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Anti-cancer therapies often exhibit only short-term effects. Tumors typically develop drug resistance causing relapses that might be tackled with drug combinations. Identification of the right combination is challenging and would benefit from high-content, high-throughput combinatorial screens directly on patient biopsies. However, such screens require a large amount of material, normally not available from patients. To address these challenges, we present a scalable microfluidic workflow, called Combi-Seq, to screen hundreds of drug combinations in picoliter-size droplets using transcriptome changes as a readout for drug effects. We devise a deterministic combinatorial DNA barcoding approach to encode treatment conditions, enabling the gene expression-based readout of drug effects in a highly multiplexed fashion. We apply Combi-Seq to screen the effect of 420 drug combinations on the transcriptome of K562 cells using only ~250 single cell droplets per condition, to successfully predict synergistic and antagonistic drug pairs, as well as their pathway activities. Current screens to assess tumour drug resistance require a large amount of material, normally not available from patients. Here the authors report CombiSeq, a scalable microfluidic workflow to screen hundreds of drug combinations in picoliter-size droplets using transcriptome changes as a readout. [ABSTRACT FROM AUTHOR]

Aims/hypothesis. Vascular disease in type 2 diabetes is associated with an up-regulation of atherogenic growth factors, which stimulate matrix synthesis including proteoglycans. We have examined the direct actions of fenofibrate on human vascular smooth muscle cells (VSMCs) and have specifically investigated proteoglycan synthesis and binding to LDL. Methods. Proteoglycans synthesised by human VSMCs treated with fenofibrate (30 μmol/l) were assessed for binding to human LDL using a gel mobility shift assay, metabolically labelled with [35S]-sulphate and quantitated by cetylpyridinium chloride. They were then assessed for electrophoretic mobility by SDSPAGE, for size by gel filtration, for sulphation pattern by fluorophore-assisted carbohydrate electrophoresis, and for glycosaminoglycan (GAG) composition by enzyme digestion. Results. Proteoglycans synthesised in the presence of fenofibrate showed an increase in the half-maximum saturation concentration of LDL from 36.8±12.4 μg/ml to 77.7±17 μg/ml under basal conditions, from 24.9±4.6 μg/ml to 39.1±6.1 μg/ml in the presence of TGF-β1, and from 9.5±4.4 μg/ml to 31.1±3.4 μg/ml in the presence of platelet-derived growth factor/insulin. Fenofibrate treatment in the presence of TGF-β1 inhibited the incorporation of [35S]-sulphate into secreted and cell-associated proteoglycans synthesised by human VSMCs by 59.2% (p<0.01) and 39.8% (p<0.01) respectively. The changes in sulphate incorporation following treatment with fenofibrate were associated with a concentration- related increase in the electrophoretic mobility due to a reduction in GAG length. There was no change in the sulphation pattern; however, there was an alteration in the disaccharide composition of the GAGs. Conclusions/interpretation. Fenofibrate modifies the structure of vascular proteoglycans by reducing the length of the GAG chains and GAG composition, resulting in reduced binding to human LDL, a mechanism which may lead to a reduction of atherosclerosis and cardiovascular disease in people with diabetes treated with fenofibrate. [ABSTRACT FROM AUTHOR]

Cardiovascular disease is the major cause of premature death in modern society, and its impact is increasing due to rising rates of obesity and type 2diabetes. Clinical studies based on targeting metabolic abnormalities and biomarkers demonstrate significant benefits, but always an element of disease remains which is resistant to treatment. Recent evidence has strongly implicated an early interaction of atherogenic lipoproteins with vascular matrix proteoglycans as the initiating step in atherogenesis. Expert commentary has pointed to the need for vascular directed therapies to provide reductions in the residual disease component. We propose that the regulation of synthesis and thus structure of glycosaminoglycans on proteoglycans provides a potential pathway to this reduction. We review existing evidence that the vascular synthesis of glycosaminoglycan chains can be regulated in a manner which reduces lipoprotein binding and the potential application of this strategy to attenuation of the current cardiovascular disease pandemic. [ABSTRACT FROM AUTHOR]