Challenges of the Human Proteome Project: 10-Year Experience of the Russian Consortium.

This manuscript collects all the efforts of the Russian Consortium, bottlenecks revealed in the course of the C-HPP realization, and ways of their overcoming. One of the main bottlenecks in the C-HPP is the insufficient sensitivity of proteomic technologies, hampering the detection of low- and ultralow-copy number proteins forming the "dark part" of the human proteome. In the frame of MP-Challenge, to increase proteome coverage we suggest an experimental workflow based on a combination of shotgun technology and selected reaction monitoring with two-dimensional alkaline fractionation. Further, to detect proteins that cannot be identified by such technologies, nanotechnologies such as combined atomic force microscopy with molecular fishing and/or nanowire detection may be useful. These technologies provide a powerful tool for single molecule analysis, by analogy with nanopore sequencing during genome analysis. To systematically analyze the functional features of some proteins (CP50 Challenge), we created a mathematical model that predicts the number of proteins differing in amino acid sequence: proteoforms. According to our data, we should expect about 100 000 different proteoforms in the liver tissue and a little more in the HepG2 cell line. The variety of proteins forming the whole human proteome significantly exceeds these results due to post-translational modifications (PTMs). As PTMs determine the functional specificity of the protein, we propose using a combination of gene-centric transcriptome-proteomic analysis with preliminary fractionation by two-dimensional electrophoresis to identify chemically modified proteoforms. Despite the complexity of the proposed solutions, such integrative approaches could be fruitful for MP50 and CP50 Challenges in the framework of the C-HPP.

Gene-centric view on the human proteome project: the example of the Russian roadmap for chromosome 18.

During the 2010 Human Proteome Organization Congress in Sydney, a gene-centric approach emerged as a feasible and tractable scaffold for assemblage of the Human Proteome Project. Bringing the gene-centric principle into practice, a roadmap for the 18th chromosome was drafted, postulating the limited sensitivity of analytical methods, as a serious bottleneck in proteomics. In the context of the sensitivity problem, we refer to the "copy number of protein molecules" as a measurable assessment of protein abundance. The roadmap is focused on the development of technology to attain the low- and ultralow -"copied" portion of the proteome. Roadmap merges the genomic, transcriptomic and proteomic levels to identify the majority of 285 proteins from 18th chromosome - master proteins. Master protein is the primary translation of the coding sequence and resembling at least one of the known isoforms, coded by the gene. The executive phase of the roadmap includes the expansion of the study of the master proteins with alternate splicing, single amino acid polymorphisms (SAPs) and post-translational modifications. In implementing the roadmap, Russian scientists are expecting to establish proteomic technologies for integrating MS and atomic force microscopy (AFM). These technologies are anticipated to unlock the value of new biomarkers at a detection limit of 10(-18) M, i.e. 1 protein copy per 1 µL of plasma. The roadmap plan is posted at www.proteome.ru/en/roadmap/ and a forum for discussion of the document is supported.

Treatment of high fracture risk patients in routine clinical practice.

A retrospective cohort study determined the high incidence of recurrent fractures in osteoporotic patients with high fracture risk during the observation. The strategy of starting treatment with more potent regimens (zoledronic acid, denosumab and/or teriparatide) seems to have the best secondary fracture prevention efficacy. OBJECTIVE: This paper describes the various medical therapy regimens prescribed to osteoporotic patients with high fracture risk and the result of treatment. METHODS: We carried out a retrospective cohort study in selected Osteoporosis Centers. Patients were considered to have high fracture risk in case of a history of a low-energy hip fracture or two or more vertebral or other site fractures. A total of 812 subjects (768 women and 44 men) aged 36-95 years were included. The observation period was 2285.1 patient-years. Demographic data, clinical findings, and BMD data obtained by DXA, as well as a history of fractures that had occurred during the follow-up, were included in the analysis. RESULTS: Overall, at baseline, there were 637 non-vertebral fractures including 104 hip fractures. A total of 590 patients had vertebral fractures; of these, 69% suffered multiple fractures. Being on treatment, 119 (14.7%) patients developed new vertebral and non-vertebral fractures. The incidence of new non-vertebral fractures and hip fractures was 39.4 and 13.1 per 1000 patient-years. The total number of vertebral fractures increased by 24.8% from 1353 to 1689. The best results of the treatment were achieved in patients who were started on zoledronic acid, denosumab, or teriparatide and had an adequate duration of treatment. Although these patients had significantly lower BMD values at the time of diagnosis compared with other patients, they showed a lower incidence of new vertebral and hip fractures, during the follow-up. CONCLUSION: Therapy of patients at high risk of fractures started with more potent treatment regimens (zoledronic acid, denosumab and/or teriparatide) of adequate duration was more effective in terms of prevention of new vertebral and hip fractures as compared with other treatment options. However, treatment appears to be challenging given the number of recurrent fractures in patients on treatment and the frequency of drug withdrawal or replacement.

Arthroscopic operations in joint tuberculosis.

We assessed the usefulness of arthroscopy in bone and joint tuberculosis (11 knees, 1 elbow and 1 ankle). In all 11 patients with knee joint involvement the arthroscopic operation were successful with a significant improvement in the range of motion (P=0.007) and knee score (P=0.003). There were no cases of reactivation or recurrence during a follow-up period of 15-42 months. In contrast, in the two patients with ankle or elbow joint involvement and bone destruction, arthroscopy had to be converted to open operation. Minor trauma, a short hospitalization time and a short post-operative rehabilitation period combined with good cosmetic and clinical results favor the use of the arthroscopic technology in the surgical treatment of tuberculous arthritis of the knee.