Proteotyping of knockout mouse strains reveals sex- and strain-specific signatures in blood plasma

We proteotyped blood plasma from 30 mouse knockout strains and corresponding wild-type mice from the International Mouse Phenotyping Consortium. We used targeted proteomics with internal standards to quantify 375 proteins in 218 samples. Our results provide insights into the manifested effects of each gene knockout at the plasma proteome level. We first investigated possible contamination by erythrocytes during sample preparation and labeled, in one case, up to 11 differential proteins as erythrocyte originated. Second, we showed that differences in baseline protein abundance between female and male mice were evident in all mice, emphasizing the necessity to include both sexes in basic research, target discovery, and preclinical effect and safety studies. Next, we identified the protein signature of each gene knockout and performed functional analyses for all knockout strains. Further, to demonstrate how proteome analysis identifies the effect of gene deficiency beyond traditional phenotyping tests, we provide in-depth analysis of two strains, C8a(-/-) and Npc2(+/-). The proteins encoded by these genes are well-characterized providing good validation of our method in homozygous and heterozygous knockout mice. Ig alpha chain C region, a poorly characterized protein, was among the differentiating proteins in C8a(-/-). In Npc2(+/-) mice, where histopathology and traditional tests failed to differentiate heterozygous from wild-type mice, our data showed significant difference in various lysosomal storage disease-related proteins. Our results demonstrate how to combine absolute quantitative proteomics with mouse gene knockout strategies to systematically study the effect of protein absence. The approach used here for blood plasma is applicable to all tissue protein extracts.Proteomic

Antitumor Responses in the Absence of Toxicity in Solid Tumors by Targeting B7-H3 via Chimeric Antigen Receptor T Cells

The high expression across multiple tumor types and restricted expression in normal tissues make B7-H3 an attractive target for immunotherapy. We generated chimeric antigen receptor (CAR) T cells targeting B7-H3 (B7-H3.CAR-Ts) and found that B7-H3.CAR-Ts controlled the growth of pancreatic ductal adenocarcinoma, ovarian cancer and neuroblastoma in vitro and in orthotopic and metastatic xenograft mouse models, which included patient-derived xenograft. We also found that 4-1BB co-stimulation promotes lower PD-1 expression in B7-H3.CAR-Ts, and superior antitumor activity when targeting tumor cells that constitutively expressed PD-L1. We took advantage of the cross-reactivity of the B7-H3.CAR with murine B7-H3, and found that B7-H3.CAR-Ts significantly controlled tumor growth in a syngeneic tumor model without evident toxicity. These findings support the clinical development of B7-H3.CAR-Ts

Detectable clonal mosaicism and its relationship to aging and cancer

In an analysis of 31,717 cancer cases and 26,136 cancer-free controls from 13 genome-wide association studies, we observed large chromosomal abnormalities in a subset of clones in DNA obtained from blood or buccal samples. We observed mosaic abnormalities, either aneuploidy or copy-neutral loss of heterozygosity, of >2 Mb in size in autosomes of 517 individuals (0.89%), with abnormal cell proportions of between 7% and 95%. In cancer-free individuals, frequency increased with age, from 0.23% under 50 years to 1.91% between 75 and 79 years (P = 4.8 × 10(-8)). Mosaic abnormalities were more frequent in individuals with solid tumors (0.97% versus 0.74% in cancer-free individuals; odds ratio (OR) = 1.25; P = 0.016), with stronger association with cases who had DNA collected before diagnosis or treatment (OR = 1.45; P = 0.0005). Detectable mosaicism was also more common in individuals for whom DNA was collected at least 1 year before diagnosis with leukemia compared to cancer-free individuals (OR = 35.4; P = 3.8 × 10(-11)). These findings underscore the time-dependent nature of somatic events in the etiology of cancer and potentially other late-onset diseases

Mouse Quantitative Proteomics Knowledgebase: reference protein concentration ranges in 20 mouse tissues using 5000 quantitative proteomics assays

Motivation Laboratory mouse is the most used animal model in biological research, largely due to its high conserved synteny with human. Researchers use mice to answer various questions ranging from determining a pathological effect of knocked out/in gene to understanding drug metabolism. Our group developed >5000 quantitative targeted proteomics assays for 20 mouse tissues and determined the concentration ranges of a total of >1600 proteins using heavy labeled internal standards. We describe here MouseQuaPro; a knowledgebase that hosts this collection of carefully curated experimental data.Results The web-based application includes protein concentrations from >700 mouse tissue samples from three common research strains, corresponding to >200k experimentally determined concentrations. The knowledgebase integrates the assay and protein concentration information with their human orthologs, functional and molecular annotations, biological pathways, related human diseases and known gene expressions. At its core are the protein concentration ranges, which provide insights into (dis)similarities between tissues, strains and sexes. MouseQuaPro implements advanced search as well as filtering functionalities with a simple interface and interactive visualization. This information-rich resource provides an initial map of protein absolute concentration in mouse tissues and allows guided design of proteomics phenotyping experiments. The knowledgebase is available on mousequapro.proteincentre.com.Proteomic

Availability and accessibility of primary mental health services for adolescents: an overview of national recommendations and services in EU.

Mental health problems in adolescence can profoundly jeopardize adolescent current and future health and functioning. We aimed to describe existing recommendations and services regarding the delivery of primary mental health care for adolescents in 31 European countries. Data on the availability and accessibility of primary mental health services were collected, as part of the Horizon 2020-funded project Models of Child Health Appraised. One expert from each country answered a closed items questionnaire during years 2017-18. All 31 participating countries had some policy or recommendations regarding the availability and accessibility of primary mental health services for adolescents, but their focus and implementation varied largely between and within countries. Only half of the participating countries had recommendations on screening adolescents for mental health issues and burdens. Merely a quarter of the countries had ambulatory facilities targeting specifically adolescents throughout the whole country. Just over half had some kind of suicide prevention programs. Same-day access to primary care in case of -health emergencies was possible in 21 countries, but often not throughout the whole country. Nineteen countries had strategies securing accessible mental health care for vulnerable adolescents. Overall, around half of European countries had strategies securing access to various primary mental health care for adolescents. They frequently did not guarantee care over the whole country and often tackled a limited number of situations. EU countries should widen the range of policies and recommendations governing the delivery of mental health care to adolescents and monitor their implementation

Plasma Protein Signatures of a Murine Venous Thrombosis Model and Slc44a2 Knockout Mice Using Quantitative-Targeted Proteomics

The plasma compartment of the blood holds important information on the risk to develop cardiovascular diseases such as venous thrombosis (VT). Mass spectrometry-based targeted proteomics with internal standards quantifies proteins in multiplex allowing generation of signatures associated with a disease or a condition. Here, to demonstrate the method, we investigate the plasma protein signatures in mice following the onset of VT, which was induced by RNA interference targeting the natural anticoagulants antithrombin and protein C. We then study mice lacking Slc44a2 , which was recently characterized as a VT-susceptibility gene in human genome-wide association studies. We use a recently developed panel of 375 multiplexed mouse protein assays measured by mass spectrometry. A strong plasma protein siganture was observed when VT was induced. Discriminators included acute phase response proteins, and proteins related to erythrocyte function. In mice lacking Slc44a2 , protein signature was primarily overruled by the difference between sexes and not by the absent gene. Upon separate analyses for males and females, we were able to establish a signature for Slc44a2 deficiency, in which glycosylation-dependent cell adhesion molecule-1 and thrombospondin-1 were shared by both sexes. The minimal impact of Slc44a2 deficiency on the measured plasma proteins suggests that the main effect of Slc44a2 on VT does not lay ultimately in the plasma compartment. This suggests further investigation into the role of this VT-susceptibility gene should perhaps also question the possible involvement in cellular mechanisms.Proteomic

Molecular phenotyping of laboratory mouse strains using 500 multiple reaction monitoring mass spectrometry plasma assays

Proteomic