HLA diversity in the Russian population assessed by next generation sequencing

Next generation sequencing is used to determine full-length sequences of HLA genes at the 4-field (allelic) resolution. The study was aimed at determining frequency and diversity of HLA alleles in a cohort of blood donors from the Registry of the National Research Center for Hematology who design ated themselves as Russians (including some not routinely typed variations in HLA gene regions). The studied population consisted of 1510 donors. HLA typing was performed by next generation sequencing. Libraries were performed with AllType NGS Amplification Kits (One Lambda, USA) and sequenced using MiSeq (Illumina, USA). Data analysis used the TypeStream Visual Software V2.0.0.68 (One Lambda, USA) and IPD-IMGT/HLA database 3.40.0.1. Arlequin 3.5 software was used for estimation of allele and haplotype frequencies, deviation from Hardy-Weinberg equilibrium. 82 HLA-A, 156 HLA-В and 85 HLA-С alleles were identified with four-field resolution. 45 HLA-DRB1 and 18 HLA-DQB1 alleles were identified with 2-3-field resolution. Considerable HLA diversity was found among the donors self-designated as Russians: the population had large numbers of distinct alleles at each HLA gene, high percentage of alleles (25-32% of HLA class I) were revealed only once. Sufficient numbers of new alleles were registered which are absent in the IPD-IMGT/HLA database. Considerable allelic diversity in Russian population is due to low-incidence alleles. Despite this diversity, the majority of HLA alleles detected at each locus were common. Significant HLA diversity of the donors was connected with a large number of alleles with rare occurrence. The novel alleles identified in our study differed from the known alleles by single nucleotide substitutions. The most common alleles at the four-field level were as follows: A*02:01:01:01 (27.1%), C*07:02:01:03 (13.1%), A*03:01:01:01 (13.0%), B*07:02:01:01 (13.0%), A*01:01:01:01 (11.6%) and C*07:01:01:01/16 (10.4%). The HLA alleles, which are common for Russian populations, are not always common or well-documented alleles in present catalogues. The data obtained in this study may be used as a reference sample for estimation of HLA allele frequencies in Russian population, for proper frequency evaluation of specific alleles when searching donors for allogeneic hematopoietic stem cell transplantation, as well as for association studies between HLA alleles and different diseases, and for research in population genetics

Identification of evolutionarily conserved DNA damage response genes that alter sensitivity to cisplatin

Ovarian, head and neck, and other cancers are commonly treated with cisplatin and other DNA damaging cytotoxic agents. Altered DNA damage response (DDR) contributes to resistance of these tumors to chemotherapies, some targeted therapies, and radiation. DDR involves multiple protein complexes and signaling pathways, some of which are evolutionarily ancient and involve protein orthologs conserved from yeast to humans. To identify new regulators of cisplatin-resistance in human tumors, we integrated high throughput and curated datasets describing yeast genes that regulate sensitivity to cisplatin and/or ionizing radiation. Next, we clustered highly validated genes based on chemogenomic profiling, and then mapped orthologs of these genes in expanded genomic networks for multiple metazoans, including humans. This approach identified an enriched candidate set of genes involved in the regulation of resistance to radiation and/or cisplatin in humans. Direct functional assessment of selected candidate genes using RNA interference confirmed their activity in influencing cisplatin resistance, degree of γH2AX focus formation and ATR phosphorylation, in ovarian and head and neck cancer cell lines, suggesting impaired DDR signaling as the driving mechanism. This work enlarges the set of genes that may contribute to chemotherapy resistance and provides a new contextual resource for interpreting next generation sequencing (NGS) genomic profiling of tumors

STUDY OF MINIMAL RESIDUAL DISEASE BY MULTICOLOR FLOW CYTOMETRY IN MULTIPLE MYELOMA AFTER AUTOLOGOUS HEMATOPOIETIC STEM CELL TRANSPLANTATION

The frequency of achieving complete remission, as well as overall and disease-free survival, in multiple myeloma (MM) had increased due to introduction in MM treatment regimens of high-dose chemotherapy with following autologous hematopoietic stem cell transplantation (ASCT). However the number of relapses remains high, caused by persistence of residual tumor cells, i.e., the presence of minimal residual disease (MRD). One of the methods for MRD study is multicolor flow cytometry (MFC) where abnormal expression of surface antigens on myeloma plasma cells (PC) is determined. The aim of our study was to investigate the MRD by MFC before and after ASCT, the frequency of MRD-negative status achievement in complete remission (CR) patients at +100 days after ASCT and the frequency of abnormal expressed antigens on myeloma plasma cells. The study included40 MMpatients in CR at +100 days after ASCT and showed that the most common aberrations of PC were: abnormal absence of CD19 and/or CD27, decreased expression of CD38 and abnormal presence of CD56. The proportion of myeloma PCs from all bone marrow cells decreased significantly after ASCT: 20 % of patients acquired MRD-negative status, 10 % had a decrease in the number of abnormal PCs by one fold. Analysis of probability of immunochemical relapse showed that the worst prognosis was in patients with MRD-positive status before and after ASCT. During the MRD monitoring within 3-18 months, MRD-relapses were detected with the subsequent development of immunochemical relapse. The detection MRD in the dynamics is more informative than the study at only one step of therapy. It may help to select more adequate treatment for patient with multiple myeloma in each specific case

INVESTIGATION OF CYTOKINE PROFILE IN PATIENTS WITH REACTIVE ARTHRITIS

Abstract. Pathogenesis of reactive arthritis (ReA) is not clear yet. Several trials suggest that increased production of proinflammatory cytokines is responsible for development of arthritis in ReA, while other studies report that Th1 cytokine response in ReA is impaired in favor of Th2 response. The aim of our study was to investigate serum levels of cytokines IL-1β, IL-4, IL-6, TNFα, IFNγ and IL-1Ra in the patients with ReA of different etiology, as compared with infection-related arthritis. The results of our study had demonstrated that serum levels of IL-1β and TNFα in the patients with ReA were significantly higher, whereas IL-1Ra, IL-4, IL-6 proved to be significantly lower than in healthy controls. Serum levels of IL-6 were significantly higher in patients with chronic ReA, as compared to the cases of acute and recurrent ReA. No significant differences in cytokine profiles were found between the patients with ReA, and the persons with infection-related arthritis. The data obtained are, generally, suggestive for proinflammatory Th1 cytokine profile in ReA patients studied, this confirming the mostly assumed pathogenetic hypothesis for reactive arthritis where an underlying cytokine imbalance is suggested. (Med. Immunol., 2008, vol. 10, N 2-3, pp 167-172)

Identification of evolutionarily conserved DNA damage response genes that alter sensitivity to cisplatin

Ovarian, head and neck, and other cancers are commonly treated with cisplatin and other DNA damaging cytotoxic agents. Altered DNA damage response (DDR) contributes to resistance of these tumors to chemotherapies, some targeted therapies, and radiation. DDR involves multiple protein complexes and signaling pathways, some of which are evolutionarily ancient and involve protein orthologs conserved from yeast to humans. To identify new regulators of cisplatin-resistance in human tumors, we integrated high throughput and curated datasets describing yeast genes that regulate sensitivity to cisplatin and/or ionizing radiation. Next, we clustered highly validated genes based on chemogenomic profiling, and then mapped orthologs of these genes in expanded genomic networks for multiple metazoans, including humans. This approach identified an enriched candidate set of genes involved in the regulation of resistance to radiation and/or cisplatin in humans. Direct functional assessment of selected candidate genes using RNA interference confirmed their activity in influencing cisplatin resistance, degree of γH2AX focus formation and ATR phosphorylation, in ovarian and head and neck cancer cell lines, suggesting impaired DDR signaling as the driving mechanism. This work enlarges the set of genes that may contribute to chemotherapy resistance and provides a new contextual resource for interpreting next generation sequencing (NGS) genomic profiling of tumors

Identification of evolutionarily conserved DNA damage response genes that alter sensitivity to cisplatin

Ovarian, head and neck, and other cancers are commonly treated with cisplatin and other DNA damaging cytotoxic agents. Altered DNA damage response (DDR) contributes to resistance of these tumors to chemotherapies, some targeted therapies, and radiation. DDR involves multiple protein complexes and signaling pathways, some of which are evolutionarily ancient and involve protein orthologs conserved from yeast to humans. To identify new regulators of cisplatin-resistance in human tumors, we integrated high throughput and curated datasets describing yeast genes that regulate sensitivity to cisplatin and/or ionizing radiation. Next, we clustered highly validated genes based on chemogenomic profiling, and then mapped orthologs of these genes in expanded genomic networks for multiple metazoans, including humans. This approach identified an enriched candidate set of genes involved in the regulation of resistance to radiation and/or cisplatin in humans. Direct functional assessment of selected candidate genes using RNA interference confirmed their activity in influencing cisplatin resistance, degree of γH2AX focus formation and ATR phosphorylation, in ovarian and head and neck cancer cell lines, suggesting impaired DDR signaling as the driving mechanism. This work enlarges the set of genes that may contribute to chemotherapy resistance and provides a new contextual resource for interpreting next generation sequencing (NGS) genomic profiling of tumors

Identification of evolutionarily conserved DNA damage response genes that alter sensitivity to cisplatin

Ovarian, head and neck, and other cancers are commonly treated with cisplatin and other DNA damaging cytotoxic agents. Altered DNA damage response (DDR) contributes to resistance of these tumors to chemotherapies, some targeted therapies, and radiation. DDR involves multiple protein complexes and signaling pathways, some of which are evolutionarily ancient and involve protein orthologs conserved from yeast to humans. To identify new regulators of cisplatin-resistance in human tumors, we integrated high throughput and curated datasets describing yeast genes that regulate sensitivity to cisplatin and/or ionizing radiation. Next, we clustered highly validated genes based on chemogenomic profiling, and then mapped orthologs of these genes in expanded genomic networks for multiple metazoans, including humans. This approach identified an enriched candidate set of genes involved in the regulation of resistance to radiation and/or cisplatin in humans. Direct functional assessment of selected candidate genes using RNA interference confirmed their activity in influencing cisplatin resistance, degree of γH2AX focus formation and ATR phosphorylation, in ovarian and head and neck cancer cell lines, suggesting impaired DDR signaling as the driving mechanism. This work enlarges the set of genes that may contribute to chemotherapy resistance and provides a new contextual resource for interpreting next generation sequencing (NGS) genomic profiling of tumors