The Dicer protein is an indispensable player in such fundamental cell pathways as miRNA biogenesis and regulation of protein expression in a cell. Most recently, both germline and somatic mutations in DICER1 have been identified in diverse types of cancers, which suggests Dicer mutations can lead to cancer progression. In addition to well-known hotspot mutations in RNAase III domains, DICER1 is characterized by a wide spectrum of variants in all the functional domains; most are of uncertain significance and unstated clinical effects. Moreover, various new somatic DICER1 mutations continuously appear in cancer genome sequencing. The latest contemporary methods of variant effect prediction utilize machine learning algorithms on bulk data, yielding suboptimal correlation with biological data. Consequently, such analysis should be conducted based on the functional and structural characteristics of each protein, using a well-grounded targeted dataset rather than relying on large amounts of unsupervised data. Domains are the functional and evolutionary units of a protein; the analysis of the whole protein should be based on separate and independent examinations of each domain by their evolutionary reconstruction. Dicer represents a hallmark example of a multidomain protein, and we confirmed the phylogenetic multidomain approach being beneficial for the clinical effect prediction of Dicer variants. Because Dicer was suggested to have a putative role in hematological malignancies, we examined variants of DICER1 occurring outside the well-known hotspots of the RNase III domain in this type of cancer using phylogenetic reconstruction of individual domain history. Examined substitutions might disrupt the Dicer function, which was demonstrated by molecular dynamic simulation, where distinct structural alterations were observed for each mutation. Our approach can be utilized to study other multidomain proteins and to improve clinical effect evaluation.

[This corrects the article DOI: 10.3389/fmolb.2024.1441180.].

The pilot clinical study presented demonstrates the possibility, safety, and effectiveness of oral microbiota transplantation from a healthy donor to a patient with neuroblastoma to prevent chemotherapy-induced oral mucositis. A 6-month-old patient with a diagnosis of retroperitoneal neuroblastoma was treated according to the NB 2004 protocol. Due to the development of severe oral mucositis, it was decided to perform oral microbiota transplantation. During the next 3 chemotherapy cycles and conditioning regimen before autologous hematopoietic cell transplantation (auto-HCT), the patient was repeatedly injected per os with donor saliva from her healthy mother. Oral microbiota transplantation was shown to effectively prevent the development of oral mucositis after chemotherapy, and only grade 1 oral mucositis developed after auto-HCT. In all loci of the oral cavity, there was a decreased abundance of bacteria from the Staphylococcaceae, Micrococcaceae, and Xanthomonadaceae families. Conversely, there was an increase in the relative abundance of Streptococcaceae and certain other bacterial taxa. In conclusion, the transplantation of maternal saliva in this patient prevented severe mucositis and was accompanied by a compositional change of the patient’s oral microbiota. No adverse events due to the transplantation of maternal saliva were noted.

BACKGROUND. Inherited bone marrow failure syndromes (IBMFS) is a heterogenous group of rare genetically determined diseases with variable hematologic and nonhematologic manifestations. The implementation of highly specific methods of genetic diagnosis advanced the understanding of IBMFS and allowed its application also beyond pediatrics. That presupposes an awareness of clinical features and reference points for recognizing IBMFS in adults. AIM. To describe the clinical profile of adult IBMFS patients. MATERIALS & METHODS. This ambispective single-center study enrolled 35 patients (10 women and 25 men) with IBMFS. Patients were aged 18–51 years (median 26 years). The following IBMFS were identified: congenital dyskeratosis (n = 10; 28 %), Diamond-Blackfan anemia (n = 9; 26 %), Fanconi anemia (n = 7; 20 %), GATA2 deficiency (n = 3; 8 %), Shwachman-Diamond syndrome (n = 1; 3 %), GATA2 deficiency (n = 1; 3 %), amegakaryocytic thrombocytopenia (n = 1; 3 %), bone marrow failure syndrome type 3 (n = 1; 3 %), severe congenital neutropenia (n = 1; 3 %), bone marrow failure with SAMD9 mutation (n = 1; 3 %). These diseases were analyzed in terms of hematologic and nonhematologic manifestations as well as main diagnosis stages and factors that contribute to recognizing IBMFS. RESULTS. Monolinear cytopenia, bilinear cytopenia, and pancytopenia were identified at hematologic onset in 18 (52 %), 6 (17 %), and 11 (31 %) patients, respectively. The median age of patients by hematologic onset was 15 years (range 0–43 years), in 14 (40 %) patients cytopenia was newly diagnosed at the age of > 18 years. In 23 (63 %) patients hypocellular bone marrow was reported, 7 (20 %) and 5 (14 %) patients had pure red cell aplasia and multilineage myelodysplasia, respectively. Chromosomal aberrations were identified in 2 patients. Paroxysmal nocturnal hemoglobinuria clone was detected in none of 27 examined patients. In 12 (34 %) patients, the criteria for non-severe aplastic anemia were met. Temporary partial or complete spontaneous hematologic recovery was observed in 6 (17 %) patients. Abnormalities with partial or complete organ dysfunctions were identified in 14 patients, whereas all patients showed minor congenital defects. All 7 Fanconi anemia patients and 9 out of 10 congenital dyskeratosis patients demonstrated organ damage specific to these diseases. Family history predominantly showing malignant neoplasms in relatives was reported in 15 (43 %) patients. Initial hematological examination yielded suspect of IBMFS in 12 (34 %) patients with the median time to diagnosis of 6 months. In 23 (66 %) patients, hematologic defects with cytopenia were erroneously accounted for by various acquired diseases, which led to a delayed correct diagnosis (median 7 years). The key factors in suspecting IBMFS were organ abnormalities and positive family history. The IBMFS diagnosis was verified by the next-generation sequencing (NGS) in 29 (83 %) patients and by other specific methods in 4 (11 %) patients. In 2 patients, the diagnosis was established on the basis of complete clinical criteria alone. CONCLUSION. IBMFS is a matter of current concern and a difficult-to-recognize clinical challenge in adult hematology patients. Differential diagnosis of acquired and congenital bone marrow failure needs to be performed irrespective of patient’s age. A detailed physical examination of patients, family history, and critical analysis of clinical profile and disease course allow for early suspicion of IBMFS. Suspected IBMFS is an indication for referral of patients to specialized centers and performing genetic diagnostics including NGS.

Humanin (HN) is a mitochondrial peptide that is encoded by the MT-RNR2 gene and can be composed of either 21 or 24 amino acids (aa), depending on whether it is expressed in the mitochondria or cytosol, respectively. It has cytoprotective, anti-inflammatory, and antioxidative properties that may involve a systemic normalization of stress-response homeostasis in many cell types. Moreover, HN analogs, such as S14G-humanin, have been synthesized and found to have strong neuroprotective effects. However, we still lack knowledge on the evolution of the MT-RNR2 gene and its cytosol-expressed products in different organisms. Using phylogenetic analysis, we have uncovered the scarcity of nuclear HN sequences in evolution and shown that true mitochondrial HN homologs are not likely to be found in non-vertebrates. Also, analyzing the diversity of HNs we have discovered that S14G substitution is presented among avian HN sequences while 21-aa HN is highly specific to mammals.

Although antagonists were found to bind different bacterial signal transduction receptors, we are still at the early stages of understanding the molecular details by which these molecules exert their inhibitory effects. Here, we provide insight into the structural changes resulting from the binding of an agonist and an antagonist to a sensor protein.

The use of Next-Generation Sequencing (NGS) has proven to be clinically beneficial, but it has also revealed a significant number of variants that we are unable to accurately define and categorize in terms of pathogenicity. These variants are known as variants of uncertain significance (VUS) which are detected en masse in each NGS run. Unlike amino acid substitutions and splice site mutations, common variants in non-coding regions have not been extensively studied and are still mostly classified as VUS. In this paper, a new concept was proposed to identify potentially tolerated variants, including variants in non-coding regions, based on the Genetic Alignment of “Pseudoreads” from Homologs (GAPH) method. We have discovered a total of 5,859,205 variants, the majority of which have never been documented in the largest population database, GnomAD, and only 0.0015% (88 variants) were classified as pathogenic according to the ClinVar database. Overall, the results of this study demonstrate the efficacy of our new method to refine a variant tolerability, many aspects of which could be further adjusted to optimize the results.

Chromosome copy number changes, also known as aneuploidy, is a ubiquitous feature of cancer cell genome.DNA sequencing studies of the tumor cells have shown that the aneuploidy patterns are nonrandom, and specific chromosome gains occur quite frequently, thus, probably, playing an oncogene-like role in tumor development.It is well known that certain copy number changes, such as 1q gain, are of predictive value in many tumor types.It has been recently shown that the 1q gain is associated with increased expression of the MDM4 oncogene located at 1q32 which is responsible for the downregulation of TP53 gene in Fanconi anemia and clonal hematopoiesis development. Case presentationHere we present a rare case of Fanconi anemia in a 31-year-old Syrian male patient at the early stage of MDS/AML transformation.His bone marrow cells showed 1q and 3q gains combined with severely suppressed TP53 gene expression, along with low WT1 and BAALC gene expression. ConclusionSince acquisition of additional 1q/MDM4 copies is common to many solid tumors and oncohematological diseases is considered a reliable marker predictive of cancer progression, this molecular feature needs further indepth study.

Актуальность. Восприимчивость к COVID-19 и тяжесть клинических проявлений зависит как от патогенных свойств SARS-CoV-2, так и от возраста пациента, его физического состояния. Кроме того, у больных обнаружен ряд генных вариантов, влияющих на тяжесть и исход COVID-19. Цель исследования – оценка ассоциаций между рядом генных вариантов, ранее выявленных при пандемии COVID-19, и клиническими параметрами вирусной инфекции у пациентов специализированной пульмонологической клиники. Материал и методы. Обследован 181 пациент от 20 до 85 лет с верифицированной инфекцией SARS-CoV-2. Маркеры воспаления включали определение в крови С-реактивного белка, интерлейкина-6, D-димера фибрина, фибриногена, ферритина. Степень тяжести определяли по совокупности клинических и лабораторных показателей. У 113 (63%) пациентов констатировано тяжелое течение, у 67 (37%) – средняя степень тяжести заболевания. В отделение реанимации и интенсивной терапии были направлены для лечения 97 (53,6%) пациентов. Доля летальных исходов среди данного контингента – 37%. Генотипирование ДНК, выделенной из лейкоцитов крови, проводили с помощью методик аллель-специфической полимеразной цепной реакции. Проводили генотипирование аллельных вариантов следующих генов: OAS1 (rs10774671); ACE1 (rs 4343); VEGF (rs3025039); TLR3 (rs3775291); IL-6 (rs1800795); IL-10 (rs1800896); триаллелльный полиморфизм гена ApoE (rs429358/rs7412). Обработку результатов проводили методами параметрической и непараметрической статистики (STATISTICA 5.0). Результаты. При первичном корреляционном анализе мы обнаружили достоверные корреляции между генотипами 7 генных вариантов и показателями тяжести заболевания только для 3 генов: OAS1, VEGF и ApoE2/3/4. При этом носительство аллеля G гена OAS1 (в том числе, в гетерозиготной форме AG) чаще отмечалось у пациентов, которым требовалась интенсивная терапия, а также при летальных исходах заболевания. Оценка частот генотипов VEGF показала, что генотип СС ассоциирован с меньшей частотой тяжелых клинических форм. Более редкий генотип ТТ не выявлен в группе выживших пациентов (0/84), а среди умерших больных его частота составила 10% (5/67, р = 0,003). Генотип ApoE 3/3 встречался несколько реже в группах пациентов с тяжелым течением заболевания (р = 0,05). Отмечена достоверная корреляция между генотипом GС IL-6 (rs1800795) и уровнями IL-6 в крови на высоте заболевания. Заключение. Полученные данные позволяют рекомендовать дальнейшее изучение OAS1 (rs10774671) и VEGF (rs3025039) в качестве кандидатных маркеров клинического течения и исхода COVID-19. HTML

A number of sequencing studies identified the prognostic impact of somatic mutations in myelodysplastic syndrome (MDS). However the majority of them focused on methylation regulation, apoptosis and proliferation genes. Despite the number of experimental studies published on the role of micro-RNA processing and checkpoint genes in the development of MDS, the clinical data about mutational landscape in these genes is limited. We performed a pilot study which evaluated mutational burden in these genes and their association with common MDS mutations. High prevalence of mutations was observed in the genes studied: 54% had mutations in DICER1, 46% had mutations in LAG3, 20% in CTLA4, 23% in B7-H3, 17% in DROSHA, 14% in PD-1 and 3% in PD-1L. Cluster analysis that included these mutations along with mutations in ASXL1, DNMT3A, EZH2, IDH1, RUNX1, SF3B1, SRSF2, TET2 and TP53 effectively predicted overall survival in the study group (HR 4.2, 95%CI 1.3-13.6, p = 0.016). The study results create the rational for incorporating micro-RNA processing and checkpoint genes in the sequencing panels for MDS and evaluate their role in the multicenter studies.

The analysis of the informativeness of traditional laboratory markers was performed by verification of individuals with the laboratory features of chronic kidney disease (blood creatinine level exceeding the reference range upper limit) among 142,494 people, who underwent laboratory tests during dispenserization and medical examination during 2015–2019. A stable growth of population groups characterized by preclinical hypercreatininaemia was observed, suggesting the increase of chronic kidney disease rate. Meanwhile the frequency of this population group, corresponding to the obtained values, is considered to be a cardiovascular cohort (primarily affected by chronic cardiac failure) in ‘Chronic kidney disease’ clinical recommendation (2019). Our results clearly demonstrate the cardiorenal continuum. Obtained data can be used in the resource support of the health care system calculation.

Untranslated gene regions (UTRs) play an important role in controlling gene expression. 3′-UTRs are primarily targeted by microRNA (miRNA) molecules that form complex gene regulatory networks. Cancer genomes are replete with non-coding mutations, many of which are connected to changes in tumor gene expression that accompany the development of cancer and are associated with resistance to therapy. Therefore, variants that occurred in 3′-UTR under cancer progression should be analysed to predict their phenotypic effect on gene expression, e.g., by evaluating their impact on miRNA target sites. Here, we analyze 3′-UTR variants in DICER1 and DROSHA genes in the context of myelodysplastic syndrome (MDS) development. The key features of this analysis include an assessment of both “canonical” and “non-canonical” types of mRNA-miRNA binding and tissue-specific profiling of miRNA interactions with wild-type and mutated genes. As a result, we obtained a list of DICER1 and DROSHA variants likely altering the miRNA sites and, therefore, potentially leading to the observed tissue-specific gene downregulation. All identified variants have low population frequency consistent with their potential association with pathology progression.

Myelodysplastic syndrome (MDS) refers to a heterogeneous group of closely related clonal hematopoietic disorders, which are characterized by accumulation of somatic mutations. The acquired mutation burden is suggested to define the pathway and consequent phenotype of the pathology. Recent studies have called attention to the role of miRNA biogenesis genes in MDS progression; in particular, the mutational pressure of the DROSHA gene was determined. Therefore, this highlights the importance of studying the impact of all collected missense mutations found within the DROSHA gene in oncohematology that might affect the functionality of the protein. In this study, the selected mutations were extensively examined by computational screening, and the most deleterious were subjected to a further molecular dynamic simulation in order to uncover the molecular mechanism of the structural damage to the protein altering its biological function. The most significant effect was found for variants I625K, L1047S, and H1170D, presumably affecting the endonuclease activity of DROSHA. Such alterations arisen during MDS progression should be taken into consideration as evoking certain clinical traits in the malignifying clonal evolution.

Introduction: There is growing evidence of a connection between tumor clonal profile and its clinical impact. However, there is a lack of a feasible and reliable method for clonal profiling in actual clinical practice. Myelodysplastic syndrome is a clonal hematopoietic stem cell disorder characterized by morphological dysplasia, cytopenia and a high risk of evolution to acute myeloid leukemia. The clinical outcome of myelodysplastic syndrome is greatly heterogeneous; therefore, specific examination of clonal profiles is needed to resolve the prognosis of patients with such complex disorders. Purpose: Development of a pipeline specifically for determining the clonal profiles in patients with myelodysplastic syndrome on the basis of target next-generation sequencing data. Results: The pipeline was developed and evaluated on a set of 35 patients with high-risk myelodysplastic syndrome. It is possible to use the target sequencing data in order to assess the heterogeneity of clonal profiles and characterize their genetic features. This approach allows you to identify the consistency between a specific individual profile and the disease prognosis, which can be critical for the treatment decision. Herein, the characterization and analysis of clonal profiles are presented. Practical relevance: The information about relation patterns between clonal profile characteristics (number of subclones, mutations-per-clone rate) and clinical outcome can be used by doctors in current practice for a more accurate therapy selection depending on the identified individual specificity of the disease.

Osteopetrosis is a group of rare inheritable disorders of the skeleton characterized by increased bone density. The disease is remarkably heterogeneous in clinical presentation and often misdiagnosed. Therefore, genetic testing and molecular pathogenicity analysis are essential for precise diagnosis and new targets for preventive pharmacotherapy. Mutations in the CLCN7 gene give rise to the complete spectrum of osteopetrosis phenotypes and are responsible for about 75% of cases of autosomal dominant osteopetrosis. In this study, we report the identification of a novel variant in the CLCN7 gene in a patient diagnosed with osteopetrosis and provide evidence for its significance (likely deleterious) based on extensive comparative genomics, protein sequence and structure analysis. A set of automated bioinformatics tools used to predict consequences of this variant identified it as deleterious or pathogenic. Structure analysis revealed that the variant is located at the same "hot spot" as the most common CLCN7 mutations causing osteopetrosis. Deep phylogenetic reconstruction showed that not only Leu614Arg, but any non-aliphatic substitutions in this position are evolutionarily intolerant, further supporting the deleterious nature of the variant. The present study provides further evidence that reconstructing a precise evolutionary history of a gene helps in predicting phenotypical consequences of variants of uncertain significance.

Aim. To assess the prognostic value of the mutation of DNA methylation genes, SF3B1, and TP53 in patients with myelodysplastic syndrome (MDS). Materials & Methods. Out of 35 MDS patients included into the trial 2 had multilineage dysplasia, 13 with excess blasts-I, 19 with excess blasts-II, and 1 had 5q-syndrome (criteria WHO 2016). In 30 patients primary MDS was identified, in 5 patients it was detected after prior chemo- or radiotherapy. 25 patients received allogeneic hematopoietic stem cell transplantation (allo-HSCT). According to IPSS-R there were 1 low-risk, 5 intermediate risk, 17 high-risk, and 12 very highrisk patients. Hypomethylating agents were administered to 28 patients. Median age of patients was 49 years (range 18-80 years). Next-generation sequencing was applied for identifying somatic mutations in DNA methylation genes (TET2, IDH1/2, ASXL1, and DNMT3A) as well as in SF3B1, TP53, and RUNX1. Time to progression (TTP) was defined as the time from the initial diagnosis to the date of acute leukemia diagnosis. Allo-HSCT- or antitumor therapy-associated death was considered as competing risk. Results. Methylation gene analysis showed no mutation in 37 % of patients, in 40 % mutation was detected only in one of the genes, in 23 % mutation was identified in > 2 genes. SF3B1 mutations were reported in 23 % and TP53 in 11 % of patients. Median follow-up was 25 months (range 5-116 months). Univariate analysis showed no considerable differences in overall survival depending on mutation status. Median TTP in the group with allo-HSCT was not achieved, in the group without allo-HSCT it was 6 months (p = 0.0001). In patients with no SF3B1 mutation median TTP was 35 months, in patients with this mutation it was not achieved (p = 0.043). With ≥ 2 mutations in methylation genes median TTP was 12 months, in other cases it was not achieved (p = 0.024). In cases of TP53 mutation median TTP was 6 months, in cases without this mutation it was 43 months (p = 0.023). Multivariate analysis confirmed unfavorable prognostic value of TP53 mutation or ≥ 2 mutations in methylation genes in terms of TTP regardless of the drug treatment or allo-HSCT performed (hazard ratio 7.1; 95% confidence interval 2.6-19.6; p = 0.0001). Conclusion. The analysis of molecular markers yields additional data concerning the MDS prognosis. Further research is required to determine the prognostic value of molecular markers in clinical practice which will enable to individualize approaches to MDS treatment.