Understanding immunity in a tissue-centric context: Combining novel imaging methods and mathematics to extract new insights into function and dysfunction.

A central question in immunology is what features allow the immune system to respond in a timely manner to a variety of pathogens encountered at unanticipated times and diverse body sites. Two decades of advanced and static dynamic imaging methods have now revealed several major principles facilitating host defense. Suborgan spatial prepositioning of distinct cells promotes time-efficient interactions upon pathogen sensing. Such pre-organization also provides an effective barrier to movement of pathogens from parenchymal tissues into the blood circulation. Various molecular mechanisms maintain effective intercellular communication among otherwise rapidly moving cells. These and related discoveries have benefited from recent increases in the number of parameters that can be measured simultaneously in a single tissue section and the extension of such multiplex analyses to 3D tissue volumes. The application of new computational methods to such imaging data has provided a quantitative, in vivo context for cell trafficking and signaling pathways traditionally explored in vitro or with dissociated cell preparations. Here, we summarize our efforts to devise and employ diverse imaging tools to probe immune system organization and function, concluding with a commentary on future developments, which we believe will reveal even more about how the immune system operates in health and disease.

Facilitators and barriers for implementation of a novel resuscitation quality improvement package in public referral hospitals of Nepal.

BACKGROUND: Improving the healthcare providers (HCP) basic resuscitation skills can reduce intrapartum related mortality in low- and middle-income countries. However, the resuscitation intervention's successful implementation is largely dependent on proper facilitation and context. This study aims to identify the facilitators and barriers for the implementation of a novel resuscitation package as part of the quality improvement project in Nepal. METHODS: The study used a qualitative descriptive design. The study sites included four purposively chosen public hospitals in Nepal, where the resuscitation package (Helping Babies Breathe [HBB] training, resuscitation equipment and NeoBeat) had been implemented as part of the quality improvement project. Twenty members of the HCP, who were trained and exposed to the package, were selected through convenience sampling to participate in the study interviews. Data were collected through semi-structured interviews conducted via telephone and video calls. Twenty interview data were analyzed with a deductive qualitative content analysis based on the core components of the i-PARiHS framework. RESULTS: The findings suggest that there was a move to more systematic resuscitation practices among the staff after the quality improvement project's implementation. This positive change was supported by a neonatal heart rate monitor (NeoBeat), which guided resuscitation and made it easier. In addition, seeing the positive outcomes of successful resuscitation motivated the HCPs to keep practicing and developing their resuscitation skills. Facilitation by the project staff enabled the change. At the same time, facilitators provided extra support to maintain the equipment, which can be a challenge in terms of sustainability, after the project. Furthermore, a lack of additional resources, an unclear leadership role, and a lack of coordination between nurses and medical doctors were barriers to the implementation of the resuscitation package. CONCLUSION: The introduction of the resuscitation package, as well as the continuous capacity building of local multidisciplinary healthcare staff, is important to continue the accelerated efforts of improving newborn care. To secure sustainable change, facilitation during implementation should focus on exploring local resources to implement the resuscitation package sustainably. TRIAL REGISTRATION: Not applicable.

IBEX: A versatile multiplex optical imaging approach for deep phenotyping and spatial analysis of cells in complex tissues.

The diverse composition of mammalian tissues poses challenges for understanding the cell-cell interactions required for organ homeostasis and how spatial relationships are perturbed during disease. Existing methods such as single-cell genomics, lacking a spatial context, and traditional immunofluorescence, capturing only two to six molecular features, cannot resolve these issues. Imaging technologies have been developed to address these problems, but each possesses limitations that constrain widespread use. Here we report a method that overcomes major impediments to highly multiplex tissue imaging. "Iterative bleaching extends multiplexity" (IBEX) uses an iterative staining and chemical bleaching method to enable high-resolution imaging of >65 parameters in the same tissue section without physical degradation. IBEX can be employed with various types of conventional microscopes and permits use of both commercially available and user-generated antibodies in an "open" system to allow easy adjustment of staining panels based on ongoing marker discovery efforts. We show how IBEX can also be used with amplified staining methods for imaging strongly fixed tissues with limited epitope retention and with oligonucleotide-based staining, allowing potential cross-referencing between flow cytometry, cellular indexing of transcriptomes and epitopes by sequencing, and IBEX analysis of the same tissue. To facilitate data processing, we provide an open-source platform for automated registration of iterative images. IBEX thus represents a technology that can be rapidly integrated into most current laboratory workflows to achieve high-content imaging to reveal the complex cellular landscape of diverse organs and tissues.

Chlorhexidine for facility-based umbilical cord care: EN-BIRTH multi-country validation study.

BACKGROUND: Umbilical cord hygiene prevents sepsis, a leading cause of neonatal mortality. The World Health Organization recommends 7.1% chlorhexidine digluconate (CHX) application to the umbilicus after home birth in high mortality contexts. In Bangladesh and Nepal, national policies recommend CHX use for all facility births. Population-based household surveys include optional questions on CHX use, but indicator validation studies are lacking. The Every Newborn Birth Indicators Research Tracking in Hospitals (EN-BIRTH) was an observational study assessing measurement validity for maternal and newborn indicators. This paper reports results regarding CHX. METHODS: The EN-BIRTH study (July 2017-July 2018) included three public hospitals in Bangladesh and Nepal where CHX cord application is routine. Clinical-observers collected tablet-based, time-stamped data regarding cord care during admission to labour and delivery wards as the gold standard to assess accuracy of women's report at exit survey, and of routine-register data. We calculated validity ratios and individual-level validation metrics; analysed coverage, quality and measurement gaps. We conducted qualitative interviews to assess barriers and enablers to routine register-recording. RESULTS: Umbilical cord care was observed for 12,379 live births. Observer-assessed CHX coverage was very high at 89.3-99.4% in all 3 hospitals, although slightly lower after caesarean births in Azimpur (86.8%), Bangladesh. Exit survey-reported coverage (0.4-45.9%) underestimated the observed coverage with substantial "don't know" responses (55.5-79.4%). Survey-reported validity ratios were all poor (0.01 to 0.38). Register-recorded coverage in the specific column in Bangladesh was underestimated by 0.2% in Kushtia but overestimated by 9.0% in Azimpur. Register-recorded validity ratios were good (0.9 to 1.1) in Bangladesh, and poor (0.8) in Nepal. The non-specific register column in Pokhara, Nepal substantially underestimated coverage (20.7%). CONCLUSIONS: Exit survey-report highly underestimated observed CHX coverage in all three hospitals. Routine register-recorded coverage was closer to observer-assessed coverage than survey reports in all hospitals, including for caesarean births, and was more accurately captured in hospitals with a specific register column. Inclusion of CHX cord care into registers, and tallied into health management information system platforms, is justified in countries with national policies for facility-based use, but requires implementation research to assess register design and data flow within health information systems.

Uterotonics for prevention of postpartum haemorrhage: EN-BIRTH multi-country validation study.

BACKGROUND: Postpartum haemorrhage (PPH) is a leading cause of preventable maternal mortality worldwide. The World Health Organization (WHO) recommends uterotonic administration for every woman after birth to prevent PPH. There are no standardised data collected in large-scale measurement platforms. The Every Newborn Birth Indicators Research Tracking in Hospitals (EN-BIRTH) is an observational study to assess the validity of measurement of maternal and newborn indicators, and this paper reports findings regarding measurement of coverage and quality for uterotonics. METHODS: The EN-BIRTH study took place in five hospitals in Bangladesh, Nepal and Tanzania, from July 2017 to July 2018. Clinical observers collected tablet-based, time-stamped data. We compared observation data for uterotonics to routine hospital register-records and women's report at exit-interview survey. We analysed the coverage and quality gap for timing and dose of administration. The register design was evaluated against gap analyses and qualitative interview data assessing the barriers and enablers to data recording and use. RESULTS: Observed uterotonic coverage was high in all five hospitals (> 99%, 95% CI 98.7-99.8%). Survey-report underestimated coverage (79.5 to 91.7%). "Don't know" replies varied (2.1 to 14.4%) and were higher after caesarean (3.7 to 59.3%). Overall, there was low accuracy in survey data for details of uterotonic administration (type and timing). Register-recorded coverage varied in four hospitals capturing uterotonics in a specific column (21.6, 64.5, 97.6, 99.4%). The average coverage measurement gap was 18.1% for register-recorded and 6.0% for survey-reported coverage. Uterotonics were given to 15.9% of women within the "right time" (1 min) and 69.8% within 3 min. Women's report of knowing the purpose of uterotonics after birth ranged from 0.4 to 64.9% between hospitals. Enabling register design and adequate staffing were reported to improve routine recording. CONCLUSIONS: Routine registers have potential to track uterotonic coverage - register data were highly accurate in two EN-BIRTH hospitals, compared to consistently underestimated coverage by survey-report. Although uterotonic coverage was high, there were gaps in observed quality for timing and dose. Standardisation of register design and implementation could improve data quality and data flow from registers into health management information reporting systems, and requires further assessment.

Barriers and enablers to routine register data collection for newborns and mothers: EN-BIRTH multi-country validation study.

BACKGROUND: Policymakers need regular high-quality coverage data on care around the time of birth to accelerate progress for ending preventable maternal and newborn deaths and stillbirths. With increasing facility births, routine Health Management Information System (HMIS) data have potential to track coverage. Identifying barriers and enablers faced by frontline health workers recording HMIS source data in registers is important to improve data for use. METHODS: The EN-BIRTH study was a mixed-methods observational study in five hospitals in Bangladesh, Nepal and Tanzania to assess measurement validity for selected Every Newborn coverage indicators. We described data elements required in labour ward registers to track these indicators. To evaluate barriers and enablers for correct recording of data in registers, we designed three interview tools: a) semi-structured in-depth interview (IDI) guide b) semi-structured focus group discussion (FGD) guide, and c) checklist assessing care-to-documentation. We interviewed two groups of respondents (January 2018-March 2019): hospital nurse-midwives and doctors who fill ward registers after birth (n = 40 IDI and n = 5 FGD); and data collectors (n = 65). Qualitative data were analysed thematically by categorising pre-identified codes. Common emerging themes of barriers or enablers across all five hospitals were identified relating to three conceptual framework categories. RESULTS: Similar themes emerged as both barriers and enablers. First, register design was recognised as crucial, yet perceived as complex, and not always standardised for necessary data elements. Second, register filling was performed by over-stretched nurse-midwives with variable training, limited supervision, and availability of logistical resources. Documentation complexity across parallel documents was time-consuming and delayed because of low staff numbers. Complete data were valued more than correct data. Third, use of register data included clinical handover and monthly reporting, but little feedback was given from data users. CONCLUSION: Health workers invest major time recording register data for maternal and newborn core health indicators. Improving data quality requires standardised register designs streamlined to capture only necessary data elements. Consistent implementation processes are also needed. Two-way feedback between HMIS levels is critical to improve performance and accurately track progress towards agreed health goals.

Prognostic significance of stem cell/ epithelial-mesenchymal transition markers in periampullary/pancreatic cancers: FGFR1 is a promising prognostic marker.

BACKGROUND: Periampullary cancers (PAC) including pancreatic, ampulla of Vater (AOV), and common bile duct (CBD) cancers are highly aggressive with a lack of useful prognostic markers beyond T stage. However, T staging can be biased due to the anatomic complexity of this region. Recently, several markers related to cancer stem cells and epithelial-mesenchymal transition (EMT) such as octamer transcription factor-4 (Oct4) and fibroblast growth factor receptor 1 (FGFR1) respectively, have been proposed as new promising markers in other solid cancers. The aim of this study was to assess the expression and prognostic significance of stem cell/EMT markers in PACs. METHODS: Formalin-fixed, paraffin-embedded tissues of surgically excised PACs from the laboratory archives from 1998 to 2014 were evaluated by immunohistochemical staining for stem cell/EMT markers using tissue microarray. The clinicopathologic parameters were documented and statistically analyzed with the immunohistochemical findings. Survival and recurrence data were collected and analyzed. RESULTS: A total of 126 PAC cases were evaluated. The average age was 63 years, with 76 male and 50 female patient samples. Age less than 74 years, AOV cancers, lower T & N stage, lower tumor size, no lymphatic, vascular, perineural invasion and histologic well differentiation, intestinal type, no fibrosis, severe inflammation were significantly associated with the better overall survival High expression levels of FGFR1 as well as CK20, CDX2, and VEGF were significantly related to better overall survival, while other stem cell markers were not related. Similar findings were observed for tumor recurrence using disease-free survival. CONCLUSIONS: In addition to other clinicopathologic parameters, severe fibrosis was related to frequent tumor recurrence, and high FGFR1 expression was associated with better overall survival. Histologic changes such as extensive fibrosis need to be investigated further in relation to EMT of PACs.

Comparative Genomic Analysis of Drechmeria coniospora Reveals Core and Specific Genetic Requirements for Fungal Endoparasitism of Nematodes.

Drechmeria coniospora is an obligate fungal pathogen that infects nematodes via the adhesion of specialized spores to the host cuticle. D. coniospora is frequently found associated with Caenorhabditis elegans in environmental samples. It is used in the study of the nematode's response to fungal infection. Full understanding of this bi-partite interaction requires knowledge of the pathogen's genome, analysis of its gene expression program and a capacity for genetic engineering. The acquisition of all three is reported here. A phylogenetic analysis placed D. coniospora close to the truffle parasite Tolypocladium ophioglossoides, and Hirsutella minnesotensis, another nematophagous fungus. Ascomycete nematopathogenicity is polyphyletic; D. coniospora represents a branch that has not been molecularly characterized. A detailed in silico functional analysis, comparing D. coniospora to 11 fungal species, revealed genes and gene families potentially involved in virulence and showed it to be a highly specialized pathogen. A targeted comparison with nematophagous fungi highlighted D. coniospora-specific genes and a core set of genes associated with nematode parasitism. A comparative gene expression analysis of samples from fungal spores and mycelia, and infected C. elegans, gave a molecular view of the different stages of the D. coniospora lifecycle. Transformation of D. coniospora allowed targeted gene knock-out and the production of fungus that expresses fluorescent reporter genes. It also permitted the initial characterisation of a potential fungal counter-defensive strategy, involving interference with a host antimicrobial mechanism. This high-quality annotated genome for D. coniospora gives insights into the evolution and virulence of nematode-destroying fungi. Coupled with genetic transformation, it opens the way for molecular dissection of D. coniospora physiology, and will allow both sides of the interaction between D. coniospora and C. elegans, as well as the evolutionary arms race that exists between pathogen and host, to be studied.

Coordinated inhibition of C/EBP by Tribbles in multiple tissues is essential for Caenorhabditis elegans development.

BACKGROUND: Tribbles proteins are conserved pseudokinases that function to control kinase signalling and transcription in diverse biological processes. Abnormal function in human Tribbles has been implicated in a number of diseases including leukaemia, metabolic syndromes and cardiovascular diseases. Caenorhabditis elegans Tribbles NIPI-3 was previously shown to activate host defense upon infection by promoting the conserved PMK-1/p38 mitogen-activated protein kinase (MAPK) signalling pathway. Despite the prominent role of Tribbles proteins in many species, our knowledge of their mechanism of action is fragmented, and the in vivo functional relevance of their interactions with other proteins remains largely unknown. RESULTS: Here, by characterizing nipi-3 null mutants, we show that nipi-3 is essential for larval development and viability. Through analyses of genetic suppressors of nipi-3 null mutant lethality, we show that NIPI-3 negatively controls PMK-1/p38 signalling via transcriptional repression of the C/EBP transcription factor CEBP-1. We identified CEBP-1's transcriptional targets by ChIP-seq analyses and found them to be enriched in genes involved in development and stress responses. Unlike its cell-autonomous role in innate immunity, NIPI-3 is required in multiple tissues to control organismal development. CONCLUSIONS: Together, our data uncover an unprecedented crosstalk involving multiple tissues, in which NIPI-3 acts as a master regulator to inhibit CEBP-1 and the PMK-1/p38 MAPK pathway. In doing so, it keeps innate immunity in check and ensures proper organismal development.

A quantitative genome-wide RNAi screen in C. elegans for antifungal innate immunity genes.

BACKGROUND: Caenorhabditis elegans has emerged over the last decade as a useful model for the study of innate immunity. Its infection with the pathogenic fungus Drechmeria coniospora leads to the rapid up-regulation in the epidermis of genes encoding antimicrobial peptides. The molecular basis of antimicrobial peptide gene regulation has been previously characterized through forward genetic screens. Reverse genetics, based on RNAi, provide a complementary approach to dissect the worm's immune defenses. RESULTS: We report here the full results of a quantitative whole-genome RNAi screen in C. elegans for genes involved in regulating antimicrobial peptide gene expression. The results will be a valuable resource for those contemplating similar RNAi-based screens and also reveal the limitations of such an approach. We present several strategies, including a comprehensive class clustering method, to overcome these limitations and which allowed us to characterize the different steps of the interaction between C. elegans and the fungus D. coniospora, leading to a complete description of the MAPK pathway central to innate immunity in C. elegans. The results further revealed a cross-tissue signaling, triggered by mitochondrial dysfunction in the intestine, that suppresses antimicrobial peptide gene expression in the nematode epidermis. CONCLUSIONS: Overall, our results provide an unprecedented system's level insight into the regulation of C. elegans innate immunity. They represent a significant contribution to our understanding of host defenses and will lead to a better comprehension of the function and evolution of animal innate immunity.

AVPdb: a database of experimentally validated antiviral peptides targeting medically important viruses.

Antiviral peptides (AVPs) have exhibited huge potential in inhibiting viruses by targeting various stages of their life cycle. Therefore, we have developed AVPdb, available online at http://crdd.osdd.net/servers/avpdb, to provide a dedicated resource of experimentally verified AVPs targeting over 60 medically important viruses including Influenza, HCV, HSV, RSV, HBV, DENV, SARS, etc. However, we have separately provided HIV inhibiting peptides in 'HIPdb'. AVPdb contains detailed information of 2683 peptides, including 624 modified peptides experimentally tested for antiviral activity. In modified peptides a chemical moiety is attached for increasing their efficacy and stability. Detailed information include: peptide sequence, length, source, virus targeted, virus family, cell line used, efficacy (qualitative/quantitative), target step/protein, assay used in determining the efficacy and PubMed reference. The database also furnishes physicochemical properties and predicted structure for each peptide. We have provided user-friendly browsing and search facility along with other analysis tools to help the users. Entering of many synthetic peptide-based drugs in various stages of clinical trials reiterate the importance for the AVP resources. AVPdb is anticipated to cater to the needs of scientific community working for the development of antiviral therapeutics.

VIRsiRNAdb: a curated database of experimentally validated viral siRNA/shRNA.

RNAi technology has been emerging as a potential modality to inhibit viruses during past decade. In literature a few siRNA databases have been reported that focus on targeting human and mammalian genes but experimentally validated viral siRNA databases are lacking. We have developed VIRsiRNAdb, a manually curated database having comprehensive details of 1358 siRNA/shRNA targeting viral genome regions. Further, wherever available, information regarding alternative efficacies of above 300 siRNAs derived from different assays has also been incorporated. Important fields included in the database are siRNA sequence, virus subtype, target genome region, cell type, target object, experimental assay, efficacy, off-target and siRNA matching with reference viral sequences. Database also provides the users with facilities of advance search, browsing, data submission, linking to external databases and useful siRNA analysis tools especially siTarAlign which align the siRNA with reference viral genomes or user defined sequences. VIRsiRNAdb contains extensive details of siRNA/shRNA targeting 42 important human viruses including influenza virus, hepatitis B virus, HPV and SARS Corona virus. VIRsiRNAdb would prove useful for researchers in picking up the best viral siRNA for antiviral therapeutics development and also for developing better viral siRNA design tools. The database is freely available at http://crdd.osdd.net/servers/virsirnadb.

AVPpred: collection and prediction of highly effective antiviral peptides.

In the battle against viruses, antiviral peptides (AVPs) had demonstrated the immense potential. Presently, more than 15 peptide-based drugs are in various stages of clinical trials. Emerging and re-emerging viruses further emphasize the efforts to accelerate antiviral drug discovery efforts. Despite, huge importance of the field, no dedicated AVP resource is available. In the present study, we have collected 1245 peptides which were experimentally checked for antiviral activity targeting important human viruses like influenza, HIV, HCV and SARS, etc. After removing redundant peptides, 1056 peptides were divided into 951 training and 105 validation data sets. We have exploited various peptides sequence features, i.e. motifs and alignment followed by amino acid composition and physicochemical properties during 5-fold cross validation using Support Vector Machine. Physiochemical properties-based model achieved maximum 85% accuracy and 0.70 Matthew's Correlation Coefficient (MCC). Performance of this model on the experimental validation data set showed 86% accuracy and 0.71 MCC which is far better than the general antimicrobial peptides prediction methods. Therefore, AVPpred-the first web server for predicting the highly effective AVPs would certainly be helpful to researchers working on peptide-based antiviral development. The web server is freely available at http://crdd.osdd.net/servers/avppred.

Spatial Patterning Analysis of Cellular Ensembles (SPACE) discovers complex spatial organization at the cell and tissue levels.

Spatial patterns of cells and other biological elements drive both physiologic and pathologic processes within tissues. While many imaging and transcriptomic methods document tissue organization, discerning these patterns is challenging, especially when they involve multiple elements in complex arrangements. To address this challenge, we present Spatial Patterning Analysis of Cellular Ensembles (SPACE), an R package for analysis of high-plex spatial data. SPACE is compatible with any data collection modality that records values (i.e., categorical cell/structure types or quantitative expression levels) at fixed spatial coordinates (i.e., 2d pixels or 3d voxels). SPACE detects not only broad patterns of co-occurrence but also context-dependent associations, quantitative gradients and orientations, and other organizational complexities. Via a robust information theoretic framework, SPACE explores all possible ensembles of tissue elements - single elements, pairs, triplets, and so on - and ranks the most strongly patterned ensembles. For single images, rankings reflect patterns that differ from random assortment. For sets of images, rankings reflect patterns that differ across sample groups (e.g., genotypes, treatments, timepoints, etc.). Further tools then thoroughly characterize the nature of each pattern for intuitive interpretation. We validate SPACE and demonstrate its advantages using murine lymph node images for which ground truth has been defined. We then use SPACE to detect new patterns across varied datasets, including tumors and tuberculosis granulomas.

VIRsiRNApred: a web server for predicting inhibition efficacy of siRNAs targeting human viruses.

BACKGROUND: Selection of effective viral siRNA is an indispensable step in the development of siRNA based antiviral therapeutics. Despite immense potential, a viral siRNA efficacy prediction algorithm is still not available. Moreover, performances of the existing general mammalian siRNA efficacy predictors are not satisfactory for viral siRNAs. Therefore, we have developed "VIRsiRNApred" a support vector machine (SVM) based method for predicting the efficacy of viral siRNA. METHODS: In the present study, we have employed a new dataset of 1725 viral siRNAs with experimentally verified quantitative efficacies tested under heterogeneous experimental conditions and targeting as many as 37 important human viruses including HIV, Influenza, HCV, HBV, SARS etc. These siRNAs were divided into training (T1380) and validation (V345) datasets. Important siRNA sequence features including mono to penta nucleotide frequencies, binary pattern, thermodynamic properties and secondary structure were employed for model development. RESULTS: During 10-fold cross validation on T1380 using hybrid approach, we achieved a maximum Pearson Correlation Coefficient (PCC) of 0.55 between predicted and actual efficacy of viral siRNAs. On V345 independent dataset, our best model achieved a maximum correlation of 0.50 while existing general siRNA prediction methods showed PCC from 0.05 to 0.18. However, using leave one out cross validation PCC was improved to 0.58 and 0.55 on training and validation datasets respectively. SVM performed better than other machine learning techniques used like ANN, KNN and REP Tree. CONCLUSION: VIRsiRNApred is the first algorithm for predicting inhibition efficacy of viral siRNAs which is developed using experimentally verified viral siRNAs. We hope this algorithm would be useful in predicting highly potent viral siRNA to aid siRNA based antiviral therapeutics development. The web server is freely available at http://crdd.osdd.net/servers/virsirnapred/.

Fabrication and Synthesis of Thiococlchicoside loaded Matrix type Transdermal Patch.

BACKGROUND: The goal of this work was to synthesize and fabricate matrix type transdermal patches based on a combination of polymers (Eudragit L100, HPMC and PVP K30), plasticizer and crosslinking agents (propylene glycol and triethyl citrate) and adhesives (Dura Tak 87-6908) to increase Thiocolchicoside (THC) absorption via topical route. This method allows avoidance of first-pass metabolism along with a consistent and extended duration of therapeutic activity. METHODS: Fabrication and casting of polymeric solutions containing THC was done either in petri plates or through lab coater to get transdermal patches. Finally, the formulated patches were studied for their physicochemical and biological evaluation using scanning electron microscopy, FTIR, DSC, XRD and ex-vivo permeation studies using pig ear skin. RESULTS: FTIR studies confirm that the THC characteristics peaks (carbonyl (Amide I) at 1525.5 cm-1, C=O stretching (tropane ring) at 1664.4 cm-1, Amide II band (N-H stretching) at 3325.9cm-1, thioether band at 2360.7cm-1, and OH group stretching band at 3400.2 cm-1) are still present in the polymer mixture even after formulation as a transdermal patch, indicating compatibility among all excipients. While on the other hand, DSC studies confirm endothermic peaks for all the polymers along with THC with the highest enthalpy of 65.979 J/g, which is an indicator of sharp endothermic peak at 198℃, leading to the melting of THC. The percentage drug content and percentage moisture uptake of all the formulation was found in the range of 96±2.04 to 98.56±1.34% and 4.13±1.16 to 8.23±0.90%, respectively. Drug release and release kinetics studies confirm that it is dependent on the composition of individual formulation. CONCLUSION: All these findings support the possibility of using suitable polymeric composition, as well as proper formulation and manufacturing circumstances, to create a one-of-a-kind technology platform for transdermal drug administration.

MYC drives platinum resistant SCLC that is overcome by the dual PI3K-HDAC inhibitor fimepinostat.

BACKGROUND: Small cell lung cancer (SCLC) is an aggressive neuroendocrine cancer with an appalling overall survival of less than 5% (Zimmerman et al. J Thor Oncol 14:768-83, 2019). Patients typically respond to front line platinum-based doublet chemotherapy, but almost universally relapse with drug resistant disease. Elevated MYC expression is common in SCLC and has been associated with platinum resistance. This study evaluates the capacity of MYC to drive platinum resistance and through screening identifies a drug capable of reducing MYC expression and overcoming resistance. METHODS: Elevated MYC expression following the acquisition of platinum resistance in vitro and in vivo was assessed. Moreover, the capacity of enforced MYC expression to drive platinum resistance was defined in SCLC cell lines and in a genetically engineered mouse model that expresses MYC specifically in lung tumors. High throughput drug screening was used to identify drugs able to kill MYC-expressing, platinum resistant cell lines. The capacity of this drug to treat SCLC was defined in vivo in both transplant models using cell lines and patient derived xenografts and in combination with platinum and etoposide chemotherapy in an autochthonous mouse model of platinum resistant SCLC. RESULTS: MYC expression is elevated following the acquisition of platinum resistance and constitutively high MYC expression drives platinum resistance in vitro and in vivo. We show that fimepinostat decreases MYC expression and that it is an effective single agent treatment for SCLC in vitro and in vivo. Indeed, fimepinostat is as effective as platinum-etoposide treatment in vivo. Importantly, when combined with platinum and etoposide, fimepinostat achieves a significant increase in survival. CONCLUSIONS: MYC is a potent driver of platinum resistance in SCLC that is effectively treated with fimepinostat.

Seroprevalence of SARS-CoV-2 infection in the general population of Nepal during the first and second generalized waves of the COVID-19 pandemic-2020-2021.

Few seroprevalence studies have been conducted on coronavirus disease (COVID-19) in Nepal. Here, we aimed to estimate seroprevalence and assess risk factors for infection in the general population of Nepal by conducting two rounds of sampling. The first round was in October 2020, at the peak of the first generalized wave of COVID-19, and the second round in July-August 2021, following the peak of the wave caused by the delta variant of SARS-CoV-2. We used cross-sectional probability-to-size (PPS)-based multistage cluster sampling to estimate the seroprevalence in the general population of Nepal at the national and provincial levels. We tested for anti-SARS-CoV-2 total antibody using the WANTAI SARS-CoV-2 Ab ELISA kit. In Round 1, the overall national seroprevalence was 14.4%, with provincial estimates ranging from 5.3% in Sudurpaschim to 27.3% in Madhesh Province. In Round 2, the estimated national seroprevalence was 70.7%, with the highest in the Madhesh Province (84.8%) and the lowest in the Gandaki Province (62.9%). Seroprevalence was comparable between males and females (Round 1, 15.8% vs. 12.2% and Round 2, 72.3% vs. 68.7%). The seroprevalence in the ecozones-Terai, hills, and mountains-was 76.3%, 65.3%, and 60.5% in Round 2 and 17.7%, 11.7%, and 4.6% in Round 1, respectively. In Nepal, COVID-19 vaccination was introduced in January 2021. At the peak of the first generalized wave of COVID-19, most of the population of Nepal remained unexposed to SARS-CoV-2. Towards the end of the second generalized wave in April 2021, two thirds of the population was exposed.

Tumor Budding in Gynecologic Cancer as a Marker for Poor Survival: A Systematic Review and Meta-Analysis of the Perspectives of Epithelial-Mesenchymal Transition.

This study aimed to assess the prognostic significance, assessment methods, and molecular features of tumor budding (TB). A literature search of Medline, EMBASE, Cochrane Library, and eleven cohort studies (seven cervical and four endometrial cancers) was conducted. Three assessment methods for TB involving 2009 patients were collected and constituted in the analysis. Our meta-analysis showed that TB was a marker of poor survival, regardless of the cancer origin site or assessment method (overall survival: hazard ratio [HR], 2.40; 95% confidence interval [CI], 1.82-3.17; disease-free survival: HR, 3.32; 95% CI, 2.46-4.48). In endometrial cancers, TB is associated with the epithelial-mesenchymal transition, microvessel density, and decreased hormone receptor expression. Thus, we suggest TB as a poor prognostic marker for all gynecologic cancers.

Recent Application of Artificial Intelligence in Non-Gynecological Cancer Cytopathology: A Systematic Review.

State-of-the-art artificial intelligence (AI) has recently gained considerable interest in the healthcare sector and has provided solutions to problems through automated diagnosis. Cytological examination is a crucial step in the initial diagnosis of cancer, although it shows limited diagnostic efficacy. Recently, AI applications in the processing of cytopathological images have shown promising results despite the elementary level of the technology. Here, we performed a systematic review with a quantitative analysis of recent AI applications in non-gynecological (non-GYN) cancer cytology to understand the current technical status. We searched the major online databases, including MEDLINE, Cochrane Library, and EMBASE, for relevant English articles published from January 2010 to January 2021. The searched query terms were: "artificial intelligence", "image processing", "deep learning", "cytopathology", and "fine-needle aspiration cytology." Out of 17,000 studies, only 26 studies (26 models) were included in the full-text review, whereas 13 studies were included for quantitative analysis. There were eight classes of AI models treated of according to target organs: thyroid (n = 11, 39%), urinary bladder (n = 6, 21%), lung (n = 4, 14%), breast (n = 2, 7%), pleural effusion (n = 2, 7%), ovary (n = 1, 4%), pancreas (n = 1, 4%), and prostate (n = 1, 4). Most of the studies focused on classification and segmentation tasks. Although most of the studies showed impressive results, the sizes of the training and validation datasets were limited. Overall, AI is also promising for non-GYN cancer cytopathology analysis, such as pathology or gynecological cytology. However, the lack of well-annotated, large-scale datasets with Z-stacking and external cross-validation was the major limitation found across all studies. Future studies with larger datasets with high-quality annotations and external validation are required.