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Article: Viral and Host Genetic and Epigenetic Biomarkers Related to SARS-CoV-2 Cell Entry, Infection Rate, and Disease Severity.

Gaspersic, Jernej / Dolzan, Vita

2022 Volume 11, Issue 2

Abstract: The rapid spread of COVID-19 outbreak lead to a global pandemic declared in March 2020. The common features of corona virus family helped to resolve structural characteristics and entry mechanism of SARS-CoV-2. However, rapid mutagenesis leads to the ... ...

Abstract	The rapid spread of COVID-19 outbreak lead to a global pandemic declared in March 2020. The common features of corona virus family helped to resolve structural characteristics and entry mechanism of SARS-CoV-2. However, rapid mutagenesis leads to the emergence of new strains that may have different reproduction rates or infectivity and may impact the course and severity of the disease. Host related factors may also play a role in the susceptibility for infection as well as the severity and outcomes of the COVID-19. We have performed a literature and database search to summarize potential viral and host-related genomic and epigenomic biomarkers, such as genetic variability, miRNA, and DNA methylation in the molecular pathway of SARS-CoV-2 entry into the host cell, that may be related to COVID-19 susceptibility and severity. Bioinformatics tools may help to predict the effect of mutations in the spike protein on the binding to the ACE2 receptor and the infectivity of the strain. SARS-CoV-2 may also target several transcription factors and tumour suppressor genes, thus influencing the expression of different host genes and affecting cell signalling. In addition, the virus may interfere with RNA expression in host cells by exploiting endogenous miRNA and its viral RNA. Our analysis showed that numerous human miRNA may form duplexes with different coding and non-coding regions of viral RNA. Polymorphisms in human genes responsible for viral entry and replication, as well as in molecular damage response and inflammatory pathways may also contribute to disease prognosis and outcome. Gene ontology analysis shows that proteins encoded by such polymorphic genes are highly interconnected in regulation of defense response. Thus, virus and host related genetic and epigenetic biomarkers may help to predict the course of the disease and the response to treatment.
Language	English
Publishing date	2022-01-23
Publishing country	Switzerland
Document type	Journal Article ; Review
ZDB-ID	2661517-4
ISSN	2079-7737
ISSN	2079-7737
DOI	10.3390/biology11020178
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Article ; Online: Potential of modern circulating cell-free DNA diagnostic tools for detection of specific tumour cells in clinical practice.

Gašperšič, Jernej / Videtič Paska, Alja

Biochemia medica

2020 Volume 30, Issue 3, Page(s) 30504

Abstract: Personalized medicine is a developing field of medicine that has gained in importance in recent decades. New diagnostic tests based on the analysis of circulating cell-free DNA (cfDNA) were developed as a tool of diagnosing different cancer types. By ... ...

Abstract	Personalized medicine is a developing field of medicine that has gained in importance in recent decades. New diagnostic tests based on the analysis of circulating cell-free DNA (cfDNA) were developed as a tool of diagnosing different cancer types. By detecting the subpopulation of mutated DNA from cancer cells, it is possible to detect the presence of a specific tumour in early stages of the disease. Mutation analysis is performed by quantitative polymerase chain reaction (qPCR) or the next generation sequencing (NGS), however, cfDNA protocols need to be modified carefully in preanalytical, analytical, and postanalytical stages. To further improve treatment of cancer the Food and Drug Administration approved more than 20 companion diagnostic tests that combine cancer drugs with highly efficient genetic diagnostic tools. Tools detect mutations in the DNA originating from cancer cells directly through the subpopulation of cfDNA, the circular tumour DNA (ctDNA) analysis or with visualization of cells through intracellular DNA probes. A large number of ctDNA tests in clinical studies demonstrate the importance of new findings in the field of cancer diagnosis. We describe the innovations in personalized medicine: techniques for detecting ctDNA and genomic DNA (gDNA) mutations approved Food and Drug Administration companion genetic diagnostics, candidate genes for assembling the cancer NGS panels, and a brief mention of the multitude of cfDNA currently in clinical trials. Additionally, an overview of the development steps of the diagnostic tools will refresh and expand the knowledge of clinics and geneticists for research opportunities beyond the development phases.
MeSH term(s)	Biomarkers, Tumor/blood ; Biomarkers, Tumor/genetics ; Cell-Free Nucleic Acids/blood ; DNA, Neoplasm/blood ; High-Throughput Nucleotide Sequencing ; Humans ; Liquid Biopsy ; Neoplasms/diagnosis ; Neoplasms/genetics ; Precision Medicine ; Real-Time Polymerase Chain Reaction
Chemical Substances	Biomarkers, Tumor ; Cell-Free Nucleic Acids ; DNA, Neoplasm
Language	English
Publishing date	2020-08-05
Publishing country	Croatia
Document type	Journal Article ; Review
ZDB-ID	1208725-7
ISSN	1846-7482 ; 1330-0962
ISSN (online)	1846-7482
ISSN	1330-0962
DOI	10.11613/BM.2020.030504
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Article: Viral and Host Genetic and Epigenetic Biomarkers Related to SARS-CoV-2 Cell Entry, Infection Rate, and Disease Severity

Gaspersic, Jernej / Dolzan, Vita

Biology. 2022 Jan. 23, v. 11, no. 2

2022

Abstract	The rapid spread of COVID-19 outbreak lead to a global pandemic declared in March 2020. The common features of corona virus family helped to resolve structural characteristics and entry mechanism of SARS-CoV-2. However, rapid mutagenesis leads to the emergence of new strains that may have different reproduction rates or infectivity and may impact the course and severity of the disease. Host related factors may also play a role in the susceptibility for infection as well as the severity and outcomes of the COVID-19. We have performed a literature and database search to summarize potential viral and host-related genomic and epigenomic biomarkers, such as genetic variability, miRNA, and DNA methylation in the molecular pathway of SARS-CoV-2 entry into the host cell, that may be related to COVID-19 susceptibility and severity. Bioinformatics tools may help to predict the effect of mutations in the spike protein on the binding to the ACE2 receptor and the infectivity of the strain. SARS-CoV-2 may also target several transcription factors and tumour suppressor genes, thus influencing the expression of different host genes and affecting cell signalling. In addition, the virus may interfere with RNA expression in host cells by exploiting endogenous miRNA and its viral RNA. Our analysis showed that numerous human miRNA may form duplexes with different coding and non-coding regions of viral RNA. Polymorphisms in human genes responsible for viral entry and replication, as well as in molecular damage response and inflammatory pathways may also contribute to disease prognosis and outcome. Gene ontology analysis shows that proteins encoded by such polymorphic genes are highly interconnected in regulation of defense response. Thus, virus and host related genetic and epigenetic biomarkers may help to predict the course of the disease and the response to treatment.
Keywords	COVID-19 infection ; DNA methylation ; Severe acute respiratory syndrome coronavirus 2 ; bioinformatics ; biomarkers ; disease severity ; epigenetics ; epigenome ; gene ontology ; genetic variation ; genomics ; humans ; microRNA ; mutagenesis ; neoplasms ; pandemic ; pathogenicity ; prognosis ; reproduction ; viruses
Language	English
Dates of publication	2022-0123
Publishing place	Multidisciplinary Digital Publishing Institute
Document type	Article
ZDB-ID	2661517-4
ISSN	2079-7737
ISSN	2079-7737
DOI	10.3390/biology11020178
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Feature Article: Metabolic Network Modelling of Chinese Hamster Ovary (CHO) Culture Bioreactors Operated as Microbial Cell Factories.

Gašperšič, Jernej / Kastelic, Miha / Novak, Uroš / Likozar, Blaž

Acta chimica Slovenica

2021 Volume 65, Issue 4, Page(s) 769–789

Abstract: Chinese hamster ovary (CHO) epithelial cells are one of the most used therapeutic medical lines for the production of different biopharmaceutical drugs. They have a high consumption rate with a fast duplication cycle that makes them an ideal biological ... ...

Abstract	Chinese hamster ovary (CHO) epithelial cells are one of the most used therapeutic medical lines for the production of different biopharmaceutical drugs. They have a high consumption rate with a fast duplication cycle that makes them an ideal biological clone. The higher accumulated amounts of toxic intracellular intermediates may lead to lower organism viability, protein productivity and manufactured biosimilar, so a careful optimal balance of medium, bioreactor operational parameters and bioprocess is needed. A precise phenomenological knowledge of metabolism's chemical transformations can predict problems that may arise during batch, semi-continuous fed batch and continuous reactor operation. For a better detailed understanding (and relations), future performance optimization and scaling, mechanistic model systems have been built. In this specific work, the main metabolic pathways in mammalian structured CHO cultures are reviewed. It starts with organic biochemical background, controlling associated phenomena and kinetics, which govern the sustaining conversion routes of biology. Then, individual turnover paths are described, overviewing standard mathematical formulations that are commonly applied in engineering. These are the core of black box modeling, which relates the substrates/products in a simplified relationship manner. Moreover, metabolic flux analysis (MFA)/flux balance analysis (FBA), that are traditionally characterizing mechanisms, are presented to a larger portion extent. Finally, similarities are discussed, illustrating the approaches for their structural design. Stated variables' equations, employed for the description of the growth in the controllable environmental conditions of a vessel, the researched reaction series of proliferating dividing CHO population, joint with the values of maximal enzymatic activity, and solutions are outlined. Processes are listed in a way so that a reader can integrate the state-of-the-art. Our particular contribution is also denoted.
Language	English
Publishing date	2021-02-05
Publishing country	Slovenia
Document type	Journal Article
ZDB-ID	2029709-9
ISSN	1580-3155 ; 1318-0207
ISSN (online)	1580-3155
ISSN	1318-0207
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Article ; Online: Erythrocytosis: genes and pathways involved in disease development.

Gašperšič, Jernej / Kristan, Aleša / Kunej, Tanja / Zupan, Irena Preložnik / Debeljak, Nataša

Blood transfusion = Trasfusione del sangue

2020 Volume 19, Issue 6, Page(s) 518–532

Abstract: Erythrocytosis is a blood disorder characterised by an increased red blood cell mass. The most common causes of erythrocytosis are acquired and caused by diseases and conditions that are accompanied by hypoxaemia or overproduction of erythropoietin. More ...

Abstract	Erythrocytosis is a blood disorder characterised by an increased red blood cell mass. The most common causes of erythrocytosis are acquired and caused by diseases and conditions that are accompanied by hypoxaemia or overproduction of erythropoietin. More rarely, erythrocytosis has a known genetic background, such as for polycythaemia vera and familial erythrocytosis. The majority of cases of polycythaemia vera are associated with acquired variants in JAK2, while familial erythrocytosis is a group of congenital disorders. Familial erythrocytosis type 1 is associated with hypersensitivity to erythropoietin (variants in EPOR), types 2-5 with defects in oxygen-sensing pathways (variants in VHL, EGLN1, EPAS1, EPO), and types 6-8 with an increased affinity of haemoglobin for oxygen (variants in HBB, HBA1, HBA2, BPGM). Due to a heterogenic genetic background, the causes of disease are not fully discovered and in more than 70% of patients the condition remains labelled idiopathic.The transfer of next-generation sequencing into clinical practice is becoming a reality enabling detection of various variants in a single rapid test. In this review, we describe the current research on erythrocytosis gene variants and the mechanisms associated with disease development, along with the currently used diagnostic tests.
MeSH term(s)	Erythropoietin/genetics ; Hemoglobins ; Humans ; Mutation ; Oxygen ; Polycythemia/genetics
Chemical Substances	Hemoglobins ; Erythropoietin (11096-26-7) ; Oxygen (S88TT14065)
Language	English
Publishing date	2020-12-16
Publishing country	Italy
Document type	Journal Article ; Research Support, Non-U.S. Gov't ; Review
ZDB-ID	2135732-8
ISSN	2385-2070 ; 0041-1787 ; 1723-2007
ISSN (online)	2385-2070
ISSN	0041-1787 ; 1723-2007
DOI	10.2450/2020.0197-20
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Article ; Online: Feature Article

Jernej Gašperšič / Miha Kastelic / Uroš Novak / Blaž Likozar

Acta Chimica Slovenica, Vol 65, Iss 4, Pp 769-

Metabolic Network Modelling of Chinese Hamster Ovary (CHO) Culture Bioreactors Operated as Microbial Cell Factories

2018 Volume 789

Abstract	Chinese hamster ovary (CHO) epithelial cells are one of the most used therapeutic medical lines for the production of different biopharmaceutical drugs. They have a high consumption rate with a fast duplication cycle that makes them an ideal biological clone. The higher accumulated amounts of toxic intracellular intermediates may lead to lower organism viability, protein productivity and manufactured biosimilar, so a careful optimal balance of medium, bioreactor operational parameters and bioprocess is needed. A precise phenomenological knowledge of metabolism’s chemical transformations can predict problems that may arise during batch, semi-continuous fed batch and continuous reactor operation. For a better detailed understanding (and relations), future performance optimization and scaling, mechanistic model systems have been built. In this specific work, the main metabolic pathways in mammalian structured CHO cultures are reviewed. It starts with organic biochemical background, controlling associated phenomena and kinetics, which govern the sustaining conversion routes of biology. Then, individual turnover paths are described, overviewing standard mathematical formulations that are commonly applied in engineering. These are the core of black box modeling, which relates the substrates/products in a simplified relationship manner. Moreover, metabolic flux analysis (MFA)/flux balance analysis (FBA), that are traditionally characterizing mechanisms, are presented to a larger portion extent. Finally, similarities are discussed, illustrating the approaches for their structural design. Stated variables’ equations, employed for the description of the growth in the controllable environmental conditions of a vessel, the researched reaction series of proliferating dividing CHO population, joint with the values of maximal enzymatic activity, and solutions are outlined. Processes are listed in a way so that a reader can integrate the state-of-the-art. Our particular contribution is also denoted.
Keywords	Microbial cell factory ; Chinese hamster ovary (CHO) cell metabolism ; Bioreactor operation modeling ; Biochemical reaction kinetics ; Metabolic flux analysis ; Biopharmaceutical and biosimilar ; Chemistry ; QD1-999
Subject code	660
Language	English
Publishing date	2018-12-01T00:00:00Z
Publisher	Slovenian Chemical Society
Document type	Article ; Online
Database	BASE - Bielefeld Academic Search Engine (life sciences selection)

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Article ; Online: High-throughput immunoaffinity enrichment and N-glycan analysis of human plasma haptoglobin.

Šimunović, Jelena / Gašperšič, Jernej / Černigoj, Urh / Vidič, Jana / Štrancar, Aleš / Novokmet, Mislav / Razdorov, Genadij / Pezer, Marija / Lauc, Gordan / Trbojević-Akmačić, Irena

Biotechnology and bioengineering

2022 Volume 120, Issue 2, Page(s) 491–502

Abstract: Haptoglobin (Hp) is a positive acute phase protein, synthesized in the liver, with four N-glycosylation sites carrying mainly complex type N-glycans. Its glycosylation is altered in different types of diseases but still has not been extensively studied ... ...

Abstract	Haptoglobin (Hp) is a positive acute phase protein, synthesized in the liver, with four N-glycosylation sites carrying mainly complex type N-glycans. Its glycosylation is altered in different types of diseases but still has not been extensively studied mainly due to analytical challenges, especially the lack of a fast, efficient, and robust high-throughput Hp isolation procedure. Here, we describe the development of a high-throughput method for Hp enrichment from human plasma, based on monolithic chromatographic support in immunoaffinity mode and downstream Hp N-glycome analysis by hydrophilic interaction ultrahigh-performance liquid chromatography with fluorescent detection (HILIC-UHPLC-FLR). Chromatographic monolithic supports in a 96-well format enable fast, efficient, and robust Hp enrichment directly from diluted plasma samples. The N-glycome analysis demonstrated that a degree of Hp deglycosylation differs depending on the conditions used for N-glycan release and on the specific glycosylation site, with Asn 241 being the most resistant to deglycosylation under tested conditions. HILIC-UHPLC-FLR analysis enables robust quantification of 28 individual chromatographic peaks, in which N-glycan compositions were determined by UHPLC coupled to electrospray ionization quadrupole time of flight mass spectrometry. The developed analytical approach enables fast evaluation of total Hp N-glycosylation and is applicable in large-scale studies.
MeSH term(s)	Humans ; Haptoglobins ; Chromatography, Liquid ; Glycosylation ; Spectrometry, Mass, Electrospray Ionization ; Polysaccharides/chemistry
Chemical Substances	Haptoglobins ; Polysaccharides
Language	English
Publishing date	2022-11-19
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	280318-5
ISSN	1097-0290 ; 0006-3592
ISSN (online)	1097-0290
ISSN	0006-3592
DOI	10.1002/bit.28280
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Article ; Online: Genetic analysis of 39 erythrocytosis and hereditary hemochromatosis-associated genes in the Slovenian family with idiopathic erythrocytosis.

Kristan, Aleša / Gašperšič, Jernej / Režen, Tadeja / Kunej, Tanja / Količ, Rok / Vuga, Andrej / Fink, Martina / Žula, Špela / Anžej Doma, Saša / Preložnik Zupan, Irena / Pajič, Tadej / Podgornik, Helena / Debeljak, Nataša

Journal of clinical laboratory analysis

2021 Volume 35, Issue 4, Page(s) e23715

Abstract: Background: Erythrocytosis is a condition with an excessive number of erythrocytes, accompanied by an elevated haemoglobin and/or haematocrit value. Congenital erythrocytosis has a diverse genetic background with several genes involved in erythropoiesis. ...

Abstract	Background: Erythrocytosis is a condition with an excessive number of erythrocytes, accompanied by an elevated haemoglobin and/or haematocrit value. Congenital erythrocytosis has a diverse genetic background with several genes involved in erythropoiesis. In clinical practice, nine genes are usually examined, but in approximately 70% of patients, no causative mutation can be identified. In this study, we screened 39 genes, aiming to identify potential disease-driving variants in the family with erythrocytosis of unknown cause. Patients and methods: Two affected family members with elevated haemoglobin and/or haematocrit and negative for acquired causes and one healthy relative from the same family were selected for molecular-genetic analysis of 24 erythrocytosis and 15 hereditary haemochromatosis-associated genes with targeted NGS. The identified variants were further analysed for pathogenicity using various bioinformatic tools and review of the literature. Results: Of the 12 identified variants, two heterozygous variants, the missense variant c.471G>C (NM_022051.2) (p.(Gln157His)) in the EGLN1 gene and the intron variant c.2572-13A>G (NM_004972.3) in the JAK2 gene, were classified as low-frequency variants in European population. None of the two variants were present in a healthy family member. Variant c.2572-13A>G has potential impact on splicing by one prediction tool. Conclusion: For the first time, we included 39 genes in the erythrocytosis clinical panel and identified two potential disease-driving variants in the Slovene family studied. Based on the reported functional in vitro studies combined with our bioinformatics analysis, we suggest further functional analysis of variant in the JAK2 gene and evaluation of a cumulative effect of both variants.
MeSH term(s)	Adult ; Aged ; Base Sequence ; Computational Biology ; Family ; Female ; Gene Frequency/genetics ; Genetic Association Studies ; Genetic Predisposition to Disease ; Hemochromatosis/genetics ; Heterozygote ; Humans ; Inheritance Patterns/genetics ; Male ; Pedigree ; Polycythemia/congenital ; Polycythemia/genetics ; Polymorphism, Single Nucleotide/genetics ; Slovenia
Language	English
Publishing date	2021-02-03
Publishing country	United States
Document type	Journal Article
ZDB-ID	645095-7
ISSN	1098-2825 ; 0887-8013
ISSN (online)	1098-2825
ISSN	0887-8013
DOI	10.1002/jcla.23715
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Article ; Online: Semi-high-throughput isolation and N-glycan analysis of human fibrinogen using monolithic supports bearing monoclonal anti-human fibrinogen antibodies.

Vidic, Urška / Trbojević-Akmačić, Irena / Černigoj, Urh / Albers, Malena / Gašperšič, Jernej / Pučić-Baković, Maja / Vidič, Jana / Štrancar, Aleš / Lauc, Gordan

Electrophoresis

2017

Abstract: Fibrinogen (FIB) is a secretory glycoprotein synthesized by hepatocytes that has a key role in blood clotting. Its glycosylation has not been studied in detail and little is known about the biological variability of FIB N-glycosylation, mainly due to the ...

Abstract	Fibrinogen (FIB) is a secretory glycoprotein synthesized by hepatocytes that has a key role in blood clotting. Its glycosylation has not been studied in detail and little is known about the biological variability of FIB N-glycosylation, mainly due to the lack of fast, simple, and robust approaches to purify FIB from blood plasma samples. In recent years, customised chromatographic monoliths have been used for a variety of biological applications due to their unique characteristics. Here we describe development and optimisation of monolithic supports bearing monoclonal anti-human fibrinogen antibodies in a single column as well as in multi-well plate formats with high FIB specificity and binding capacity for fast immunoaffinity purification of FIB from human blood samples. The developed semi-high-throughput workflow has been successfully applied for FIB immunoaffinity isolation and subsequent ultra performance liquid chromatography N-glycosylation analysis in ten healthy human individuals, demonstrating the potential of monolithic supports in glycomics studies.
Language	English
Publishing date	2017-05-30
Publishing country	Germany
Document type	Journal Article
ZDB-ID	619001-7
ISSN	1522-2683 ; 0173-0835
ISSN (online)	1522-2683
ISSN	0173-0835
DOI	10.1002/elps.201700140
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Article ; Online: Titanium dioxide nanoparticle coating of polymethacrylate-based chromatographic monoliths for phosphopetides enrichment.

Černigoj, Urh / Gašperšič, Jernej / Fichtenbaum, Andreas / Lendero Krajnc, Nika / Vidič, Jana / Mitulović, Goran / Štrancar, Aleš

Analytica chimica acta

2016 Volume 942, Page(s) 146–154

Abstract: Metal oxide affinity chromatography has been one of the approaches for specific enrichment of phosphopeptides from complex samples, based on specific phosphopeptide adsorption forming bidentate chelates between phosphate anions and the surface of a metal ...

Abstract	Metal oxide affinity chromatography has been one of the approaches for specific enrichment of phosphopeptides from complex samples, based on specific phosphopeptide adsorption forming bidentate chelates between phosphate anions and the surface of a metal oxide, such as TiO
Language	English
Publishing date	2016-10-26
Publishing country	Netherlands
Document type	Journal Article
ZDB-ID	1483436-4
ISSN	1873-4324 ; 0003-2670
ISSN (online)	1873-4324
ISSN	0003-2670
DOI	10.1016/j.aca.2016.08.044
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