Quantifying the effects of risk-stratified breast cancer screening when delivered in real time as routine practice versus usual screening: the BC-Predict non-randomised controlled study (NCT04359420).

BACKGROUND: Risk stratification as a routine part of the NHS Breast Screening Programme (NHSBSP) could provide a better balance of benefits and harms. We developed BC-Predict, to offer women when invited to the NHSBSP, which collects standard risk factor information; mammographic density; and in a sub-sample, a Polygenic Risk Score (PRS). METHODS: Risk prediction was estimated primarily from self-reported questionnaires and mammographic density using the Tyrer-Cuzick risk model. Women eligible for NHSBSP were recruited. BC-Predict produced risk feedback letters, inviting women at high risk (≥8% 10-year) or moderate risk (≥5-<8% 10-year) to have appointments to discuss prevention and additional screening. RESULTS: Overall uptake of BC-Predict in screening attendees was 16.9% with 2472 consenting to the study; 76.8% of those received risk feedback within the 8-week timeframe. Recruitment was 63.2% with an onsite recruiter and paper questionnaire compared to <10% with BC-Predict only (P < 0.0001). Risk appointment attendance was highest for those at high risk (40.6%); 77.5% of those opted for preventive medication. DISCUSSION: We have shown that a real-time offer of breast cancer risk information (including both mammographic density and PRS) is feasible and can be delivered in reasonable time, although uptake requires personal contact. Preventive medication uptake in women newly identified at high risk is high and could improve the cost-effectiveness of risk stratification. TRIAL REGISTRATION: Retrospectively registered with clinicaltrials.gov (NCT04359420).

Bone marrow cells in murine colitis: multi-signal analysis confirms pericryptal myofibroblast engraftment without epithelial involvement.

BACKGROUND: The contribution of bone marrow-derived cells to epithelial tissues in the inflamed gut remains controversial. Recent reports have suggested that cell fusion between bone marrow-derived cells and the intestinal epithelium takes place in inflammatory conditions. METHODS: In attempts to confirm this, we have undertaken gender mis-matched bone marrow (BM) transplants from male Swiss Webster (SWR) mice to B and T cell-deficient female Rag2 KO mice which, 4 weeks later, were given 5% dextran sodium sulphate in drinking water to induce acute colitis. A further BM-treated group of animals with a graft versus host-like condition was also studied. We developed a new method to combine up to three brightfield or fluorescent lectin- or immuno-histochemical signals with fluorescent in situ hybridisation for the Y and X chromosomes to enable us unequivocally to identify BM-derived male cells which presented as different cell types in the gastrointestinal tract. PRINCIPAL FINDINGS: In rolled preparations of whole intestines we scanned around 1.5 million crypts at many tissue levels. In no instance did we see a Y chromosome-positive cell in the epithelial compartment, which was not also CD45-positive. We saw no evidence of cell fusion, based on combined X and Y chromosome analysis. Levels of CD45-positive stromal and lymphoid cells and pericryptal myfibroblasts (positive for α-smooth muscle actin) increased with time up to a plateau, which resembled the level seen in untreated control grafted animals. We saw very few Y chromosome-positive endothelial cells in intestinal stromal vessels. CONCLUSIONS: We conclude that whole BM transplantation does not result in intestinal epithelial engraftment in this model. Our new methods can usefully assist in multi-signal analyses of cell phenotypes following BM transplant and in models of chimaerism and regenerative medicine.

4-(1,3-Thiazol-2-yl)morpholine derivatives as inhibitors of phosphoinositide 3-kinase.

4-(1,3-Thiazol-2-yl)morpholine derivatives have been identified as potent and selective inhibitors of phosphoinositide 3-kinase. The SAR data of selected examples are presented and the in vivo profiling of compound 18 is shown to demonstrate the utility of this class of compounds in xenograft models of tumor growth.

Analysis of endogenous S1P and LPA receptor expression in CHO-K1 cells.

The CHO-K1 cell line is commonly used for studies of recombinantly expressed proteins, including proteins of the G protein-coupled receptor (GPCR) family. This laboratory has used CHO-K1 cells for the functional characterization of Edg family GPCRs. However, parental CHO-K1 cells respond to lysophospholipids in in-vitro functional assays, which suggests expression of endogenous Edg family GPCRs. To determine the repertoire of Edg family receptor expression in this cell line, alignments of human and rodent sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA) receptor sequences were used to design semi-redundant oligonucleotide pairs. A portion of each receptor gene coding sequence was amplified from Chinese hamster genomic DNA and the resultant gene fragments sequenced. Species-specific oligonucleotide pairs were designed using this novel sequence information and used to detect expression of S1P(1,2,4) and LPA(1) transcripts in CHO-K1 cells by RT-PCR.

Metalloprotease-disintegrin ADAM8: expression analysis and targeted deletion in mice.

ADAM8 (a disintegrin and metalloprotease 8, also referred to as MS2/CD156a) is a membrane-anchored metalloprotease that was first identified in a macrophage cell line and has been implicated in neurodegenerative diseases. Here, we evaluated the expression of ADAM8 during mouse development and generated mice lacking ADAM8 (Adam8-/- mice). During early mouse development, ADAM8 is expressed by maternal cells in the decidua and by trophoblast derivatives of the embryo but not in the derivatives of the inner cell mass. At later stages, prominent expression of ADAM8 is seen in the embryo proper, in the gonadal ridge, thymus, developing cartilage and bone, brain and spinal cord, and in the mesenchyme in close proximity to the branch point between the jugular vein and developing lymphatic vessels. Examination of Adam8-/- mice, however, revealed no major defects in these or other structures during development or in adult tissues and no evident pathological phenotypes.

A single amino acid determines preference between phospholipids and reveals length restriction for activation of the S1P4 receptor.

BACKGROUND: Sphingosine-1-phosphate and lysophosphatidic acid (LPA) are ligands for two related families of G protein-coupled receptors, the S1P and LPA receptors, respectively. The lysophospholipid ligands of these receptors are structurally similar, however recognition of these lipids by these receptors is highly selective. A single residue present within the third transmembrane domain (TM) of S1P receptors is thought to determine ligand selectivity; replacement of the naturally occurring glutamic acid with glutamine (present at this position in the LPA receptors) has previously been shown to be sufficient to change the specificity of S1P1 from S1P to 18:1 LPA. RESULTS: We tested whether mutation of this "ligand selectivity" residue to glutamine could confer LPA-responsiveness to the related S1P receptor, S1P4. This mutation severely affected the response of S1P4 to S1P in a [35S]GTP gamma S binding assay, and imparted sensitivity to LPA species in the order 14:0 LPA > 16:0 LPA > 18:1 LPA. These results indicate a length restriction for activation of this receptor and demonstrate the utility of using LPA-responsive S1P receptor mutants to probe binding pocket length using readily available LPA species. Computational modelling of the interactions between these ligands and both wild type and mutant S1P4 receptors showed excellent agreement with experimental data, therefore confirming the fundamental role of this residue in ligand recognition by S1P receptors. CONCLUSIONS: Glutamic acid in the third transmembrane domain of the S1P receptors is a general selectivity switch regulating response to S1P over the closely related phospholipids, LPA. Mutation of this residue to glutamine confers LPA responsiveness with preference for short-chain species. The preference for short-chain LPA species indicates a length restriction different from the closely related S1P1 receptor.