Publications search

Tumor environment influences anticancer therapy response but which extracellular nutrients affect drug sensitivity is largely unknown. Using functional genomics, we determine modifiers of l-asparaginase (ASNase) response and identify thiamine pyrophosphate kinase 1 as a metabolic dependency under ASNase treatment. While thiamine is generally not limiting for cell proliferation, a DNA-barcode competition assay identifies leukemia cell lines that grow suboptimally under low thiamine and are characterized by low expression of solute carrier family 19 member 2 (SLC19A2), a thiamine transporter. SLC19A2 is necessary for optimal growth and ASNase resistance, when standard medium thiamine is lowered similar to 100-fold to human plasma concentrations. In addition, humanizing blood thiamine content of mice through diet sensitizes SLC19A2-low leukemia cells to ASNase in vivo. Together, our work reveals that thiamine utilization is a determinant of ASNase response for some cancer cells and that oversupplying vitamins may affect therapeutic response in leukemia.

A correction to this paper has been published and can be accessed via a link at the top of the paper.

Protein synthesis is required in distinct populations of inhibitory neurons in the mouse amygdala to store memories of danger and safety. To survive in a dynamic environment, animals need to identify and appropriately respond to stimuli that signal danger(1). Survival also depends on suppressing the threat-response during a stimulus that predicts the absence of threat (safety)(2-5). An understanding of the biological substrates of emotional memories during a task in which animals learn to flexibly execute defensive responses to a threat-predictive cue and a safety cue is critical for developing treatments for memory disorders such as post-traumatic stress disorder(5). The centrolateral amygdala is an important node in the neuronal circuit that mediates defensive responses(6-9), and a key brain area for processing and storing threat memories. Here we applied intersectional chemogenetic strategies to inhibitory neurons in the centrolateral amygdala of mice to block cell-type-specific translation programs that are sensitive to depletion of eukaryotic initiation factor 4E (eIF4E) and phosphorylation of eukaryotic initiation factor 2 alpha (p-eIF2 alpha). We show that de novo translation in somatostatin-expressing inhibitory neurons in the centrolateral amygdala is necessary for the long-term storage of conditioned-threat responses, whereas de novo translation in protein kinase C delta-expressing inhibitory neurons in the centrolateral amygdala is necessary for the inhibition of a conditioned response to a safety cue. Our results provide insight into the role of de novo protein synthesis in distinct inhibitory neuron populations in the centrolateral amygdala during the consolidation of long-term memories.

Working memory is a form of short-term memory that involves maintaining and updating task-relevant information toward goal-directed pursuits. Classical models posit persistent activity in prefrontal cortex (PFC) as a primary neural correlate, but emerging views suggest additional mechanisms may exist. We screened similar to 200 genetically diverse mice on a working memory task and identified a genetic locus on chromosome 5 that contributes to a substantial proportion (17%) of the phenotypic variance. Within the locus, we identified a gene encoding an orphan G-protein-coupled receptor. Gpr12, which is sufficient to drive substantial and bidirectional changes in working memory. Molecular, cellular, and imaging studies revealed that Gpr12 enables high thalamus-PFC synchrony to support memory maintenance and choice accuracy. These findings identify an orphan receptor as a potent modifier of short-term memory and supplement classical PFC-based models with an emerging thalamus-centric framework for the mechanistic understanding of working memory.

Burkitt lymphoma (BL) is an aggressive B-cell lymphoma characterized by translocation and deregulation of the proto-oncogene c-MYC. Transcription factor 3 (TCF3) has also been shown to be involved in BL pathogenesis. In BL, TCF3 is constitutively active, and/or expression of its transcriptional targets are altered as a result of BL-associated mutations. Here, we found that BL-relatedTCF3mutations affect TCF3 alternative splicing, in part by reducing binding of the splicing regulator hnRNPH1 to exon 18b. This leads to greater exon 18b inclusion, thereby generating more of the mutated E47 isoform of TCF3. Interestingly, upregulation of E47 dysregulates the expression of TCF3 targetsPTPN6, and perhapsCCND3, which are known to be involved in BL pathogenesis. Our findings thus reveal a mechanism by whichTCF3somatic mutations affect multilayered gene regulation underlying BL pathogenesis.

Addiction to prescription opioid, such as oxycodone, has affected millions of adolescents and young adults. Kappa opioid receptor (KOP-r) agonist can counterbalance the euphoria effects of mu opioid agonists like oxycodone. Nalfurafine is a KOP-r agonist. The current study examined how nalfurafine affected the reinforcing-effect of oxycodone in adolescent male and female mice using intravenous self-administration (SA) and conditioned place preference (CPP) paradigms. Adolescent mice (5 week-old) first received surgery for catheter implantation. After recovery, mice were then placed into the SA chambers and allowed to self-administer oxycodone, 2 h per day for 14 days. Following 14 day oxycodone SA, mice were injected with saline and a single dose of nalfurafine (10, 20, 30, 40 mu g/kg, s.c.) 10 min before each oxycodone SA session for 5 consecutive days. The mice were then injected with Nor-BNI (10 mg/kg, i.p.) 24 h before oxycodone SA following injection of nalfurafine (40 mu g/kg, s.c.). Separate groups of male and female adolescent mice underwent oxycodone CPP or hot plate test with or without nalfurafine preinjection. Nalfurafine decreased oxycodone SA in a dose dependent manner. Nor-BNI blocked the effect of nalfurafine on oxycodone SA. Nalfurafine significantly attenuated the oxycodone-induced hyperlocomotor activities and CPP, but enhanced oxycodone-induced analgesia. In conclusion, nalfurafine reduced the reinforcing effects of oxycodone in male and female adolescent mice. Nalfurafine also increased oxycodone-induced antinociception.

There are currently no therapies proven to promote early recovery of consciousness in patients with severe brain injuries in the intensive care unit (ICU). For patients whose families face time-sensitive, life-or-death decisions, treatments that promote recovery of consciousness are needed to reduce the likelihood of premature withdrawal of life-sustaining therapy, facilitate autonomous self-expression, and increase access to rehabilitative care. Here, we present the Connectome-based Clinical Trial Platform (CCTP), a new paradigm for developing and testing targeted therapies that promote early recovery of consciousness in the ICU. We report the protocol for STIMPACT (Stimulant Therapy Targeted to Individualized Connectivity Maps to Promote ReACTIvation of Consciousness), a CCTP-based trial in which intravenous methylphenidate will be used for targeted stimulation of dopaminergic circuits within the subcortical ascending arousal network (ClinicalTrials.gov NCT03814356). The scientific premise of the CCTP and the STIMPACT trial is that personalized brain network mapping in the ICU can identify patients whose connectomes are amenable to neuromodulation. Phase 1 of the STIMPACT trial is an open-label, safety and dose-finding study in 22 patients with disorders of consciousness caused by acute severe traumatic brain injury. Patients in Phase 1 will receive escalating daily doses (0.5-2.0 mg/kg) of intravenous methylphenidate over a 4-day period and will undergo resting-state functional magnetic resonance imaging and electroencephalography to evaluate the drug's pharmacodynamic properties. The primary outcome measure for Phase 1 relates to safety: the number of drug-related adverse events at each dose. Secondary outcome measures pertain to pharmacokinetics and pharmacodynamics: (1) time to maximal serum concentration; (2) serum half-life; (3) effect of the highest tolerated dose on resting-state functional MRI biomarkers of connectivity; and (4) effect of each dose on EEG biomarkers of cerebral cortical function. Predetermined safety and pharmacodynamic criteria must be fulfilled in Phase 1 to proceed to Phase 2A. Pharmacokinetic data from Phase 1 will also inform the study design of Phase 2A, where we will test the hypothesis that personalized connectome maps predict therapeutic responses to intravenous methylphenidate. Likewise, findings from Phase 2A will inform the design of Phase 2B, where we plan to enroll patients based on their personalized connectome maps. By selecting patients for clinical trials based on a principled, mechanistic assessment of their neuroanatomic potential for a therapeutic response, the CCTP paradigm and the STIMPACT trial have the potential to transform the therapeutic landscape in the ICU and improve outcomes for patients with severe brain injuries.

Background Preclinical testing of new locoregional therapies for pancreatic cancer has been challenging, due to the lack of a suitable large animal model. Purpose To develop and characterize a porcine model of pancreatic cancer. Unlike small animals, pigs have similar physiology, drug dosing, and immune response to humans. Locoregional therapy in pigs can be performed using the same size catheters and devices as in humans. Methods The Oncopig is a transgenic pig with Cre-inducibleTP53(R167H)andKRAS(G12D)mutations. In 12 Oncopigs, CT-guided core biopsy of the pancreas was performed. The core biopsy was incubated with an adenoviral vector carrying the Cre recombinase gene. The transformed core biopsy was injected back into the pancreas (head, tail, or both). The resulting tumors (n= 19) were characterized on multi-phase contrast-enhanced CT, and on pathology, including immunohistochemistry. Angiographic characterization of the tumors was performed in 3 pigs. Results Pancreatic tumors developed at 19 out of 22 sites (86%) that were inoculated. Average tumor size was 3.0 cm at 1 week (range: 0.5-5.1 cm). H&E and immunohistochemical stains revealed undifferentiated carcinomas, similar to those of the pancreatobiliary system in humans. Neoplastic cells were accompanied by a major inflammatory component. 1 of 12 pigs only had inflammatory nodules without evidence of neoplasia. On multiphase CT, tumors were hypovascular compared to the normal pancreas. There was no pancreatic duct dilation. In 3 pigs, angiography was performed, and in all 3 cases, the artery supplying the pancreatic tumor could be catheterized using a 2.4 F microcatheter. Selective angiography showed the pancreatic tumor, without extra-pancreatic perfusion. Conclusion Pancreatic cancer can be induced in a transgenic pig. Intra-arterial procedures using catheters designed for human interventions were technically feasible in this large animal model.

Symmetry breaking is essential for polarization of cells and generation of left-right body asymmetry. Here the authors investigate the arrangement of hair cells in zebrafish and show that mirror-symmetric patterns arise from a combination of biochemical and mechanical symmetry-breaking events. Actively regulated symmetry breaking, which is ubiquitous in biological cells, underlies phenomena such as directed cellular movement and morphological polarization. Here, we investigate how an organ-level polarity pattern emerges through symmetry breaking at the cellular level during the formation of a mechanosensory organ. Combining theory, genetic perturbations and in vivo imaging, we study the development and regeneration of the fluid-motion sensors in the zebrafish's lateral line. We find that two interacting symmetry-breaking events-one mediated by biochemical signalling and the other by cellular mechanics-give rise to precise rotations of cell pairs, which produce a mirror-symmetric polarity pattern in the receptor organ.

Metabolic reprogramming is a key feature of many cancers, but how and when it contributes to tumorigenesis remains unclear. Here we demonstrate that metabolic reprogramming induced by mitochondrial fusion can be rate-limiting for immortalization of tumor-initiating cells (TICs) and trigger their irreversible dedication to tumorigenesis. Using single-cell transcriptomics, we find that Drosophila brain tumors contain a rapidly dividing stem cell population defined by upregulation of oxidative phosphorylation (OxPhos). We combine targeted metabolomics and in vivo genetic screening to demonstrate that OxPhos is required for tumor cell immortalization but dispensable in neural stem cells (NSCs) giving rise to tumors. Employing an in vivo NADH/NAD+ sensor, we show that NSCs precisely increase OxPhos during immortalization. Blocking OxPhos or mitochondrial fusion stalls TICs in quiescence and prevents tumorigenesis through impaired NAD+ regeneration. Our work establishes a unique connection between cellular metabolism and immortalization of tumor-initiating cells.