Publications by authors named "Tomasz Wróbel"

153 Publications

Immunosuppression as a Hallmark of Critical COVID-19: Prospective Study.

Cells 2021 05 23;10(6). Epub 2021 May 23.

Department and Clinic of Hematology, Blood Neoplasms, and Bone Marrow Transplantation, Wroclaw Medical University, 50-367 Wroclaw, Poland.

The dysregulation of both the innate and adaptive responses to SARS-CoV-2 have an impact on the course of COVID-19, and play a role in the clinical outcome of the disease. Here, we performed a comprehensive analysis of peripheral blood lymphocyte subpopulations in 82 patients with COVID-19, including 31 patients with a critical course of the disease. In COVID-19 patients who required hospitalization we analyzed T cell subsets, including Treg cells, as well as TCRα/β and γ/δ, NK cells, and B cells, during the first two weeks after admission to hospital due to the SARS-CoV-2 infection, with marked reductions in leukocytes subpopulations, especially in critically ill COVID-19 patients. We showed decreased levels of Th, Ts cells, Treg cells (both naïve and induced), TCRα/β and γ/δ cells, as well as CD16+CD56+NK cells in ICU compared to non-ICU COVID-19 patients. We observed impaired function of T and NK cells in critically ill COVID-19 patients with extremely low levels of secreted cytokines. We found that the IL-2/INFγ ratio was the strongest indicator of a critical course of COVID-19, and was associated with fatal outcomes. Our findings showed markedly impaired innate and adaptive responses in critically ill COVID-19 patients, and suggest that the immunosuppressive state in the case of a critical course of SARS-CoV-2 infection might reflect subsequent clinical deterioration and predict a fatal outcome.
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http://dx.doi.org/10.3390/cells10061293DOI Listing
May 2021

IDH2 mutations in patients with normal karyotype AML predict favorable responses to daunorubicin, cytarabine and cladribine regimen.

Sci Rep 2021 May 11;11(1):10017. Epub 2021 May 11.

Department of Cancer Prevention, School of Public Health, Silesian Medical University, Katowice, Poland.

Mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) genes occur in about 20% patients with acute myeloid leukemia (AML), leading to DNA hypermethylation and epigenetic deregulation. We assessed the prognostic significance of IDH1/2 mutations (IDH1/2) in 398 AML patients with normal karyotype (NK-AML), treated with daunorubicine + cytarabine (DA), DA + cladribine (DAC), or DA + fludarabine. IDH2 mutation was an independent favorable prognostic factor for 4-year overall survival (OS) in total NK-AML population (p = 0.03, censoring at allotransplant). We next evaluated the effect of addition of cladribine to induction regimen on the patients' outcome according to IDH1/2 mutation status. In DAC group, 4-year OS was increased in IDH2 patients, compared to IDH-wild type group (54% vs 33%; p = 0.0087, censoring at allotransplant), while no difference was observed for DA-treated subjects. In multivariate analysis, DAC independently improved the survival of IDH2 patients (HR = 0.6 [0.37-0.93]; p = 0.024; censored at transplant), indicating that this group specifically benefits from cladribine-containing therapy. In AML cells with R140Q or R172K IDH2 mutations, cladribine restrained mutations-related DNA hypermethylation. Altogether, DAC regimen produces better outcomes in IDH2 NK-AML patients than DA, and this likely results from the hypomethylating activity of cladribine. Our observations warrant further investigations of induction protocols combining cladribine with IDH1/2 inhibitors in IDH2-mutant.
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http://dx.doi.org/10.1038/s41598-021-88120-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8113255PMC
May 2021

Platelet Reactivity and Response to Aspirin and Clopidogrel in Patients with Platelet Count Disorders.

Cardiol Res Pract 2021 17;2021:6637799. Epub 2021 Apr 17.

Department of Hematology, Blood Neoplasms, and Bone Marrow Transplantation, Wroclaw Medical University, Wrocław, Poland.

Background: Platelet reactivity and response to antiplatelet drugs, acetylsalicylic acid (ASA) and clopidogrel, in patients with thrombocytopenia and thrombocythemia can have a potentially important effect on the outcome. The effectiveness and safety of antiplatelet drugs in such patients has not been well examined. Measuring the effect of ASA and clopidogrel on platelets could help guide the therapy. Nevertheless, platelet response to antiplatelet drugs is not routinely measured in platelet count disorders and relevant evidence is scarce.

Aims: The study aimed to measure platelet reactivity and response to ASA and clopidogrel in patients with platelet count disorders.

Materials And Methods: This was a cross-sectional study of consecutive patients hospitalized in cardiology and hematology departments in the years 2018-2019. The study included patients with thrombocytopenia (PLT < 150 G/L) and thrombocythemia (PLT > 450 G/L) on ASA or dual antiplatelet therapy (DAPT; ASA plus clopidogrel). Controls included patients on antiplatelet drugs with normal platelet count. Platelet reactivity was measured in whole blood (Multiplate aggregometer, Roche, Switzerland) using arachidonic acid (AA), adenosine-5'-diphosphate (ADP), and thrombin receptor agonist peptide-6 (TRAP) as agonists. Platelet aggregation was expressed in arbitrary units (AU). AA-induced aggregation was used as a measure of response to ASA with a cut-off above 30 AU showing high on-treatment platelet reactivity to ASA (HTPR-A). ADP-induced aggregation measured response to clopidogrel with a cut-off above 48 AU for high on-treatment platelet reactivity to clopidogrel (HTPR-C). TRAP-induced aggregation measured baseline platelet reactivity not affected by oral antiplatelet drugs.

Results: The study included 174 patients. There were 64 patients with thrombocytopenia, 30 patients with chronic thrombocythemia, and 80 controls. All patients were on 75 mg of ASA and 32% of them additionally on 75 mg of clopidogrel due to a history of recent coronary artery angioplasty. AA- and ADP-induced aggregation was comparable between thrombocytopenic patients and controls (median (IQR) 19 (7-28) vs. 23 (15-38) for AA AU and 32 (16-44) vs. 50 (32-71) for ADP AU, respectively), while it was significantly higher in thrombocythemic patients (median (IQR) 80 (79-118) for AA AU and 124 (89-139) for ADP AU). TRAP-induced aggregation showed significantly lowest aggregation in thrombocytopenic (median (IQR) 41 (34-60) for TRAP AU) and highest in thrombocythemic patients (median (IQR) 137 (120-180) for TRAP AU). HTPR-A was frequent in thrombocythemic patients in comparison with thrombocytopenic patients and controls (60% vs. 4% vs. 15%, respectively; < 0.0002). HTPR-C was highly common in thrombocythemic patients and least common in thrombocytopenic ones in comparison with controls (80% vs. 8% vs. 40%, respectively; < 0.001).

Conclusion: Chronic thrombocytopenia does not significantly affect platelet reactivity and response to ASA and clopidogrel in comparison with controls. Thrombocytosis significantly increases platelet reactivity and attenuates response to both ASA and clopidogrel.
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http://dx.doi.org/10.1155/2021/6637799DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8068533PMC
April 2021

The Impact of Preprocessing Methods for a Successful Prostate Cell Lines Discrimination Using Partial Least Squares Regression and Discriminant Analysis Based on Fourier Transform Infrared Imaging.

Cells 2021 Apr 20;10(4). Epub 2021 Apr 20.

Institute of Nuclear Physics Polish Academy of Sciences, PL-31342 Krakow, Poland.

Fourier transform infrared spectroscopy (FT-IR) is widely used in the analysis of the chemical composition of biological materials and has the potential to reveal new aspects of the molecular basis of diseases, including different types of cancer. The potential of FT-IR in cancer research lies in its capability of monitoring the biochemical status of cells, which undergo malignant transformation and further examination of spectral features that differentiate normal and cancerous ones using proper mathematical approaches. Such examination can be performed with the use of chemometric tools, such as partial least squares discriminant analysis (PLS-DA) classification and partial least squares regression (PLSR), and proper application of preprocessing methods and their correct sequence is crucial for success. Here, we performed a comparison of several state-of-the-art methods commonly used in infrared biospectroscopy (denoising, baseline correction, and normalization) with the addition of methods not previously used in infrared biospectroscopy classification problems: Mie extinction extended multiplicative signal correction, Eiler's smoothing, and probabilistic quotient normalization. We compared all of these approaches and their effect on the data structure, classification, and regression capability on experimental FT-IR spectra collected from five different prostate normal and cancerous cell lines. Additionally, we tested the influence of added spectral noise. Overall, we concluded that in the case of the data analyzed here, the biggest impact on data structure and performance of PLS-DA and PLSR was caused by the baseline correction; therefore, much attention should be given, especially to this step of data preprocessing.
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http://dx.doi.org/10.3390/cells10040953DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8073124PMC
April 2021

Temozolomide Induces the Acquisition of Invasive Phenotype by O6-Methylguanine-DNA Methyltransferase (MGMT) Glioblastoma Cells in a Snail-1/Cx43-Dependent Manner.

Int J Mol Sci 2021 Apr 16;22(8). Epub 2021 Apr 16.

Department of Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University in Kraków, ul. Gronostajowa 7, 30-387 Kraków, Poland.

Glioblastoma multiforme (GBM) recurrences after temozolomide (TMZ) treatment result from the expansion of drug-resistant and potentially invasive GBM cells. This process is facilitated by O6-Methylguanine-DNA Methyltransferase (MGMT), which counteracts alkylating TMZ activity. We traced the expansion of invasive cell lineages under persistent chemotherapeutic stress in MGMT (U87) and MGMT (T98G) GBM populations to look into the mechanisms of TMZ-induced microevolution of GBM invasiveness. TMZ treatment induced short-term, pro-invasive phenotypic shifts of U87 cells, in the absence of Snail-1 activation. They were illustrated by a transient induction of their motility and followed by the hypertrophy and the signs of senescence in scarce U87 sub-populations that survived long-term TMZ stress. In turn, MGMT T98G cells reacted to the long-term TMZ treatment with the permanent induction of invasiveness. Ectopic Snail-1 down-regulation attenuated this effect, whereas its up-regulation augmented T98G invasiveness. MGMT and MGMT cells both reacted to the long-term TMZ stress with the induction of Cx43 expression. However, only in MGMT T98G populations, Cx43 was directly involved in the induction of invasiveness, as manifested by the induction of T98G invasiveness after ectopic Cx43 up-regulation and by the opposite effect after Cx43 down-regulation. Collectively, Snail-1/Cx43-dependent signaling participates in the long-term TMZ-induced microevolution of the invasive GBM front. High MGMT activity remains a prerequisite for this process, even though MGMT-related GBM chemoresistance is not necessary for its initiation.
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http://dx.doi.org/10.3390/ijms22084150DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8073161PMC
April 2021

Successful treatment of COVID-19 in a patient with severe haemophilia A on emicizumab prophylaxis in the intensive care unit.

Haemophilia 2021 Apr 29. Epub 2021 Apr 29.

Department of Haematology, Blood Neoplasms and Bone Marrow Transplantation, Wroclaw Medical University, Wroclaw, Poland.

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http://dx.doi.org/10.1111/hae.14326DOI Listing
April 2021

Co-occurrence of unclassified myeloproliferative neoplasm and giant cell arteritis in a patient treated with allogeneic hematopoietic stem cell transplantation: a case report and literature review.

Cent Eur J Immunol 2021 18;46(1):121-126. Epub 2021 Apr 18.

Department of Hematology, Wroclaw Medical University, Wroclaw, Poland.

Myeloproliferative neoplasms (MPNs) are a group of hematologic disorders characterized by clonal proliferation of myeloid lineage cells. The diagnostic criteria are based on morphological features of bone marrow and peripheral blood cells but also include specific genomic mutations. In some patients, co-occurrence of hematologic and rheumatic diseases could be observed. To date, most of the reported cases concerned patients with myelodysplastic syndrome (MDS) or essential thrombocythemia (ET). In this paper, we present a case of a patient with a complicated diagnostic process leading to the diagnosis of unclassified MPN and giant cell arteritis (GCA). Routine tests did not reveal any mutations typical for MPNs such as JAK-2, CALR, MPL or BCR-ABL. Targeted next-generation sequencing (NGS) helped to confirm the diagnosis by demonstrating the presence of heterozygous ASXL1, TET2, SRSF2, and CBL mutations. The second important issue was the overlapping of symptoms of MPN and seronegative rheumatic disease, which finally was diagnosed as GCA. Leukocytosis and musculoskeletal pain, which were present at the time of diagnosis, resolved after allogeneic hematopoietic stem cell transplantation but recurred after a few months along with decreasing donor cell chimerism. Differentiation of the causes of recurrence of the symptoms was an important issue. This case shows the diagnostic challenge posed by co-incidence of MPN and rheumatic disease, especially its atypical variants.
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http://dx.doi.org/10.5114/ceji.2019.83140DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8056354PMC
April 2021

Early Administration of Convalescent Plasma Improves Survival in Patients with Hematological Malignancies and COVID-19.

Viruses 2021 03 8;13(3). Epub 2021 Mar 8.

Department and Clinic of Haematology, Blood Neoplasms, and Bone Marrow Transplantation, Wroclaw Medical University, Pasteura Street 4, 50-367 Wroclaw, Poland.

The use of convalescent plasma in the treatment of COVID-19 may lead to a milder course of infection and has been associated with improved outcomes. Determining optimal treatments in high risk populations is crucial, as is the case in those with hematological malignancies. We analyzed a cohort of 23 patients with hematological malignancies and COVID-19 who had received plasma 48-72 h after the diagnosis of infection and compared it with a historical group of 22 patients who received other therapy. Overall survival in those who received convalescent plasma was significantly higher than in the historical group ( = 0.03460). The plasma-treated group also showed a significantly milder course of infection ( = 0.03807), characterized by less severe symptoms and faster recovery ( = 0.00001). In conclusion, we have demonstrated that convalescent plasma is an effective treatment and its early administration leads to clinical improvement, increased viral clearance and longer overall survival in patients with hematological malignancies and COVID-19. To our knowledge, this is the first report to analyze the efficacy of convalescent plasma in a cohort of patients with hematological malignancies.
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http://dx.doi.org/10.3390/v13030436DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8001057PMC
March 2021

What factors determine the pregnancy outcome in patients with essential thrombocythemia?

J Matern Fetal Neonatal Med 2021 Apr 1:1-5. Epub 2021 Apr 1.

Department and Clinic of Angiology, Hypertension, and Diabetology, Wroclaw Medical University, Wrocław, Poland.

Treatment of essential thrombocythemia (ET) is particularly challenging in pregnancy due to the increased risk of thromboembolic complications. Therefore, the use of antithrombotic regimens are recommended in pregnant women with ET. The study included 52 pregnancies in 27 patients diagnosed with ET, who were treated in Department of Haematology. The influence of anticoagulant, antiplatelet and cytoreductive therapy on the course and outcome of pregnancy was analysed. This study also examined if there was any correlation between molecular and clinical features such as mutational profile, blood count, presence of acquired von Willebrand syndrome (AvWS), the International Prognostic Score for Essential Thrombocythemia (IPSET) risk group and the IPSET-thrombosis risk group and pregnancy outcome. Study participants who received antithrombotic therapy were significantly more likely to give birth to a healthy child. The best outcomes were observed in patients who received low dose acetylsalicylic acid (ASA) together with low-molecular-weight heparin (LMWH). There was a statistically significant correlation between classification to the high-risk group according to the IPSET-thrombosis score and incidence of miscarriage. Cytoreductive treatment with interferon-α2, as well as the presence of AvWS did not increase the likelihood of pregnancy loss. Blood counts and presence of specific gene mutations profile were also not found to be significant determinants of pregnancy outcome. To our best knowledge, this is the first clinical study investigating the correlation between risk group (according to IPSET and IPSET-thrombosis) and pregnancy outcome in women with ET.
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http://dx.doi.org/10.1080/14767058.2020.1863362DOI Listing
April 2021

Exploring subcellular responses of prostate cancer cells to clinical doses of X-rays by Raman microspectroscopy.

Spectrochim Acta A Mol Biomol Spectrosc 2021 Jul 8;255:119653. Epub 2021 Mar 8.

Institute of Nuclear Physics Polish Academy of Sciences, Radzikowskiego 152, 31-342 Krakow, Poland.

Modern techniques of radiotherapy such as fractioned radiotherapy require applications of low doses of ionizing radiation (up to 10 Gy) for effective patient treatment. It is, therefore, crucial to understand the response mechanisms in cancer cells irradiated with low (clinical) doses. The cell's response to irradiation depends on a dose and post-irradiation time. Both factors should be considered when studying the influence of ionizing radiation on cancer cells. Thus, in the present study, PC-3 prostate cancer cells were irradiated with clinical doses of X-rays to determine dose- and time-dependent response to the irradiation. Raman spectroscopy and biological methods (MTT and comet assays) were applied for the analysis of biochemical changes in the cells induced by low doses of X-ray irradiation at 0 h and 24 h post-irradiation timepoints. Due to a limited view of the biochemical changes at the subcellular level given by single spectrum Raman measurements, Raman mapping of the whole cell area was performed. The results were compared with those obtained for cell irradiation with high doses. The analysis was based on the Partial Least Squares Regression (PLSR) method for the cytoplasmic and nuclear regions separately. Additionally, for the first time, irradiation classification was performed to confirm Raman spectroscopy as a powerful tool for studies on cancer cells treated with clinical doses of ionizing radiation.
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http://dx.doi.org/10.1016/j.saa.2021.119653DOI Listing
July 2021

Stem cell mobilization in multiple myeloma patients relapsing after previous autologous hematopoietic stem cell transplantation: A multicenter report by the Polish Myeloma Study Group.

J Clin Apher 2021 Jun 16;36(3):443-453. Epub 2021 Feb 16.

Department of Hematology, Transplantation and Internal Diseases, Medical University of Warsaw, Warsaw, Poland.

Background: Salvage autologous hematopoietic stem cell transplantation (autoHSCT) may be used to treat relapse of multiple myeloma occurring after previous autoHSCT. When insufficient number of hematopoietic stem cells was stored from the initial harvest, remobilization of stem cells is necessary.

Purpose: The analysis of stem cell remobilization after previous autoHSCT.

Patients And Methods: Fifty-eight patients, 60% males, median 59 years, were included. Median time interval between autoHSCT and remobilization was 42 months. The first remobilization was performed mostly after chemotherapy: cyclophosphamide (33%), cytarabine (43%), and etoposide (19%).

Results: The first remobilization was successful in 67% patients. About 19% patients required plerixafor rescue, among whom it allowed for successful harvesting in 14%. Use of cyclophosphamide, cytarabine, and etoposide allowed for successful remobilization in 53%, 84%, and 55% patients, respectively. Patients treated with cytarabine had the highest yield of CD34+ cells (median 7.5 × 10 /kg vs 5.8 and 2.4 for etoposide and cyclophosphamide, P = .001). Higher percentage of patients was able to collect ≥2 × 10 CD34+ cells/kg during one leukapheresis after cytarabine (76% vs 21% for cyclophosphamide vs 36% for etoposide, P = .001). Cytarabine use was associated with lower risk of remobilization failure OR = 0.217, P = .02. Toxicity comprised mostly hematological toxicity (thrombocytopenia and neutropenia). One patient succumbed to septic shock.

Conclusion: Remobilization after previous autoHSCT is feasible only in a proportion of patients. Cytarabine is associated with the highest rate of successful mobilization and the highest yield of mobilized CD34+ cells. The toxicity requires careful surveillance of these patients.
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http://dx.doi.org/10.1002/jca.21885DOI Listing
June 2021

Efficacy and safety of oral panobinostat plus subcutaneous bortezomib and oral dexamethasone in patients with relapsed or relapsed and refractory multiple myeloma (PANORAMA 3): an open-label, randomised, phase 2 study.

Lancet Oncol 2021 01 7;22(1):142-154. Epub 2020 Dec 7.

Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, USA.

Background: Improved therapeutic options are needed for patients with relapsed or relapsed and refractory multiple myeloma. Subcutaneous bortezomib has replaced intravenous bortezomib as it is associated with a more favourable toxicity profile. We investigated the activity and safety of three different dosing regimens of oral panobinostat in combination with subcutaneous bortezomib and oral dexamethasone for this indication.

Methods: PANORAMA 3 is an open-label, randomised, phase 2 study being done at 71 sites (hospitals and medical centres) across 21 countries. Patients aged 18 years or older with relapsed or relapsed and refractory multiple myeloma (as per International Myeloma Working Group 2014 criteria), who had received one to four previous lines of therapy (including an immunomodulatory agent), and had an Eastern Cooperative Oncology Group performance status of 2 or lower, were randomly assigned (1:1:1) to receive oral panobinostat 20 mg three times weekly, 20 mg twice weekly, or 10 mg three times weekly, plus subcutaneous bortezomib and oral dexamethasone. All study drugs were administered in 21-day cycles. Randomisation was done by an interactive response technology provider, and stratified by number of previous treatment lines and age. The primary endpoint was overall response rate after up to eight treatment cycles (analysed in all randomly assigned patients by intention to treat). Safety analyses included all patients who received at least one dose of any study drug. No statistical comparisons between groups were planned. This trial is ongoing and registered with ClinicalTrials.gov, NCT02654990.

Findings: Between April 27, 2016, and Jan 17, 2019, 248 patients were randomly assigned (82 to panobinostat 20 mg three times weekly, 83 to panobinostat 20 mg twice weekly, and 83 to 10 mg panobinostat three times weekly). Median duration of follow-up across all treatment groups was 14·7 months (IQR 7·8-24·1). The overall response rate after up to eight treatment cycles was 62·2% (95% CI 50·8-72·7; 51 of 82 patients) for the 20 mg three times weekly group, 65·1% (53·8-75·2; 54 of 83 patients) for the 20 mg twice weekly group, and 50·6% (39·4-61·8; 42 of 83 patients) for the 10 mg three times weekly group. Grade 3-4 adverse events occurred in 71 (91%) of 78 patients in the 20 mg three times weekly group, 69 (83%) of 83 patients in the 20 mg twice weekly group, and 60 (75%) of 80 patients in the 10 mg three times weekly group; the most common (≥20% patients in any group) grade 3-4 adverse events were thrombocytopenia (33 [42%] of 78, 26 [31%] of 83, and 19 [24%] of 83 patients) and neutropenia (18 [23%], 13 [16%], and six [8%]). Serious adverse events occurred in 42 (54%) of 78 patients in the 20 mg three times weekly group, 40 (48%) of 83 patients in the 20 mg twice weekly group, and 35 (44%) of 83 patients in the 10 mg three times weekly group; the most common serious adverse event (≥10% patients in any group) was pneumonia (nine [12%] of 78, ten [12%] of 83, and nine [11%] of 80 patients). There were 14 deaths during the study (five [6%] of 78 patients in the 20 mg three times weekly group, three [4%] of 83 in the 20 mg twice weekly group, and six [8%] of 80 in the 10 mg three times weekly group); none of these deaths was deemed treatment related.

Interpretation: The safety profile of panobinostat 20 mg three times weekly was more favourable than in previous trials of this regimen with intravenous bortezomib, suggesting that subcutaneous bortezomib improves the tolerability of the panobinostat plus bortezomib plus dexamethasone regimen. The overall response rate was highest in the 20 mg three times weekly and 20 mg twice weekly groups, with 10 mg three times weekly best tolerated.

Funding: Novartis Pharmaceuticals and Secura Bio.
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http://dx.doi.org/10.1016/S1470-2045(20)30680-XDOI Listing
January 2021

In search of the correlation between nanomechanical and biomolecular properties of prostate cancer cells with different metastatic potential.

Arch Biochem Biophys 2021 01 6;697:108718. Epub 2020 Dec 6.

Institute of Nuclear Physics Polish Academy of Sciences, Krakow, PL-31342, Poland.

Nanomechanical properties of living cells, as measured with atomic force microscopy (AFM), are increasingly recognized as criteria that differentiate normal and pathologically altered cells. Locally measured cell elastic properties, described by the parameter known as Young's modulus, are currently proposed as a new diagnostic parameter that can be used at the early stage of cancer detection. In this study, local mechanical properties of normal human prostate (RWPE-1) cells and a range of malignant (22Rv1) and metastatic prostate cells (LNCaP, Du145 and PC3) were investigated. It was found that non-malignant prostate cells are stiffer than cancer cells while the metastatic cells are much softer than malignant cells from the primary tumor site. Next, the biochemical properties of the cells were measured using confocal Raman (RS) and Fourier-transform infrared (FT-IR) spectroscopies to reveal these cells' biochemical composition as malignant transformation proceeds. Nanomechanical and biochemical profiles of five different prostate cell lines were subsequently analyzed using partial least squares regression (PLSR) in order to identify which spectral features of the RS and FT-IR spectra correlate with the cell's elastic properties. The PLSR-based model could predict Young's modulus values based on both RS and FT-IR spectral information. These outcomes show not only that AFM, RS and FT-IR techniques can be used for discrimination between normal and cancer cells, but also that a linear correlation between mechanical response and biomolecular composition of the cells that undergo malignant transformation can be found. This knowledge broadens our understanding of how prostate cancer cells evolve thorough the multistep process of tumor pathogenesis.
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http://dx.doi.org/10.1016/j.abb.2020.108718DOI Listing
January 2021

The Antipsychotic D2AAK1 as a Memory Enhancer for Treatment of Mental and Neurodegenerative Diseases.

Int J Mol Sci 2020 Nov 23;21(22). Epub 2020 Nov 23.

Department of Synthesis and Chemical Technology of Pharmaceutical Substances with Computer Modeling Laboratory, Faculty of Pharmacy, Medical University of Lublin, 4A Chodzki St., 20-093 Lublin, Poland.

The treatment of memory impairments associated with the central nervous system diseases remains an unmet medical need with social and economic implications. Here we show, that a multi-target ligand of aminergic G protein-coupled receptors with antipsychotic activity in vivo (D2AAK1) stimulates neuron growth and survival and promotes neuron integrity. We focused on the multilevel evaluation of the D2AAK1-related effects on neurons in terms of behavioral, cellular, molecular, and biochemical features in vivo and in vitro, such as memory-related responses, locomotor activity, tissue sections analysis, metabolic activity, proliferation level, neurons morphology, and proteins level involved in intracellular signaling pathways. studies indicate that activation of calcium/calmodulin-dependent protein kinase I (CaMKI) may underline some of the observed activities of the compound. Furthermore, the compound increases hippocampal neuron proliferation via the activation of neurotrophic factors and cooperating signals responsible for cell growth and proliferation. D2AAK1 improves memory and learning processes in mice after both acute and chronic administration. D2AAK1 also causes an increase in the number of hippocampal pyramidal neurons after chronic administration. Because of its neuroprotective properties and pro-cognitive activity in behavioral studies D2AAK1 has the potential for the treatment of memory disturbances in neurodegenerative and mental diseases.
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http://dx.doi.org/10.3390/ijms21228849DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7700684PMC
November 2020

Influence of interference effects on the spectral quality and histological classification by FT-IR imaging in transflection geometry.

Analyst 2021 Jan 18;146(2):646-654. Epub 2020 Nov 18.

Solaris National Synchrotron Radiation Centre, Jagiellonian University, Czerwone Maki 98, 30-392 Krakow, Poland.

Infrared (IR) imaging can be used for fast, accurate and non-destructive pathology recognition of biopsies when supported by machine learning algorithms. Transflection mode of measurements has the potential to be translated into the clinic due to economic reasons of large-scale imaging with the need for inexpensive substrates. Unfortunately, in this mode spectral distortions originating from light interference appear. Due to this fact transmission measurement mode is more frequently used in pathology recognition. Nevertheless, this measurement mode also is not devoid of spectral distortion effects like scattering. However, this effect is better understood and there are preprocessing algorithms to minimize it. In this work, we investigated the influence of interference effects on spectral quality of pancreatic tissues measured in transmission and transflection mode with Fourier tranform IR (FT-IR) microscopy using samples embedded with and without paraffin. The removal of paraffin leads to an altered magnitude of interference in transflection and provides a platform for a detailed analysis of its effect on the spectra of biological material, since the same sample is measured with different interference conditions. Moreover, the potential of transflection mode measurements in histological classification of analyzed samples was investigated and compared with classification results for transmission mode.
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http://dx.doi.org/10.1039/d0an01565bDOI Listing
January 2021

Lymphocyte subsets in haematological patients with COVID-19: Multicentre prospective study.

Transl Oncol 2021 Jan 11;14(1):100943. Epub 2020 Nov 11.

Department and Clinic of Haematology, Blood Neoplasms, and Bone Marrow Transplantation, Wroclaw Medical University, Pasteura Street 4, 50-367 Wroclaw, Poland.

The role of immune dysregulation in the course and prognosis of COVID-19 is not clearly established. In particular, immune status in specific populations such as haematological patients, who have an impaired immunological system, has not been described so far. Here, we performed a comprehensive analysis of peripheral blood lymphocyte subsets in 27 SARS-CoV-2-infected patients, including 16 patients with haematological malignancies. We identified T cell subpopulations, B cells, NK cells and TCR α/ß and ɣ/ƍ-expressing T cells during COVID-19 infection, with significant changes observed in immune profiles during the course of disease, especially in haematological patients. We observed an increase in activated T lymphocytes (CD3+HLA-DR+ and CD3+CD8+HLA-DR+) in the early stages of SARS-CoV-2 infection with a concomitant decrease in the CD4/CD8 ratio in haematological patients compared to non-haematological patients affected by COVID-19. We also found a decrease in ɣ/ƍ T cells in both studied groups of patients, with lower numbers of CD25+ T cells and CD16+CD56+ NK cells in haematological patients compared to non-haematological patients with COVID-19. Our findings demonstrate, for the first time, impaired adaptive immunity in patients with haematological malignancies infected with COVID-19, resulting in impaired cellular immune responses to SARS-CoV-2. This warrants further investigation of this disease group in COVID-19 patient cohorts.
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http://dx.doi.org/10.1016/j.tranon.2020.100943DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7657028PMC
January 2021

Long-Term Infection Switches Gastric Epithelium Reprogramming Towards Cancer Stem Cell-Related Differentiation Program in -Activated Gastric Fibroblast-TGFβ Dependent Manner.

Microorganisms 2020 Oct 2;8(10). Epub 2020 Oct 2.

Department of Physiology, the Faculty of Medicine, Jagiellonian University Medical College, 31-531 Cracow, Poland.

(-induced inflammatory reaction leads to a persistent disturbance of gastric mucosa and chronic gastritis evidenced by deregulation of tissue self-renewal and local fibrosis with the crucial role of epithelial-mesenchymal transition (EMT) in this process. As we reported before, activated gastric fibroblasts into cells possessing cancer-associated fibroblast properties (CAFs), which secreted factors responsible for EMT process initiation in normal gastric epithelial RGM1 cells. Here, we showed that the long-term incubation of RGM1 cells in the presence of -activated gastric fibroblast (-AGF) secretome induced their shift towards plastic LGR5/Oct4/Sox-2/c-Myc/Klf4 phenotype (l.t.EMTRGM1 cells), while -non-infected gastric fibroblast (GF) secretome prompted a permanent epithelial-myofibroblast transition (EMyoT) of RGM1 cells favoring LGR/Oct4/Sox2/c-Myc/Klf4 phenotype (l.t.EMTRGM1 cells). TGFβ1 rich secretome from -reprogrammed fibroblasts prompted phenotypic plasticity and EMT of gastric epithelium, inducing pro-neoplastic expansion of post-EMT cells in the presence of low TGFβR1 and TGFβR2 activity. In turn, TGFβR1 activity along with GF-induced TGFβR2 activation in l.t.EMTRGM1 cells prompted their stromal phenotype. Collectively, our data show that infected and non-infected gastric fibroblast secretome induces alternative differentiation programs in gastric epithelium at least partially dependent on TGFβ signaling. infection-activated fibroblasts can switch gastric epithelium microevolution towards cancer stem cell-related differentiation program that can potentially initiate gastric neoplasm.
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http://dx.doi.org/10.3390/microorganisms8101519DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7599721PMC
October 2020

CD44 cells determine fenofibrate-induced microevolution of drug-resistance in prostate cancer cell populations.

Stem Cells 2020 Sep 27. Epub 2020 Sep 27.

Department of Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland.

Combinations of metabolic blockers (incl. fenofibrate) with chemotherapeutic drugs interfere with the drug-resistance of prostate cancer cells. However, their effect on cancer stem cells-dependent microevolution of prostate cancer malignancy remains unaddressed. Here, we hypothesize that the combined docetaxel/fenofibrate treatment prompts the selective expansion of cancer stem cells that affects the microevolution of their progenies. Accordingly, we adapted a combined in vitro/in vivo approach to identify biological and therapeutic consequences of this process. Minute subpopulations of docetaxel-resistant CD133 and/or CD44 cancer stem cell-like (SCL) cells were found in prostate cancer DU145 and PC3 cell populations. When pretreated with docetaxel, they readily differentiated into docetaxel-resistant CD44 "bulk" cells, thus accounting for the microevolution of drug-resistant cell lineages. Combined docetaxel/fenofibrate treatment induced the generation of poly(morpho)nuclear giant cells and drug-resistant CD44 SCL cells. However, the CD44 offspring of docetaxel- and docetaxel/fenofibrate-treated SCLs remained relatively sensitive to the combined treatment, while retaining enhanced resistance to docetaxel. Long-term propagation of drug-resistant SCL-derived lineages in the absence of docetaxel/fenofibrate resulted in their reverse microevolution toward the drug-sensitivity and invasive phenotype. Consequently, prostate tumors were able to recover from the combined docetaxel/fenofibrate stress after the initial arrest of their expansion in vivo. In conclusion, we have confirmed the potential of fenofibrate for the metronomic treatment of drug-resistant prostate tumors. However, docetaxel/fenofibrate-induced selective expansion of hyper-resistant CD44 SCL prostate cells and their "bulk" progenies prompts the microevolution of prostate tumor drug-resistance. This process can limit the implementation of metabolic chemotherapy in prostate cancer treatment.
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http://dx.doi.org/10.1002/stem.3281DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7756969PMC
September 2020

Platelet polyphosphate level is elevated in patients with chronic primary thrombocytopenia: A preliminary study.

Adv Clin Exp Med 2020 Sep;29(9):1051-1056

Department of Pharmaceutical Biochemistry, Wroclaw Medical University, Poland.

Background: Platelets are key players in hemostasis. These blood cells contain different types of granules. Recently, there has been a growing interest in the role of inorganic polyphosphate (polyP) structures stored in dense granules of platelets and secreted during platelet activation.

Objectives: To measure platelet polyP levels in patients with thrombocytopenia and thrombocythemia, and to examine the relationship of this indicator with platelet aggregation.

Material And Methods: The study included 36 patients with hematological disorders (26 with primary chronic thrombocytopenia and 10 with essential thrombocythemia (ET)) and 40 healthy subjects. Platelet reactivity was measured using whole blood impedance aggregometry. The polyP levels were isolated from lysed platelets, which were obtained from citrated platelet-rich plasma. The procedure included inactivating endogenous phosphatases, removing phosphate units derived from DNA and proteins, and finally hydrolyzing them into monophosphate units. A colorimetric assay using malachite green and ammonium molybdate was performed in order to quantify polyP levels.

Results: The polyP concentrations were significantly higher in the patients with thrombocytopenia than in the patients with thrombocythemia or the controls. The polyP level was not correlated with the level of aggregation.

Conclusions: The higher polyP levels observed in the patients with low platelet counts may indicate the existence of a compensatory mechanism that prevents excessive bleeding in such patients. Our study provides evidence of an essential role of polyP in platelet function and the coagulation process.
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http://dx.doi.org/10.17219/acem/125430DOI Listing
September 2020

Ibrutinib does not have clinically relevant interactions with oral contraceptives or substrates of CYP3A and CYP2B6.

Pharmacol Res Perspect 2020 10;8(5):e00649

Clinical Research Oncology, Janssen Research & Development LLC, Beerse, Belgium.

Ibrutinib may inhibit intestinal CYP3A4 and induce CYP2B6 and/or CYP3A. Secondary to potential induction, ibrutinib may reduce the exposure and effectiveness of oral contraceptives (OCs). This phase I study evaluated the effect of ibrutinib on the pharmacokinetics of the CYP2B6 substrate bupropion, CYP3A substrate midazolam, and OCs ethinylestradiol (EE) and levonorgestrel (LN). Female patients (N = 22) with B-cell malignancies received single doses of EE/LN (30/150 μg) and bupropion/midazolam (75/2 mg) during a pretreatment phase on days 1 and 3, respectively (before starting ibrutinib on day 8), and again after ibrutinib 560 mg/day for ≥ 2 weeks. Intestinal CYP3A inhibition was assessed on day 8 (single-dose ibrutinib plus single-dose midazolam). Systemic induction was assessed at steady-state on days 22 (EE/LN plus ibrutinib) and 24 (bupropion/midazolam plus ibrutinib). The geometric mean ratios (GMRs; test/reference) for maximum plasma concentration (C ) and area under the plasma concentration-time curve (AUC) were derived using linear mixed-effects models (90% confidence interval within 80%-125% indicated no interaction). On day 8, the GMR for midazolam exposure with ibrutinib coadministration was ≤ 20% lower than the reference, indicating lack of intestinal CYP3A4 inhibition. At ibrutinib steady-state, the C and AUC of EE were 33% higher than the reference, which was not considered clinically relevant. No substantial changes were noted for LN, midazolam, or bupropion. No unexpected safety findings were observed. A single dose of ibrutinib did not inhibit intestinal CYP3A4, and repeated administration did not induce CYP3A4/2B6, as assessed using EE, LN, midazolam, and bupropion.
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http://dx.doi.org/10.1002/prp2.649DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7506988PMC
October 2020

Macromolecular Orientation in Biological Tissues Using a Four-Polarization Method in FT-IR Imaging.

Anal Chem 2020 10 11;92(19):13313-13318. Epub 2020 Sep 11.

Solaris National Synchrotron Radiation Centre, Jagiellonian University, Czerwone Maki 98, Krakow 30-392, Poland.

Fourier transform infrared spectroscopy has emerged as a powerful tool for tissue specimen investigation. Its nondestructive and label-free character enables direct determination of biochemical composition of samples. Furthermore, the introduction of polarization enriches this technique by the possibility of molecular orientation study apart from purely quantitative analysis. Most of the molecular orientation studies focused on polymer samples with a well-defined molecular axis. Here, a four-polarization approach for Herman's in-plane orientation function and azimuthal angle determination was applied to a human tissue sample investigation for the first time. Attention was focused on fibrous tissues rich in collagen because of their cylindrical shape and established amide bond vibrations. Despite the fact that the tissue specimen contains a variety of molecules, the presented results of molecular ordering and orientation agree with the theoretical prediction based on sample composition and vibration directions.
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http://dx.doi.org/10.1021/acs.analchem.0c02591DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7547855PMC
October 2020

Physicochemical damage and early-stage biological response to X-ray radiation studied in prostate cancer cells by Raman spectroscopy.

J Biophotonics 2020 12 18;13(12):e202000252. Epub 2020 Sep 18.

Institute of Nuclear Physics, Polish Academy of Sciences, Krakow, Poland.

Exposure to ionizing radiation significantly affects biochemistry of cancer cells. The effect of irradiation can be divided into two stages, that is, the physicochemical stage and the biological response. Both effects induce different biochemical changes in the cells and should be analyzed as two separate phenomena. Thus, in the current study, Raman spectroscopy of prostate cancer cells fixed before (the physicochemical damage model) and just after (the biological response model) irradiation was undertaken to compare biochemical composition of irradiated cancer cells at both stages. Spectroscopic analysis of the cells was performed separately for cytoplasmic and nuclear regions. Biochemical changes of irradiated cells were analyzed using partial least squares regression (PLSR) method on the basis of the collected Raman spectra. Regression coefficients were therefore used to describe differences and similarities between biochemical composition of cancer cells undergoing the physicochemical stage and biological response. Additionally, PLSR models of both phenomena were compared for linear dose-dependence and a cross prediction.
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http://dx.doi.org/10.1002/jbio.202000252DOI Listing
December 2020

Nosocomial outbreak of SARS-CoV-2 infection in a haematological unit - High mortality rate in infected patients with haematologic malignancies.

J Clin Virol 2020 Sep 1;130:104574. Epub 2020 Aug 1.

Department and Clinic of Haematology, Blood Neoplasms, and Bone Marrow Transplantation, Wroclaw Medical University, Pasteura Street 4, 50-367, Poland. Electronic address:

Background: Here we report nosocomial outbreak of COVID-19 among patients in a haematological unit. To our knowledge this is the first report from Central Europe comparing morbidity and mortality in infected and non-infected patients after exposure to SARS-CoV-2.

Methods: The outbreak involved 39 individuals: 19 patients and 20 health care workers. The SARS-CoV-2 test by nasopharyngeal swabs was performed by real-time RT-PCR. Exposed patients were divided into two groups: quarantine patients with and without COVID-19. All patients were prospectively examined at the following time points: 0, 7 days, 14 days, 21 days and 28 days after confirmation or exclusion of SARS-CoV-2.

Results: Infection was confirmed in a total of 5/20 health care workers and 10/19 patients. Among the patients positive for SARS-CoV-2 infection, the mortality rate was 36.8 %. The probability of death in patients infected with SARS-CoV-2 increased 8-fold (p = 0.03). Bacterial, fungal, and viral co-infection significantly decreased survival in these patients (p < 0.05). Additionally, the probability of death was much higher in patients older than 40 years of age (p = 0.032).

Conclusion: This study showed significantly higher mortality rate in COVID-19 patients with haematologic diseases compared to the non-infected patient group. Haematologic patients with COVID-19 have 50 % less chance of survival.
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http://dx.doi.org/10.1016/j.jcv.2020.104574DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395815PMC
September 2020

Discovery of Novel Non-Steroidal Cytochrome P450 17A1 Inhibitors as Potential Prostate Cancer Agents.

Int J Mol Sci 2020 Jul 9;21(14). Epub 2020 Jul 9.

Department of Drug Design and Pharmacology, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen, Denmark.

The current study presents the design, synthesis, and evaluation of novel cytochrome P450 17A1 (CYP17A1) ligands. CYP17A1 is a key enzyme in the steroidogenic pathway that produces androgens among other steroids, and it is implicated in prostate cancer. The obtained compounds are potent enzyme inhibitors (sub µM) with antiproliferative activity in prostate cancer cell lines. The binding mode of these compounds is also discussed.
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http://dx.doi.org/10.3390/ijms21144868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7402352PMC
July 2020

Profound Nanoscale Structural and Biomechanical Changes in DNA Helix upon Treatment with Anthracycline Drugs.

Int J Mol Sci 2020 Jun 10;21(11). Epub 2020 Jun 10.

Department of Biomedical Engineering, Faculty of Fundamental Problems of Technology, Wroclaw University of Science and Technology, 27 Wybrzeze Wyspianskiego, 50-370 Wroclaw, Poland.

In our study, we describe the outcomes of the intercalation of different anthracycline antibiotics in double-stranded DNA at the nanoscale and single molecule level. Atomic force microscopy analysis revealed that intercalation results in significant elongation and thinning of dsDNA molecules. Additionally, using optical tweezers, we have shown that intercalation decreases the stiffness of DNA molecules, that results in greater susceptibility of dsDNA to break. Using DNA molecules with different GC/AT ratios, we checked whether anthracycline antibiotics show preference for GC-rich or AT-rich DNA fragments. We found that elongation, decrease in height and decrease in stiffness of dsDNA molecules was highest in GC-rich dsDNA, suggesting the preference of anthracycline antibiotics for GC pairs and GC-rich regions of DNA. This is important because such regions of genomes are enriched in DNA regulatory elements. By using three different anthracycline antibiotics, namely doxorubicin (DOX), epirubicin (EPI) and daunorubicin (DAU), we could compare their detrimental effects on DNA. Despite their analogical structure, anthracyclines differ in their effects on DNA molecules and GC-rich region preference. DOX had the strongest overall effect on the DNA topology, causing the largest elongation and decrease in height. On the other hand, EPI has the lowest preference for GC-rich dsDNA. Moreover, we demonstrated that the nanoscale perturbations in dsDNA topology are reflected by changes in the microscale properties of the cell, as even short exposition to doxorubicin resulted in an increase in nuclei stiffness, which can be due to aberration of the chromatin organization, upon intercalation of doxorubicin molecules.
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http://dx.doi.org/10.3390/ijms21114142DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7312087PMC
June 2020

Fecal microbiota transplantation in the treatment of intestinal steroid-resistant graft-versus-host disease: two case reports and a review of the literature.

J Int Med Res 2020 Jun;48(6):300060520925693

Department and Clinic of Haematology, Blood Neoplasms, and Bone Marrow Transplantation, Wroclaw Medical University, Wroclaw, Poland.

Acute graft-versus-host disease (aGvHD) reduces the efficiency and safety of allogeneic hematopoietic stem cell transplantation (allo-HSCT). In recent years, attempts have been made to transplant fecal microbiota from healthy donors to treat intestinal GvHD. This study presented two cases of patients undergoing allo-HSCT who were later selected for fecal microbiota transplantation (FMT). In the first patient, FMT resulted in the complete resolution of symptoms, whereas therapeutic efficacy was not achieved in the second patient. FMT eliminated drug-resistant pathogens, namely very drug-resistant spp., but not multidrug-resistant or spp. Further research is needed, particularly on the safety of FMT in patients with intestinal steroid-resistant GvHD and on the distant impact of transplanted microflora on the outcomes of allo-HSCT. FMT appears promising for the treatment of patients with steroid-resistant GvHD.
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http://dx.doi.org/10.1177/0300060520925693DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7294377PMC
June 2020

Lipid droplets in prostate cancer cells and effect of irradiation studied by Raman microspectroscopy.

Biochim Biophys Acta Mol Cell Biol Lipids 2020 09 3;1865(9):158753. Epub 2020 Jun 3.

Institute of Nuclear Physics Polish Academy of Sciences, PL-31342 Krakow, Poland.

Lipid droplets (LDs) are key organelles in cancer cells proliferation, growth, and response to stress. These nanometric structures can aggregate to reach the size of microns becoming important cell components. Although it is known that LDs contain various lipids, their chemical composition is still under investigation. Moreover, their function in cell's response to exogenous factors is also not fully understood. Raman spectroscopy, together with chemometrics, has been shown to be a powerful tool for analytical analyses of cancer cell components on the subcellular level. It provides the opportunity to analyse LDs in a label-free manner in live cells. In the current study, this method was applied to investigate LDs composition in untreated and irradiated with X-ray beams prostate cancer cells. Raman mapping technique proved lipids accumulation in PC-3 cells and allowed visualization of LDs spatial distribution in cytoplasm. A heterogeneous composition of LDs was revealed by detailed analysis of Raman spectra. Interestingly, PC-3 cells were found to accumulate either triacylglycerols or cholesteryl esters. Finally, effect of X-ray radiation on the cells was investigated using Raman spectroscopy and fluorescence staining. Significant influence of LDs in the process of cell response was confirmed and time dependence of this phenomenon was determined.
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http://dx.doi.org/10.1016/j.bbalip.2020.158753DOI Listing
September 2020

Tuning Down the Pain - An Overview of Allosteric Modulation of Opioid Receptors: Mechanisms of Modulation, Allosteric Sites, Modulator Syntheses.

Curr Top Med Chem 2020 ;20(31):2852-2865

Department of Synthesis and Chemical Technology of Pharmaceutical Substances with Computer Modelling Lab, Medical University of Lublin, Lublin, Poland.

Opioid signaling plays a central role in pain perception. As such, it remains the main target in the development of antinociceptive agents, despite serious side effects involved. In recent years, hopes for improved opioid painkillers are rising, together with our understanding of allosterism and biased signaling mechanisms. In this review, we focus on recently discovered allosteric modulators of opioid receptors, insights into phenomena underlying their action, as well as on how they extend our understanding of mechanisms of previously known compounds. A brief overlook of their synthesis is also presented.
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http://dx.doi.org/10.2174/1568026620666200601155451DOI Listing
January 2020

Epidermal Growth Factor (EGF) Augments the Invasive Potential of Human Glioblastoma Multiforme Cells via the Activation of Collaborative EGFR/ROS-Dependent Signaling.

Int J Mol Sci 2020 May 20;21(10). Epub 2020 May 20.

Department of Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Kraków, Poland.

Abnormal secretion of epidermal growth factor (EGF) by non-neuronal cells (e.g., glioma-associated microglia) establishes a feedback loop between glioblastoma multiforme (GBM) invasion and a functional disruption of brain tissue. Considering the postulated significance of this vicious circle for GBM progression, we scrutinized mechanisms of EGF-dependent pro-invasive signaling in terms of its interrelations with energy metabolism and reactive oxygen species (ROS) production. The effects of EGF on the invasiveness of human glioblastoma T98G cells were estimated using time-lapse video microscopy, immunocytochemistry, cell cycle assay, immunoblot analyses, and Transwell® assay. These techniques were followed by quantification of the effect of EGFR (Epidermal Growth Factor Receptor) and ROS inhibitors on the EGF-induced T98G invasiveness and intracellular ROS, ATP, and lactate levels and mitochondrial metabolism. The EGF remarkably augmented the proliferation and motility of the T98G cells. Responses of these cells were accompanied by cellular rear-front polarization, translocation of vinculin to the leading lamellae, and increased promptness of penetration of micropore barriers. Erlotinib (the EGFR inhibitor) significantly attenuated the EGF-induced T98G invasiveness and metabolic reprogramming of the T98G cells, otherwise illustrated by the increased mitochondrial activity, glycolysis, and ROS production in the EGF-treated cells. In turn, ROS inhibition by N-acetyl-L-cysteine (NAC) had no effect on T98G morphology, but considerably attenuated EGF-induced cell motility. Our data confirmed the EGFR/ROS-dependent pro-neoplastic and pro-invasive activity of EGF in human GBM. These EGF effects may depend on metabolic reprogramming of GBM cells and are executed by alternative ROS-dependent/-independent pathways. The EGF may thus preserve bioenergetic homeostasis of GBM cells in hypoxic regions of brain tissue.
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http://dx.doi.org/10.3390/ijms21103605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7279139PMC
May 2020

Translation of an esophagus histopathological FT-IR imaging model to a fast quantum cascade laser modality.

J Biophotonics 2020 08 1;13(8):e202000122. Epub 2020 Jun 1.

Solaris National Synchrotron Radiation Centre, Jagiellonian University, Krakow, Poland.

The technical progress in fast quantum cascade laser (QCL) microscopy offers a platform where chemical imaging becomes feasible for clinical diagnostics. QCL systems allow the integration of previously developed FT-IR-based pathology recognition models in a faster workflow. The translation of such models requires a systematic approach, focusing only on the spectral frequencies that carry crucial information for discrimination of pathologic features. In this study, we optimize an FT-IR-based histopathological method for esophageal cancer detection to work with a QCL system. We explore whether the classifier's performance is affected by paraffin presence from tissue blocks compared to removing it chemically. Working with paraffin-embedded samples reduces preprocessing time in the lab and allows samples to be archived after analysis. Moreover, we test, whether the creation of a QCL model requires a preestablished FTIR model or can be optimized using solely QCL measurements.
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http://dx.doi.org/10.1002/jbio.202000122DOI Listing
August 2020