Fee Arrangements and Fee Shifting: Lessons From the Experience in Ontario

About one-third of oestrogen receptor alpha-positive breast cancer patients treated with tamoxifen relapse. Here we identify the nuclear receptor retinoic acid receptor alpha as a marker of tamoxifen resistance. Using quantitative mass spectrometry-based proteomics, we show that retinoic acid receptor alpha protein networks and levels differ in a tamoxifen-sensitive (MCF7) and a tamoxifen-resistant (LCC2) cell line. High intratumoural retinoic acid receptor alpha protein levels also correlate with reduced relapse-free survival in oestrogen receptor alpha-positive breast cancer patients treated with adjuvant tamoxifen solely. A similar retinoic acid receptor alpha expression pattern is seen in a comparable independent patient cohort. An oestrogen receptor alpha and retinoic acid receptor alpha ligand screening reveals that tamoxifen-resistant LCC2 cells have increased sensitivity to retinoic acid receptor alpha ligands and are less sensitive to oestrogen receptor alpha ligands compared with MCF7 cells. Our data indicate that retinoic acid receptor alpha may be a novel therapeutic target and a predictive factor for oestrogen receptor alpha-positive breast cancer patients treated with adjuvant tamoxifen

Nuclear poly(ADP-ribose) activity is a therapeutic target in amyotrophic lateral sclerosis

Abstract Amyotrophic lateral sclerosis (ALS) is a devastating and fatal motor neuron disease. Diagnosis typically occurs in the fifth decade of life and the disease progresses rapidly leading to death within ~ 2–5 years of symptomatic onset. There is no cure, and the few available treatments offer only a modest extension in patient survival. A protein central to ALS is the nuclear RNA/DNA-binding protein, TDP-43. In > 95% of ALS patients, TDP-43 is cleared from the nucleus and forms phosphorylated protein aggregates in the cytoplasm of affected neurons and glia. We recently defined that poly(ADP-ribose) (PAR) activity regulates TDP-43-associated toxicity. PAR is a posttranslational modification that is attached to target proteins by PAR polymerases (PARPs). PARP-1 and PARP-2 are the major enzymes that are active in the nucleus. Here, we uncovered that the motor neurons of the ALS spinal cord were associated with elevated nuclear PAR, suggesting elevated PARP activity. Veliparib, a small-molecule inhibitor of nuclear PARP-1/2, mitigated the formation of cytoplasmic TDP-43 aggregates in mammalian cells. In primary spinal-cord cultures from rat, Veliparib also inhibited TDP-43-associated neuronal death. These studies uncover that PAR activity is misregulated in the ALS spinal cord, and a small-molecular inhibitor of PARP-1/2 activity may have therapeutic potential in the treatment of ALS and related disorders associated with abnormal TDP-43 homeostasis

Phosphoproteomic Profiling of NSCLC Cells Reveals that Ephrin B3 Regulates Pro-survival Signaling through Akt1-Mediated Phosphorylation of the EphA2 Receptor

The ephrin and Eph signaling circuit has been reported as deregulated in a number of tumor types including nonsmall cell lung cancer (NSCLC). Here we show that suppression of the ephrin-familly member ephrin B3 decreases NSCLC cell proliferation and has profound effects on cell morphology. To reveal which signaling networks ephrin B3 utilize to regulate such effects on growth and morphology, differential regulation of phosphorylated proteins was analyzed in the NSCLC cell line U-1810. Using strong cat ion exchange (SCX) and TiO(2)-based fractionation followed by nano-LC and mass spectrometry analysis, we identified 1083 unique phosphorylated proteins. Out of these, 150 proteins were found only when ephrin B3 is expressed, whereas 66 proteins were found exclusively in U-1810 cells with silenced ephrin B3. Network analysis of changes in the phosphoproteome with regard to the presence or absence of ephrin B3 expression generated a hypothesis that the site specific phosphorylation on Ser-897 detected on the erythropoietin-producing hepatocellular receptor tyrosine kinase class A2 (EphA2) is critical for the survival of NSCLC cells. Upstream of the EphA2 phosphorylation, activation of Akt1 on Ser 129 was also revealed as part of the ephrin B3-mediated signaling pathway. Phosphorylation of these sites was further confirmed by immune-based strategies in combination with mass spectrometry. Moreover, by further stepwise pathway walking, annotating the phosphorylated sites and their corresponding kinases upstream, our data support the process in which a Heat shock protein 90 isoform (HSP90AA1) acts as a protector of EphA2, thereby saving it from degradation. In addition, protein kinase CK2 (CK2) is suggested as a dominant kinase, activating downstream substrates to generate the effects on NSCLC proliferation and morphology

Therapeutic cancer vaccination with immunopeptidomics-discovered antigens confers protective antitumor efficacy

Knowledge of clinically targetable tumor antigens is becoming vital for broader design and utility of therapeutic cancer vaccines. This information is obtained reliably by directly interro-gating the MHC-I presented peptide ligands, the immunopeptidome, with state-of-the-art mass spectrometry. Our manuscript describes direct identification of novel tumor antigens for an aggres-sive triple-negative breast cancer model. Immunopeptidome profiling revealed 2481 unique anti-gens, among them a novel ERV antigen originating from an endogenous retrovirus element. The clinical benefit and tumor control potential of the identified tumor antigens and ERV antigen were studied in a preclinical model using two vaccine platforms and therapeutic settings. Prominent control of established tumors was achieved using an oncolytic adenovirus platform designed for flexi-ble and specific tumor targeting, namely PeptiCRAd. Our study presents a pipeline integrating im-munopeptidome analysis-driven antigen discovery with a therapeutic cancer vaccine platform for improved personalized oncolytic immunotherapy