Micronutrients, genetics and COVID-19.

PURPOSE OF REVIEW: Marked inter-individual differences in the clinical manifestation of coronavirus disease 2019 (COVID-19) has initiated studies in the field of genetics. This review evaluates recent genetic evidence (predominantly in the last 18 months) related to micronutrients (vitamins and trace elements) and COVID-19. RECENT FINDINGS: In patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), altered circulating levels of micronutrients may serve as prognostic markers of disease severity. Mendelian randomization (MR) studies did not find significant effect of variable genetically predicted levels of micronutrients on COVID-19 phenotypes, however, recent clinical studies on COVID-19 point out to vitamin D and zinc supplementation as a nutritional strategy to reduce disease severity and mortality. Recent evidence also points to variants in vitamin D receptor ( VDR ) gene, most notably rs2228570 (FokI) "f" allele and rs7975232 (ApaI) "aa" genotype as poor prognostic markers. SUMMARY: Since several micronutrients were included in the COVID-19 therapy protocols, research in the field of nutrigenetics of micronutrients is in progress. Recent findings from MR studies prioritize genes involved in biological effect, such as the VDR gene, rather than micronutrient status in future research. Emerging evidence on nutrigenetic markers may improve patient stratification and inform nutritional strategies against severe COVID-19.

Genetic predictors of celiac disease, lactose intolerance, and vitamin D function and presence of peptide morphins in urine of children with neurodevelopmental disorders.

Gastrointestinal disturbances, nutritional deficiencies, and food intolerances are frequently observed in children with neurodevelopmental disorders (NDD). To reveal possible association of celiac disease risk variants (HLA-DQ), lactose intolerance associated variant (LCT-13910C>T) as well as variant associated with vitamin D function (VDR FokI) with NDD, polymerase chain reaction-based methodology was used. Additionally, intestinal peptide permeability was estimated in NDD patients and healthy children by measuring the level of peptides in urine using high-performance liquid chromatography. Levels of opioid peptides, casomorphin 8, and gluten exorphin C were significantly elevated in urine samples of NDD patients (P = 0.004 and P = 0.005, respectively), but no association of genetic risk variants for celiac disease and lactose intolerance with NDD was found. Our results indicate that increased intestinal peptide permeability observed in analyzed NDD patients is not associated with genetic predictors of celiac disease or lactose intolerance. We have also found that FF genotype of VDR FokI and lower serum levels of vitamin D (25-OH) showed association with childhood autism (CHA), a subgroup of NDD. We hypothesize that vitamin D might be important for the development of CHA.

Importance of TLR9-IL23-IL17 axis in inflammatory bowel disease development: Gene expression profiling study.

BACKGROUND AND AIMS: Mucosal gene expression have not been fully enlightened in inflammatory bowel disease (IBD). Aim of this study was to define IL23A, IL17A, IL17F and TLR9 expression in different IBD phenotypes. METHODS: Evaluation of mRNA levels was performed in paired non-inflamed and inflamed mucosal biopsies of newly diagnosed 50 Crohn's disease (CD) and 54 ulcerative colitis (UC) patients by quantitative real-time PCR analysis. RESULTS: IL17A and IL17F expression levels were significantly increased in inflamed IBD mucosa. Inflamed CD ileal and UC mucosa showed increased IL23A, while only inflamed CD ileal samples showed increased TLR9 mRNA level. Correlation between analysed mRNAs levels and endoscopic and clinical disease activity were found in UC, but only with clinical activity in CD. CONCLUSION: Both CD and UC presented expression of Th17-associated genes. Nevertheless, expression profiles between different disease forms varies which should be taken into account for future research and therapeutics strategies.

Novel genetic risk variants for pediatric celiac disease.

BACKGROUND: Celiac disease is a complex chronic immune-mediated disorder of the small intestine. Today, the pathobiology of the disease is unclear, perplexing differential diagnosis, patient stratification, and decision-making in the clinic. METHODS: Herein, we adopted a next-generation sequencing approach in a celiac disease trio of Greek descent to identify all genomic variants with the potential of celiac disease predisposition. RESULTS: Analysis revealed six genomic variants of prime interest: SLC9A4 c.1919G>A, KIAA1109 c.2933T>C and c.4268_4269delCCinsTA, HoxB6 c.668C>A, HoxD12 c.418G>A, and NCK2 c.745_746delAAinsG, from which NCK2 c.745_746delAAinsG is novel. Data validation in pediatric celiac disease patients of Greek (n = 109) and Serbian (n = 73) descent and their healthy counterparts (n = 111 and n = 32, respectively) indicated that HoxD12 c.418G>A is more prevalent in celiac disease patients in the Serbian population (P < 0.01), while NCK2 c.745_746delAAinsG is less prevalent in celiac disease patients rather than healthy individuals of Greek descent (P = 0.03). SLC9A4 c.1919G>A and KIAA1109 c.2933T>C and c.4268_4269delCCinsTA were more abundant in patients; nevertheless, they failed to show statistical significance. CONCLUSIONS: The next-generation sequencing-based family genomics approach described herein may serve as a paradigm towards the identification of novel functional variants with the aim of understanding complex disease pathobiology.

Women's Experiences of Childbirth in Serbian Public Healthcare Institutions: a Qualitative Study.

PURPOSE: The aim of the present study was to explore how women, users of public maternity healthcare services in Serbia, experience birth and what the most problematic relational aspects of institutional context associated with negative or even traumatic aspects of birth experiences are, as described by the women themselves. METHOD: An exploratory qualitative study was undertaken using semi-structured interviews with 15 primiparous women aged 26 to 49 who have recently given birth in different public healthcare institutions in Belgrade. The basic framework for the analysis of interview transcripts was the interpretative phenomenological analysis (IPA). RESULTS: Through the analysis, four relatively broad and partly interrelated themes emerged: (a) feelings of isolation and abandonment, (b) lack of communication, (c) lack of a caring relationship, and (d) lack of control and agency. The aspects of institutional environment that were considered particularly distressing in most of the childbirth experiences are related to distant and cold relationship with healthcare providers which adds to the feelings of isolation and abandonment, in addition to the lack of insight into and control over the process of birth that is managed in the hospital context without relying on women's subjective involvement in any relevant way. CONCLUSION: The present study emphasized a supportive and caring relationship with medical practitioners, as well as allowing women to be more involved into their birthing process, as crucial for positive experience of birth, which might have profound and long-lasting psychosocial consequences. Recommendations for policy makers and future research are offered.

Situated technology in reproductive health care: Do we need a new theory of the subject to promote person-centred care?

Going through reproductive experiences (especially pregnancy and childbirth) in contemporary Western societies almost inevitably involves interaction with medical practitioners and various medical technologies in institutional context. This has important consequences for women as embodied subjects. A critical appraisal of these consequences-coming dominantly from feminist scholarship-relied on a problematic theory of both technology and the subject, which are in contemporary approaches no longer considered as given, coherent and well individualized wholes, but as complex constellations that are locally situated and that can only be described empirically. In this study, we will be relying on the developments in phenomenological theory to reconceptualize women as technologically mediated embodied subjects and on the new paradigms in philosophy of technology and STS to reconstruct medical technology as situated-with the aim of reconceptualizing their relationship and exploring different possibilities for the mediating role of medical technology. It will be argued that technologization of female reproductive processes and alienating consequences for women are not necessary or directly interrelated. The role of technology varies from case to case and depends mainly on the nontechnological and relational aspects of institutional context, in which medical practitioners play a decisive role.

Frequencies of EGFR single nucleotide polymorphisms in non-small cell lung cancer patients and healthy individuals in the Republic of Serbia: a preliminary study.

The purpose of this study was to determine the frequencies of EGFR -216G>T, -191C>A, and 181946C>T in Serbian non-small cell lung cancer (NSCLC) patients, as well as to compare it with healthy individuals, in order to assess their potential importance for lung cancer in Serbia. The study involved 56 NSCLC patients and 53 unrelated healthy volunteers, and genotyping was performed on DNA samples obtained from formalin-fixed paraffin-embedded lung tumor tissue and blood, respectively. This was the first time to show genotype frequencies of those single nucleotide polymorphisms for this study group from the territory of the Republic of Serbia. There was very strong evidence of association between age and death due to lung cancer (Pearson chi-square = 43.439, df = 2, p < 0,001), as well as between ever smoking and death due to lung cancer (Pearson chi-square = 31.727, df = 1, p < 0.001). When dominant genetic model (GG vs. GT+TT) was used for -216G>T, we have found significant association (p = 0.012) between -216GG genotype and NSCLC patients within smokers' subgroup. So, carriers of -216GG genotype had higher risk (OR = 4.33, 95 % CI = 1.324-14.179) than noncarriers (GT and TT) for developing non-small cell lung cancer in our patients.

Functional analysis of a novel KLF1 gene promoter variation associated with hereditary persistence of fetal hemoglobin.

Hereditary persistence of fetal hemoglobin (HPFH) is a rare hereditary condition resulting in elevated levels of fetal hemoglobin (HbF) in adults. Typical HPFH is associated with promoter mutations or large deletions affecting the human fetal globin (HBG1 and HBG2) genes, while genetic defects in other genes involved in human erythropoiesis, e.g. KLF1, also result in atypical HPFH. Here, we report the first KLF1 gene promoter mutation (KLF1:g.-148G > A) that is associated with increased HbF level. This mutation was shown to result in drastically reduced CAT reporter gene expression in K562 cells, compared to the wild-type sequence (p = 0.009) and also in reduced KLF1 gene expression in vivo. Furthermore, consistent with in silico analysis, electrophoretic mobility shift analysis showed that the KLF1:g.-148G > A mutation resides in a Sp1 binding site and further that this mutation leads to the ablation of Sp1 binding in vitro. These data suggest that the KLF1:g-148G > A mutation could play a role in increasing HbF levels in adults and further underlines the role of KLF1 as one of the key transcription factors involved in human fetal globin gene switching.

-174G/C interleukin-6 gene promoter polymorphism predicts therapeutic response to etanercept in rheumatoid arthritis.

To examine whether -174G/C interleukin-6 (IL-6) gene polymorphism, previously reported to correlate with IL-6 level, influences response to etanercept therapy in patients with rheumatoid arthritis. Seventy-seven patients with active RA were studied, at baseline and 6- and 12-month follow-up after etanercept therapy. Treatment response was estimated according to the European League Against Rheumatism response criteria. RA patients were genotyped for -174G/C IL-6 gene polymorphism by the PCR-RFLP method, and influence of genotype at this polymorphism to clinical response to etanercept was assessed. After 12 months of treatment, the percentage of responders (patients who had DAS28 improvement >1.2) was significantly increased in patients carrying the IL-6 -174G/G genotype (95.7 %) compared with those with the G/C (75.6 %) or CC (44.4 %) genotype (p = 0.006 by Chi-square test). No significant difference in the mean values of DAS28 improvement was observed between groups with different genotype. RA patients with an IL-6 -174GG genotype respond to etanercept better than patients with GC or CC genotype. This finding, if confirmed in future studies, suggests that the -174G/C IL-6 polymorphism may be a genetic marker of responsiveness to tumor necrosis factor-alpha (TNF-α) blockers in RA.

Genome-Wide Association Study of COVID-19 Outcomes Reveals Novel Host Genetic Risk Loci in the Serbian Population.

Host genetics, an important contributor to the COVID-19 clinical susceptibility and severity, currently is the focus of multiple genome-wide association studies (GWAS) in populations affected by the pandemic. This is the first study from Serbia that performed a GWAS of COVID-19 outcomes to identify genetic risk markers of disease severity. A group of 128 hospitalized COVID-19 patients from the Serbian population was enrolled in the study. We conducted a GWAS comparing (1) patients with pneumonia (n = 80) against patients without pneumonia (n = 48), and (2) severe (n = 34) against mild disease (n = 48) patients, using a genotyping array followed by imputation of missing genotypes. We have detected a significant signal associated with COVID-19 related pneumonia at locus 13q21.33, with a peak residing upstream of the gene KLHL1 (p = 1.91 × 10-8). Our study also replicated a previously reported COVID-19 risk locus at 3p21.31, identifying lead variants in SACM1L and LZTFL1 genes suggestively associated with pneumonia (p = 7.54 × 10-6) and severe COVID-19 (p = 6.88 × 10-7), respectively. Suggestive association with COVID-19 pneumonia has also been observed at chromosomes 5p15.33 (IRX, NDUFS6, MRPL36, p = 2.81 × 10-6), 5q11.2 (ESM1, p = 6.59 × 10-6), and 9p23 (TYRP1, LURAP1L, p = 8.69 × 10-6). The genes located in or near the risk loci are expressed in neural or lung tissues, and have been previously associated with respiratory diseases such as asthma and COVID-19 or reported as differentially expressed in COVID-19 gene expression profiling studies. Our results revealed novel risk loci for pneumonia and severe COVID-19 disease which could contribute to a better understanding of the COVID-19 host genetics in different populations.

Genetic Aspects of Micronutrients Important for Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD), Crohn's disease (CD) and ulcerative colitis (UC) are complex diseases whose etiology is associated with genetic and environmental risk factors, among which are diet and gut microbiota. To date, IBD is an incurable disease and the main goal of its treatment is to reduce symptoms, prevent complications, and improve nutritional status and the quality of life. Patients with IBD usually suffer from nutritional deficiency with imbalances of specific micronutrient levels that contribute to the further deterioration of the disease. Therefore, along with medications usually used for IBD treatment, therapeutic strategies also include the supplementation of micronutrients such as vitamin D, folic acid, iron, and zinc. Micronutrient supplementation tailored according to individual needs could help patients to maintain overall health, avoid the triggering of symptoms, and support remission. The identification of individuals' genotypes associated with the absorption, transport and metabolism of micronutrients can modify future clinical practice in IBD and enable individualized treatment. This review discusses the personalized approach with respect to genetics related to micronutrients commonly used in inflammatory bowel disease treatment.

Machine Learning Modeling from Omics Data as Prospective Tool for Improvement of Inflammatory Bowel Disease Diagnosis and Clinical Classifications.

Research of inflammatory bowel disease (IBD) has identified numerous molecular players involved in the disease development. Even so, the understanding of IBD is incomplete, while disease treatment is still far from the precision medicine. Reliable diagnostic and prognostic biomarkers in IBD are limited which may reduce efficient therapeutic outcomes. High-throughput technologies and artificial intelligence emerged as powerful tools in search of unrevealed molecular patterns that could give important insights into IBD pathogenesis and help to address unmet clinical needs. Machine learning, a subtype of artificial intelligence, uses complex mathematical algorithms to learn from existing data in order to predict future outcomes. The scientific community has been increasingly employing machine learning for the prediction of IBD outcomes from comprehensive patient data-clinical records, genomic, transcriptomic, proteomic, metagenomic, and other IBD relevant omics data. This review aims to present fundamental principles behind machine learning modeling and its current application in IBD research with the focus on studies that explored genomic and transcriptomic data. We described different strategies used for dealing with omics data and outlined the best-performing methods. Before being translated into clinical settings, the developed machine learning models should be tested in independent prospective studies as well as randomized controlled trials.

Association of Vitamin D, Zinc and Selenium Related Genetic Variants With COVID-19 Disease Severity.

Background: COVID-19 pandemic has proved to be an unrelenting health threat for more than a year now. The emerging amount of data indicates that vitamin D, zinc and selenium could be important for clinical presentation of COVID-19. Here, we investigated association of genetic variants related to the altered level and bioavailability of vitamin D, zinc and selenium with clinical severity of COVID-19. Methods: We analyzed variants in genes significant for the status of vitamin D (DHCR7/NADSYN1 rs12785878, GC rs2282679, CYP2R1 rs10741657, and VDR rs2228570), zinc (PPCDC rs2120019) and selenium (DMGDH rs17823744) in 120 Serbian adult and pediatric COVID-19 patients using allelic discrimination. Furthermore, we carried out comparative population genetic analysis among European and other worldwide populations to investigate variation in allelic frequencies of selected variants. Results: Study showed that DHCR7/NADSYN rs12785878 and CYP2R1 rs10741657 variants were associated with severe COVID-19 in adults (p = 0.03, p = 0.017, respectively); carriers of DHCR7/NADSYN TG+GG and CYP2R1 GG genotypes had 0.21 and 5.9 the odds for developing severe disease, OR 0.21 (0.05-0.9) and OR 5.9 (1.4-25.2), respectively. There were no associations between selected genetic variants and disease severity in pediatric patients. Comparative population genetic analysis revealed that Serbian population had the lowest frequency of CYP2R1 rs10741657 G allele compared to other non-Finish Europeans (0.58 compared to 0.69 and 0.66 in Spanish and Italian population, respectively), suggesting that other populations should also investigate the relationship of CYP2R1 variant and the COVID-19 disease course. Conclusion: The results of the study indicated that vitamin D related genetic variants were implicated in severe COVID-19 in adults. This could direct prevention strategies based on population specific nutrigenetic profiles.

The FKBP5 genotype and childhood trauma effects on FKBP5 DNA methylation in patients with psychosis, their unaffected siblings, and healthy controls.

Hypothalamic-pituitary-adrenal (HPA) axis activity mediates the relationship between childhood trauma (CT) and psychosis. The FKBP5 gene, one of the key regulators of HPA axis activity after stress exposure, has been found associated with psychosis. Allele-specific and CT related FKBP5 demethylation in intron 7 was revealed in different psychiatric disorders. However, no studies have investigated FKBP5 methylation in subjects with different genetic liability for psychosis. A total of 144 participants were included in the study: 48 patients with psychotic disorders, 50 unaffected siblings, and 46 healthy controls. CT was assessed by Childhood Trauma Questionnaire. The FKBP5 rs1360780 was genotyped and FKBP5 methylation analyses were performed using bisulfite conversion followed by Sanger sequencing at three CpG sites in intron 7. Mixed linear model was used to assess group differences depending on rs1360780 T allele and CT. Results showed a significant T allele-dependent decrease of FKBP5 methylation in patients compared to unaffected siblings and controls. Effect of interaction between T allele and CT exposure on FKBP5 demethylation was found in controls. No effect of both risk factors (T allele and CT) on FKBP5 methylation level was found in unaffected siblings. We confirmed previous evidence of the association between the FKBP5 rs1360780 T allele, CT, and decreased FKBP5 methylation in intron 7. Allele-specific FKBP5 demethylation found in patients could shed a light on altered HPA axis activity in a subgroup of patients related to stress-induced psychosis. FKBP5 methylation and potential protective mechanisms in unaffected siblings after trauma exposure require further investigation.

Thalassemia syndromes in Serbia: an update.

Thalassemia syndromes constitute a group of genetic disorders, widespread throughout the world. The present study contains data on thalassemia syndromes and their chromosomal environment obtained in Serbia over a period of 10 years. Ten different ß-thalassemia (ß-thal) mutations and two hemoglobin (Hb) variants were detected in 127 members of 68 families. Hb Lepore-Boston-Washington (Lepore-BW) (δ87Gln-ß-IVS-II-8), a thalassemic Hb variant, was shown to be the most common cause of thalassemia in Serbia. Haplotype analyses of the ß-globin gene clusters of healthy individuals as well as of individuals affected with ß-thal showed that haplotype I was the most frequent haplotype in the Serbian population, followed by haplotypes II and IX. Two novel haplotypes were detected. Haplotype analyses showed the association between certain haplotypes and the most common thalassemic mutations. Results presented in this paper will update the Serbian national mutation database and contribute to a better understanding of genographic history of South European and Balkan populations.

Functional prediction and comparative population analysis of variants in genes for proteases and innate immunity related to SARS-CoV-2 infection.

New coronavirus SARS-CoV-2 is capable to infect humans and cause a novel disease COVID-19. Aiming to understand a host genetic component of COVID-19, we focused on variants in genes encoding proteases and genes involved in innate immunity that could be important for susceptibility and resistance to SARS-CoV-2 infection. Analysis of sequence data of coding regions of FURIN, PLG, PRSS1, TMPRSS11a, MBL2 and OAS1 genes in 143 unrelated individuals from Serbian population identified 22 variants with potential functional effect. In silico analyses (PolyPhen-2, SIFT, MutPred2 and Swiss-Pdb Viewer) predicted that 10 variants could impact the structure and/or function of proteins. These protein-altering variants (p.Gly146Ser in FURIN; p.Arg261His and p.Ala494Val in PLG; p.Asn54Lys in PRSS1; p.Arg52Cys, p.Gly54Asp and p.Gly57Glu in MBL2; p.Arg47Gln, p.Ile99Val and p.Arg130His in OAS1) may have predictive value for inter-individual differences in the response to the SARS-CoV-2 infection. Next, we performed comparative population analysis for the same variants using extracted data from the 1000 Genomes project. Population genetic variability was assessed using delta MAF and Fst statistics. Our study pointed to 7 variants in PLG, TMPRSS11a, MBL2 and OAS1 genes with noticeable divergence in allelic frequencies between populations worldwide. Three of them, all in MBL2 gene, were predicted to be damaging, making them the most promising population-specific markers related to SARS-CoV-2 infection. Comparing allelic frequencies between Serbian and other populations, we found that the highest level of genetic divergence related to selected loci was observed with African, followed by East Asian, Central and South American and South Asian populations. When compared with European populations, the highest divergence was observed with Italian population. In conclusion, we identified 4 variants in genes encoding proteases (FURIN, PLG and PRSS1) and 6 in genes involved in the innate immunity (MBL2 and OAS1) that might be relevant for the host response to SARS-CoV-2 infection.

Clinical Application of Thiopurine Pharmacogenomics in Pediatrics.

BACKGROUND: Thiopurine drugs are used for the treatment of pediatric diseases. Inter-individual differences in the metabolism of these drugs greatly influence the risk of thiopurine induced toxicity and therapy failure. These differences are the consequence of genomic, epigenomic and transcriptomic variability among patients. Pharmacogenomics aims to individualize therapy according to the specific genetic signature of a patient. Treatment protocols based on thiopurine drugs have already been improved by applying pharmacogenomics in pediatric clinical practice. OBJECTIVE: The aim of this review was to summarize the application of thiopurine pharmacogenomics in pediatric patients suffering from acute leukemias, different types of autoimmune and inflammatory diseases, as well as in posttransplant care. METHODS: We searched PubMed/Medline database to identify thiopurine pharmacogenomic markers clinically relevant in pediatric diseases. RESULTS: TPMT and NUDT15 pharmacogenomic testing is done in pediatric care, contributing to the reduction of thiopurine induced toxicity. Data on numerous novel potential pharmacogenomic markers relevant for optimization of thiopurine treatment are still controversial (ITPA, ABCC4, NT5C2, PRPS1, GSTM1, FTO gene variants). Majority of evidences regarding thiopurine pharmacogenomics in pediatrics have been acquired by studying acute lymphoblastic leukemia and inflammatory bowel disease. For other pediatric diseases, namely acute myeloid leukemia, non-Hodgkin lymphoma, juvenile idiopathic arthritis, atopic dermatitis, juvenile autoimmune hepatitis and renal allograft transplantation, data are still scarce. CONCLUSION: Thiopurine pharmacogenomics has shown to be one of the best examples of successful application of pharmacogenomics in pediatrics.

Pharmacogenomics landscape of COVID-19 therapy response in Serbian population and comparison with worldwide populations.

BACKGROUND: Since there are no certified therapeutics to treat COVID-19 patients, drug repurposing became important. With lack of time to test individual pharmacogenomics markers, population pharmacogenomics could be helpful in predicting a higher risk of developing adverse reactions and treatment failure in COVID-19 patients. Aim of our study was to identify pharmacogenes and pharmacogenomics markers associated with drugs recommended for COVID-19 treatment, chloroquine/hydroxychloroquine, azithromycin, lopinavir and ritonavir, in population of Serbia and other world populations. METHODS: Genotype information of 143 individuals of Serbian origin was extracted from database previously obtained using TruSight One Gene Panel (Illumina). Genotype data of individuals from different world populations were extracted from the 1000 Genome Project. Fisher's exact test was used for comparison of allele frequencies. RESULTS: We have identified 11 potential pharmacogenomics markers in 7 pharmacogenes relevant for COVID-19 treatment. Based on high alternative allele frequencies in population and the functional effect of the variants, ABCB1 rs1045642 and rs2032582 could be relevant for reduced clearance of azithromycin, lopinavir and ritonavir drugs and UGT1A7 rs17868323 for hyperbilirubinemia in ritonavir treated COVID-19 patients in Serbian population. SLCO1B1 rs4149056 is a potential marker of lopinavir response, especially in Italian population. Our results confirmed that pharmacogenomics profile of African population is different from the rest of the world. CONCLUSIONS: Considering population specific pharmacogenomics landscape, preemptive testing for pharmacogenes relevant for drugs used in COVID-19 treatment could contribute to better understanding of the inconsistency in therapy response and could be applied to improve the outcome of the COVID-19 patients.

Pharmacogenomic Markers of Methotrexate Response in the Consolidation Phase of Pediatric Acute Lymphoblastic Leukemia Treatment.

Methotrexate (MTX) is one of the staples of pediatric acute lymphoblastic leukemia (ALL) treatment. MTX targets the folate metabolic pathway (FMP). Abnormal function of the enzymes in FMP, due to genetic aberrations, leads to adverse drug reactions. The aim of this study was to investigate variants in pharmacogenes involved in FMP and their association with MTX pharmacokinetics (MTX elimination profile) and toxicity in the consolidation therapy phase of pediatric ALL patients. Eleven variants in the thymidylate synthetase (TYMS), methylenetetrahydrofolate reductase (MTHFR), dihydrofolate reductase (DHFR), SLC19A1 and SLCO1B genes were analyzed in 148 patients, using PCR- and sequencing-based methodology. For the Serbian and European control groups, data on allele frequency distribution were extracted from in-house and public databases. Our results show that the A allele of SLC19A1 c.80 variant contributes to slow MTX elimination. Additionally, the AA genotype of the same variant is a predictor of MTX-related hepatotoxicity. Patients homozygous for TYMS 6bp deletion were more likely to experience gastrointestinal toxicity. No allele frequency dissimilarity was found for the analyzed variants between Serbian and European populations. Statistical modelling did not show a joint effect of analyzed variants. Our results indicate that SLC19A1 c.80 variant and TYMS 6bp deletion are the most promising pharmacogenomic markers of MTX response in pediatric ALL patients.

Exploring inflammatory and apoptotic signatures in distinct Crohn's disease phenotypes: Way towards molecular stratification of patients and targeted therapy.

BACKGROUND: Crohn's disease (CD) is chronic inflammatory bowel disease with different phenotypic characteristics influencing disease prognosis and therapeutic strategies. The aim of this pilot study was to analyze selected inflammatory and apoptotic markers in non-inflamed and inflamed samples of ileal mucosa of non-stricturing/non-penetrating (NS/NP) and stricturing (S) CD mucosal phenotypes in order to characterize their distinct profiles. METHODS: From twenty CD patients (9 NS/NP, 11 S) paired non-inflamed and inflamed ileal biopsies were collected and used for analysis of cytokine (TNF and IL6) and apoptotic (Bcl2, Bax, Fas and FasL) genes' expression levels by real-time PCR, while NFκB transcriptional potency was assessed by electromobility gel shift assay. RESULTS: Our results demonstrated significant upregulation of TNF and IL6 in inflamed area of both NS/NP (p = 0.03, p = 0.01) and S phenotypes (p = 0.04, p = 0.04), respectively. However, TNF increase was more prominent in NS/NP compared to S inflamed mucosa (p = 0.02). Also, level of proapoptotic Bax was significantly higher in NS/NP compared to S inflamed mucosa (p = 0.01). Opposing transcription potency of NFκB has been detected between two phenotypes: being decreased in NS/NP (p = 0.07) and increased in S (p = 0.1) inflamed compared to non-inflamed mucosa, demonstrating trend towards statistical significance. CONCLUSIONS: We found that two distinct CD phenotypes have specific molecular signatures. Obtained results could direct improvement of current and development of new therapeutic strategies based on more specific molecular stratification of CD patients.