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BackgroundChronic hemodynamic overloading leads to heart failure (HF) due to incompletely understood mechanisms. To gain deeper insight into the molecular pathophysiology of volume overload-induced HF and to identify potential markers and targets for novel therapies, we performed proteomic and mRNA expression analysis comparing myocardium from Wistar rats with HF induced by a chronic aorto-caval fistula (ACF) and sham-operated rats harvested at the advanced, decompensated stage of HF.MethodsWe analyzed control and failing myocardium employing iTRAQ labeling, two-dimensional peptide separation combining peptide IEF and nano-HPLC with MALDI-MS/MS. For the transcriptomic analysis we employed Illumina RatRef-12v1 Expression BeadChip.ResultsIn the proteomic analysis we identified 2030 myocardial proteins, of which 66 proteins were differentially expressed. The mRNA expression analysis identified 851 differentially expressed mRNAs.ConclusionsThe differentially expressed proteins confirm a switch in the substrate preference from fatty acids to other sources in the failing heart. Failing hearts showed downregulation of the major calcium transporters SERCA2 and ryanodine receptor 2 and altered expression of creatine kinases. Decreased expression of two NADPH producing proteins suggests a decreased redox reserve. Overexpression of annexins supports their possible potential as HF biomarkers. Most importantly, among the most up-regulated proteins in ACF hearts were monoamine oxidase A and transglutaminase 2 that are both potential attractive targets of low molecular weight inhibitors in future HF therapy.

BackgroundMantle cell lymphoma (MCL) is an aggressive type of B-cell non-Hodgkin lymphoma associated with poor prognosis. Implementation of high-dose cytarabine (araC) into induction therapy became standard-of-care for all newly diagnosed younger MCL patients. However, many patients relapse even after araC-based regimen. Molecular mechanisms responsible for araC resistance in MCL are unknown and optimal treatment strategy for relapsed/refractory MCL patients remains elusive.MethodsFive araC-resistant (R) clones were derived by long-term culture of five MCL cell lines (CTRL) with increasing doses of araC up to 50 microM. Illumina BeadChip and 2-DE proteomic analysis were used to identify gene and protein expression changes associated with araC resistance in MCL. In vitro cytotoxicity assays and experimental therapy of MCL xenografts in immunodeficient mice were used to analyze their relative responsiveness to a set of clinically used anti-MCL drugs. Primary MCL samples were obtained from patients at diagnosis and after failure of araC-based therapies.ResultsMarked downregulation of deoxycytidine-kinase (DCK) mRNA and protein expression was identified as the single most important molecular event associated with araC-resistance in all tested MCL cell lines and in 50% primary MCL samples. All R clones were highly (20-1000x) cross-resistant to all tested nucleoside analogs including gemcitabine, fludarabine and cladribine. In vitro sensitivity of R clones to other classes of clinically used anti-MCL agents including genotoxic drugs (cisplatin, doxorubicin, bendamustine) and targeted agents (bortezomib, temsirolimus, rituximab) remained unaffected, or was even increased (ibrutinib). Experimental therapy of immunodeficient mice confirmed the anticipated loss of anti-tumor activity (as determined by overall survival) of the nucleoside analogs gemcitabine and fludarabine in mice transplanted with R clone compared to mice transplanted with CTRL cells, while the anti-tumor activity of cisplatin, temsirolimus, bortezomib, bendamustine, cyclophosphamide and rituximab remained comparable between the two cohorts.ConclusionsAcquired resistance of MCL cells to araC is associated with downregulation of DCK, enzyme of the nucleotide salvage pathway responsible for the first phosphorylation (=activation) of most nucleoside analogs used in anti-cancer therapy. The data suggest that nucleoside analogs should not be used in the therapy of MCL patients, who relapse after failure of araC-based therapies.

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