an MPS1 Inhibitor (BAY 1217389) in Combination with Paclitaxel Using a Novel Randomized Continual Reassessment Method for Dose Escalation

Purpose: Monopolar spindle 1 (MPS1) kinase inhibitor, BAY 1217389 (BAY) synergizes with paclitaxel. This phase I study assessed the combination of BAY with paclitaxel using a novel randomized continuous reassessment method (rCRM) to improve dose determination. Patients and Methods: Patients with solid tumors were randomized to receive oralBAY(twice daily 2-days-on/5-days-off) withweekly paclitaxel (90 mg/m2) or paclitaxel monotherapy in cycle 1. Dose escalation was guided by CRM modeling. Primary objectives were to assess safety, establish the MTD of BAY, and to evaluate the pharmacokinetic profiles for both compounds. Simulations were performed to determine the contribution of the rCRM for dose determination. Results: In total, 75 patients were enrolled. The main doselimiting toxicities were hematologic toxicities (55.6%). The MTD of BAY was established at 64 mg twice daily with paclitaxel. Inclusion of a control arm enabled the definitive attribution of grade ≥3 neutropenia to higher BAY exposure [AUC0-12 (P< 0.001)]. After determining the MTD, we included 19 patients with breast cancer at this dose for dose expansion. Other common toxicities were nausea (45.3%), fatigue (41.3%), and diarrhea (40.0%). Overall confirmed responses were seen in 31.6% of evaluable patients. Simulations showed that rCRM outperforms traditional designs in determining the true MTD. Conclusions: The combination of BAY with paclitaxel was associated with considerable toxicity without a therapeutic window. However, the use of the rCRM design enabled us to determine the exposure-toxicity relation for BAY. Therefore, we propose that the rCRM could improve dose determination in phase I trials that combine agents with overlapping toxicities. _2021 American Association for Cancer Research