Clinical impact of mutated JAK2 allele burden reduction in polycythemia vera and essential thrombocythemia.

The variant allele frequency (VAF) of driver mutations (JAK2, CALR) in myeloproliferative neoplasms is associated with features of advanced disease and complications. Ruxolitinib and interferon were reported to variably reduce the mutant VAF, but the long-term impact of molecular responses (MR) remains debated. We prospectively measured changes in JAK2 and CALR VAF in 77 patients with polycythemia vera and essential thrombocythemia, treated with ruxolitinib for a median of 8 years, and assessed correlation with complete clinical and hematological response (CCHR) and outcomes. At last observation time, JAK2 VAF reduced overall from a median of 68% (range, 20%-99%) to 3.5% (0%-98%). A profound and durable MR (DMR; defined as a VAF stably ≤2%), including complete MR in 8%, was achieved in 20% of the patients, a partial MR (PMR; VAF reduction >50% of the baseline level) in 25%, and 56% had no molecular response (NMR). A CCHR was reached by 69% overall, independently of any degree of MR achieved; conversely, a DMR correlated with longer duration of CCHR and, most importantly, with reduced rate of progression to myelofibrosis and with longer myelofibrosis-free, event-free and progression-free survival. Achievement of PMR also had some favorable impact on outcomes, compared to NMR. A baseline JAK2 VAF <50%, and a VAF reduction of ≥35% after 2 years of treatment, predicted for the achievement of DMR and reduced progression to myelofibrosis. Overall, these findings support the clinical value of achieving profound, durable MR and its consideration as surrogate endpoint in future clinical trials.

Shared and Unique Disease Pathways in Amyotrophic Lateral Sclerosis and Parkinson's Disease Unveiled in Peripheral Blood Mononuclear Cells.

Recent evidence supports an association between amyotrophic lateral sclerosis (ALS) and Parkinson's disease (PD). Indeed, prospective population-based studies demonstrated that about one-third of ALS patients develop parkinsonian (PK) signs, even though different neuronal circuitries are involved. In this context, proteomics represents a valuable tool to identify unique and shared pathological pathways. Here, we used two-dimensional electrophoresis to obtain the proteomic profile of peripheral blood mononuclear cells (PBMCs) from PD and ALS patients including a small cohort of ALS patients with parkinsonian signs (ALS-PK). After the removal of protein spots correlating with confounding factors, we applied a sparse partial least square discriminant analysis followed by recursive feature elimination to obtain two protein classifiers able to discriminate (i) PD and ALS patients (30 spots) and (ii) ALS-PK patients among all ALS subjects (20 spots). Functionally, the glycolysis pathway was significantly overrepresented in the first signature, while extracellular interactions and intracellular signaling were enriched in the second signature. These results represent molecular evidence at the periphery for the classification of ALS-PK as ALS patients that manifest parkinsonian signs, rather than comorbid patients suffering from both ALS and PD. Moreover, we confirmed that low levels of fibrinogen in PBMCs is a characteristic feature of PD, also when compared with another movement disorder. Collectively, we provide evidence that peripheral protein signatures are a tool to differentially investigate neurodegenerative diseases and highlight altered biochemical pathways.

Exploring the Impact of PARK2 Mutations on the Total and Mitochondrial Proteome of Human Skin Fibroblasts.

Mutations in PARK2 gene are the most frequent cause of familial forms of Parkinson's disease (PD). This gene encodes Parkin, an E3 ubiquitin ligase involved in several cellular mechanisms, including mitophagy. Parkin loss-of-function is responsible for the cellular accumulation of damaged mitochondria, which in turn determines an increment of reactive oxygen species (ROS) levels, lower ATP production, and apoptosis activation. Given the importance of mitochondrial dysfunction and mitophagy impairment in PD pathogenesis, the aim of the present study was to investigate both total and mitochondrial proteome alterations in human skin fibroblasts of PARK2-mutated patients. To this end, both total and mitochondria-enriched protein fractions from fibroblasts of five PARK2-mutated patients and five control subjects were analyzed by quantitative shotgun proteomics to identify proteins specifically altered by Parkin mutations (mass spectrometry proteomics data have been submitted to ProteomeXchange with the identifier PXD015880). Both the network-based and gene set enrichment analyses pointed out pathways in which Rab GTPase proteins are involved. To have a more comprehensive view of the mitochondrial alterations due to PARK2 mutations, we investigated the impact of Parkin loss on mitochondrial function and network morphology. We unveiled that the mitochondrial membrane potential was reduced in PARK2-mutated patients, without inducing PINK1 accumulation, even when triggered with the ionophore carbonyl cyanide m-chlorophenylhydrazone (CCCP). Lastly, the analysis of the mitochondrial network morphology did not reveal any significant alterations in PARK2-mutated patients compared to control subjects. Thus, our results suggested that the network morphology was not influenced by the mitochondrial depolarization and by the lack of Parkin, revealing a possible impairment of fission and, more in general, of mitochondrial dynamics. In conclusion, the present work highlighted new molecular factors and pathways altered by PARK2 mutations, which will unravel possible biochemical pathways altered in the sporadic form of PD.

Proteomics turns functional.

Proteomics is acquiring a pivotal role in the comprehensive understanding of human biology. Biochemical processes involved in complex diseases, such as neurodegenerative diseases, diabetes and cancer, can be identified by combining proteomics analysis and bioinformatics tools. In the last ten years, the main output of differential proteomics investigations evolved from long lists of proteins to the generation of new hypotheses and their functional verification. The Journal of Proteomics participated to this progress, reporting more and more biologically-oriented papers with functional interpretation of proteomics data. This change in the field was due to both technological development and novel strategies in exploiting the deep characterization of proteomes. In this review, we explore several approaches that allow proteomics to turn functional. In particular, systems biology tools for data analysis are now routinely used to interpret results, thus defining the biological meaning of differentially abundant proteins. Moreover, by considering the importance of protein-protein interactions and the composition of macromolecular complexes, interactomics is complementing the information given by differential quantitative proteomics. Eventually, terminomics is unveiling new functions for cleaved proteoforms, by analyzing the effect of proteolysis globally. SIGNIFICANCE: Proteomics is rapidly evolving not only technologically but also strategically. The correct interpretation of proteomics data can reveal new functions of proteins in several biological backgrounds. Systems biology tools allow researchers to formulate new hypotheses to be further functionally tested. Interactomics is shedding new light on protein complexes truly involved in biochemical pathways and how their alteration can lead to dysfunctionality (in disease pathogenesis, for example). Terminomics is revealing the function of new discovered proteoforms and attributing a novel role to proteolysis. This review would provide the biologist important insights into current applications of several proteomic approaches that could offer new strategies to investigate biological systems.

Network Analysis Identifies Disease-Specific Pathways for Parkinson's Disease.

Neurodegenerative diseases are characterized by the progressive loss of specific neurons in selected regions of the central nervous system. The main clinical manifestation (movement disorders, cognitive impairment, and/or psychiatric disturbances) depends on the neuron population being primarily affected. Parkinson's disease is a common movement disorder, whose etiology remains mostly unknown. Progressive loss of dopaminergic neurons in the substantia nigra causes an impairment of the motor control. Some of the pathogenetic mechanisms causing the progressive deterioration of these neurons are not specific for Parkinson's disease but are shared by other neurodegenerative diseases, like Alzheimer's disease and amyotrophic lateral sclerosis. Here, we performed a meta-analysis of the literature of all the quantitative proteomic investigations of neuronal alterations in different models of Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis to distinguish between general and Parkinson's disease-specific pattern of neurodegeneration. Then, we merged proteomics data with genetics information from the DisGeNET database. The comparison of gene and protein information allowed us to identify 25 proteins involved uniquely in Parkinson's disease and we verified the alteration of one of them, i.e., transaldolase 1 (TALDO1), in the substantia nigra of 5 patients. By using open-source bioinformatics tools, we identified the biological processes specifically affected in Parkinson's disease, i.e., proteolysis, mitochondrion organization, and mitophagy. Eventually, we highlighted four cellular component complexes mostly involved in the pathogenesis: the proteasome complex, the protein phosphatase 2A, the chaperonins CCT complex, and the complex III of the respiratory chain.