Tradeoff between metabolic i-proteasome addiction and immune evasion in triple-negative breast cancer

In vitro studies have suggested proteasome inhibitors could be effective in triple-negative breast cancer (TNBC). We found that bortezomib and carfilzomib induce proteotoxic stress and apoptosis via the unfolded protein response (UPR) in TNBC cell lines, with sensitivity correlated with expression of immuno-(PSMB8/9/10) but not constitutive-(PSMB5/6/7) proteasome subunits. Equally, the transcriptomes of i-proteasome-high human TNBCs are enriched with UPR gene sets, and the genomic copy number landscape reflects positive selection pressure favoring i-proteasome activity, but in the setting of adjuvant treatment, this is actually associated with favorable prognosis. Tumor expression of PSMB8 protein (β5i) is associated with levels of MHC-I, interferon-γ-inducible proteasome activator PA28β, and the densities of stromal antigen-presenting cells and lymphocytes (TILs). Crucially, TILs were protective among TNBCs that maintain high β5i but did not stratify survival amongst β5i-low TNBCs. Moreover, β5i expression was lower in brain metastases than in patient-matched primary breast tumors (n = 34; P = 0.007), suggesting that suppression contributes to immune evasion and metastatic progression. Hence, inhibiting proteasome activity could be counterproductive in the adjuvant treatment setting because it potentiates anti-TNBC immunity.Alaknanda Adwal, Priyakshi Kalita-de Croft, Reshma Shakya, Malcolm Lim, Emarene Kalaw, Lucinda D Taege, Amy E McCart Reed, Sunil R Lakhani, David F Callen, Jodi M Saunu

Quantifying the habitat and zoogeomorphic capabilities of spawning European barbel Barbus barbus, a lithophilous cyprinid

Suitable gravel availability is critical for the spawning success of lithophilous fishes, including redd builders. Redd construction during spawning can alter substrate characteristics, thereby influencing hydraulic conditions and sediment transport, highlighting the importance of spawning as a zoogeomorphic activity. Here, interactions between redd‐building fish and their spawning environment were investigated for European barbel Barbus barbus with a comparative approach across three English rivers: Teme (western), Great Ouse (eastern) and Idle (central). Sediment characteristics of spawning habitats were similar across the rivers, including subsurface fine sediment (<2 mm) content (≈20% dry weight), but elevated subsurface silt content and coarser surface sediments were found in the river Teme. Water velocities were similar at spawning sites despite differences in channel width and depth. Redds were characterized by a pit and tailspill, with no differences in surface grain‐size characteristics between these and the surrounding riverbed, but with topographic alteration (dimensions and tailspill amplitude) in line with those of salmonids. Estimates of the fraction of the bed that spawning barbel were capable of moving exceeded 97% in all rivers. Estimated reproductive potential varied significantly between the rivers Idle and Teme (3,098 to 9,715 eggs/m2), which was largely due to differences in barbel lengths affecting fecundity. Larger barbel, capable of producing and depositing more eggs, but in more spatially extensive redds, meaning fewer redds per given surface area of riverbed. Predictions of barbel egg mortality based on sand content were low across both rivers. The effects of silt on barbel egg and larvae development are unknown, but the levels detected here would significantly impact salmon egg mortality. Similarities in fish length to redd area and the size of moveable grains by spawning barbel and salmon suggest they have similar geomorphic effects on sediments, although fine sediment tolerance is highly divergent

Enhancement of Vaccinia Virus Based Oncolysis with Histone Deacetylase Inhibitors

Histone deacetylase inhibitors (HDI) dampen cellular innate immune response by decreasing interferon production and have been shown to increase the growth of vesicular stomatitis virus and HSV. As attenuated tumour-selective oncolytic vaccinia viruses (VV) are already undergoing clinical evaluation, the goal of this study is to determine whether HDI can also enhance the potency of these poxviruses in infection-resistant cancer cell lines. Multiple HDIs were tested and Trichostatin A (TSA) was found to potently enhance the spread and replication of a tumour selective vaccinia virus in several infection-resistant cancer cell lines. TSA significantly decreased the number of lung metastases in a syngeneic B16F10LacZ lung metastasis model yet did not increase the replication of vaccinia in normal tissues. The combination of TSA and VV increased survival of mice harbouring human HCT116 colon tumour xenografts as compared to mice treated with either agent alone. We conclude that TSA can selectively and effectively enhance the replication and spread of oncolytic vaccinia virus in cancer cells

Sexual conflict maintains variation at an insecticide resistance locus

Background: The maintenance of genetic variation through sexually antagonistic selection is controversial, partly because specific sexually-antagonistic alleles have not been identified. The Drosophila DDT resistance allele (DDT-R) is an exception. This allele increases female fitness, but simultaneously decreases male fitness, and it has been suggested that this sexual antagonism could explain why polymorphism was maintained at the locus prior to DDT use. We tested this possibility using a genetic model and then used evolving fly populations to test model predictions. Results: Theory predicted that sexual antagonism is able to maintain genetic variation at this locus, hence explaining why DDT-R did not fix prior to DDT use despite increasing female fitness, and experimentally evolving fly populations verified theoretical predictions. Conclusions: This demonstrates that sexually antagonistic selection can maintain genetic variation and explains the DDT-R frequencies observed in nature

The Stem Cell Marker CD133 Associates with Enhanced Colony Formation and Cell Motility in Colorectal Cancer

CD133 is a membrane molecule that has been, controversially, reported as a CSC marker in colorectal cancer (CRC). In this study, we sought to clarify the expression and role of CD133 in CRC. Initially the size of the CD133−expressing (CD133+) population in eight well-described CRC cell lines was measured by flow cytometry and was found to range from 0% to >95%. The cell line HT29 has a CD133+ population of >95% and was chosen for functional evaluation of CD133 after gene knockdown by RNA interference. A time course assay showed that CD133 inhibition had no significant effect on cell proliferation or apoptosis. However, CD133 knockdown did result in greater susceptibility to staurosporine-induced apoptosis (p = 0.01) and reduction in cell motility (p<0.04). Since gene knockdown may cause “off-target” effects, the cell line SW480 (which has a CD133+ population of 40%) was sorted into pure CD133+ and CD133− populations to allow functional comparison of isogenic populations separated only by CD133 expression. In concordance with the knockdown experiments, a time course assay showed no significant proliferative differences between the CD133+/CD133− populations. Also greater resistance to staurosporine-induced apoptosis (p = 0.008), greater cell motility (p = 0.03) and greater colony forming efficiency was seen in the CD133+ population than the CD133− population in both 2D and 3D culture (p<0.0001 and p<0.003 respectively). Finally, the plasticity of CD133 expression in tumour cells was tested. Quantitative PCR analysis showed there was transcriptional repression in the CD133− population of SW480. Prolonged culture of a pure CD133− population resulted in re-emergence of CD133+ cells. We conclude that CD133 expression in CRCs is associated with some features attributable to stemness and that there is plasticity of CD133 expression. Further studies are necessary to delineate the mechanistic basis of these features

Cross-Sectional Study into the Costs and Impact on Family Functioning of 4-Year-Old Children with Aggressive Behavior

Early-onset aggressive behavior is known for its negative developmental consequences, and the associated high costs for families, the health care system and wider society. Although the origins of aggressive behavior are to be found in early childhood, the costs incurred by aggressive behavior of young children have not been studied extensively. The present study aimed to investigate whether preschool children with a high level of aggressive behavior already differ in the generated amount of costs and impact on family functioning from children with lower levels of aggressive behavior. A population-based sample of 317 preschool children was divided into four groups with different levels of aggression (moderate, borderline, clinical). Parents filled out questionnaires to assess service use (lifetime and past 3 months) and impact on family functioning. Over the past 3 months as well as over the first 4 years of life, children with a clinical level of aggression were more costly than children with a low level of aggression (mean total costs over the past 3 months: low = €167,05 versus clinical € = 1034,83 and mean lifetime costs: low € = 817,37 versus clinical € = 1433,04), due to higher costs of services used by the child. In addition, families of children with a borderline or clinical level of aggressive behavior reported more impairment in their daily functioning than families of children with lower levels of aggression. The findings demonstrate that a high level of aggressive behavior results in high costs and impaired family functioning in the preschool years already

Pleiotropic Effects of DDT Resistance on Male Size and Behaviour

Understanding the evolution and spread of insecticide resistance requires knowing the relative fitness of resistant organisms. In the absence of insecticides, resistance is predicted to be costly. The Drosophila melanogaster DDT resistance allele (DDT-R) is associated with a male mating cost. This could be because resistant males are generally smaller, but DDT-R may also alter courtship behaviours. Here we tested for body size and courtship effects of DDT-R on mating success in competitive and non-competitive mating trials respectively. We also assessed relative aggression in resistant and susceptible males because aggression can also influence mating success. While the effect of DDT-R on male size partly contributed to reduced mating success, resistant males also had lower rates of courtship and were less aggressive than susceptible males. These differences contribute to the observed DDT-R mating costs. Additionally, these pleiotropic effects of DDT-R are consistent with the history and spread of resistance alleles in nature

A Selectable and Excisable Marker System for the Rapid Creation of Recombinant Poxviruses

Genetic manipulation of poxvirus genomes through attenuation, or insertion of therapeutic genes has led to a number of vector candidates for the treatment of a variety of human diseases. The development of recombinant poxviruses often involves the genomic insertion of a selectable marker for purification and selection purposes. The use of marker genes however inevitably results in a vector that contains unwanted genetic information of no therapeutic value.Here we describe an improved strategy that allows for the creation of marker-free recombinant poxviruses of any species. The Selectable and Excisable Marker (SEM) system incorporates a unique fusion marker gene for the efficient selection of poxvirus recombinants and the Cre/loxP system to facilitate the subsequent removal of the marker. We have defined and characterized this new methodological tool by insertion of a foreign gene into vaccinia virus, with the subsequent removal of the selectable marker. We then analyzed the importance of loxP orientation during Cre recombination, and show that the SEM system can be used to introduce site-specific deletions or inversions into the viral genome. Finally, we demonstrate that the SEM strategy is amenable to other poxviruses, as demonstrated here with the creation of an ectromelia virus recombinant lacking the EVM002 gene.The system described here thus provides a faster, simpler and more efficient means to create clinic-ready recombinant poxviruses for therapeutic gene therapy applications