A high throughput protein biomarker discovery tool has been developed based on multiplexed proximity ligation assays in a homogeneous format in the sense of no washing steps. The platform consists of four 24-plex panels profiling 74 putative biomarkers with sub-pm sensitivity each consuming only 1 μl of human plasma sample. The system uses either matched monoclonal antibody pairs or the more readily available single batches of affinity purified polyclonal antibodies to generate the target specific reagents by covalently linking with unique nucleic acid sequences. These paired sequences are united by DNA ligation upon simultaneous target binding forming a PCR amplicon. Multiplex proximity ligation assays thereby converts multiple target analytes into real-time PCR amplicons that are individually quantified using microfluidic high capacity qPCR in nano liter volumes. The assay shows excellent specificity, even in multiplex, by its dual recognition feature, its proximity requirement, and most importantly by using unique sequence specific reporter fragments on both antibody-based probes. To illustrate the potential of this protein detection technology, a pilot biomarker research project was performed using biobanked plasma samples for the detection of colorectal cancer using a multivariate signature.

ColoLipidGene: signature of lipid metabolism-related genes to predict prognosis in stage-II colon cancer patients ColoLipidGene: signature of lipid metabolism-related genes to predict prognosis in stage-II colon cancer patients

Colorectal and pancreatic cancers remain important contributors to cancer mortality burden and, therefore, new therapeutic approaches are urgently needed. Rosemary (Rosmarinus officinalis L.) extracts and its components have been reported as natural potent antiproliferative agents against cancer cells. However, to potentially apply rosemary as a complementary approach for cancer therapy, additional information regarding the most effective composition, its antitumor effect in vivo and its main molecular mediators is still needed. In this work, five carnosic acid-rich supercritical rosemary extracts with different chemical compositions have been assayed for their antitumor activity both in vivo (in nude mice) and in vitro against colon and pancreatic cancer cells. We found that the antitumor effect of carnosic acid together with carnosol was higher than the sum of their effects separately, which supports the use of the rosemary extract as a whole. In addition, gene and microRNA expression analyses have been performed to ascertain its antitumor mechanism, revealing that up-regulation of the metabolic-related gene GCNT3 and down-regulation of its potential epigenetic modulator miR-15b correlate with the antitumor effect of rosemary. Moreover, plasmatic miR-15b down-regulation was detected after in vivo treatment with rosemary. Our results support the use of carnosic acid-rich rosemary extract as a complementary approach in colon and pancreatic cancer and indicate that GCNT3 expression may be involved in its antitumor mechanism and that miR-15b might be used as a non-invasive biomarker to monitor rosemary anticancer effect.

Consumption of the long-chain ω-3 (n–3) polyunsaturated fatty acid docosahexaenoic acid (DHA) is associated with a reduced risk of cardiovascular dis

Abnormal acyl-CoA synthetase/ stearoyl-CoA desaturase (ACSL/SCD) lipid network fuels colon cancer progression endowing cells with invasive and migratory properties. Therapies against this metabolic network may be useful to improve clinical outcomes. Since miRNAs (miRNAs/miR) are important epigenetic regulators, we investigated novel miRNAs targeting this pro-tumorigenic axis; and hence to be used both as therapeutic or prognostic miRNAs. Thirty-one putative common miRNAs were predicted to target simultaneously the three enzymes comprising ACSL/SCD network. Target validation by RT-QPCR, Western blotting and luciferase assays showed miR-544a, miR-142 and miR-19b-1 as major regulators of the metabolic axis ACSL/SCD. Importantly, lower miR-19b-1 expression associates with a decreased survival rate in CRC patients, accordingly with ACSL/SCD involvement in patient relapse. Finally, miR-19b-1 regulates the pro-tumorigenic axis ACSL/SCD being able to inhibit invasion in colon cancer cells. Since its expression correlates with an increased survival rate in CRC patients, we propose miR-19b-1 as a potential noninvasive biomarker of disease-free survival (DFS) and a promising therapeutic miRNA in CRC.