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Background and Objectives: Workplace burnout syndrome is often as sociated with particular aspects of certain job positions, especially those that entail working with people with special needs. The burnout syndrome in healthcare jobs is a serious problem that has grown into an epidemic among healthcare workers and associates. The aim of this research is to assess the presence of stress and burnout syndrome at work with healthcare workers, expert workers, professional associates, and associates in social service institutions in Belgrade. Materials and Methods: This research was conducted in the form of a cross-sectional study of a representative sample in social institutions in Belgrade. It was conducted from March to the end of June of 2023. The sample of the study had 491 participants. The questionnaires used were a structured instrument with social–demographic and social–economic characteristics, workplace characteristics, lifestyle characteristics, and the following questionnaires: DASS-21, Copenhagen, Brief Resilience Scale, and Brief Resilient Coping Scale. Results: The end results indicate the following to be significant risk factors for the occurrence of workplace burnout syndrome: overtime (OR = 2.62; CI = 1.50–4.56), BRS average score (OR = 0.28; CI = 0.17–0.44), DASS21 D heightened depression (OR = 2.09; CI = 1.1–4.04), DASS21 A heightened anxiety (OR = 2.38; CI = 1.34–4.21), and DASS21 S heightened stress (OR = 2.08; CI = 1.11–3.89). The only protective risk factor that stood out was the self-assessment of health levels (OR = 0.60; CI = 0.42–0.85). Conclusion: Overtime is a significant factor associated with workplace burnout. Apart from it, other significant factors associated with workplace burnout were heightened depression, anxiety, and stress levels.

Inhibitors of dihydrofolate reductase (DHFR), an enzyme that catalyzes 5,6,7,8-tetrahydrofolate synthesis, have been used as antimicrobial as well as antimetabolite drugs for a long time. Although structurally belonging to different classes, the majority of DHFR inhibitors contain 2,4-diamino substitution in pyrimidine ring. With the aim to introduce pyrimido-pyrimidines as a novel class of bacterial DHFR (bDHFR) inhibitors, 42 compounds belonging to that class have been tested and compared with 18 pteridines using cell and enzyme models and docking studies. A few pyrimido-pyrimidine compounds showed high potency (IC50≤0.05 μM) and selectivity as inhibitors of bDHFR. These properties seem to be dependent on the stringent structure freedom and flexibility, based on the specific combination of prerequisite structural motifs that enable pyrimido-pyrimidine compounds to fit into bDHFR active site in a relatively specific manner. The presented results will help to set the basis for designing new small molecules, inhibitors of DHFR, with interesting and potentially selective antibacterial properties.

Background: By 2021, the FDA approved the use of the drugs pembrolizumab and atezolizumab in the first-line treatment of patients with high positivity of programmed death ligand-1 (PD-L1) in locally advanced and metastatic non-small-cell-lung cancer (NSCLC). This approval was the result of statistically significant evidence from international, multicentric clinical studies that all reported increasing progression-free survival (PFS) and overall survival (OS) in these patients.Methods: In our study, we reported the demographic and clinical characteristics of 79 patients diagnosed with NSCLC with expression of PD-L1 ≥50% from January 2019 to December 2022 at the Institute for Pulmonary Diseases of Vojvodina, who received pembrolizumab therapy as the first-line treatment. Patients were divided according to the histological type of lung cancer as adenocarcinoma (ADC) or squamous cell carcinoma (SCC) of the lung. In 52 of the 79 patients, PFS and in 32 of them overall survival (censored OS) was shown according to the histological type of tumor, the tumor proportion score (TPS) of PDL-1 expression, and the metastatic status within the Tumor Nodes Metastasis (TNM) disease classification. Independent factors of death outcome were shown by multivariable proportional hazard regression analysis.Results: The study included 79 patients diagnosed with NSCLC with an expression of PD-L1 ≥50%, 50 (63.3%) patients with ADC, and 29 (36.7%) patients with SCC, whose 55 (69.6%) PDL-1 expression was obtained from broncho biopsy (BB). The majority of patients, 49 (62%), had a TPS PD-L1 score of 51%–79%. Median, PFS for adenocarcinoma was 22 months and censored OS was 27 months, while for squamous cell carcinoma, median PFS was 12 months, and censored OS was 21 months. M1b disease stage, which was the most common in patients, had a PFS of 16 months and a censored OS of 18 months.Conclusion: Pembrolizumab monotherapy in patients with NSCLC in the fourth stage of the disease and with the positivity of the immune checkpoint protein TPS PD-L1 above 50% represents a safe therapy that allows a satisfactory period without disease progression and overall survival with acceptable treatment complications.

Background and Objectives: The primary objective of this study is to assess the effectiveness of 18F-FDG–PET–CT in preoperative staging of cervical cancer, focusing on determining surgical operability and exploring the correlation between its quantitative parameters and clinicopathological characteristics. Materials and Methods: This retrospective study included 62 cervical cancer patients treated at the Department of Gynecology, Clinic for Operative Oncology at the Institute of Oncology Vojvodina between January 2016 and January 2020, where preoperative clinical examinations and 18F-FDG–PET–CT were performed to assess the extent of cancer, followed by intraoperative and pathohistological examinations of surgically removed specimens to provide a comprehensive evaluation. Results: The mean tumor size measured by 18F-FDG–PET–CT was slightly greater than that obtained through clinical examination (26.4 mm vs. 26.0 mm), with a strong linear correlation (r = 0.678, p < 0.001) observed between the two measurement methods. The overall prediction accuracy of 18F-FDG–PET–CT for primary tumors is 88.7% (55/62) [sensitivity 86.8%, specificity 100.0%, PPV 100.0%, NPV 56.2%] and for intraoperative examination is 88.7% (55/62) [sensitivity 98.1%, specificity 33.3%, PPV 89.7%, NPV 75.0%]. The agreement with histopathological examination was good for 18F-FDG–PET–CT and moderate for intraoperative examination for primary tumors. Regarding lymph nodes, the overall prediction accuracy of 18F-FDG–PET–CT is 82.2% (51/62) [sensitivity 53.8%, specificity 89.8%, PPV 58.3%, NPV 88.8%] and for intraoperative examination 66.1% (41/62) [sensitivity 76.9%, specificity 63.3%, PPV 35.7%, NPV 91.2%]. The agreement with histopathological examination was moderate for 18F-FDG–PET–CT and poor for intraoperative examination for lymph node metastasis, highlighting that the overall accuracy of 18F-FDG–PET–CT (82.1%) was significantly higher than that of intraoperative examination (66.1%) (p = 0.002). Conclusions: In conclusion, 18F-FDG–PET–CT provides high accuracy in detecting primary tumors and superior predictive value for lymph node metastases compared to intraoperative examination, highlighting the importance of incorporating this imaging modality into the preoperative evaluation process to enhance diagnostic precision and inform treatment decisions.

Background and Objectives: Advanced lung cancer is usually manifested by endoluminal tumor propagation, resulting in central airway obstruction. The objective of this study is to compare the high dose rate brachytherapy treatment outcomes in non-small-cell lung cancer (NSCLC) depending on the treatment planning pattern—two-dimension (2D) or three-dimension (3D) treatment planning. Materials and Methods: The study was retrospective and two groups of patients were compared in it (a group of 101 patients who underwent 2D planned high-dose-rate endobronchial brachytherapy (HDR-EBBT) in 2017/18 and a group of 83 patients who underwent 3D planned HDR-EBBT between January 2021 and June 2023). Results: In the group of 3D planned brachytherapy patients, there was a significant improvement in terms of loss of symptoms of bronchial obstruction (p = 0.038), but no improvement in terms of ECOG PS (European Cooperative Oncology Group Performance Status) of the patient (p = 0.847) and loss of lung atelectasis (if there was any at the beginning of the disease) (p = 0.781). Two-year overall survival and time-to-progression periods were similar for both groups of patients (p = 0.110 and 0.154). Fewer treatment complications were observed, and 91.4% were in 3D planned brachytherapy (BT) patients. Conclusions: Three-dimensionally planned HDR-EBBT is a suggestive, effective palliative method for the disobstruction of large airways caused by endobronchial lung tumor growth. Independent or more often combined with other types of specific oncological treatment, it certainly leads to the loss of symptoms caused by bronchial obstruction and the improvement of the quality of life of patients with advanced NSCLC. Complications of the procedure with 3D planning are less compared to 2D planned HDR-EBBT.

Abstract Prostate cancer is the second most frequent cancer diagnosed in men worldwide. Localized disease can be successfully treated, but advanced cases are more problematic. After initial effectiveness of androgen deprivation therapy, resistance quickly occurs. Therefore, we aimed to investigate the role of Hedgehog-GLI (HH-GLI) signaling in sustaining androgen-independent growth of prostate cancer cells. We found various modes of HH-GLI signaling activation in prostate cancer cells depending on androgen availability. When androgen was not deprived, we found evidence of non-canonical SMO signaling through the SRC kinase. After short-term androgen deprivation canonical HH-GLI signaling was activated, but we found little evidence of canonical HH-GLI signaling activity in androgen-independent prostate cancer cells. We show that in androgen-independent cells the pathway ligand, SHH-N, non-canonically binds to the androgen receptor through its cholesterol modification. Inhibition of this interaction leads to androgen receptor signaling downregulation. This implies that SHH-N activates the androgen receptor and sustains androgen-independence. Targeting this interaction might prove to be a valuable strategy for advanced prostate cancer treatment. Also, other non-canonical aspects of this signaling pathway should be investigated in more detail and considered when developing potential therapies.

Dipeptidyl peptidase III (DPP III), a zinc exopeptidase, is involved in the final steps of intercellular protein degradation and has a marked affinity for opioid peptides such as enkephalins and endomorphins. Recently, we characterized a number of neuropeptides as potential substrates and inhibitors of human DPP III and provided an explanation for their differential behavior. These studies prompted us to investigate the influence of the conserved R399 and R669 on neuropeptides binding to DPP III. Measuring kinetic parameters in inhibitory assays, we found that mutation of R669 to Ala or Met significantly reduced the inhibitory properties of the slow substrates tynorphin and valorphin, whereas the effects on binding of the good substrates Arg2-2NA and Leu-enkephalin were small. Molecular dynamics simulations of wild-type (WT) and mutant DPP III complexes with Leu-enkephalin, tynorphin, valorphin, and Arg2-2NA in conjunction with calculations of binding free energies revealed that the lower inhibitory potency of slow substrates in the R669A mutant can be explained by the lower binding affinity of tynorphin and the higher propensity of valorphin to hydrolyze in the mutant than in WT. The R399A mutation was shown to affect the binding and/or hydrolysis of both good and slow substrates, with the effects on Leu-enkephalin being the most pronounced.

Interactions of an array of nucleic acid structures with a small series of benzothiazole ligands (bis-benzothiazolyl-pyridines—group 1, 2-thienyl/2-benzothienyl-substituted 6-(2-imidazolinyl)benzothiazoles—group 2, and three 2-aryl/heteroaryl-substituted 6-(2-imidazolinyl)benzothiazoles—group 3) were screened by competition dialysis. Due to the involvement of DNA:RNA hybrids and triplex helices in many essential functions in cells, this study’s main aim is to detect benzothiazole-based moieties with selective binding or spectroscopic response to these nucleic structures compared to regular (non-hybrid) DNA and RNA duplexes and single-stranded forms. Complexes of nucleic acids and benzothiazoles, selected by this method, were characterized by UV/Vis, fluorescence and circular dichroism (CD) spectroscopy, isothermal titration calorimetry, and molecular modeling. Two compounds (1 and 6) from groups 1 and 2 demonstrated the highest affinities against 13 nucleic acid structures, while another compound (5) from group 2, despite lower affinities, yielded higher selectivity among studied compounds. Compound 1 significantly inhibited RNase H. Compound 6 could differentiate between B- (binding of 6 dimers inside minor groove) and A-type (intercalation) helices by an induced CD signal, while both 5 and 6 selectively stabilized ATT triplex in regard to AT duplex. Compound 3 induced strong condensation-like changes in CD spectra of AT-rich DNA sequences.

Novel dyes were prepared by simple “click CuAAC” attachment of a triarylborane–alkyne to the azide side chain of an amino acid yielding triarylborane dye 1 which was conjugated with pyrene (dye 2) forming a triarylborane–pyrene FRET pair. In contrast to previous cationic triarylboranes, the novel neutral dyes interact only with proteins, while their affinity to DNA/RNA is completely abolished. Both the reference triarylborane amino acid and triarylborane–pyrene conjugate bind to BSA and the hDPP III enzyme with high affinities, exhibiting a strong (up to 100-fold) fluorescence increase, whereby the triarylborane–pyrene conjugate additionally retained FRET upon binding to the protein. Furthermore, the triarylborane dyes, upon binding to the hDPP III enzyme, did not impair its enzymatic activity under a wide range of experimental conditions, thus being the first non-covalent fluorimetric markers for hDPP III, also applicable during enzymatic reactions with hDPP III substrates.

Dipeptidyl peptidase III (DPP III), a zinc-dependent exopeptidase, is a member of the metalloproteinase family M49 with distribution detected in almost all forms of life. Although the physiological role of human DPP III (hDPP III) is not yet fully elucidated, its involvement in pathophysiological processes such as mammalian pain modulation, blood pressure regulation, and cancer processes, underscores the need to find new hDPP III inhibitors. In this research, five series of structurally different coumarin derivatives were studied to provide a relationship between their inhibitory profile toward hDPP III combining an in vitro assay with an in silico molecular modeling study. The experimental results showed that 26 of the 40 tested compounds exhibited hDPP III inhibitory activity at a concentration of 10 µM. Compound 12 (3-benzoyl-7-hydroxy-2H-chromen-2-one) proved to be the most potent inhibitor with IC50 value of 1.10 μM. QSAR modeling indicates that the presence of larger substituents with double and triple bonds and aromatic hydroxyl groups on coumarin derivatives increases their inhibitory activity. Docking predicts that 12 binds to the region of inter-domain cleft of hDPP III while binding mode analysis obtained by MD simulations revealed the importance of 7-OH group on the coumarin core as well as enzyme residues Ile315, Ser317, Glu329, Phe381, Pro387, and Ile390 for the mechanism of the binding pattern and compound 12 stabilization. The present investigation, for the first time, provides an insight into the inhibitory effect of coumarin derivatives on this human metalloproteinase.

Kelch-like ECH associated protein 1 (Keap1) is a cellular sensor for oxidative stress and a negative regulator of the transcription factor Nrf2. Keap1 and Nrf2 control expression of nearly 500 genes with diverse cytoprotective functions and the Nrf2-Keap1 signaling pathway is a major regulator of cytoprotective responses to oxidative and electrophilic stress. It was found that the metallopeptidase dipeptidyl peptidase III (DPP III) contributes to Nrf2 activation by binding to Keap1, probably by binding to the Kelch domain, and thereby influences Nrf2 activity in cancer. We here first determined that the KD of the DPP III-Kelch domain complex is in the submicromolar range. In order to elucidate the molecular details of the DPP III – Kelch interaction we then built models of the complex between human DPP III and the Keap1 Kelch domain and performed coarse-grained and atomistic simulations of the complexes. In the most stable complexes, the ETGE motif in the DPP III flexible loop binds near the central pore of the six-blade β-propeller Kelch domain. According to the preliminary HD exchange experiments DPP III binds to the more unstructured end of Kelch domain. According to the results of MD simulations DPP III binding to the Kelch domain does not influence the overall DPP III structure or the long-range domain fluctuations. We can conclude that DPP III forms the stable complexes with the Keap1 Kelch domain by inserting the flexible loop into the entrance to the central pore of the six blade β-propeller Kelch domain at its more unstructured, N-terminus. This work is licensed under a Creative Commons Attribution 4.0 International License.

A molecular modeling study is performed on series of benzimidazol-based inhibitors of human dipeptidyl peptidase III (DPP III). An eight novel compounds were synthesized in excellent yields using green chemistry approach. This study is aimed to elucidate the structural features of benzimidazole derivatives required for antagonism of human DPP III activity using Quantitative Structure-Activity Relationship (QSAR) analysis, and to understand the mechanism of one of the most potent inhibitor binding into the active site of this enzyme, by molecular dynamics (MD) simulations. The best model obtained includes S3K and RDF045m descriptors which have explained 89.4 % of inhibitory activity. Depicted moiety for strong inhibition activity matches to the structure of most potent compound. MD simulation has revealed importance of imidazolinyl and phenyl groups in the mechanism of binding into the active site of human DPP III.

Dipeptidyl peptidase III (DPP III) isolated from the thermophilic bacteria Caldithrix abyssi (Ca) is a two-domain zinc exopeptidase, a member of the M49 family. Like other DPPs III, it cleaves dipeptides from the N-terminus of its substrates but differently from human, yeast and Bacteroides thetaiotaomicron (mesophile) orthologs, it has the pentapeptide zinc binding motif (HEISH) in the active site instead of the hexapeptide (HEXXGH). The aim of our study was to investigate structure, dynamics and activity of CaDPP III, as well as to find possible differences with already characterized DPPs III from mesophiles, especially B. thetaiotaomicron. The enzyme structure was determined by X-ray diffraction, while stability and flexibility were investigated using MD simulations. Using molecular modeling approach we determined the way of ligands binding into the enzyme active site and identified the possible reasons for the decreased substrate specificity compared to other DPPs III. The obtained results gave us possible explanation for higher stability, as well as higher temperature optimum of CaDPP III. The structural features explaining its altered substrate specificity are also given. The possible structural and catalytic significance of the HEISH motive, unique to CaDPP III, was studied computationally, comparing the results of long MD simulations of the wild type enzyme with those obtained for the HEISGH mutant. This study presents the first structural and biochemical characterization of DPP III from a thermophile.

Porphyromonas gingivalis, an asaccharolytic Gram-negative oral anaerobe, is a major pathogen associated with adult periodontitis, a chronic infective disease that a significant percentage of the human population suffers from. It preferentially utilizes dipeptides as its carbon source, suggesting the importance of dipeptidyl peptidase (DPP) types of enzyme for its growth. Until now DPP IV, DPP5, 7 and 11 have been extensively investigated. Here, we report the characterization of DPP III using molecular biology, biochemical, biophysical and computational chemistry methods. In addition to the expected evolutionarily conserved regions of all DPP III family members, PgDPP III possesses a C-terminal extension containing an Armadillo (ARM) type fold similar to the AlkD family of bacterial DNA glycosylases, implicating it in alkylation repair functions. However, complementation assays in a DNA repair-deficient Escherichia coli strain indicated the absence of alkylation repair function for PgDPP III. Biochemical analyses of recombinant PgDPP III revealed activity similar to that of DPP III from Bacteroides thetaiotaomicron, and in the range between activities of human and yeast counterparts. However, the catalytic efficiency of the separately expressed DPP III domain is ~1000-fold weaker. The structure and dynamics of the ligand-free enzyme and its complex with two different diarginyl arylamide substrates was investigated using small angle X-ray scattering, homology modeling, MD simulations and hydrogen/deuterium exchange (HDX). The correlation between the experimental HDX and MD data improved with simulation time, suggesting that the DPP III domain adopts a semi-closed or closed form in solution, similar to that reported for human DPP III. The obtained results reveal an atypical DPP III with increased structural complexity: its superhelical C-terminal domain contributes to peptidase activity and influences DPP III interdomain dynamics. Overall, this research reveals multifunctionality of PgDPP III and opens direction for future research of DPP III family proteins.