A Next Generation Connectivity Map: L1000 Platform and the First 1,000,000 Profiles.

We previously piloted the concept of a Connectivity Map (CMap), whereby genes, drugs, and disease states are connected by virtue of common gene-expression signatures. Here, we report more than a 1,000-fold scale-up of the CMap as part of the NIH LINCS Consortium, made possible by a new, low-cost, high-throughput reduced representation expression profiling method that we term L1000. We show that L1000 is highly reproducible, comparable to RNA sequencing, and suitable for computational inference of the expression levels of 81% of non-measured transcripts. We further show that the expanded CMap can be used to discover mechanism of action of small molecules, functionally annotate genetic variants of disease genes, and inform clinical trials. The 1.3 million L1000 profiles described here, as well as tools for their analysis, are available at https://clue.io.

Aneuploidy renders cancer cells vulnerable to mitotic checkpoint inhibition.

Selective targeting of aneuploid cells is an attractive strategy for cancer treatment1. However, it is unclear whether aneuploidy generates any clinically relevant vulnerabilities in cancer cells. Here we mapped the aneuploidy landscapes of about 1,000 human cancer cell lines, and analysed genetic and chemical perturbation screens2-9 to identify cellular vulnerabilities associated with aneuploidy. We found that aneuploid cancer cells show increased sensitivity to genetic perturbation of core components of the spindle assembly checkpoint (SAC), which ensures the proper segregation of chromosomes during mitosis10. Unexpectedly, we also found that aneuploid cancer cells were less sensitive than diploid cells to short-term exposure to multiple SAC inhibitors. Indeed, aneuploid cancer cells became increasingly sensitive to inhibition of SAC over time. Aneuploid cells exhibited aberrant spindle geometry and dynamics, and kept dividing when the SAC was inhibited, resulting in the accumulation of mitotic defects, and in unstable and less-fit karyotypes. Therefore, although aneuploid cancer cells could overcome inhibition of SAC more readily than diploid cells, their long-term proliferation was jeopardized. We identified a specific mitotic kinesin, KIF18A, whose activity was perturbed in aneuploid cancer cells. Aneuploid cancer cells were particularly vulnerable to depletion of KIF18A, and KIF18A overexpression restored their response to SAC inhibition. Our results identify a therapeutically relevant, synthetic lethal interaction between aneuploidy and the SAC.

Genetic and transcriptional evolution alters cancer cell line drug response.

Human cancer cell lines are the workhorse of cancer research. Although cell lines are known to evolve in culture, the extent of the resultant genetic and transcriptional heterogeneity and its functional consequences remain understudied. Here we use genomic analyses of 106 human cell lines grown in two laboratories to show extensive clonal diversity. Further comprehensive genomic characterization of 27 strains of the common breast cancer cell line MCF7 uncovered rapid genetic diversification. Similar results were obtained with multiple strains of 13 additional cell lines. Notably, genetic changes were associated with differential activation of gene expression programs and marked differences in cell morphology and proliferation. Barcoding experiments showed that cell line evolution occurs as a result of positive clonal selection that is highly sensitive to culture conditions. Analyses of single-cell-derived clones demonstrated that continuous instability quickly translates into heterogeneity of the cell line. When the 27 MCF7 strains were tested against 321 anti-cancer compounds, we uncovered considerably different drug responses: at least 75% of compounds that strongly inhibited some strains were completely inactive in others. This study documents the extent, origins and consequences of genetic variation within cell lines, and provides a framework for researchers to measure such variation in efforts to support maximally reproducible cancer research.

Patient and pathological predictors of management strategy for malignant polyps following polypectomy: a systematic review and meta-analysis.

PURPOSE: Malignant polyps present a treatment dilemma for clinicians and patients. This meta-analysis sought to identify the factors that predicted the management strategy for patients diagnosed with a malignant polyp. METHODS: A literature search was performed following the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) and the Cochrane Collaboration prognostic studies guidelines. Reports from 1985 onwards were included, data on patient and pathological factors were extracted and random effects meta-analysis models were used. RESULTS: Fifteen studies were included. Seven studies evaluated lymphovascular invasion (LVI). The odds of surgery were significantly higher in malignant polyps with LVI (OR 2.20, 95% CI 1.36-3.55). Ten studies revealed the odds of surgery were significantly higher with positive polypectomy margins (OR 8.09, 95% CI 4.88-13.40). Tumour differentiation was compared in eight studies. There were significantly lower odds of surgery in malignant polyps with well/moderate differentiation compared with poor differentiation (OR 0.31, 95% CI 0.21-0.46). There were non-significant trends favouring surgical resection in younger patients, males and Haggitt 4/Kikuchi Sm3 lesions. There was considerable heterogeneity in the meta-analyses for the variables age, gender, polyp morphology and Haggitt/Kikuchi level (I2 > 75%). CONCLUSION: This meta-analysis has demonstrated that LVI, positive polypectomy resection margins, and poor tumour differentiation significantly predict malignant polypectomy patients who underwent subsequent surgery. Age and gender were important factors predicting management, but not consistently across studies, whilst polyp morphology and Haggitt/Kikuchi levels did not significantly predict the management strategy. Further research may assist in understanding the management preferences.

Preclinical Evaluation of a Fluorescent Probe Targeting Receptor CDCP1 for Identification of Ovarian Cancer.

Optimal cytoreduction for ovarian cancer is often challenging because of aggressive tumor biology and advanced stage. It is a critical issue since the extent of residual disease after surgery is the key predictor of ovarian cancer patient survival. For a limited number of cancers, fluorescence-guided surgery has emerged as an effective aid for tumor delineation and effective cytoreduction. The intravenously administered fluorescent agent, most commonly indocyanine green (ICG), accumulates preferentially in tumors, which are visualized under a fluorescent light source to aid surgery. Insufficient tumor specificity has limited the broad application of these agents in surgical oncology including for ovarian cancer. In this study, we developed a novel tumor-selective fluorescent agent by chemically linking ICG to mouse monoclonal antibody 10D7 that specifically recognizes an ovarian cancer-enriched cell surface receptor, CUB-domain-containing protein 1 (CDCP1). 10D7ICG has high affinity for purified recombinant CDCP1 and CDCP1 that is located on the surface of ovarian cancer cells in vitro and in vivo. Our results show that intravenously administered 10D7ICG accumulates preferentially in ovarian cancer, permitting visualization of xenograft tumors in mice. The data suggest CDCP1 as a rational target for tumor-specific fluorescence-guided surgery for ovarian cancer.

Thermal compensation of molded silicone optics.

Optical grade silicone has various properties that make it attractive for solar concentrators, such as excellent transmission across the solar spectrum and flexible moldability for freeform profiles. In this study, a glass-silicone lens structure is proposed to reduce the optothermal effect on the silicone lens. Experimental measurements and simulation modeling results demonstrate that the focal length sensitivity of the glass-silicone lens with respect to temperature can be reduced by a factor of 10 when compared to a silicone lens alone. This model has been extended to the simulation of a proposed two-stage silicone solar concentrator, consisting of an array of acylindrical lenslets and rows of waveguides that focus light onto microphotovoltaic cells. The optical efficiency of the solar concentration system showed a change of less than 10% compared to the efficiency at room temperature for temperature changes from -10∘C to 70°C.

Silicone optical elements for cost-effective freeform solar concentration.

The use of silicone optical elements is demonstrated for a concentrated photovoltaic system. These components show over 96% transmission through most of the solar spectrum and excellent temperature stability. Unique moldability enables the use of complex freeform designs. A light, compact, and cost-effective concentrated photovoltaic system based on silicone optics is proposed. In this system, air-plasma treatment is utilized to overcome the mechanical properties of silicone and difficulties with coating to reduce Fresnel loss. Lens arrays and waveguides are fabricated by injection molding following freeform optical design by LightTools. First-order characterizations are also performed.

Knowledge and risk factors for foot-and-mouth disease among small-scale dairy farmers in an endemic setting.

Foot-and-mouth disease (FMD) is a highly contagious viral infection of cloven-hoofed animals. In Kenya, the disease is endemic with outbreaks typically occurring throughout the year. A cross-sectional study was undertaken in Nakuru County to investigate farmer knowledge and risk factors for clinical disease. Semi-structured interviews were conducted on 220 smallholder farmers, selected using random spatial sampling. The majority of respondents (207/220 [94.1%]) knew of FMD and 166/207 (80.2%) of them could correctly identify the disease based on their knowledge of the clinical signs. Forty-five out of 220 farmers (20.4%) vaccinated their livestock against FMD in the previous 6 months, although of those who knew of FMD only 96/207 (46.4%) perceived it as a preventive measure undertaken to reduce the risk of disease in their farm. FMD had occurred in 5.9% of the surveyed farms within the previous 6 months (from May to November 2016). Using multivariate analysis, the use of a shared bull (OR = 9.7; p = 0.014) and the number of sheep owned (for each additional sheep owned OR = 1.1; p = 0.066) were associated with an increased likelihood of a farm experiencing a case of FMD in the previous 6 months, although the evidence for the latter was weak. This study reports risk factors associated with clinical FMD at the farm level in a densely populated smallholder farming area of Kenya. These results can be used to inform the development of risk-based strategic plans for FMD control and as a baseline for evaluating interventions and control strategies.

Cdk1-dependent destruction of Eco1 prevents cohesion establishment after S phase.

Accurate genome segregation depends on cohesion mechanisms that link duplicated sister chromatids, thereby allowing their tension-dependent biorientation in metaphase. In Saccharomyces cerevisiae, cohesion is established during DNA replication when Eco1 acetylates the cohesin subunit Smc3. Cohesion establishment is restricted to S phase of the cell cycle, but the molecular basis of this regulation is unknown. Here, we show that Eco1 is negatively regulated by the protein kinase Cdk1. Phosphorylation of Eco1 after S phase targets it to SCF(Cdc4) for ubiquitination and subsequent degradation. A nonphosphorylatable mutant of Eco1 establishes cohesion after DNA replication, suggesting that Cdk1-dependent phosphorylation of Eco1 is a key factor limiting establishment to S phase. We also show that deregulation of Eco1 results in chromosome separation defects in anaphase. We conclude that this regulatory mechanism helps optimize the level of sister chromatid cohesion, ensuring a robust and efficient anaphase.

Infrared Fingerprint Engineering: A Molecular-Design Approach to Long-Wave Infrared Transparency with Polymeric Materials.

Optical technologies in the long-wave infrared (LWIR) spectrum (7-14 µm) offer important advantages for high-resolution thermal imaging in near or complete darkness. The use of polymeric transmissive materials for IR imaging offers numerous cost and processing advantages but suffers from inferior optical properties in the LWIR spectrum. A major challenge in the design of LWIR-transparent organic materials is that nearly all organic molecules absorb in this spectral window which lies within the so-called IR-fingerprint region. We report on a new molecular-design approach to prepare high refractive index polymers with enhanced LWIR transparency. Computational methods were used to accelerate the design of novel molecules and polymers. Using this approach, we have prepared chalcogenide hybrid inorganic/organic polymers (CHIPs) with enhanced LWIR transparency and thermomechanical properties via inverse vulcanization of elemental sulfur with new organic co-monomers.

Kin discrimination between sympatric Bacillus subtilis isolates.

Kin discrimination, broadly defined as differential treatment of conspecifics according to their relatedness, could help biological systems direct cooperative behavior toward their relatives. Here we investigated the ability of the soil bacterium Bacillus subtilis to discriminate kin from nonkin in the context of swarming, a cooperative multicellular behavior. We tested a collection of sympatric conspecifics from soil in pairwise combinations and found that despite their history of coexistence, the vast majority formed distinct boundaries when the swarms met. Some swarms did merge, and most interestingly, this behavior was only seen in the most highly related strain pairs. Overall the swarm interaction phenotype strongly correlated with phylogenetic relatedness, indicative of kin discrimination. Using a subset of strains, we examined cocolonization patterns on plant roots. Pairs of kin strains were able to cocolonize roots and formed a mixed-strain biofilm. In contrast, inoculating roots with pairs of nonkin strains resulted in biofilms consisting primarily of one strain, suggestive of an antagonistic interaction among nonkin strains. This study firmly establishes kin discrimination in a bacterial multicellular setting and suggests its potential effect on ecological interactions.

Impact of foot-and-mouth disease on mastitis and culling on a large-scale dairy farm in Kenya.

Foot and mouth disease (FMD) is a highly transmissible viral infection of cloven hooved animals associated with severe economic losses when introduced into FMD-free countries. Information on the impact of the disease in FMDV-endemic countries is poorly characterised yet essential for the prioritisation of scarce resources for disease control programmes. A FMD (virus serotype SAT2) outbreak on a large-scale dairy farm in Nakuru County, Kenya provided an opportunity to evaluate the impact of FMD on clinical mastitis and culling rate. A cohort approach followed animals over a 12-month period after the commencement of the outbreak. For culling, all animals were included; for mastitis, those over 18 months of age. FMD was recorded in 400/644 cattle over a 29-day period. During the follow-up period 76 animals were culled or died whilst in the over 18 month old cohort 63 developed clinical mastitis. Hazard ratios (HR) were generated using Cox regression accounting for non-proportional hazards by inclusion of time-varying effects. Univariable analysis showed FMD cases were culled sooner but there was no effect on clinical mastitis. After adjusting for possible confounders and inclusion of time-varying effects there was weak evidence to support an effect of FMD on culling (HR = 1.7, 95% confidence intervals [CI] 0.88-3.1, P = 0.12). For mastitis, there was stronger evidence of an increased rate in the first month after the onset of the outbreak (HR = 2.9, 95%CI 0.97-8.9, P = 0.057).

Reactive Oxygen Species as Mediators of Disease Progression and Therapeutic Response in Colorectal Cancer.

Significance: Reactive oxygen species (ROS) are critical to normal cellular function with redox homeostasis achieved by balancing ROS production with removal through detoxification mechanisms. Many of the conventional chemotherapies used to treat colorectal cancer (CRC) derive a proportion of their cytotoxicity from ROS generation, and resistance to chemotherapy is associated with elevated detoxification mechanisms. Furthermore, cancer stem cells demonstrate elevated detoxification mechanisms making definitive treatment with existing chemotherapy challenging. In this article, we review the roles of ROS in normal and malignant colonic cell biology and how existing and emerging therapies might harness ROS for therapeutic benefit. Recent Advances: Recent publications have elucidated the contribution of ROS to the cytotoxicity of conventional chemotherapy alongside the emerging approaches of photodynamic therapy (PDT), sonodynamic therapy (SDT), and radiodynamic therapy (RDT), in which ROS are generated in response to excitatory light, sound, or X-ray stimuli to promote cancer cell apoptosis. Critical Issues: The majority of patients with metastatic CRC have a very poor prognosis with a 5-year survival of ∼13% making the need for new or more effective treatments an imperative. Future Directions: Modulation of ROS through a combination of new and emerging therapies may improve the efficacy of current chemotherapy providing novel approaches to treat the otherwise resistant disease. Antioxid. Redox Signal. 39, 186-205.

Comparison of vaccination schedules for foot-and-mouth disease among cattle and sheep in Mongolia.

Vaccines are a critical tool for the control strategy for foot-and-mouth disease (FMD) in Mongolia where sporadic outbreaks regularly occur. A two-dose primary vaccination course is recommended for most commercial vaccines though this can be logistically challenging to deliver among nomadic pastoralist systems which predominate in the country. Although there is evidence that very high potency vaccines can provide prolonged duration of immunity, this has not been demonstrated under field conditions using commercially available vaccines. This study compared neutralizing titres to a O/ME-SA/Panasia strain over a 6-month period following either a two-dose primary course or a single double-dose vaccination among Mongolian sheep and cattle using a 6.0 PD50 vaccine. Titers were not significantly different between groups except in sheep at six-months post vaccination when the single double-dose group had significantly lower titers. These results indicate the single double-dose regimen may be a cost-effective approach for vaccination campaigns supporting FMD control in Mongolia.

Identification of small molecules as novel anti-adipogenic compounds based on Connectivity Map.

Several physiological and pathological conditions such as aging, obesity, diabetes, anorexia nervosa are associated with increased adipogenesis in the bone marrow. A lack of effective drugs hinder the improved treatment for aberrant accumulation of bone marrow adipocytes. Given the higher costs, longer duration and sometimes lack of efficacy in drug discovery, computational and experimental strategies have been used to identify previously approved drugs for the treatment of diseases, also known as drug repurposing. Here, we describe the method of small molecule-prioritization by employing adipocyte-specific genes using the connectivity map (CMap). We then generated transcriptomic profiles using human mesenchymal stromal cells under adipogenic differentiation with the treatment of prioritized compounds, and identified emetine and kinetin-riboside to have a potent inhibitory effect on adipogenesis. Overall, we demonstrated a proof-of-concept method to identify repurposable drugs capable of inhibiting adipogenesis, using the Connectivity Map.

Genotyping of foot-and-mouth disease viruses collected in Sudan between 2009 and 2018.

Foot-and-mouth disease (FMD) is widely distributed in Sudan where outbreaks occur on an annual basis especially during the winter months (December-February). This study aimed to increase our understanding of the epidemiological patterns of FMD in Sudan and connections to neighbouring countries by characterizing the genetic sequences of FMD viruses (FMDV) collected from samples collected in 10 Sudanese states over a 10-year period (between 2009 and 2018). FMDV was detected in 91 of the 265 samples using an antigen-detection ELISA. Three serotypes were detected: O (46.2%), A (34.0%), and SAT 2 (19.8%). Fifty-two of these samples were submitted for sequence analyses, generating sequences that were characterized as belonging to O/EA-3 (n = 17), A/AFRICA/G-IV (n = 23) and SAT 2/VII/Alx-12 (n = 12) viral lineages. Phylogenetic analyses provided evidence that FMDV lineages were maintained within Sudan, and also highlighted epidemiological connections to FMD outbreaks reported in neighbouring countries in East and North Africa (such as Ethiopia and Egypt). This study motivates continued FMD surveillance in Sudan to monitor the circulating viral lineages and broader initiatives to improve our understanding of the epidemiological risks in the region.

CUB Domain-Containing Protein 1 (CDCP1) is a rational target for the development of imaging tracers and antibody-drug conjugates for cancer detection and therapy.

Rationale: An antibody-drug conjugate (ADC) is a targeted therapy consisting of a cytotoxic payload that is linked to an antibody which targets a protein enriched on malignant cells. Multiple ADCs are currently used clinically as anti-cancer agents significantly improving patient survival. Herein, we evaluated the rationale of targeting the cell surface oncoreceptor CUB domain-containing protein 1 (CDCP1) using ADCs and assessed the efficacy of CDCP1-directed ADCs against a range of malignant tumors. Methods: CDCP1 mRNA expression was evaluated using large transcriptomic datasets of normal/tumor samples for 23 types of cancer and 15 other normal organs, and CDCP1 protein expression was examined in 34 normal tissues, >300 samples from six types of cancer, and in 49 cancer cell lines. A recombinant human/mouse chimeric anti-CDCP1 antibody (ch10D7) was labelled with 89Zirconium or monomethyl auristatin E (MMAE) and tested in multiple pre-clinical cancer models including 36 cancer cell lines and three mouse xenograft models. Results: Analysis of CDCP1 expression indicates elevated CDCP1 expression in the majority of the cancers and restricted expression in normal human tissues. Antibody ch10D7 demonstrates a high affinity and specificity for CDCP1 inducing cell signalling via Src accompanied by rapid internalization of ch10D7/CDCP1 complexes in cancer cells. 89Zirconium-labelled ch10D7 accumulates in CDCP1 expressing cells enabling detection of pancreatic cancer xenografts in mice by PET imaging. Cytotoxicity of MMAE-labelled ch10D7 against kidney, colorectal, lung, ovarian, pancreatic and prostate cancer cells in vitro, correlates with the level of CDCP1 on the plasma membrane. ch10D7-MMAE displays robust anti-tumor effects against mouse xenograft models of pancreatic, colorectal and ovarian cancer. Conclusion: CDCP1 directed imaging agents will be useful for selecting cancer patients for personalized treatment with cytotoxin-loaded CDCP1 targeting agents including antibody-drug conjugates.

Development of anti-Crimean-Congo hemorrhagic fever virus Gc and NP-specific ELISA for detection of antibodies in domestic animal sera.

Crimean-Congo hemorrhagic fever (CCHF) is a priority emerging disease. CCHF, caused by the CCHF virus (CCHFV), can lead to hemorrhagic fever in humans with severe cases often having fatal outcomes. CCHFV is maintained within a tick-vertebrate-tick cycle, which includes domestic animals. Domestic animals infected with CCHFV do not show clinical signs of the disease and the presence of antibodies in the serum can provide evidence of their exposure to the virus. Current serological tests are specific to either one CCHFV antigen or the whole virus antigen. Here, we present the development of two in-house ELISAs for the detection of serum IgG that is specific for two different CCHFV antigens: glycoprotein Gc (CCHFV Gc) and nucleoprotein (CCHFV NP). We demonstrate that these two assays were able to detect anti-CCHFV Gc-specific and anti-CCHFV NP-specific IgG in sheep from endemic CCHFV areas with high specificity, providing new insight into the heterogeneity of the immune response induced by natural infection with CCHFV in domestic animals.

A multi-omic analysis of MCF10A cells provides a resource for integrative assessment of ligand-mediated molecular and phenotypic responses.

The phenotype of a cell and its underlying molecular state is strongly influenced by extracellular signals, including growth factors, hormones, and extracellular matrix proteins. While these signals are normally tightly controlled, their dysregulation leads to phenotypic and molecular states associated with diverse diseases. To develop a detailed understanding of the linkage between molecular and phenotypic changes, we generated a comprehensive dataset that catalogs the transcriptional, proteomic, epigenomic and phenotypic responses of MCF10A mammary epithelial cells after exposure to the ligands EGF, HGF, OSM, IFNG, TGFB and BMP2. Systematic assessment of the molecular and cellular phenotypes induced by these ligands comprise the LINCS Microenvironment (ME) perturbation dataset, which has been curated and made publicly available for community-wide analysis and development of novel computational methods ( synapse.org/LINCS_MCF10A ). In illustrative analyses, we demonstrate how this dataset can be used to discover functionally related molecular features linked to specific cellular phenotypes. Beyond these analyses, this dataset will serve as a resource for the broader scientific community to mine for biological insights, to compare signals carried across distinct molecular modalities, and to develop new computational methods for integrative data analysis.

Economic Considerations for Advancement Through the Progressive Control Pathway: Cost-Benefit Analysis of an FMD Disease-Free Zone in Punjab Province, Pakistan.

Foot-and-mouth disease (FMD) is a priority disease of livestock in Pakistan, which was classified in stage 2 of the Progressive Control Pathway (PCP-FMD) in 2015, aiming to reduce disease impact. Further progression requires efforts to reduce viral circulation that may ultimately result in being awarded official disease-free status by the World Organisation for Animal Health [Office International des Epizooties (OIE)]. Typically, FMD control is reliant on the extensive use of vaccines, requiring careful consideration of the costs and benefits to ensure investment is likely to provide a positive return. This study conducted a cost-benefit analysis (CBA) for a proposed zone within Punjab Province, Pakistan. Benefits were assumed to come from averted production losses and treatment costs and the costs based on typical measures required for establishing a disease-free zone. To estimate the impact of FMD at the farm level, models were created to estimate effects on milk production, offtakes, and changes in herd value over a 5-year period with different parameters used to represent the production systems present. Control strategy costs incorporated aspects of vaccination, surveillance, sanitary measures, program management, stakeholder engagement, preparatory studies, training, and capacity building. The results indicated a median benefit-cost ratio of 1.03 (90% central range 0.37, 1.63) with a median net present value of 1.99 billion Pakistan Rupees (90% central range -37.7, 37.0). The greatest cost was due to vaccination at 56%, followed by sanitary measures (including implementing and maintaining an animal ID system and quarantine stations around the zone) at 41%. Although the median benefit-cost ratio and net present value indicated that investment is likely to generate a positive return, the large variation indicates caution in interpreting the results and it is possible that an increase in animal value through new export markets will be required. Further refinement in our knowledge of disease impact and the details of the control strategy are needed. Moreover, there are implications regarding vaccine security, since the strategy is reliant on the steady provision of quality vaccines in order to achieve the anticipated benefits, raising important issues on vaccine availability for countries to maintain lucrative export markets for FMD.