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Diamond-Blackfan anemia (DBA) is a ribosomopathy that is characterized by macrocytic anemia, congenital malformations, and early onset during childhood. Genetic studies have demonstrated that most patients carry mutations in one of the 20 related genes, most of which encode ribosomal proteins (RP). Treatment of DBA includes corticosteroid therapy, chronic red blood cell transfusion, and other forms of immunosuppression. Currently, hematopoietic stem cell transplantation is the only cure for DBA. Interestingly, spontaneous remissions occur in 10-20% of transfusion-dependent DBA patients. However, there is no consistent association between specific mutations and clinical manifestations. In the past decades, researchers have made significant progress in understanding the pathogenesis of DBA, but it remains unclear how the ubiquitous RP haploinsufficiency causes the erythroid-specific defect in hematopoiesis in DBA patients, and why there is a difference in penetrance and spontaneous remission among individuals who carry identical mutations. In this paper, we provide a comprehensive review of the development of DBA animal models and discuss the future research directions for these important experimental systems.

Diamond-Blackfan Anemia (DBA) is an inherited bone marrow failure syndrome that is associated with anemia, congenital anomalies, and cancer predisposition. It is categorized as a ribosomopathy, because more than 80% or patients have haploinsufficiency of either a small or large subunit-associated ribosomal protein (RP). The erythroid pathology is due predominantly to a block and delay in early committed erythropoiesis with reduced megakaryocyte/erythroid progenitors (MEPs). To understand the molecular pathways leading to pathogenesis of DBA, we performed RNA sequencing on mRNA and miRNA from RPS19-deficient human hematopoietic stem and progenitor cells (HSPCs) and compared existing database documenting transcript fluctuations across stages of early normal erythropoiesis. We determined the chromatin regulator, SATB1 was prematurely downregulated through the coordinated action of upregulated miR-34 and miR-30 during differentiation in ribosomal insufficiency. Restoration of SATB1 rescued MEP expansion, leading to a modest improvement in erythroid and megakaryocyte expansion in RPS19 insufficiency. However, SATB1 expression did not affect expansion of committed erythroid progenitors, indicating ribosomal insufficiency affects multiple stages during erythroid differentiation.

Supplementary Figure 3 from The Multitargeted Receptor Tyrosine Kinase Inhibitor Linifanib (ABT-869) Induces Apoptosis through an Akt and Glycogen Synthase Kinase 3β–Dependent Pathway

Supplementary Figure 1 from The Multitargeted Receptor Tyrosine Kinase Inhibitor Linifanib (ABT-869) Induces Apoptosis through an Akt and Glycogen Synthase Kinase 3β–Dependent Pathway

Supplementary Figure 2 from The Multitargeted Receptor Tyrosine Kinase Inhibitor Linifanib (ABT-869) Induces Apoptosis through an Akt and Glycogen Synthase Kinase 3β–Dependent Pathway

The FMS-like receptor tyrosine kinase 3 (FLT3) plays an important role in controlling differentiation and proliferation of hematopoietic cells. Activating mutations in FLT3 occur in patients with acute myeloid leukemia (AML; 15%–35%), resulting in abnormal cell proliferation. Furthermore, both adult and pediatric patients with AML harboring the FLT3 internal tandem duplication (ITD) mutation have a poor prognosis. Several inhibitors have been developed to target mutant FLT3 for the treatment of AML, yet the molecular pathways affected by drug inhibition of the mutated FLT3 receptor alone have not been characterized as yet. Linifanib (ABT-869) is a multitargeted tyrosine kinase receptor inhibitor that suppresses FLT3 signaling. In this article, we show that treatment with linifanib inhibits proliferation and induces apoptosis in ITD mutant cells in vitro and in vivo. We show that treatment with linifanib reduces phosphorylation of Akt and glycogen synthase kinase 3β (GSK3β). In addition, we show that inhibition of GSK3β decreases linifanib-induced apoptosis. This study shows the importance of GSK3 as a potential target for AML therapy, particularly in patients with FLT3 ITD mutations. Mol Cancer Ther; 10(6); 949–59. ©2011 AACR.

Neuroblastoma is a deadly pediatric solid tumor with infrequent recurrent somatic mutations. Particularly, the pathophysiology of tumors without MYCN amplification remains poorly defined. Utilizing an unbiased approach, we performed gene set enrichment analysis of RNA-sequencing data from 498 patients with neuroblastoma and revealed a differentially overexpressed gene signature in MYCN nonamplified neuroblastomas with telomerase reverse transcriptase (TERT) gene overexpression and coordinated activation of oncogenic signaling pathways, including E2Fs, Wnt, Myc, and the DNA repair pathway. Promoter rearrangement of the TERT gene juxtaposes the coding sequence to strong enhancer elements, leading to TERT overexpression and poor prognosis in neuroblastoma, but TERT-associated oncogenic signaling remains unclear. ChIP-seq analysis of the human CLB-GA neuroblastoma cells harboring TERT rearrangement uncovered genome-wide chromatin co-occupancy of Brd4 and H3K27Ac and robust enrichment of H3K36me3 in TERT and multiple TERT-associated genes. Brd4 and cyclin-dependent kinases (CDK) had critical regulatory roles in the expression and chromatin activation of TERT and multiple TERT-associated genes. Epigenetically targeting Brd4 or CDKs with their respective inhibitors suppressed the expression of TERT and multiple TERT-associated genes in neuroblastoma with TERT overexpression or MYCN amplification. ChIP-seq and ChIP-qPCR provided evidence that the CDK inhibitor directly inhibited Brd4 recruitment to activate chromatin globally. Therefore, inhibiting Brd4 and CDK concurrently with AZD5153 and dinaciclib would be most effective in tumor growth suppression, which we demonstrated in neuroblastoma cell lines, primary human cells, and xenografts. In summary, we describe a unique mechanism in neuroblastoma with TERT overexpression and an epigenetically targeted novel therapeutic strategy.Significance:Epigenetically cotargeting Brd4 and Cdks suppresses human neuroblastoma with TERT overexpression by inhibiting the TERT-associated gene expression networks.

Dasatinib is a multi-tyrosine kinase inhibitor approved for treatment of Ph+ acute lymphoblastic leukemia (ALL), but its efficacy is limited by resistance. Recent preclinical studies suggest that dasatinib may be a candidate therapy in additional ALL subtypes including pre-BCR+ ALL. Here we utilized shRNA library screening and global transcriptomic analysis to identify several novel genes and pathways that may enhance dasatinib efficacy or mitigate potential resistance in human pre-BCR+ ALL. Depletion of the transcriptional coactivator CBP increased dasatinib sensitivity by downregulating transcription of the pre-BCR signaling pathway previously associated with dasatinib sensitivity. Acquired resistance was due, in part, to upregulation of alternative pathways including WNT through a mechanism, suggesting transcriptional plasticity. Small molecules that disrupt CBP interactions with the CREB KID domain or β-catenin showed promising preclinical efficacy in combination with dasatinib. These findings highlight novel modulators of sensitivity to targeted therapies in human pre-BCR+ ALL, which can be reversed by small-molecule inhibitors. They also identify promising therapeutic approaches to ameliorate dasatinib sensitivity and prevent resistance in ALL.Significance: These findings reveal mechanisms that modulate sensitivity to dasatinib and suggest therapeutic strategies to improve the outcome of patients with acute lymphoblastic leukemia.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/22/6497/F1.large.jpg. Cancer Res; 78(22); 6497–508. ©2018 AACR.

Unbiased shRNA library screening and global transcriptome analyses reveal mechanisms that modulate dasatinib sensitivity and suggest therapeutic strategies to improve outcome of patients with acute lymphoblastic leukemia.

Supplementary Fig. S1. Elevated mRNA expression of TERT and TERT-associated genes in NB with TERT gene rearrangement. Supplementary Fig. S2. Effects of dinaciclib on genome-wide chromatin occupancy of epigenetic modulators in CLB-GA cells. Supplementary Fig. S3: C-Myc and MYCN protein expression in four NB cell lines. Supplementary Fig. S4: Effects of dinaciclib on the chromatin occupancy of epigenetic modulators at Dvl2, AURKA, and AURKB and in CLB-GA cells. Supplementary Fig. S5. Association of overexpression of FEN1, BIRC5, and UHRF1 with poor prognosis and effects of dinaciclib on chromatin occupancy of epigenetic modulators in CLB-GA cells. Supplementary Fig. S6. JQ1 sensitive Brd4 binding profiles in OCI-AML3 cells. Supplementary Fig. S7. JQ1 and dinaciclib are synergistic or additive in inducing cytotoxicity in NB cell lines. Supplementary Fig. S8. Combination index (CI) values for synergy experiment combining Dinaciclib with Brd4 inhibitor JQ1. Supplementary Fig. S9. Effects of dinaciclib on phosphorylation of Rb in CLB-GA and Kelly cells. Supplementary table S1. Primer sequences used in ChIP-qPCR and real-time RT-PCR analysis. Supplementary table 2. Gene sets enriched in NB patients with TERT gene rearrangement (n=32) versus without TERT gene rearrangement (n=278) within MYCN non-amplified NB of Tumor Neuroblastoma (TERT)-Fischer-394-custom. Supplementary table 3. Gene sets enriched in NB patients with high versus low TERT within Tumor Neuroblastoma Gene-TARGET-161. Supplementary table 4: Information on patient samples used in this study.

Correction: Cancer Res 2008;68:2557-60 from Role of the Aggresome Pathway in Cancer: Targeting Histone Deacetylase 6–Dependent Protein Degradation

Misfolded or aggregated proteins have two fates: they are either refolded with the help of chaperones or degraded by the proteasome. Cells also have an alternative pathway that involves intracellular “storage bins” for misfolded intracellular proteins known as aggresomes. Aggresomes recruit motor proteins that transport misfolded or aggregated proteins to chaperones and proteasomes for subsequent destruction. There is emerging evidence that inhibiting the aggresome pathway leads to accumulation of misfolded proteins and apoptosis in tumor cells through autophagy. We discuss the role of aggresomes in cancer and the potential to target this pathway for therapy. [Cancer Res 2008;68(8):2557–60]

Diamond Blackfan anemia (DBA) is an inherited bone marrow failure syndrome associated with severe anemia, congenital malformations, and an increased risk of developing cancer. The chromatin-binding special AT-rich sequence-binding protein-1 (SATB1) is downregulated in megakaryocyte/erythroid progenitors (MEPs) in patients and cell models of DBA, leading to a reduction in MEP expansion. Here we demonstrate that SATB1 expression is required for the upregulation of the critical erythroid factors heat shock protein 70 (HSP70) and GATA1 which accompanies MEP differentiation. SATB1 binding to specific sites surrounding the HSP70 genes promotes chromatin loops that are required for the induction of HSP70, which, in turn, promotes GATA1 induction. This demonstrates that SATB1, although gradually downregulated during myelopoiesis, maintains a biological function in early myeloid progenitors.

Cryogenic electron microscopy (cryo-EM) has emerged as a viable structural tool for molecular therapeutics development against human diseases. However, it remains a challenge to determine structures of proteins that are flexible and smaller than 30 kDa. The 11 kDa KIX domain of CREB-binding protein (CBP), a potential therapeutic target for acute myeloid leukemia and other cancers, is a protein which has defied structure-based inhibitor design. Here, we develop an experimental approach to overcome the size limitation by engineering a protein double-shell to sandwich the KIX domain between apoferritin as the inner shell and maltose-binding protein as the outer shell. To assist homogeneous orientations of the target, disulfide bonds are introduced at the target-apoferritin interface, resulting in a cryo-EM structure at 2.6 Å resolution. We used molecular dynamics simulations to design peptides that block the interaction of the KIX domain of CBP with the intrinsically disordered pKID domain of CREB. The double-shell design allows for fluorescence polarization assays confirming the binding between the KIX domain in the double-shell and these interacting peptides. Further cryo-EM analysis reveals a helix-helix interaction between a single KIX helix and the best peptide, providing a possible strategy for developments of next-generation inhibitors.

Elucidation of biological phenomena requires imaging of microenvironments in vivo. Although the seamless visualization of in vivo hypoxia from the level of whole-body to single-cell has a great potential to discover unknown phenomena in biological and medical fields, no methodology for achieving it has been established thus far. Here, we for the first time report the whole-body and whole-organ imaging of hypoxia, an important microenvironment, at single-cell resolution using activatable covalent fluorescent probes compatible with tissue clearing. We initially focused on overcoming the incompatibility of fluorescent dyes and refractive index matching solutions (RIMSs), which has greatly hindered the development of fluorescent molecular probes in the field of tissue clearing. The fluorescent dyes compatible with RIMS were then incorporated into the development of activatable covalent fluorescent probes for hypoxia. We combined the probes with tissue clearing, achieving comprehensive single-cell-resolution imaging of hypoxia in a whole mouse body and whole organs.

Microbial source tracking (MST) identifies sources of fecal contamination in the environment using fecal host-associated markers. While there are numerous bacterial MST markers, there are few viral markers. Here we design and test novel viral MST markers based on tomato brown rugose fruit virus (ToBRFV) genomes. We assembled eight nearly complete genomes of ToBRFV from wastewater and stool samples from the San Francisco Bay Area in the United States of America. Next, we developed two novel probe-based RT-PCR assays based on conserved regions of the ToBRFV genome, and tested the markers’ sensitivities and specificities using human and non-human animal stool as well as wastewater. TheToBRFV markers are sensitive and specific; in human stool and wastewater, they are more prevalent and abundant than a currently used marker, the pepper mild mottle virus (PMMoV) coat protein (CP) gene. We applied the assays to detect fecal contamination in urban stormwater samples and found that the ToBRFV markers matched cross-assembly phage (crAssphage), an established viral MST marker, in prevalence across samples. Taken together, ToBRFV is a promising viral human-associated MST marker.ImportanceHuman exposure to fecal contamination in the environment can cause transmission of infectious diseases. Microbial source tracking (MST) can identify sources of fecal contamination so that contamination can be remediated and human exposures can be reduced. MST requires the use of fecal host-associated MST markers. Here we design and test novel MST markers from genomes of tomato brown rugose fruit virus (ToBRFV). The markers are sensitive and specific to human stool, and highly abundant in human stool and wastewater samples.

Chronic myeloid leukemia (CML) is effectively treated with long-term tyrosine kinase inhibitor (TKI) therapy, yet little is known about risks of prolonged TKI exposure in young patients, and long-term effect monitoring is not standardized. We surveyed North American pediatric oncologists (n = 119) to evaluate perceived risk of and surveillance practices for potential toxicities associated with prolonged TKI exposure in children and adolescents/young adults (AYAs) with CML. Survey domains included general and specific risk perceptions and surveillance practices for asymptomatic patients on chronic TKI therapy. We analyzed data descriptively and explored relationships between risk perceptions and surveillance. Risk perceptions varied among oncologists but were similar across six categories (thyroid, cardiac, vascular, metabolic, fertility, psychologic), with less than one-third rating each risk as moderate or high in pediatric and AYA patients. More oncologists perceived moderate or high risk of growth abnormalities in children (62% pediatric, 14% AYA) and financial toxicity in all patients (60% pediatric, 64% AYA). A greater proportion of oncologists with moderate or high perceived risk of thyroid abnormalities reported testing thyroid function compared to those with lower perceived risk; patterns for metabolic risk/lipid tests and cardiac risk/tests were similar. In summary, we found that pediatric oncologists had variable risk perceptions and surveillance practices for potential toxicities associated with prolonged TKI exposure. Standardizing surveillance would help quantify risks and refine recommendations. Supplemental data for this article is available online at https://doi.org/10.1080/08880018.2021.2017085 .

B- and T-cell acute lymphoblastic leukemia (B/T-ALL) may be refractory or recur after therapy by suppressing host anticancer immune surveillance mediated specifically by natural killer (NK) cells. We delineated the phenotypic and functional defects in NK cells from high-risk patients with B/T-ALL using mass cytometry, flow cytometry, and in silico cytometry, with the goal of further elucidating the role of NK cells in sustaining acute lymphoblastic leukemia (ALL) regression. We found that, compared with their normal counterparts, NK cells from patients with B/T-ALL are less cytotoxic but exhibit an activated signature that is characterized by high CD56, high CD69, production of activated NK cell–origin cytokines, and calcium (Ca2+) signaling. We demonstrated that defective maturation of NK cells into cytotoxic effectors prevents NK cells from ALL from lysing NK cell–sensitive targets as efficiently as do normal NK cells. Additionally, we showed that NK cells in ALL are exhausted, which is likely caused by their chronic activation. We found that increased frequencies of activated cytokine-producing NK cells are associated with increased disease severity and independently predict poor clinical outcome in patients with ALL. Our studies highlight the benefits of developing NK cell profiling as a diagnostic tool to predict clinical outcome in patients with ALL and underscore the clinical potential of allogeneic NK cell infusions to prevent ALL recurrence.

Background: It is critical for surgeons to have a full understanding of the complex courses and ramifications of the human internal iliac artery and its parietal branches. Although numerous anatomical studies have been performed, not all variations at this site are currently understood. Therefore, we characterised these blood vessels in foetal pigs to provide additional insight from a comparative anatomical perspective. Materials and methods: Eighteen half-pelvis specimens from foetal pigs were dissected and examined on macroscopic scale. Results: Among our findings, we identified the internal iliac artery as a descending branch of the abdominal aorta. A very thick umbilical artery arose from the internal iliac artery. The superior gluteal, inferior gluteal, and internal pudendal arteries formed the common arterial trunk. Although the superior gluteal artery emerged from the common trunk from inside the pelvis, the inferior gluteal and internal pudendal arteries bifurcated at deep layer within the gluteus muscles after leaving pelvic cavity. We were unable to detect an typical obturator artery emerging from the internal iliac artery. A branch supplying the hip adductors was identified as arising from the inferior epigastric artery which itself was derived from the distal end of the external iliac artery. Conclusions: We identified the anatomic characteristics of the internal iliac artery and its parietal branches in the foetal pig. Our findings provide new insight into the comparative anatomy of the internal iliac artery and will promote understanding of related morphogenetic processes.

Clinical pathways are evidence-based tools that have been integrated into many aspects of pediatric hospital medicine and have proven effective at reducing in-hospital complications from a variety of diseases. Adaptation of similar tools for specific, high-risk patient populations in pediatric oncology has been slower, in part due to patient complexities and variations in management strategies. There are few published studies of clinical pathways for pediatric oncology patients. Pediatric patients with a new diagnosis of leukemia or lymphoma often present with one or more “oncologic emergencies” that require urgent intervention and deliberate multidisciplinary care to prevent significant consequences. Here, we present two clinical pathways that have recently been developed using a multidisciplinary approach at a single institution, intended for the care of patients who present with hyperleukocytosis or an anterior mediastinal mass. These clinical care pathways have provided a critical framework for the immediate care of these patients who are often admitted to the pediatric intensive care unit for initial management. The goal of the pathways is to facilitate multidisciplinary collaborations, expedite diagnosis, and streamline timely treatment initiation. Standardizing the care of high-risk pediatric oncology patients will ultimately decrease morbidity and mortality associated with these diseases to increase the potential for excellent outcomes.

Ticks are able to transmit the highest number of pathogen species of any blood-feeding arthropod and represent a growing threat to public health and agricultural systems worldwide. While there are numerous and varied causes and effects of changes to tick-borne disease (re)emergence, three primary challenges to tick control were identified in this review from a U.S. borders perspective. (1) Climate change is implicated in current and future alterations to geographic ranges and population densities of tick species, pathogens they can transmit, and their host and reservoir species, as highlighted by Ixodes scapularis and its expansion across southern Canada. (2) Modern technological advances have created an increasingly interconnected world, contributing to an increase in invasive tick species introductions through the increased speed and frequency of trade and travel. The introduction of the invasive Haemaphysalis longicornis in the eastern U.S. exemplifies the challenges with control in a highly interconnected world. (3) Lastly, while not a new challenge, differences in disease surveillance, control, and management strategies in bordering countries remains a critical challenge in managing ticks and tick-borne diseases. International inter-agency collaborations along the U.S.–Mexico border have been critical in control and mitigation of cattle fever ticks (Rhipicephalus spp.) and highlight the need for continued collaboration and research into integrated tick management strategies. These case studies were used to identify challenges and opportunities for tick control and mitigation efforts through a One Health framework.

Ticks are important vectors of pathogenic viruses, bacteria, and protozoans to humans, wildlife, and domestic animals. Due to their life cycle, ticks face significant challenges related to water homeostasis. When blood-feeding they must excrete water and ions, but when off-host (for stretches lasting several months) they must conserve water to avoid desiccation. Aquaporins (AQPs), a family of membrane-bound water channels, are key players in osmoregulation in many animals but remain poorly characterized in ticks. Here, we bioinformatically identified AQP-like genes from the deer tick Ixodes scapularis and used phylogenetic approaches to map the evolution of the aquaporin gene family in arthropods. We find that most arachnid AQP-like sequences (including those of I. scapularis) form a monophyletic group clustered within aquaglycerolporins (GLPs) from bacteria to vertebrates. This gene family is absent from insects, revealing divergent evolutionary paths for AQPs in different hematophagous arthropods. We next sequenced the full-length cDNA of I. scapularis aquaporin 1 (IsAQP1) and expressed it heterologously in Xenopus oocytes to functionally characterize its permeability to water and solutes. We additionally examined IsAQP1 expression across different life stages and adult female tissues. We found IsAQP1 to be an efficient water channel with salivary gland-specific expression, implying its function may be closely tied to osmoregulation during blood-feeding. Its functional properties were unique: unlike most GLPs, IsAQP1 has low glycerol permeability, and unlike most AQPs, it is insensitive to mercury. Together, our results suggest IsAQP1 plays an important role in tick water balance during blood feeding and may hold promise as a target for novel vector control efforts.

BackgroundType I interferons (IFN-Is), secreted by hematopoietic cells, drive immune surveillance of solid tumors. However, the mechanisms of suppression of IFN-I-driven immune responses in hematopoietic malignancies including B-cell acute lymphoblastic leukemia (B-ALL) are unknown.MethodsUsing high-dimensional cytometry, we delineate the defects in IFN-I production and IFN-I-driven immune responses in high-grade primary human and mouse B-ALLs. We develop natural killer (NK) cells as therapies to counter the intrinsic suppression of IFN-I production in B-ALL.ResultsWe find that high expression of IFN-I signaling genes predicts favorable clinical outcome in patients with B-ALL, underscoring the importance of the IFN-I pathway in this malignancy. We show that human and mouse B-ALL microenvironments harbor an intrinsic defect in paracrine (plasmacytoid dendritic cell) and/or autocrine (B-cell) IFN-I production and IFN-I-driven immune responses. Reduced IFN-I production is sufficient for suppressing the immune system and promoting leukemia development in mice prone to MYC-driven B-ALL. Among anti-leukemia immune subsets, suppression of IFN-I production most markedly lowers the transcription of IL-15 and reduces NK-cell number and effector maturation in B-ALL microenvironments. Adoptive transfer of healthy NK cells significantly prolongs survival of overt ALL-bearing transgenic mice. Administration of IFN-Is to B-ALL-prone mice reduces leukemia progression and increases the frequencies of total NK and NK-cell effectors in circulation. Ex vivo treatment of malignant and non-malignant immune cells in primary mouse B-ALL microenvironments with IFN-Is fully restores proximal IFN-I signaling and partially restores IL-15 production. In B-ALL patients, the suppression of IL-15 is the most severe in difficult-to-treat subtypes with MYC overexpression. MYC overexpression promotes sensitivity of B-ALL to NK cell-mediated killing. To counter the suppressed IFN-I-induced IL-15 production in MYChighhuman B-ALL, we CRISPRa-engineered a novel human NK-cell line that secretes IL-15. CRISPRa IL-15-secreting human NK cells kill high-grade human B-ALL in vitro and block leukemia progression in vivo more effectively than NK cells that do not produce IL-15.ConclusionWe find that restoration of the intrinsically suppressed IFN-I production in B-ALL underlies the therapeutic efficacy of IL-15-producing NK cells and that such NK cells represent an attractive therapeutic solution for the problem of drugging MYC in high-grade B-ALL.

We have investigated the generation region of intermittent plasma structures, which could be due to the blob-like cross field transport around the divertor simulation experimental module (D-module) in the tandem mirror device GAMMA 10/PDX. A positive skewness of the ion saturation current was clearly seen when the electrode of the movable probe was located at the radial edge and just in front of the entrance limiter of the D-module. Fourier analysis and conditional averaging clarified that positive spikes intermittently appeared in the same region. This is the first indication that the phenomenon producing the high-density isolated plasma structures occurred in the upstream of the D-module. A negative skewness was also found, and large amplitude fluctuation was detected between regions showing positive and negative skewness. Furthermore, this and light emission fluctuations become stronger during the transient state from attached to detached state on the V-shaped target.

This study aimed to clarify the practices of skincare such as bathing, face-washing, and using of skincare products, among older adults living in nursing homes in Japan. From July to August 2017, questionnaire surveys were mailed to 343 long-term care facilities in one prefecture in Japan, and 105 facilities returned the survey. Management policies on the use of facial skincare products was significantly associated with the number of resident/caregiver ratio. Meanwhile, residents in nursing homes with higher numbers of caregivers washed their faces and bathed at lower rates. There were no correlations between the number of resident/caregiver ratio, face-washing, and bathing times. When older residents could not independently perform their skin cleansing and care, that care is likely to happen less often. Sufficient manpower for the support of older residents’ skincare will improve the quality of life in long-term nursing facilities.

Oxygen isotope ratios (δ18O) of tree-ring cellulose are a novel proxy for summer hydroclimate in monsoonal Asia. In central Japan, we collected 67 conifer wood samples, mainly Chamaecyparis obtusa, with ages encompassing the past 2600 years. The samples were taken from living trees, archeological wood, architectural wood, and buried logs. We analyzed stable isotope ratios of oxygen (δ18O) and hydrogen (δ2H) in tree-ring cellulose in these samples (more than 15 000 rings in total) without using a pooling method and constructed a statistically reliable tree-ring cellulose δ18O time series for the past 2500 years. However, there were distinct age trends and level offsets in the δ18O record, and cellulose δ18O values showed a gradual decrease as an individual tree matures. This suggested it is difficult to establish a cellulose δ18O chronology for low-frequency signals by simple averaging of all the δ18O time series data. In addition, there were opposite age trends in the cellulose δ2H, and δ2H gradually increased with tree age. There were clear positive correlations in the short-periodicity variations between δ18O and δ2H, probably indicating a common climate signal. A comparison of the δ18O and δ2H time series in individual trees with tree-ring width suggested that the opposite age trends of δ18O and δ2H are caused by temporal changes in the degree of post-photosynthetic isotope exchange with xylem water (physiological effect), accompanied by changes in stem growth rate that are influenced by human activity in the forests of central Japan. Based on the assumptions that cellulose δ18O and δ2H vary positively and negatively with constant proportional coefficients due to climatological and physiological effects, respectively, we solved simultaneous equations for the climatological and physiological components of variations in tree-ring cellulose δ18O and δ2H in order to remove the age trend. This enabled us to evaluate the climatic record from cellulose δ18O variations. The extracted climatological component in the cellulose δ18O for the past 2600 years in central Japan was well correlated with numerous instrumental, historical, and paleoclimatological records of past summer climate at various spatial and temporal scales. This indicates that integration of tree-ring cellulose δ18O and δ2H data is a promising method to reconstruct past summer climate variations on annual to millennial timescales, irrespective of the growth environment. However, analytical and statistical methods need to be improved for further development of this climate proxy.

In this paper, we discuss how to improve the quality of classification results when deep learning is applied for the filtering of airborne LiDAR point cloud. We introduce the baseline method which utilizes convolutional neural network (CNN) based on voxelization, and then we propose three methods to improve the quality of classification result. The first method is data pre-processing that aims to exclude data in advance that is clearly not on the ground surface in order to efficiently extract the ground surface data. Data pre-processing can greatly reduce the number of target points and the subsequent processing can be performed efficiently. It also has the effect of preventing noise-like points floating in the air from being misclassified as the ground surface, as compared to the case without pre-processing. The second method is changing the network structure. In recent years, various networks have been proposed for classifying point clouds. In our study, the baseline is using very simple networks. In order to improve the classification result of the baseline method, the layer depth and the range size of convolution are changed, and we investigated about the improvements of the results. The current discussion can be used as a guidance when considering new networks. The third method is the integration of classification results from multiple networks. We integrated individual results from multiple networks with varying layer depths and convolution sizes, starting with the baseline, and investigated whether the results improved. We observed that even if the individual results were similar, the classification results can be improved by integrating the results.

The historical castles (castellated walls), which are cultural heritages in Japan, require regular maintenance, and it is necessary to record the arrangement of individual wall stones in the maintenance work. Recently, image processing techniques are practiced to optimize maintenance and management of the infrastructure assets. In the previous study, we proposed an automatic method for efficiently extracting individual wall stone polygons by improved multiscale image segmentation technique. However, the problem has remained that wall stone polygons could not be extracted properly when there were no clear gaps or boundaries between stones. To address this problem, we improved the multiscale image segmentation technique used in our previous studies. The first improvement is that in the region growing process, selecting the best combination of a plurality of objects instead of two. The second improvement is the modification of the shape criterion to be used. Besides, we proposed three-stage Stacked cGAN for wall stone edge detection that enables us to complement areas with weak or broken boundaries of stone edges. This approach is composed of a coarse-to-fine based image-to-edges translation network. The edge images derived from this method are used as the additional channel in multiscale image segmentation with a higher weight compared to the other RGB channels. It was confirmed that the separation performance of individual wall stone polygons was improved by the proposed method. Furthermore, the proposed method is highly effective to reduce the difficulty in setting of the scale parameter, which is usually sensitive to segmentation results and requires trial and error.

Neuroblastoma is a deadly pediatric solid tumor with infrequent recurrent somatic mutations. Particularly, the pathophysiology of tumors without MYCN amplification remains poorly defined. Utilizing an unbiased approach, we performed gene set enrichment analysis of RNA-sequencing data from 498 patients with neuroblastoma and revealed a differentially overexpressed gene signature in MYCN nonamplified neuroblastomas with telomerase reverse transcriptase (TERT) gene overexpression and coordinated activation of oncogenic signaling pathways, including E2Fs, Wnt, Myc, and the DNA repair pathway. Promoter rearrangement of the TERT gene juxtaposes the coding sequence to strong enhancer elements, leading to TERT overexpression and poor prognosis in neuroblastoma, but TERT-associated oncogenic signaling remains unclear. ChIP-seq analysis of the human CLB-GA neuroblastoma cells harboring TERT rearrangement uncovered genome-wide chromatin co-occupancy of Brd4 and H3K27Ac and robust enrichment of H3K36me3 in TERT and multiple TERT-associated genes. Brd4 and cyclin-dependent kinases (CDK) had critical regulatory roles in the expression and chromatin activation of TERT and multiple TERT-associated genes. Epigenetically targeting Brd4 or CDKs with their respective inhibitors suppressed the expression of TERT and multiple TERT-associated genes in neuroblastoma with TERT overexpression or MYCN amplification. ChIP-seq and ChIP-qPCR provided evidence that the CDK inhibitor directly inhibited Brd4 recruitment to activate chromatin globally. Therefore, inhibiting Brd4 and CDK concurrently with AZD5153 and dinaciclib would be most effective in tumor growth suppression, which we demonstrated in neuroblastoma cell lines, primary human cells, and xenografts. In summary, we describe a unique mechanism in neuroblastoma with TERT overexpression and an epigenetically targeted novel therapeutic strategy. Significance: Epigenetically cotargeting Brd4 and Cdks suppresses human neuroblastoma with TERT overexpression by inhibiting the TERT-associated gene expression networks.

The increasing use of mass cytometry for analyzing clinical samples offers the possibility to perform comparative analyses across public datasets. However, challenges in batch normalization and data integration limit the comparison of datasets not intended to be analyzed together. Here, we present a data integration strategy, CytofIn, using generalized anchors to integrate mass cytometry datasets from the public domain. We show that low-variance controls, such as healthy samples and stable channels, are inherently homogeneous, robust against stimulation, and can serve as generalized anchors for batch correction. Single-cell quantification comparing mass cytometry data from 989 leukemia files pre- and post normalization with CytofIn demonstrates effective batch correction while recapitulating the gold-standard bead normalization. CytofIn integration of public cancer datasets enabled the comparison of immune features across histologies and treatments. We demonstrate the ability to integrate public datasets without necessitating identical control samples or bead standards for fast and robust analysis using CytofIn.

The inability to extinguish a traumatic memory is a key aspect of post-traumatic stress disorder (PTSD). While PTSD affects 10–20% of individuals who experience a trauma, women are particularly susceptible to developing the disorder. Despite this notable female vulnerability, few studies have investigated this particular resistance to fear extinction observed in females. Similar to humans, rodent models of Pavlovian fear learning and extinction show a wide range of individual differences in fear learning and extinction, although female rodents are considerably understudied. Therefore, the present study examined individual differences in fear responses, including freezing behavior and ultrasonic vocalizations (USVs), of female Long–Evans rats during acquisition of fear conditioning and cued fear extinction. Similar to prior studies in males, female rats displayed individual variation in freezing during cued fear extinction and were divided into extinction competent (EC) and extinction resistant (ER) phenotypes. Differences in freezing between ER and EC females were accompanied by shifts in rearing during extinction, but no darting was seen in any trial. Freezing behavior during fear learning did not differ between the EC and ER females. Vocalizations emitted in the 22 and 50 kHz ranges during fear learning and extinction were also examined. Unlike vocalizations seen in previous studies in males, very few 22 kHz distress vocalizations were emitted by female rats during fear acquisition and extinction, with no difference between ER and EC groups. Interestingly, all female rats produced significant levels of 50 kHz USVs, and EC females emitted significantly more 50 kHz USVs than ER rats. This difference in 50 kHz USVs was most apparent during initial exposure to the testing environment. These results suggest that like males, female rodents show individual differences in both freezing and USVs during fear extinction, although females appear to vocalize more in the 50 kHz range, especially during initial periods of exposure to the testing environment, and emit very few of the 22 kHz distress calls that are typically observed in males during fear learning or extinction paradigms. Overall, these findings show that female rodents display fear behavior repertoires divergent from males.

Disruption of cyclic adenosine monophosphate response element binding protein (CREB) provides a potential new strategy to address acute leukemia, a disease associated with poor prognosis, and for which conventional treatment options often carry a significant risk of morbidity and mortality. We describe the structure-activity relationships (SAR) for a series of XX-650-23 derived from naphthol AS-E phosphate that disrupts binding and activation of CREB by the CREB-binding protein (CBP). Through the development of this series, we identified several salicylamides that are potent inhibitors of acute leukemia cell viability through inhibition of CREB-CBP interaction. Among them, a biphenyl salicylamide, compound 71, was identified as a potent inhibitor of CREB-CBP interaction with improved physicochemical properties relative to previously described derivatives of naphthol AS-E phosphate.

The Government of Nepal, with the support of the Japan International Cooperation Agency (JICA), conducted a benchmarking activity with performance indicators, water quality tests, and a consumer survey for 26 larger and older town water supply services in the country. Performance indicators varied widely between the different towns: number of staff/1,000 connection (staff ratio), for example, ranged from 2.8 to 15.3, whereas water supply coverage ranged from 13.8% to 98.4%, and operating ratios which indicate financial performance ranged from 0.24 to 2.8. Critically poor biological water quality was found in water quality tests that 55% of tap water samples were Escherichia coli-contaminated. Overall, customer satisfaction ranged between 14% and 97%. Analysis showed that the operating ratio was significantly correlated with the staff ratio, the number of water supply connections, the water coverage, and the metered ratio. Results of water quality tests and the consumer survey revealed risk in direct drinking of tap water and a need to improve tap water quality by managing free residual chlorine and leakage reduction. Customers' satisfaction with water sufficiency was found to be significantly correlated with their perception of the reliability of supply time rather than the number of supply hours and water consumption rate. This article has been made Open Access thanks to the generous support of a global network of libraries as part of the Knowledge Unlatched Select initiative.

Ribosomal S6 Kinases (RSKs) are a group of serine/threonine kinases that function downstream of the Ras/Raf/MEK/ERK signaling pathway. Four RSK isoforms are directly activated by ERK1/2 in response to extracellular stimuli including growth factors, hormones, and chemokines. RSKs phosphorylate many cytosolic and nuclear targets resulting in the regulation of diverse cellular processes such as cell proliferation, survival, and motility. In hematological malignancies such as acute myeloid leukemia (AML), RSK isoforms are highly expressed and aberrantly activated resulting in poor outcomes and resistance to chemotherapy. Therefore, understanding RSK function in leukemia could lead to promising therapeutic strategies. This review summarizes the current information on human RSK isoforms and discusses their potential roles in the pathogenesis of AML and mechanism of pharmacological inhibitors.

Background Targeted anticancer therapies such as BCR-ABL tyrosine kinase inhibitors (TKIs) have improved outcomes for chronic myeloid leukemia (CML) and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph + ALL). However, little is known about long-term risks of TKIs in children. Exposure-based survivorship guidelines do not include TKIs, thus surveillance practices may be variable. Methods We retrospectively examined surveillance for cardiac and endocrine late effects in children receiving TKIs for Ph + leukemias, diagnosed at Results 66 patients (CML n = 44; Ph + ALL n = 22) met inclusion criteria. Among patients with CML, ≥1 evaluation was done: ECHO (50.0%), EKG (48.8%), TSH (43.9%), DXA (2.6%), bone age (7.4%). Among patients with Ph + ALL, ≥1 evaluation was done: ECHO (86.4%), EKG (68.2%), TSH (59.1%), DXA (63.6%), bone age (44.4%). Over a median 6.3 and 5.7 years of observation, respectively, 2% of patients with CML and 57% with Ph + ALL attended a survivorship clinic. Conclusions Despite common exposure to TKIs in survivors of Ph + leukemias, patterns of surveillance for late effects differed in CML and Ph + ALL, with the latter receiving more surveillance likely due to concomitant chemotherapy exposures. Targeted therapies such as TKIs are revolutionizing cancer treatment, but surveillance for late effects and referral to survivorship clinics are variable despite the chronicity of exposure. Evidence based guidelines and longer follow-up are needed.

Highlights • Transitional high-energy diets reduce peripheral blood lymphocytes in dairy cows. • High-energy diets upregulate IL-1β and IL-2 and downregulate IL-10 expression. • Functional lactobacillus plantarum LP1 restores normal levels of lymphocytes subset. • Lactobacillus plantarum LP1-added diets reduce inflammatory cytokine expression. • LP1 mitigates immune response imbalances caused by transitional high energy diets., High-yield dairy cows need high energy feed during periods of increased milk production. The transitional feeding to high energy feed increases the risk of developing a variety of metabolic disorders. Here, five Holstein cows were fed a four-stage feeding protocol (3 weeks for each stage) ranging from 54.9 to 73.7% total digestive nutrients (TDN). The purpose of the study was to investigate the effect of lactic acid bacteria on high-energy-fed cows associated with transitional feeding, and to evaluate the effects of probiotics on intestinal bacterial changes and inflammatory responses. Three feed transition periods were established for five cows, and Lactobacillus plantarum RGU-LP1 (LP1) was fed as a probiotic during the high-energy feeding period. The number of lymphocyte subsets such as CD3-, CD4-, and CD8 positive cells decreased in response to the high energy feed. Lipopolysaccharide (LPS)-induced cytokine (IL-1β and IL-2) gene expression in peripheral blood mononuclear cells (PBMCs) was shown to increase in those animals receiving the high energy feed. However, supplementation with LP1 resulted in an increase in the number of lymphocyte subsets and the expression of IL-1β and IL-2 were returned to the level at low energy diet. These results suggest that high energy diets induce inflammatory cytokine responses following LPS stimulation, and that the addition of LP1 mitigates these results by regulating the LPS-induced inflammatory reaction. Therefore, the functional lactic acid bacteria LP1 is expected to regulate inflammation resulting from high energy feeding, and this probiotic could be applied to support inflammatory regulation in high-yield dairy cows.

Chronic myelogenous leukemia (CML) is rare in children, requiring extrapolation from treatment of adults. In this review, we explore similarities and differences between adult and pediatric CML with a focus on therapeutic advances and emerging clinical questions. Pediatric CML is effectively treated with long-term targeted therapy using tyrosine kinase inhibitors (TKIs). Newly diagnosed pediatric patients in chronic phase can now be treated with imatinib, dasatinib, or nilotinib without allogeneic hematopoietic stem cell transplantation. While treatment-free remission is possible in adults in chronic phase with optimal response to therapy, data are currently insufficient to support stopping TKI in pediatrics outside of a clinical trial. Knowledge gaps remain regarding long-term and late effects of TKIs in pediatric CML. Targeted therapy has markedly improved outcomes for pediatric CML, while raising a number of clinical questions, including the possibility of treatment-free remission and long-term health implications of prolonged TKI exposure at a young age.

Chronic myelogenous leukemia (CML) is rare in children, requiring extrapolation from treatment of adults. In this review, we explore similarities and differences between adult and pediatric CML with a focus on therapeutic advances and emerging clinical questions.Pediatric CML is effectively treated with long-term targeted therapy using tyrosine kinase inhibitors (TKIs). Newly diagnosed pediatric patients in chronic phase can now be treated with imatinib, dasatinib, or nilotinib without allogeneic hematopoietic stem cell transplantation. While treatment-free remission is possible in adults in chronic phase with optimal response to therapy, data are currently insufficient to support stopping TKI in pediatrics outside of a clinical trial. Knowledge gaps remain regarding long-term and late effects of TKIs in pediatric CML. Targeted therapy has markedly improved outcomes for pediatric CML, while raising a number of clinical questions, including the possibility of treatment-free remission and long-term health implications of prolonged TKI exposure at a young age.

A divertor simulation experimental module (D-module) in the tandem mirror GAMMA 10/PDX was used for the study of plasma detachment. In previous studies, it was difficult to measure far-upstream plasma parameters in the D-module, and only electrostatic probes on the target plate were used to perform electron temperature and density measurements. To study the detached plasma structure, a Thomson scattering (TS) system and a microwave interferometer system have been installed to measure the inside plasma parameters of the D-module, and a movable electrostatic probe has been placed at the inlet of the D-module to measure the inlet plasma density and temperature. The TS system in the central cell observed the electron temperature and density of the core plasma simultaneously. These measurements revealed the entire density and temperature structure from the core plasma to the divertor plate. The line average electron density measured by the microwave interferometer showed a rollover behavior during detachment. The results indicated that the ionization region was located around the center of the D-module, and it appears to move upstream along the axis. See for correction on original figures 1&5, Publisher's note AIP Advances 12, 029902 (2022); DOI: https://doi.org/10.1063/5.0085087

This chapter focuses on the development of therapeutics that work to induce the degradation of nuclear receptors. These therapeutics, unlike traditional small molecule inhibitors, work by redirecting the cellular waste disposal machinery to remove a therapeutic target. They are advantageous because they work through a catalytic mechanism, raising the possibility of drugs with enhanced potency. Current work in the field is toward identifying novel molecular “glues,” in the hopes of bringing proteolysis targeting chimeras (PROTACs) to the clinic. However, translating it from a concept dreamed up in a laboratory to a clinically viable tool took almost 20 years of basic research.

BackgroundAnti-melanoma differentiation-associated gene 5–positive dermatomyositis (MDA5 DM) is a rare but distinct subtype of idiopathic inflammatory myopathy (IIM) that is characterized by high mortality due to rapid progressive interstitial lung disease (ILD). MDA5 is a cytosolic protein and a family of retinoic-acid inducible gene-I (RIG-I) like receptor, which functions as a virus RNA sensor and induces the production of type I interferons (IFN-I) and proinflammatory cytokines. This leads to the death of infected cells and the activation of the antigen-specific antiviral immune response. Although the pathogenesis of MDA5 DM is largely unknown, a hypothesis that pathogenic involvement of anti-MDA5 antibodies has been proposed. Recently, similarities have been noted between multifaceted features of COVID-19 and MDA5 DM, which suggests shared underlying autoinflammatory mechanisms.ObjectivesTo detect the critical actors in the pathogenesis of MDA5 DM by gene expression analysis of peripheral blood.MethodsTotal of 31 DM cases were investigated, including anti-aminoacyl-tRNA synthetase positive (ARS) DM (n=12), MDA5 DM (n=7, survivor=3) and others (n=12). Peripheral blood was drawn at baseline and 2 to 3 months after treatments. Total RNAs were then extracted with using PAXgene miRNA kit. After quantifying the expressions of transcripts by multiplex sequencing. And then, hierarchical clustering analysis, enrichment analysis using gene ontology (GO) terms, single sample gene set enrichment analysis (ssGSEA) and weighted gene co-expression network analysis (WGCNA) were performed.ResultsThe hierarchical clustering with expression profiles of peripheral blood at baseline showed major 3 clusters. Interestingly, ARS DM cases were segregated into right side of the 3rd cluster while MDA5 DM cases fell into 1st and 2nd clusters. ARS and MDA5 DM were clearly discriminated if differentially expressed genes (DEGs) between these subtypes of DM were analyzed. By GO enrichment analysis, the terms, such as related to “defense response to virus” including “type1 interferon signaling pathway” were found in the DEGs. In the MDA5 DM cases, ssGSEA revealed that genes of “Fcγ receptor mediated phagocytosis pathway” or “complement and coagulation cascade” were significantly enriched and WGCNA showed that pathways of “T-cell antigen receptor signaling” or “lung fibrosis” were significantly upregulated. Next, we also investigated the DEGs of peripheral blood at 2-3 months after treatment between survival and fatal cases in MDA5 DM. We found that suppressing RIG-I like receptor and type 1 and type 2 interferon (IFN) signaling were the keys for survival.ConclusionMDA5 is a key sensor of several RNA viruses including coronavirus families and then activate antiviral gene transcription such as type 1 IFN genes, leading to establish an antiviral host response. As the pulmonary damage of COVID-19 is known to be difficult to distinguish from the ILD associated with anti-MDA5 DM, the life-threating ILD of MDA5 DM may be caused by the over-activation of RIG-I like receptor signaling via MDA5. The hypothesis is supported by our findings that the defining features of MDA5 DM are activation of “type 1 IFN pathways” and antigen-specific antiviral immune responses including “Fcγ receptor mediated phagocytosis pathway” or “T-cell antigen receptor signaling”. As the levels of anti-MDA5 antibodies reported to be important prognostic parameter, it may be involved in pathogenesis of MDA5 DM. As we found that suppression of type 1 and type 2 IFN signaling were the keys for survival, it seems to be reasonable to use inhibitors of Janus Kinases (JAK) for treatment of MDA5 DM.Disclosure of InterestsYoshinobu Koyama Speakers bureau: Abbvie, Asahikasei, Ayumi, BMS, Eli-Lilly, Mitsubishi Tanabe, Grant/research support from: Abbvie, GSK, Yoshiharu Sato: None declared, Moe Sakamoto: None declared

Introduction Data on the risk factors for symptomatic radiation pneumonitis (RP) in non–small-cell lung cancer (NSCLC) patients treated with concurrent chemoradiotherapy (CCRT) and consolidation durvalumab are limited; we aimed to investigate these risk factors. Materials and Methods This multicenter retrospective study, conducted at 15 institutions in Japan, included patients who were ≥20 years of age; who started definitive CCRT for NSCLC between July 1, 2018, and July 31, 2019; and who then received durvalumab. The primary endpoint was grade 2 or worse (grade 2+) RP. Results In the 146 patients analyzed, the median follow-up period was 16 months. A majority of the patients had stage III disease (86%), received radiation doses of 60 to 66 Gy equivalent in 2-Gy fractions (93%) and carboplatin and paclitaxel/nab-paclitaxel (77%), and underwent elective nodal irradiation (71%) and 3-dimensional conformal radiotherapy (75%). RP grade 2 was observed in 44 patients (30%); grade 3, in four patients (3%); grade 4, in one patient (1%); and grade 5, in one patient (1%). In the multivariable analysis, lung V20 was a significant risk factor, whereas age, sex, smoking history, irradiation technique, and chemotherapy regimen were not. The 12-month grade 2+ RP incidence was 34.4% (95% confidence interval [CI], 26.7%-42.1%); the values were 50.0% (95% CI, 34.7%-63.5%) and 27.1% (95% CI, 18.8%-36.2%) in those with lung V20 ≥ 26% and Conclusion The incidence of grade 2+ RP was relatively high in this multicenter real-world study, and its risk increased remarkably at elevated lung V20. Our findings can aid in RP risk prediction and the safe radiotherapy treatment planning.