Metallaphotoredox aryl and alkyl radiomethylation for PET ligand discovery.

Positron emission tomography (PET) radioligands (radioactively labelled tracer compounds) are extremely useful for in vivo characterization of central nervous system drug candidates, neurodegenerative diseases and numerous oncology targets1. Both tritium and carbon-11 radioisotopologues are generally necessary for in vitro and in vivo characterization of radioligands2, yet there exist few radiolabelling protocols for the synthesis of either, inhibiting the development of PET radioligands. The synthesis of such radioligands also needs to be very rapid owing to the short half-life of carbon-11. Here we report a versatile and rapid metallaphotoredox-catalysed method for late-stage installation of both tritium and carbon-11 into the desired compounds via methylation of pharmaceutical precursors bearing aryl and alkyl bromides. Methyl groups are among the most prevalent structural elements found in bioactive molecules, and so this synthetic approach simplifies the discovery of radioligands. To demonstrate the breadth of applicability of this technique, we perform rapid synthesis of 20 tritiated and 10 carbon-11-labelled complex pharmaceuticals and PET radioligands, including a one-step radiosynthesis of the clinically used compounds [11C]UCB-J and [11C]PHNO. We further outline the direct utility of this protocol for preclinical PET imaging and its translation to automated radiosynthesis for routine radiotracer production in human clinical imaging. We also demonstrate this protocol for the installation of other diverse and pharmaceutically useful isotopes, including carbon-14, carbon-13 and deuterium.

Total-body imaging of mu-opioid receptors with [11C]carfentanil in non-human primates.

PURPOSE: Mu-opioid receptors (MORs) are widely expressed in the central nervous system (CNS), peripheral organs, and immune system. This study measured the whole body distribution of MORs in rhesus macaques using the MOR selective radioligand [11C]carfentanil ([11C]CFN) on the PennPET Explorer. Both baseline and blocking studies were conducted using either naloxone or GSK1521498 to measure the effect of the antagonists on MOR binding in both CNS and peripheral organs. METHODS: The PennPET Explorer was used for MOR total-body PET imaging in four rhesus macaques using [11C]CFN under baseline, naloxone pretreatment, and naloxone or GSK1521498 displacement conditions. Logan distribution volume ratio (DVR) was calculated by using a reference model to quantitate brain regions, and the standard uptake value ratios (SUVRs) were calculated for peripheral organs. The percent receptor occupancy (%RO) was calculated to establish the blocking effect of 0.14 mg/kg naloxone or GSK1521498. RESULTS: The %RO in MOR-abundant brain regions was 75-90% for naloxone and 72-84% for GSK1521498 in blocking studies. A higher than 90% of %RO were observed in cervical spinal cord for both naloxone and GSK1521498. It took approximately 4-6 min for naloxone or GSK1521498 to distribute to CNS and displace [11C]CFN from the MOR. A smaller effect was observed in heart wall in the naloxone and GSK1521498 blocking studies. CONCLUSION: [11C]CFN total-body PET scans could be a useful approach for studying mechanism of action of MOR drugs used in the treatment of acute and chronic opioid use disorder and their effect on the biodistribution of synthetic opioids such as CFN. GSK1521498 could be a potential naloxone alternative to reverse opioid overdose.

Implementing the FAIR Data Principles in precision oncology: review of supporting initiatives.

Compelling research has recently shown that cancer is so heterogeneous that single research centres cannot produce enough data to fit prognostic and predictive models of sufficient accuracy. Data sharing in precision oncology is therefore of utmost importance. The Findable, Accessible, Interoperable and Reusable (FAIR) Data Principles have been developed to define good practices in data sharing. Motivated by the ambition of applying the FAIR Data Principles to our own clinical precision oncology implementations and research, we have performed a systematic literature review of potentially relevant initiatives. For clinical data, we suggest using the Genomic Data Commons model as a reference as it provides a field-tested and well-documented solution. Regarding classification of diagnosis, morphology and topography and drugs, we chose to follow the World Health Organization standards, i.e. ICD10, ICD-O-3 and Anatomical Therapeutic Chemical classifications, respectively. For the bioinformatics pipeline, the Genome Analysis ToolKit Best Practices using Docker containers offer a coherent solution and have therefore been selected. Regarding the naming of variants, we follow the Human Genome Variation Society's standard. For the IT infrastructure, we have built a centralized solution to participate in data sharing through federated solutions such as the Beacon Networks.

PolyGA targets the ER stress-adaptive response by impairing GRP75 function at the MAM in C9ORF72-ALS/FTD.

ER stress signaling is linked to the pathophysiological and clinical disease manifestations in amyotrophic lateral sclerosis (ALS). Here, we have investigated ER stress-induced adaptive mechanisms in C9ORF72-ALS/FTD, focusing on uncovering early endogenous neuroprotective mechanisms and the crosstalk between pathological and adaptive responses in disease onset and progression. We provide evidence for the early onset of ER stress-mediated adaptive response in C9ORF72 patient-derived motoneurons (MNs), reflected by the elevated increase in GRP75 expression. These transiently increased GRP75 levels enhance ER-mitochondrial association, boosting mitochondrial function and sustaining cellular bioenergetics during the initial stage of disease, thereby counteracting early mitochondrial deficits. In C9orf72 rodent neurons, an abrupt reduction in GRP75 expression coincided with the onset of UPR, mitochondrial dysfunction and the emergence of PolyGA aggregates, which co-localize with GRP75. Similarly, the overexpression of PolyGA in WT cortical neurons or C9ORF72 patient-derived MNs led to the sequestration of GRP75 within PolyGA inclusions, resulting in mitochondrial calcium (Ca2+) uptake impairments. Corroborating these findings, we found that PolyGA aggregate-bearing human post-mortem C9ORF72 hippocampal dentate gyrus neurons not only display reduced expression of GRP75 but also exhibit GRP75 sequestration within inclusions. Sustaining high GRP75 expression in spinal C9orf72 rodent MNs specifically prevented ER stress, normalized mitochondrial function, abrogated PolyGA accumulation in spinal MNs, and ameliorated ALS-associated behavioral phenotype. Taken together, our results are in line with the notion that neurons in C9ORF72-ALS/FTD are particularly susceptible to ER-mitochondrial dysfunction and that GRP75 serves as a critical endogenous neuroprotective factor. This neuroprotective pathway, is eventually targeted by PolyGA, leading to GRP75 sequestration, and its subsequent loss of function at the MAM, compromising mitochondrial function and promoting disease onset.

Longitudinal minimal residual disease assessment in multiple myeloma patients in complete remission - results from the NMSG flow-MRD substudy within the EMN02/HO95 MM trial.

BACKGROUND: Multiple myeloma remains an incurable disease with multiple relapses due to residual myeloma cells in the bone marrow of patients after therapy. Presence of small number of cancer cells in the body after cancer treatment, called minimal residual disease, has been shown to be prognostic for progression-free and overall survival. However, for multiple myeloma, it is unclear whether patients attaining minimal residual disease negativity may be candidates for treatment discontinuation. We investigated, if longitudinal flow cytometry-based monitoring of minimal residual disease (flow-MRD) may predict disease progression earlier and with higher sensitivity compared to biochemical assessments. METHODS: Patients from the Nordic countries with newly diagnosed multiple myeloma enrolled in the European-Myeloma-Network-02/Hovon-95 (EMN02/HO95) trial and undergoing bone marrow aspiration confirmation of complete response, were eligible for this Nordic Myeloma Study Group (NMSG) substudy. Longitdudinal flow-MRD assessment of bone marrow samples was performed to identify and enumerate residual malignant plasma cells until observed clinical progression. RESULTS: Minimal residual disease dynamics were compared to biochemically assessed changes in serum free light chain and M-component. Among 20 patients, reaching complete response or stringent complete response during the observation period, and with ≥3 sequential flow-MRD assessments analysed over time, increasing levels of minimal residual disease in the bone marrow were observed in six cases, preceding biochemically assessed disease and clinical progression by 5.5 months and 12.6 months (mean values), respectively. Mean malignant plasma cells doubling time for the six patients was 1.8 months (95% CI, 1.4-2.3 months). Minimal malignant plasma cells detection limit was 4 × 10-5. CONCLUSIONS: Flow-MRD is a sensitive method for longitudinal monitoring of minimal residual disease dynamics in multiple myeloma patients in complete response. Increasing minimal residual disease levels precedes biochemically assessed changes and is an early indicator of subsequent clinical progression. TRIAL REGISTRATION: NCT01208766.

Optical Probing of Crystal Lattice Configurations in Single CsPbBr3 Nanoplatelets.

Quantum-confined nanostructures of CsPbBr3 with luminescence quantum efficiencies approaching unity have shown tremendous potential for lighting and quantum light applications. In contrast to CsPbBr3 quantum dots, where the fine structure of the emissive exciton state has been intensely discussed, the relationship among lattice orientation, shape anisotropy, and exciton fine structure in lead halide nanoplatelets has not yet been established. In this work, we investigate the fine structure of the bright triplet exciton of individual CsPbBr3 nanoplatelets by polarization-resolved micro- and magnetophotoluminescence spectroscopy at liquid helium temperature and find a large zero-field splitting of up to 2.5 meV. A unique relation between the crystal structure and the photoluminescence emission confirms the existence of two distinct crystal configurations in such nanoplatelets with different alignments of the crystal axes with respect to the nanoplatelet facets. Polarization-resolved experiments eventually allow us to determine the absolute orientation of an individual nanoplatelet on the substrate purely by optical means.

Massively parallel gene expression variation measurement of a synonymous codon library.

BACKGROUND: Cell-to-cell variation in gene expression strongly affects population behavior and is key to multiple biological processes. While codon usage is known to affect ensemble gene expression, how codon usage influences variation in gene expression between single cells is not well understood. RESULTS: Here, we used a Sort-seq based massively parallel strategy to quantify gene expression variation from a green fluorescent protein (GFP) library containing synonymous codons in Escherichia coli. We found that sequences containing codons with higher tRNA Adaptation Index (TAI) scores, and higher codon adaptation index (CAI) scores, have higher GFP variance. This trend is not observed for codons with high Normalized Translation Efficiency Index (nTE) scores nor from the free energy of folding of the mRNA secondary structure. GFP noise, or squared coefficient of variance (CV2), scales with mean protein abundance for low-abundant proteins but does not change at high mean protein abundance. CONCLUSIONS: Our results suggest that the main source of noise for high-abundance proteins is likely not originating at translation elongation. Additionally, the drastic change in mean protein abundance with small changes in protein noise seen from our library implies that codon optimization can be performed without concerning gene expression noise for biotechnology applications.

Dynamic control in metabolic engineering: Theories, tools, and applications.

Metabolic engineering has allowed the production of a diverse number of valuable chemicals using microbial organisms. Many biological challenges for improving bio-production exist which limit performance and slow the commercialization of metabolically engineered systems. Dynamic metabolic engineering is a rapidly developing field that seeks to address these challenges through the design of genetically encoded metabolic control systems which allow cells to autonomously adjust their flux in response to their external and internal metabolic state. This review first discusses theoretical works which provide mechanistic insights and design choices for dynamic control systems including two-stage, continuous, and population behavior control strategies. Next, we summarize molecular mechanisms for various sensors and actuators which enable dynamic metabolic control in microbial systems. Finally, important applications of dynamic control to the production of several metabolite products are highlighted, including fatty acids, aromatics, and terpene compounds. Altogether, this review provides a comprehensive overview of the progress, advances, and prospects in the design of dynamic control systems for improved titer, rate, and yield metrics in metabolic engineering.

Correcting time-intensity curves in dynamic contrast-enhanced breast MRI for inhomogeneous excitation fields at 7T.

PURPOSE: Inhomogeneous excitation at ultrahigh field strengths (7T and above) compromises the reliability of quantified dynamic contrast-enhanced breast MRI. This can hamper the introduction of ultrahigh field MRI into the clinic. Compensation for this non-uniformity effect can consist of both hardware improvements and post-acquisition corrections. This paper investigated the correctable radiofrequency transmit ( B1+ ) range post-acquisition in both simulations and patient data for 7T MRI. METHODS: Simulations were conducted to determine the minimum B1+ level at which corrections were still beneficial because of noise amplification. Two correction strategies leading to differences in noise amplification were tested. The effect of the corrections on a 7T patient data set (N = 38) with a wide range of B1+ levels was investigated in terms of time-intensity curve types as well as washin, washout and peak enhancement values. RESULTS: In simulations assuming a common amount of T1 saturation, the lowest B1+ level at which the SNR of the corrected images was at least that of the original precontrast image was 43% of the nominal angle. After correction, time-intensity curve types changed in 24% of included patients, and the distribution of curve types corresponded better to the distribution found in literature. Additionally, the overlap between the distributions of washin, washout, and peak enhancement values for grade 1 and grade 2 tumors was slightly reduced. CONCLUSION: Although the correctable range varies with the amount of T1 saturation, post-acquisition correction for inhomogeneous excitation was feasible down to B1+ levels of 43% of the nominal angle in vivo.

Shortening of apparent transverse relaxation time of inorganic phosphate as a breast cancer biomarker.

Phosphorus MRS offers a non-invasive tool for monitoring cell energy and phospholipid metabolism and can be of additional value in diagnosing cancer and monitoring cancer therapy. In this study, we determined the transverse relaxation times of a number of phosphorous metabolites in a group of breast cancer patients by adiabatic multi-echo spectroscopic imaging at 7 T. The transverse relaxation times of phosphoethanolamine, phosphocholine, inorganic phosphate (Pi ), glycerophosphocholine and glycerophosphatidylcholine were 184 ± 8 ms, 203 ± 17 ms, 87 ± 8 ms, 240 ± 56 ms and 20 ± 10 ms, respectively. The transverse relaxation time of Pi in breast cancer tissue was less than half that of healthy fibroglandular tissue. This effect is most likely caused by an up-regulation of glycolysis in breast cancer tissue that leads to interaction of Pi with the GAPDH enzyme, which forms part of the reversible pathway of exchange of Pi with gamma-adenosine tri-phosphate, thus shortening its apparent transverse relaxation time. As healthy breast tissue shows very little glycolytic activity, the apparent T2 shortening of Pi due to malignant transformation could possibly be used as a biomarker for cancer.

Convenient Access to Chiral Cyclobutanes with Three Contiguous Stereocenters from Verbenone by Directed C(sp3 )-H arylation.

This work demonstrates how a series of complex, chiral cyclobutane derivatives can be accessed in four steps from the terpene verbenone through the application of a directed C-H functionalization approach. The developed synthetic route involved an 8-aminoquinoline-directed C(sp3 )-H arylation as the key step, and this reaction could be carried out with a wide range of aryl and heteroaryl iodides to furnish a variety of cyclobutane products with three contiguous stereocenters. Moreover, it was shown that the 8-aminoquinoline auxiliary could be effectively removed from the cyclobutane derivatives using an ozonolysis-based cleavage method.

Synthesis and evaluation of an AZD2461 [18F]PET probe in non-human primates reveals the PARP-1 inhibitor to be non-blood-brain barrier penetrant.

Poly(ADP-ribose)polymerase-1 inhibitor (PARPi) AZD2461 was designed to be a weak P-glycoprotein (P-gp) analogue of FDA approved olaparib. With this chemical property in mind, we utilized the AZD2461 ligand architecture to develop a CNS penetrant and PARP-1 selective imaging probe, in order to investigate PARP-1 mediated neuroinflammation and neurodegenerative diseases, such as Alzheimer's and Parkinson's. Our work led to the identification of several high-affinity PARPi, including AZD2461 congener 9e (PARP-1 IC50 = 3.9 ±â€¯1.2 nM), which was further evaluated as a potential 18F-PET brain imaging probe. However, despite the similar molecular scaffolds of 9e and AZD2461, our studies revealed non-appreciable brain-uptake of [18F]9e in non-human primates, suggesting AZD2461 to be non-CNS penetrant.

Current-Induced Magnetic Polarons in a Colloidal Quantum-Dot Device.

Electrical spin manipulation remains a central challenge for the realization of diverse spin-based information processing technologies. Motivated by the demonstration of confinement-enhanced sp-d exchange interactions in colloidal diluted magnetic semiconductor (DMS) quantum dots (QDs), such materials are considered promising candidates for future spintronic or spin-photonic applications. Despite intense research into DMS QDs, electrical control of their magnetic and magneto-optical properties remains a daunting goal. Here, we report the first demonstration of electrically induced magnetic polaron formation in any DMS, achieved by embedding Mn2+-doped CdSe/CdS core/shell QDs as the active layer in an electrical light-emitting device. Tracing the electroluminescence from cryogenic to room temperatures reveals an anomalous energy shift that reflects current-induced magnetization of the Mn2+ spin sublattice, that is, excitonic magnetic polaron formation. These electrically induced magnetic polarons exhibit an energy gain comparable to their optically excited counterparts, demonstrating that magnetic polaron formation is achievable by current injection in a solid-state device.

The myeloma stem cell concept, revisited: from phenomenology to operational terms.

The concept of the myeloma stem cell may have important therapeutic implications, yet its demonstration has been hampered by a lack of consistency in terms and definitions. Here, we summarize the current documentation and propose single-cell in vitro studies for future translational studies. By the classical approach, a CD19-/CD45low/-/CD38high/CD138+ malignant plasma cell, but not the CD19+/CD38low/- memory B cell compartment, is enriched for tumorigenic cells that initiate myeloma in xenografted immunodeficient mice, supporting that myeloma stem cells are present in the malignant PC compartment. Using a new approach, analysis of c-DNA libraries from CD19+/CD27+/CD38- single cells has identified clonotypic memory B cell, suggested to be the cell of origin. This is consistent with multiple myeloma being a multistep hierarchical process before or during clinical presentation. We anticipate that further characterization will require single cell geno- and phenotyping combined with clonogenic assays. To implement such technologies, we propose a revision of the concept of a myeloma stem cell by including operational in vitro assays to describe the cellular components of origin, initiation, maintenance, and evolution of multiple myeloma. These terms are in accordance with recent (2012) consensus statements on the definitions, assays, and nomenclature of cancer stem cells, which is technically precise without completely abolishing established terminology. We expect that this operational model will be useful for future reporting of parameters used to identify and characterize the multiple myeloma stem cells. We strongly recommend that these parameters include validated standard technologies, reproducible assays, and, most importantly, supervised prospective sampling of selected biomaterial which reflects clinical stages, disease spectrum, and therapeutic outcome. This framework is key to the characterization of the cellular architecture of multiple myeloma and its use in precision medicine.

Identification of gene knockdown targets conferring enhanced isobutanol and 1-butanol tolerance to Saccharomyces cerevisiae using a tunable RNAi screening approach.

Improving yeast tolerance to 1-butanol and isobutanol is a step toward enabling high-titer production. To identify previously unknown genetic targets leading to increased tolerance, we establish a tunable RNA interference (RNAi) screening approach. Specifically, we optimized the efficiency and tunability of RNA interference library screening in yeast, ultimately enabling downregulation efficiencies from 0 to 94 %. Using this system, we identified the Hsp70 family as a key regulator of isobutanol tolerance in a single round of screening, with downregulation of these genes conferring up to 64 % increased growth in 12 g/L isobutanol. For 1-butanol, we find through two rounds of iterative screening that the combined downregulation of alcohol dehydrogenase and enolase improves growth up to 3100 % in 10 g/L 1-butanol. Collectively, this work improves the tunability of RNAi in yeast as demonstrated by the discovery of novel effectors for these complex phenotypes.

Design and Characterization of a Three-Axis Hall Effect-Based Soft Skin Sensor.

This paper presents an easy means to produce a 3-axis Hall effect-based skin sensor for robotic applications. It uses an off-the-shelf chip and is physically small and provides digital output. Furthermore, the sensor has a soft exterior for safe interactions with the environment; in particular it uses soft silicone with about an 8 mm thickness. Tests were performed to evaluate the drift due to temperature changes, and a compensation using the integral temperature sensor was implemented. Furthermore, the hysteresis and the crosstalk between the 3-axis measurements were evaluated. The sensor is able to detect minimal forces of about 1 gf. The sensor was calibrated and results with total forces up to 1450 gf in the normal and tangential directions of the sensor are presented. The test revealed that the sensor is able to measure the different components of the force vector.

Improving IMES Localization Accuracy by Integrating Dead Reckoning Information.

Indoor positioning remains an open problem, because it is difficult to achieve satisfactory accuracy within an indoor environment using current radio-based localization technology. In this study, we investigate the use of Indoor Messaging System (IMES) radio for high-accuracy indoor positioning. A hybrid positioning method combining IMES radio strength information and pedestrian dead reckoning information is proposed in order to improve IMES localization accuracy. For understanding the carrier noise ratio versus distance relation for IMES radio, the signal propagation of IMES radio is modeled and identified. Then, trilateration and extended Kalman filtering methods using the radio propagation model are developed for position estimation. These methods are evaluated through robot localization and pedestrian localization experiments. The experimental results show that the proposed hybrid positioning method achieved average estimation errors of 217 and 1846 mm in robot localization and pedestrian localization, respectively. In addition, in order to examine the reason for the positioning accuracy of pedestrian localization being much lower than that of robot localization, the influence of the human body on the radio propagation is experimentally evaluated. The result suggests that the influence of the human body can be modeled.

Predicting response to multidrug regimens in cancer patients using cell line experiments and regularised regression models.

BACKGROUND: Patients suffering from cancer are often treated with a range of chemotherapeutic agents, but the treatment efficacy varies greatly between patients. Based on recent popularisation of regularised regression models the goal of this study was to establish workflows for pharmacogenomic predictors of response to standard multidrug regimens using baseline gene expression data and origin specific cell lines. The proposed workflows are tested on diffuse large B-cell lymphoma treated with R-CHOP first-line therapy. METHODS: First, B-cell cancer cell lines were tested successively for resistance towards the chemotherapeutic components of R-CHOP: cyclophosphamide (C), doxorubicin (H), and vincristine (O). Second, baseline gene expression data were obtained for each cell line before treatment. Third, regularised multivariate regression models with cross-validated tuning parameters were used to generate classifier and predictor based resistance gene signatures (REGS) for the combination and individual chemotherapeutic drugs C, H, and O. Fourth, each developed REGS was used to assign resistance levels to individual patients in three clinical cohorts. RESULTS: Both classifier and predictor based REGS, for the combination CHO, were of prognostic value. For patients classified as resistant towards CHO the risk of progression was 2.33 (95% CI: 1.6, 3.3) times greater than for those classified as sensitive. Similarly, an increase in the predicted CHO resistance index of 10 was related to a 22% (9%, 36%) increased risk of progression. Furthermore, the REGS classifier performed significantly better than the REGS predictor. CONCLUSIONS: The regularised multivariate regression models provide a flexible workflow for drug resistance studies with promising potential. However, the gene expressions defining the REGSs should be functionally validated and correlated to known biomarkers to improve understanding of molecular mechanisms of drug resistance.

Insights into Structure-Activity Relationships of Bacterial RNA Polymerase Inhibiting Corallopyronin Derivatives.

The new compound precorallopyronin A is a stable precursor in the biosynthesis of the antibiotic corallopyronin A. This natural product was isolated from the producer strain Corallococcus coralloides B035. Together with various semisynthetically obtained corallopyronin A derivatives its antibacterial effects were evaluated. In combination with an X-ray crystallization model limitations of derivatization possibilities were revealed. The antibiotic potential of the novel precorallopyronin A is comparable to that of the structurally more complex corallopyronin A, which highlights that the additional chiral center is not essential for activity.