Quantification of cytosine modifications in the aged mouse brain.

Quantifying cytosine modifications in various brain regions provides important insights into the gene expression regulation and pathophysiology of neuropsychiatric disorders. In this study, we quantified 5-methylcytosine (5-mC), 5-hydroxymethylation (5-hmC), and 5-formylcytosine (5-fC) levels in five brain regions (the frontal lobe, cerebral cortical region without frontal lobe, hippocampus, basal ganglia, and the cerebellum) and the heart at three developmental periods (12, 48, and 101 weeks). We observed significant regional variations in cytosine modification. Notably, regional variations were generally maintained throughout development, suggesting that epigenetic regulation is unique to each brain region and remains relatively stable with age. The 5-mC and 5-hmC levels were positively correlated, although the extent of the correlations seemed to differ in different brain regions. On the contrary, 5-fC levels did not correlate with 5-mC or 5-hmC levels. Additionally, we observed an age-dependent decrease in 5-fC levels in the basal ganglia, suggesting a unique epigenetic regulation mechanism. Further high-resolution studies using animal models of neuropsychiatric disorders as well as postmortem brain evaluation are warranted.

Structure and Function of TET Enzymes.

Mammalian DNA methylation mainly occurs at the carbon-C5 position of cytosine (5mC). TET enzymes were discovered to successively oxidize 5mC to 5-hydromethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). Ten-eleven translocation (TET) enzymes and oxidized 5mC derivatives play important roles in various biological and pathological processes, including regulation of DNA demethylation, gene transcription, embryonic development, and oncogenesis. In this chapter, we will discuss the discovery of TET-mediated 5mC oxidation and the structure, function, and regulation of TET enzymes. We start with brief descriptions of the mechanisms of TET-mediated 5mC oxidation and TET-dependent DNA demethylation. We then discuss the TET-mediated epigenetic reprogramming in pluripotency maintenance and embryogenesis, as well as in tumorigenesis and neural system. We further describe the structural basis for substrate recognition and preference in TET-mediated 5mC oxidation. Finally, we summarize the chemical molecules and interacting proteins that regulate TET's activity.

A soluble DR5-Fc chimeric protein attenuates inflammatory responses induced by coronavirus MHV-A59 and SARS-CoV-2.

Mortality in coronavirus disease 2019 (COVID-19) patients has been linked to the presence of a "cytokine storm" induced by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, which involves elevated levels of circulating cytokines and immune-cell hyperactivation. Targeting cytokines during the management of COVID-19 patients has the potential to improve survival rates and reduce mortality. Although cytokine blockers and immune-host modulators are currently being tested in severely ill COVID-19 patients to cope with the overwhelming systemic inflammation, there is not too many successful cases, thus finding new cytokine blockers to attenuate the cytokine storm syndrome is meaningful. In this paper, we significantly attenuated the inflammatory responses induced by mouse hepatitis viruses A59 and SARS-CoV-2 through a soluble DR5-Fc (sDR5-Fc) chimeric protein that blocked the TNF-related apoptosis-inducing ligand-death receptor 5 (TRAIL-DR5) interaction. Our findings indicates that blocking the TRAIL-DR5 pathway through the sDR5-Fc chimeric protein is a promising strategy to treat COVID-19 severe patients requiring intensive care unit  admission or with chronic metabolic diseases.

Distribution and regulatory roles of oxidized 5-methylcytosines in DNA and RNA of the basidiomycete fungi Laccaria bicolor and Coprinopsis cinerea.

The formation of three oxidative DNA 5-methylcytosine (5mC) modifications (oxi-mCs)-5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC)-by the TET/JBP family of dioxygenases prompted intensive studies of their functional roles in mammalian cells. However, the functional interplay of these less abundant modified nucleotides in other eukaryotic lineages remains poorly understood. We carried out a systematic study of the content and distribution of oxi-mCs in the DNA and RNA of the basidiomycetes Laccaria bicolor and Coprinopsis cinerea, which are established models to study DNA methylation and developmental and symbiotic processes. Quantitative liquid chromatography-tandem mass spectrometry revealed persistent but uneven occurrences of 5hmC, 5fC and 5caC in the DNA and RNA of the two organisms, which could be upregulated by vitamin C. 5caC in RNA (5carC) was predominantly found in non-ribosomal RNA, which potentially includes non-coding, messenger and small RNA species. Genome-wide mapping of 5hmC and 5fC using the single CG analysis techniques hmTOP-seq and foTOP-seq pointed at involvement of oxi-mCs in the regulation of gene expression and silencing of transposable elements. The implicated diverse roles of 5mC and oxi-mCs in the two fungi highlight the epigenetic importance of the latter modifications, which are often neglected in standard whole-genome bisulfite analyses.

Humoral immunity in hepatitis B virus infection: Rehabilitating the B in HBV.

Insights into the immunopathogenesis of chronic HBV infections are fundamental in the quest for novel treatment approaches aimed at a functional cure. While much is known about the ineffective HBV-specific T-cell responses that characterise persistent HBV replication, B cells have been left largely understudied. However, an important role for humoral immunity during the natural history of HBV infections, as well as after functional cure, has been inadvertently revealed by the occurrence of HBV flares following B cell-depleting treatments. Herein, we review our current understanding of the role of the humoral immune response in chronic HBV, both at the level of HBV-specific antibody production and at the phenotypic and broader functional level of B cells. The recent development of fluorescently labelled HBV proteins has given us unprecedented insights into the phenotype and function of HBsAg- and HBcAg-specific B cells. This should fuel novel research into the mechanisms behind dysfunctional HBsAg-specific and fluctuating, possibly pathogenic, HBcAg-specific B-cell responses in chronic HBV. Finally, novel immunomodulatory treatments that partly target B cells are currently in clinical development, but a detailed assessment of their impact on HBV-specific B-cell responses is lacking. We plead for a rehabilitation of B-cell studies related to both the natural history of HBV and treatment development programmes.

Affibody-conjugated 5-fluorouracil prodrug system preferentially targets and inhibits HER2-expressing cancer cells.

Overexpression of HER2 is associated with cancer phenotypes, such as proliferation, survival, metastasis and angiogenesis, and has been validated as a therapeutic target. However, only a portion of patients benefited from anti-HER2 treatments, and many would develop resistance. A more effective HER2 targeted therapeutics is needed. Here, we adopted a prodrug system that uses 5-fluorocytosine (5-FC) and a HER2-targeting scaffold protein, ZHER2:2891, fused with yeast cytosine deaminase (Fcy) to target HER2-overexpressing cancer cells and to convert 5-FC to a significantly more toxic chemotherapeutic, 5-fluorouracil (5-FU). We cloned the coding gene of ZHER2:2891 and fused with those of ABD (albumin-binding domain) and Fcy. The purified ZHER2:2891-ABD-Fcy fusion protein specifically binds to HER2 with a Kd value of 1.6 nM ZHER2:2891-ABD-Fcy binds to MDA-MB-468, SKOV-3, BT474, and MC38-HER2 cells, which overexpress HER2, whereas with a lower affinity to HER2 non-expresser, MC38. Correspondingly, the viability of HER2-expressing cells was suppressed by relative low concentrations of ZHER2:2891-ABD-Fcy in the presence of 5-FC, and the IC50 values of ZHER2:2891-ABD-Fcy for HER2 high-expresser cells were approximately 10-1000 fold lower than those of non-HER2-targeting Fcy, and ABD-Fcy. This novel prodrug system, ZHER2:2891-ABD-Fcy/5-FC, might become a promising addition to the existing class of therapeutics specifically target HER2-expressing cancers.

New perspective into mesenchymal stem cells: Molecular mechanisms regulating osteosarcoma.

Mesenchymal stem cells (MSCs) are multipotent stem cells with significant potential for regenerative medicine. The tumorigenesis of osteosarcoma is an intricate system and MSCs act as an indispensable part of this, interacting with the tumor microenvironment (TME) during the process. MSCs link to cells by acting on each component in the TME via autocrine or paracrine extracellular vesicles for cellular communication. Because of their unique characteristics, MSCs can be modified and processed into good biological carriers, loaded with drugs, and transfected with anticancer genes for the targeted treatment of osteosarcoma. Previous high-quality reviews have described the biological characteristics of MSCs; this review will discuss the effects of MSCs on the components of the TME and cellular communication and the prospects for clinical applications of MSCs.

Quantitative evaluation and comparison of two prodrug-activating suicide gene therapies on oral squamous cell carcinoma.

Prodrug-activating suicide gene therapy (PA suicide gene therapy for short) for cancer is to introduce cancer cells with suicide genes. The enzyme encoded by suicide gene is not toxic but is able to kill cancer cells by converting a non-toxic prodrug into a toxic compound. This approach is a promising cancer gene therapy that could reduce non-specific toxicity to normal tissue. However, there is no quantitative method to evaluate efficacy of suicide gene therapy in preclinical study. The aim of this study is to develop a new method to quantitatively evaluate and compare prodrug-activating suicide gene therapies. This study was carried out on an oral squamous cell carcinoma (OSCC) cell line CAL-27. Suicide genes were integrated into ROSA26 locus of CAL-27 by CRISPR-Cas9. CAL-27 cell lines stably expressing herpes simplex virus-thymidine kinase (TK) or yeast cytosine deaminase (CD) were used to evaluate and compare PA suicide gene therapies. The efficacies of PA suicide gene therapies were quantitatively evaluated from three aspects: effective prodrug concentration, prodrug treatment time, and bystander effect. This method also could be used for different types of suicide gene therapies and different types of cancer. When the prodrug concentration, treatment time, and rate of suicide gene-positive cells (related to bystander effect) are fixed, anti-cancer effects could be quantitatively measured. This information is important for suicide gene therapy preclinical development.

Anti-Tumor Effects of MAPK-Dependent Tumor-Selective Oncolytic Vaccinia Virus Armed with CD/UPRT against Pancreatic Ductal Adenocarcinoma in Mice.

Engineered vaccinia virus serves as an oncolytic virus for cancer virotherapy. We evaluated the oncolytic characteristics of VGF- and O1-deleted recombinant mitogen-activated protein kinase (MAPK)-dependent vaccinia virus (MDRVV). We found that compared with viruses with the deletion of either gene alone, MDRVV is more attenuated in normal cells and can replicate in cancer cells that exhibit constitutive ERK1/2 activation in the MAPK pathway. We armed MDRVV with a bifunctional fusion gene encoding cytosine deaminase and uracil phosphoribosyltransferase (CD/UPRT), which converts 5-fluorocytosine (5-FC) into chemotherapeutic agents, and evaluated its oncolytic activity alone or in combination with 5-FC in human pancreatic cancer cell lines, tumor mouse models of peritoneal dissemination and liver metastasis, and ex vivo-infected live pancreatic cancer patient-derived tissues. CD/UPRT-armed MDRVV alone could efficiently eliminate pancreatic cancers, and its antitumor effects were partially enhanced in combination with 5-FC in vitro and in vivo. Moreover, the replication of MDRVV was detected in tumor cells of patient-derived, surgically resected tissues, which showed enlarged nuclei and high expression of pERK1/2 and Ki-67, and not in stromal cells. Our findings suggest that systemic injections of CD/UPRT-armed MDRVV alone or in combination with 5-FC are promising therapeutic strategies for pancreatic ductal adenocarcinoma.

Functionally distinct roles for TET-oxidized 5-methylcytosine bases in somatic reprogramming to pluripotency.

Active DNA demethylation via ten-eleven translocation (TET) family enzymes is essential for epigenetic reprogramming in cell state transitions. TET enzymes catalyze up to three successive oxidations of 5-methylcytosine (5mC), generating 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), or 5-carboxycytosine (5caC). Although these bases are known to contribute to distinct demethylation pathways, the lack of tools to uncouple these sequential oxidative events has constrained our mechanistic understanding of the role of TETs in chromatin reprogramming. Here, we describe the first application of biochemically engineered TET mutants that unlink 5mC oxidation steps, examining their effects on somatic cell reprogramming. We show that only TET enzymes proficient for oxidation to 5fC/5caC can rescue the reprogramming potential of Tet2-deficient mouse embryonic fibroblasts. This effect correlated with rapid DNA demethylation at reprogramming enhancers and increased chromatin accessibility later in reprogramming. These experiments demonstrate that DNA demethylation through 5fC/5caC has roles distinct from 5hmC in somatic reprogramming to pluripotency.

Immunochemical Detection of Modified Cytosine Species in Mammalian Preimplantation Embryos.

DNA methylation undergoes dynamic changes at the genome-wide scale during the early steps of mammalian embryo development. Immunochemical detection of 5-methylcytosine (5mC) in the zygote has led to the discovery that a global loss of DNA methylation takes place soon after fertilization, occurring rapidly in the paternal pronucleus. Using the same method employed above, which detects modified bases in the denatured single stranded DNA, we showed that this active DNA "demethylation" in the paternal pronucleus involves oxidation of 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxycytosine (5caC) by the TET3 enzyme. By immunostaining of genetically altered zygotes we revealed that the maternal pronucleus is protected from TET3-mediated oxidation by histone H3K9 methyltransferase enzymes, EHMT2 and SETDB1. The same assays are also applicable for visualizing the temporal and spatial distribution of the modified cytosine residues in preimplantation embryos. Here, we provide a detailed protocol for detecting 5mC, 5hmC, 5fC, and 5caC in mouse zygotes and preimplantation-stage embryos using antibodies raised against modified cytosine species.

Recent progress in the research of suicide gene therapy for malignant glioma.

Malignant glioma, which is characterized by diffuse infiltration into the normal brain parenchyma, is the most aggressive primary brain tumor with dismal prognosis. Over the past 40 years, the median survival has only slightly improved. Therefore, new therapeutic modalities must be developed. In the 1990s, suicide gene therapy began attracting attention for the treatment of malignant glioma. Some clinical trials used a viral vector for suicide gene transduction; however, it was found that viral vectors cannot cover the large invaded area of glioma cells. Interest in this therapy was recently revived because some types of stem cells possess a tumor-tropic migratory capacity, which can be used as cellular delivery vehicles. Immortalized, clonal neural stem cell (NSC) line has been used for patients with recurrent high-grade glioma, which showed safety and efficacy. Embryonic and induced pluripotent stem cells may be considered as sources of NSC because NSC is difficult to harvest, and ethical issues have been raised. Mesenchymal stem cells are alternative candidates for cellular vehicle and are easily harvested from the bone marrow. In addition, a new type of nonlytic, amphotropic retroviral replicating vector encoding suicide gene has shown efficacy in patients with recurrent high-grade glioma in a clinical trial. This replicating viral capacity is another possible candidate as delivery vehicle to tackle gliomas. Herein, we review the concept of suicide gene therapy, as well as recent progress in preclinical and clinical studies in this field.

Epigenetics in hepatocellular carcinoma development and therapy: The tip of the iceberg.

Hepatocellular carcinoma (HCC) is a deadly tumour whose causative agents are generally well known, but whose pathogenesis remains poorly understood. Nevertheless, key genetic alterations are emerging from a heterogeneous molecular landscape, providing information on the tumorigenic process from initiation to progression. Among these molecular alterations, those that affect epigenetic processes are increasingly recognised as contributing to carcinogenesis from preneoplastic stages. The epigenetic machinery regulates gene expression through intertwined and partially characterised circuits involving chromatin remodelers, covalent DNA and histone modifications, and dedicated proteins reading these modifications. In this review, we summarise recent findings on HCC epigenetics, focusing mainly on changes in DNA and histone modifications and their carcinogenic implications. We also discuss the potential drugs that target epigenetic mechanisms for HCC treatment, either alone or in combination with current therapies, including immunotherapies.

Inflammatory Mediators in Glioma Microenvironment Play a Dual Role in Gliomagenesis and Mesenchymal Stem Cell Homing: Implication for Cellular Therapy.

Glioblastoma is the most aggressive malignant primary brain tumor, with a dismal prognosis and a devastating overall survival. Despite aggressive surgical resection and adjuvant treatment, average survival remains approximately 14.6 months. The brain tumor microenvironment is heterogeneous, comprising multiple populations of tumor, stromal, and immune cells. Tumor cells evade the immune system by suppressing several immune functions to enable survival. Gliomas release immunosuppressive and tumor-supportive soluble factors into the microenvironment, leading to accelerated cancer proliferation, invasion, and immune escape. Mesenchymal stem cells (MSCs) isolated from bone marrow, adipose tissue, or umbilical cord are a promising tool for cell-based therapies. One crucial mechanism mediating the therapeutic outcomes often seen in MSC application is their tropism to sites of injury. Furthermore, MSCs interact with host immune cells to regulate the inflammatory response, and data points to the possibility of using MSCs to achieve immunomodulation in solid tumors. Interleukin 1ß, interleukin 6, tumor necrosis factor α, transforming growth factor ß, and stromal cell-derived factor 1 are notably up-regulated in glioblastoma and dually promote immune and MSC trafficking. Mesenchymal stem cells have widely been regarded as hypoimmunogenic, enabling this cell-based administration across major histocompatibility barriers. In this review, we will highlight (1) the bidirectional communication of glioma cells and tumor-associated immune cells, (2) the inflammatory mediators enabling leukocytes and transplantable MSC migration, and (3) review preclinical and human clinical trials using MSCs as delivery vehicles. Mesenchymal stem cells possess innate abilities to migrate great distances, cross the blood-brain barrier, and communicate with surrounding cells, all of which make them desirable "Trojan horses" for brain cancer therapy.

Effects of sDR5-Fc fusion protein on infant mice with ulcerative colitis via the TRAIL-DR5 pathway.

To explore effects of the sDR5-Fc fusion protein on ulcerative colitis of infant mice via the TRAIL-DR5 pathway, 50 female mice were randomly divided into 5 groups, i.e., control group (group A), dextran sulfate sodium group (group B), hIgG group (group C), 10 mg/kg sDR5-Fc group (group D), and 20 mg/ kg sDR5-Fc group (group E). The acute ulcerative colitis models were established. The weights and disease activity index (DAI) of each group were monitored daily. In addition, the pathological changes of colon tissues were observed by Hematoxylin-Eosin staining. The number of macrophages in colon tissues was detected by immunohistochemistry assay. Changes in the expression of inflammatory factors in colon tissues were detected by quantitative real-time polymerase chain reaction (PCR). Lipopolysaccharide (LPS) of different concentrations was utilized alone or in combination with TRAIL to stimulate the NCM460 cells. The activation of NLRP3 inflammasomes was detected by Western blot. The apoptosis of NCM460 cells was detected by flow cytometry. The results showed that in groups B and C, the body weights decreased, the DAI increased, the colon epithelial cells were injured, the inflammatory cells were infiltrated, and the macrophages in colon tissues increased significantly. In groups D and E, the body weights increased, the DAI decreased, the inflammation was significantly improved, the macrophages decreased significantly, and the gene expression levels of NLRP3, Caspase-1, and IL-1ß decreased significantly. Thus, sDR5-Fc could inhibit the activation of NLRP3 inflammasomes induced by TRAIL, thereby decreasing the apoptosis of NCM460 cells. In conclusion, the sDR5-Fc fusion protein could block the TRAIL-DR5 pathway to reduce the expression of NLRP3 inflammasomes, thereby improving ulcerative colitis.

The enhanced efficacy of herpes simplex virus by lentivirus mediated VP22 and cytosine deaminase gene therapy against glioma.

Despite the comprehensive treatment of surgery following by concurrent radiochemotherapy, the prognosis of malignant glioma remains unsatisfactory with an awful median survival time of only 12-18 months. This might be improved by the development of oncolytic herpes simplex virus. However, the biggest challenge of recombinant herpes simplex virus (rHSV) lies in their limited therapeutic efficiency in clinical trials. This study aims to enhance the efficacy of rHSV against glioma by a HSV-1 tegument protein VP22 modified cytosine deaminase/5-flurocytosine (CD/5-FC) suicide gene system. Stable glioma cell lines carrying CD or VP22-CD fusion gene were successfully obtained by lentiviral infection following Fluorescence-activated cell sorting. The lethal effect of the prodrug 5-FC was significantly increased in the transduced VP22-CD glioma cells compared to the transduced CD glioma cells. Interestingly, VP22 was found to elicit the enhanced efficacy of rHSV-1 against glioma. Furthermore, we detected a synergistic efficacy of rHSV-1 combined with lentivirus mediated VP22 and CD suicide gene therapy in the orthotopic glioma xenograft models. In conclusion, we successfully established the stable cell lines carrying VP22-CD fusion genes. The incorporation of VP22 further induced an enhanced efficacy of rHSV-1 as well as CD suicide gene therapy respectively and synthetically in vitro. Also, we demonstrated an increased therapeutic benefit of rHAV-1 by VP22 modified CD gene therapy against glioma in vivo.

Time to embrace access programmes for medicines: lessons from the South African flucytosine access programme.

BACKGROUND: Cryptococcal meningitis (CM) is estimated to cause 181 000 deaths annually, with the majority occurring in Sub-Saharan Africa. Flucytosine is recommended by the World Health Organization as part of the treatment for CM. Widespread use of flucytosine could reduce mortality in hospital by as much as 40% compared to the standard of care, yet due to market failure, quality-assured flucytosine remains unregistered and largely inaccessible throughout Africa. METHODS: The recently established South African flucytosine clinical access programme is an attempt to address the market failure that led to a lack of public sector access to flucytosine for CM, by making the medicine freely available to tertiary hospitals in South Africa. RESULTS: Between November 2018 and September 2019, 327 CM patients received flucytosine through this programme, with efforts to support sustainable national scale-up presently ongoing. We describe why this programme was needed, its catalytic potential, what is still required to ensure widespread access to flucytosine, and observations from this experience that may have wider relevance. CONCLUSIONS: The South African flucytosine access programme illustrates how access programmes may be one part of the solution to addressing the vicious cycle of perceived low demand, limiting manufacturer interest in specific product markets.

The clinical values of dysregulated DNA methylation and demethylation intermediates in acute lymphoblastic leukemia.

Objectives: DNA methylation is a well-known epigenetic modification, and it can be iteratively oxidized to 5-hydroxymethylcytosine (5-hmC), 5-formylcytosine (5-fC) and 5-carboxylcytosine (5-caC). Acute lymphoblastic leukemia (ALL) is a severe hematological disease, and it is essential to find out new biomarkers to better diagnose and cure ALL due to the development of chemo-resistance and low cure rate in adult ALL. This study aims to describe the role of global methylation and demethylation intermediates in ALL. Methods: The levels of global methylation and its oxidation products in the peripheral blood (PB) of ALL patients and healthy controls were determined by Enzyme-Linked Immunosorbent Assay (ELISA). Results: In this study, we described that global 5-mC, 5-hmC and 5-fC levels were dysregulated in ALL, and they were associated with clinical characteristics and genetic abnormalities of ALL patients. Interestingly, 5-mC and 5-hmC were closely related to inflammation, and the levels of 5-hmC were inversely correlated with C-Reactive protein (CRP) and ferritin. Finally, 5-mC and 5-hmC were associated with complete remission (CR), and 5-hmC was revealed as an independent prognostic indicator for ALL. Conclusion: This study described a novel role for global methylation and demethylation intermediates in ALL detection and prognosis, and provided new clue to distinguish high-risk patients and improve the curative effect on ALL patients.

Mesenchymal stromal cells for bone sarcoma treatment: Roadmap to clinical practice.

Over the past few decades, there has been growing interest in understanding the molecular mechanisms of cancer pathogenesis and progression, as it is still associated with high morbidity and mortality. Current management of large bone sarcomas typically includes the complex therapeutic approach of limb salvage or sacrifice combined with pre- and postoperative multidrug chemotherapy and/or radiotherapy, and is still associated with high recurrence rates. The development of cellular strategies against specific characteristics of tumour cells appears to be promising, as they can target cancer cells selectively. Recently, Mesenchymal Stromal Cells (MSCs) have been the subject of significant research in orthopaedic clinical practice through their use in regenerative medicine. Further research has been directed at the use of MSCs for more personalized bone sarcoma treatments, taking advantage of their wide range of potential biological functions, which can be augmented by using tissue engineering approaches to promote healing of large defects. In this review, we explore the use of MSCs in bone sarcoma treatment, by analyzing MSCs and tumour cell interactions, transduction of MSCs to target sarcoma, and their clinical applications on humans concerning bone regeneration after bone sarcoma extraction.

Performance of revised STO(1M)-3G basis set for prediction of 5-fluorocytosine chemical shifts.

Nuclear shieldings and chemical shifts of 5-fluorocytosine (5FC) were predicted in the gas phase and DMSO solution modeled by polarizable continuum model using B3LYP density functional and revised STO(1M)-3G basis set. For comparison, eight arbitrary selected basis sets including STO-3G and medium-size Pople-type and larger dedicated Jensen-type ones were applied. The former basis sets were significantly smaller, but the calculated structural parameters, harmonic vibrational frequencies, were very accurate and close to those obtained with larger, polarization-consistent ones. The predicted 13 C and 1 H chemical shieldings of 5FC and cytosine, selected as parent molecule, were acceptable (root mean square for 13 C chemical shifts in DMSO of about 5 ppm and less) though less accurate than those calculated with large basis sets, dedicated for prediction of nuclear magnetic resonance parameters.