Publications by authors named "Anna Żaczek"

64 Publications

Novel Isoniazid-Carborane Hybrids Active in Vitro Against .

Pharmaceuticals (Basel) 2020 Dec 15;13(12). Epub 2020 Dec 15.

Institute of Medical Biology, Polish Academy of Sciences, 106 Lodowa St., 93-232 Lodz, Poland.

Tuberculosis (TB) is a severe infectious disease with high mortality and morbidity. The emergence of drug-resistant TB has increased the challenge to eliminate this disease. Isoniazid (INH) remains the key and effective component in the therapeutic regimen recommended by World Health Organization (WHO). A series of isoniazid-carborane derivatives containing 1,2-dicarba--dodecaborane, 1,7-dicarba--dodecaborane, 1,12-dicarba--dodecaborane, or 7,8-dicarba--undecaborate anion were synthesized for the first time. The compounds were tested against the () H37Rv strain and its mutant (D) defective in the synthesis of catalase-peroxidase (KatG). '-((7,8-dicarba--undecaboranyl)methylidene)isonicotinohydrazide () showed the highest activity against the wild-type strain. All hybrids could inhibit the growth of the Δ mutant in lower concentrations than INH. '-([(1,12-dicarba--dodecaboran-1yl)ethyl)isonicotinohydrazide () exhibited more than 60-fold increase in activity against D as compared to INH. This compound was also found to be noncytotoxic up to a concentration four times higher than the minimum inhibitory concentration 99% (MIC) value.
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http://dx.doi.org/10.3390/ph13120465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7765321PMC
December 2020

Functional Disassociation Between the Protein Domains of MSMEG_4305 of () .

Front Microbiol 2020 19;11:2008. Epub 2020 Aug 19.

Laboratory of Genetics and Physiology of Mycobacterium, Institute of Medical Biology, Polish Academy of Sciences, łLódź, Poland.

MSMEG_4305 is a two-domain protein of () (). The N-terminal domain of MSMEG_4305 encodes an RNase H type I. The C-terminal domain is a presumed CobC, predicted to be involved in the aerobic synthesis of vitamin B12. Both domains reach their maximum at distinct pH, approximately 8.5 and 4.5, respectively. The presence of the CobC domain influenced RNase activity in homolog Rv2228c. Here, we analyzed the role of MSMEG_4305 in vitamin B12 synthesis and the functional association between both domains in . We used knock-out mutant of , deficient in MSMEG_4305. Whole-cell lysates of the mutants strain contained a lower concentration of vitamin B12, as it determined with immunoenzimatic assay. We observed growth deficits, related to vitamin B12 production, on media containing sulfamethazine and propionate. Removal of the CobC domain of MSMEG_4305 in Δ background hardly affected the growth rate of . The strain carrying truncation showed no fitness deficit in the competitive assay and it did not show increased level of RNA/DNA hybrids in its genome. We show that homologs of MSMEG_4305 are present only in the phylogenetic branch (according to the old classification system). The domains of MSMEG_4305 homologs accumulate mutations at a different rate, while the linker region is highly variable. We conclude that MSMEG_4305 is a multidomain protein that most probably was fixed in the phylogenetic tree of life due to genetic drift.
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http://dx.doi.org/10.3389/fmicb.2020.02008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7466739PMC
August 2020

EGFR as a stable marker of prostate cancer dissemination to bones.

Br J Cancer 2020 Dec 9;123(12):1767-1774. Epub 2020 Sep 9.

Laboratory of Translational Oncology, Institute of Medical Biotechnology and Experimental Oncology, Medical University of Gdańsk, Gdańsk, Poland.

Background: Prostate cancer (PCa) is among the most commonly diagnosed malignancies in men. Although 5-year survival in patients with localised disease reaches nearly 100%, metastatic disease still remains incurable. Therefore, there is a need for markers indicating metastatic dissemination.

Methods: EGFR overexpression (EGFR) was tracked in 1039 primary tumours, circulating tumour cells from 39 d'Amico high-risk patients and metastatic samples from 21 castration-resistant PCa cases. EGFR status was compared to clinical parameters and multiple molecular factors were assessed using immunohistochemistry and gene ontology analysis. The functional aspect of EGFR was evaluated by plating PC-3 cells on soft and rigid matrices.

Results: EGFR was found in 14% of primary tumours, where it was associated with shorter metastasis-free survival and was an independent indicator of worse overall survival. EGFR correlated with a pro-migratory and pro-metastatic phenotype of tumour cells as well as rich collagen fibre content. All circulating tumour cells (detected in 13% of cases) were positive for EGFR, independent of their EMT-related phenotype. EGFR was more prevalent in castration-resistant bone metastases (29% of patients) and supported growth of human PCa cells on rigid matrices mimicking bone stiffness.

Conclusions: EGFR is a stable, EMT-independent marker of PCa disseminating to rigid organs, preferentially bones.
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http://dx.doi.org/10.1038/s41416-020-01052-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7722745PMC
December 2020

microRNA Expression Profile in Single Hormone Receptor-Positive Breast Cancers is Mainly Dependent on HER2 Status-A Pilot Study.

Diagnostics (Basel) 2020 Aug 20;10(9). Epub 2020 Aug 20.

Department of Oncology and Radiotherapy, Medical University of Gdansk, 80-214 Gdansk, Poland.

Estrogen (ER) and progesterone (PgR) receptors and HER2 are crucial in the assessment of breast cancer specimens due to their prognostic and predictive significance. Single hormone receptor-positive breast cancers are less common and their clinical course is less favorable than ER(+)/PgR(+) tumors. Their molecular features, especially microRNA (miRNA) profiles, have not been investigated to date. Tumor specimens from 36 chemonaive breast cancer patients with known ER and PgR status (18 ER(+)/PgR(-) and 18 ER(-)/PgR(+) cases) were enrolled to the study. The expression of 829 miRNAs was evaluated with nCounter Human v3 miRNA expression Assay (NanoString). miRNAs differentiating between ER/PgR/HER2 phenotypes were selected based on fold change (FC) calculated for the mean normalized counts of each probe in compared groups. The differences were estimated with Student's -test or Two-Way ANOVA (considering also the HER2 status). The results were validated using The Cancer Genome Atlas (TCGA) dataset. Following quality control of raw data, fourcases were excluded due to low sample quality, leaving 14 ER(+)/PgR(-) and 18 ER(-)/PgR(+) cases. After correction for multiple comparisons, we did not find miRNA signature differentiating between ER(-)/PgR(+) and ER(+)/PgR(-) breast cancers. However, a trend for differing expression (-value ≤ 0.05; FDR > 0.2; ANOVA) in eight miRNAs was observed. The ER(+)/PgR(-) group demonstrated elevated levels of four miRNAs-miR-30a-5p, miR-29c-3p, miR-141-3p and miR-423-5p-while the ER(-)/PgR(+) tumors were enriched in another four miRNAs-miR-514b-5p, miR-424-5p, miR-495-3p, and miR-92a-3p. For one of the miRNAs-miR-29c-3p-the association with the ER(+)/PgR(-) phenotype was confirmed in the TCGA cohort (-value = 0.024; -test). HER2 amplification/overexpression in the NanoString cohort was related to significant differences observed in 33 miRNA expression levels (FDR ≤ 0.2; ANOVA). The association with HER2 status was confirmed in the TCGA cohort for four miRNAs (miR-1180-3p, miR-223-3p, miR-30d-5p, and miR-195-5p). The main differences in miRNA expression amongst single hormone receptor-positive tumors were identified according to their HER2 status. However, ER(+)/PgR(-) cases tended to express higher levels of miRNAs associated with ER-positivity (miR-30a-5p, miR-29c-3p, miR-141-3p), whereas ER(-)/PgR(+) cancers showed elevated levels of miRNAs characteristic for double- and triple-negative tumors (miR-92a-3p, miR-424-5p). Further studies are necessary to comprehensively analyze miRNA signatures characteristic of ER(-)/PgR(+) and ER(+)/PgR(-) tumors.
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http://dx.doi.org/10.3390/diagnostics10090617DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7555149PMC
August 2020

Circulating Tumor Cells in Early and Advanced Breast Cancer; Biology and Prognostic Value.

Int J Mol Sci 2020 Feb 29;21(5). Epub 2020 Feb 29.

Department of Molecular and Translational Oncology, Maria Sklodowska-Curie National Research Institute of Oncology, Roentgena 5, 02-781 Warsaw, Poland.

Breast cancer metastasis is the leading cause of cancer deaths in women and is difficult to combat due to the long periods in which disseminated cells retain a potential to be re-activated and start the relapse. Assessing the number and molecular profile of circulating tumor cells (CTCs) in breast cancer patients, especially in early breast cancer, should help in identifying the possibility of relapse in time for therapeutic intervention to prevent or delay recurrence. While metastatic breast cancer is considered incurable, molecular analysis of CTCs still have a potential to define particular susceptibilities of the cells representing the current tumor burden, which may differ considerably from the cells of the primary tumor, and offer more tailored therapy to the patients. In this review we inspect the routes to metastasis and how they can be linked to specific features of CTCs, how CTC analysis may be used in therapy, and what is the current status of the research and efforts to include CTC analysis in clinical practice.
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http://dx.doi.org/10.3390/ijms21051671DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7084781PMC
February 2020

The Prognostic Significance of Eukaryotic Translation Initiation Factors (eIFs) in Endometrial Cancer.

Int J Mol Sci 2019 Dec 6;20(24). Epub 2019 Dec 6.

Area 2 Cancer, Center for Biomarker Research in Medicine, Stiftingtalstraße 5, 8010 Graz, Austria.

Whilst the role of eukaryotic translation initiation factors (eIFs) has already been investigated in several human cancers, their role in endometrial cancer (EC) is relatively unknown. In the present retrospective study, 279 patients with EC (1180 samples) were included (mean age: 63.0 years, mean follow-up: 6.1 years). Samples were analysed for expression of 7 eIFs subunits (eIF2α, eIF3c, eIF3h, eIF4e, eIF4g, eIF5, eIF6) through immunohistochemistry and western blotting. Fifteen samples of healthy endometrium served as controls. Density and intensity were assessed and mean combined scores (CS) calculated for each patient. Upon immunohistochemistry, median eIF5 CS were significantly higher in EC as compared with non-neoplastic tissue (NNT, < 0.001), whilst median eIF6 CS were significantly lower in EC ( < 0.001). Moreover, eIF5 ( = 0.002), eIF6 ( = 0.032) and eIF4g CS ( = 0.014) were significantly different when comparing NNT with EC grading types. Median eIF4g CS was higher in type II EC ( = 0.034). Upon western blot analysis, eIF4g ( < 0.001), peIF2α ( < 0.001) and eIF3h ( < 0.05) were significantly overexpressed in EC, while expression of eIF3c was significantly reduced in EC as compared with NNT ( < 0.001). The remaining eIFs were non-significant. Besides tumour stage ( < 0.001) and patient's age ( < 0.001), high eIF4g CS-levels were independently associated with poor prognosis (HR: 1.604, 95%CI: 1.037-2.483, = 0.034). The other eIFs had no prognostic significance. Notably, the independent prognostic significance of eIF4g was lost when adding tumour type. Considering the difficulties in differentiating EC type I and II, eIF4g may serve as a novel prognostic marker indicating patient outcome.
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http://dx.doi.org/10.3390/ijms20246169DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6941158PMC
December 2019

NF-kappa B Signaling-Related Signatures Are Connected with the Mesenchymal Phenotype of Circulating Tumor Cells in Non-Metastatic Breast Cancer.

Cancers (Basel) 2019 Dec 6;11(12). Epub 2019 Dec 6.

Laboratory of Translational Oncology, Intercollegiate Faculty of Biotechnology, Medical University of Gdansk, 80-211 Gdansk, Poland.

The role of circulating tumor cells (CTCs), tumor microenvironment (TME), and the immune system in the formation of metastasis is evident, yet the details of their interactions remain unknown. This study aimed at exploring the immunotranscriptome of primary tumors associated with the status of CTCs in breast cancer (BCa) patients. The expression of 730 immune-related genes in formalin-fixed paraffin-embedded samples was analyzed using the multigenomic NanoString technology and correlated with the presence and the phenotype of CTCs. Upregulation of 37 genes and downregulation of 1 gene were observed in patients characterized by a mesenchymal phenotype of CTCs when compared to patients with epithelial CTCs. The upregulated genes were involved in NF-kappa B signaling and in the production of type I interferons. The clinical significance of the differentially expressed genes was evaluated using The Cancer Genome Atlas (TCGA) data of a breast invasive carcinoma (BRCA) cohort. Five of the upregulated genes-, , , and -were independent prognostic factors in terms of overall and disease-free survival. To conclude, our data identify a group of genes that are upregulated in BCa patients with mesenchymal CTCs and reveal their prognostic potential, thus indicating that they merit further investigation.
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http://dx.doi.org/10.3390/cancers11121961DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6966426PMC
December 2019

Liquid biopsy for minimally invasive heart transplant monitoring: a pilot study.

J Clin Pathol 2020 Aug 5;73(8):507-510. Epub 2019 Dec 5.

Department of Cardiac & Vascular Surgery, Faculty of Medicine, Medical University of Gdańsk, Gdańsk, Poland.

Background: Heart transplantation allows for a long-term management of patients with end-stage heart failure. After the surgery, organ rejection is monitored with endomyocardial biopsy, which is an invasive, but not always informative procedure. Therefore, there is a pressing need for a new, safe, yet reliable, diagnostic method. Here, we present a pilot study confronting liquid biopsy based on donor-specific cell-free DNA with the protocol endomyocardial biopsy.

Methods: The study was performed on 21 blood samples matched with endomyocardial biopsy (graded according to acute cellular rejection scale) from nine patients after heart transplantation. Genotyping was performed on genomic DNA from donors and recipients for 10 single-nucleotide polymorphisms (SNPs). Cell-free DNA isolated from plasma was analysed with digital droplet PCR to detect donor-specific alleles.

Results: From 21 analysed endomyocardial biopsies, 4 were graded as 0R and 17 as 1R. Liquid biopsy was successfully performed in each sample for all informative SNPs (median of 3 per patient). We observed a high homogeneity of the results between SNPs in each sample (interclass correlation coefficient of >0.9).

Conclusions: There is a undeniable need for an alternative, non-invasive diagnostic procedure of early transplant rejection and investigation of donor-derived cell-free DNA seems to be the promising choice. The very high sensitivity is particularly enticing to consider liquid biopsy as a potential screening tool. Its minimal invasiveness may allow for more frequent examination and, thus, tighter monitoring. The reliable assessment of its clinical utility requires an adequately powered and properly designed multicentre study.
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http://dx.doi.org/10.1136/jclinpath-2019-205926DOI Listing
August 2020

Sensitive detection of caspase-3 enzymatic activities and inhibitor screening by mass spectrometry with dual maleimide labelling quantitation.

Analyst 2019 Nov;144(22):6751-6759

Institute of Functional Nano and Soft Materials (FUNSOM), Jiangsu Key Laboratory for Carbon-Based Functional Materials and Devices, Joint International Research Laboratory of Carbon-Based Materials and Devices, Soochow University, Suzhou, Jiangsu 215123, China.

There is a great need to develop sensitive and specific methods for quantitative analysis of caspase-3 activities in cell apoptosis. Herein, we report a new method for sensitive detection of caspase-3 enzyme activities and inhibitor screening based on dual maleimide (DuMal) labeling quantitation and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Evaluation of caspase-3 activities is performed using MS analysis of the enzymatic product of the peptide probe, which fuses a caspase-3 cleavable peptide segment (DEVD) and a quantifiable "ID tag" (a peptide segment of FRGLRGFKC labeled by maleimide). The DuMal labeling technique features non-isotopic tagging, rapid reactions, and reproducible quantitation. We have achieved quantitative analysis of the enzyme activities with a limit of detection (LOD) and limit of quantitation (LOQ) of caspase-3 down to 0.11 nM and 0.29 nM respectively and a proof-of-concept demonstration of its inhibitor screening. Our method has further been tested for caspase-3 activities in a Parkinson's disease cellular model, suggesting a useful tool for protease activity research.
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http://dx.doi.org/10.1039/c9an01458fDOI Listing
November 2019

High-grade non-small cell lung carcinoma: a comparative analysis of the phenotypic profile in small biopsies with the corresponding postoperative material.

Pol J Pathol 2019 ;70(2):100-108

Department of Pathomorphology, Medical University of Gdañsk, Poland.

The most recent classification of the lung cancer expanded the diagnostic criteria of its histological subtypes and included its immunophenotypic profile. We performed the study to compare the reliability of selected markers in high-grade non-small cell lung carcinoma (NSCLC) in the oligobiopsies with the matched postoperative samples. We evaluated expression of p40, p63, TTF1, cytokeratin 5/6, cytokeratin 7, napsin A, desmoglein 3, desmocollin 3 and mucin secretion as detected by mucicarmine staining. The study cohort included 123 cases of poorly-differentiated NSCLC. The tissue oligobiopsy material was available in 38 cases. Tissue microarrays (TMAs) from all postoperative cases were constructed. Comparing the immunophenotype between postsurgical samples and oligobiopsies we found an almost perfect agreement for most of performed IHC reactions. The highest concordance of results was found for desmoglein 3, CK7, and p40, whereas the lowest - for desmocollin 3. Immunoprofile of the oligobiopsies corresponded well to that in the resection specimens. The most useful markers in poorly differentiated ADs are: TTF1 and napsin A, and for non-keratinizing SCCs: p40, p63, CK5/6 and desmoglein 3.
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http://dx.doi.org/10.5114/pjp.2019.84087DOI Listing
October 2019

Low Numbers of Vascular Vessels Correlate to Progression in Hormone-Naïve Prostate Carcinomas Undergoing Radical Prostatectomy.

Cancers (Basel) 2019 Sep 12;11(9). Epub 2019 Sep 12.

Laboratory of Cell Biology, Department of Medical Biotechnology, Medical University of Gdańsk, Gdańsk 80-211, Poland.

Vascularization influences tumor development by supporting the nutrition and dissemination of tumor cells. On the other hand, a low number of vascular vessels (VV) may induce hypoxia, accounting for selection of resistant clone(s) of tumor cells. This study aimed to evaluate the prognostic significance of vascular (VV) and lymphatic vessels (LV) in prostate cancer (PCa). Tumor samples from 400 PCa patients undergoing radical prostatectomy (RP) were prepared in duplex as tissue microarrays. Numbers of VV and LV were evaluated using immunohistochemistry detecting CD34 and podoplanin, respectively, and correlated to clinical data, biochemical recurrence (BR), and proteins analyzed in tumor cells. VV and LV were found in 32% and 43% of patients with informative PCa samples, respectively. VV correlated with a shorter time to BR 3, 5, and 10 years after RP in hormone-naïve patients ( = 0.028, = 0.027 and = 0.056, respectively). It was also shown to be an independent prognostic factor 5 years after surgery (multivariate analysis, = 0.046). Tumors characterized by VV expressed the epithelial cell adhesion molecule, EpCAM, less frequently ( = 0.016) and revealed a borderline correlation to increased levels of tumor cell invasion marker Loxl-2 ( = 0.059). No correlations were found for LV. In summary, VV in hormone-naïve patients undergoing RP has prognostic potential and seems to be related to an aggressive phenotype of tumor cells.
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http://dx.doi.org/10.3390/cancers11091356DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6770894PMC
September 2019

Clinical and Biological Significance of Gene Alteration and Estrogen Receptors Isoforms Expression in Breast Cancer Patients.

Int J Mol Sci 2019 Apr 16;20(8). Epub 2019 Apr 16.

Department of Medical Biotechnology, Intercollegiate Faculty of Biotechnology, University of Gdansk and Medical University of Gdansk, 80-211 Gdansk, Poland.

The amplification of estrogen receptor alpha (ERα) encoded by the gene has been described as having a prognostic role in breast cancer patients. However, increased dosage of the gene (tested by real-time PCR) is also observed in ER-negative breast cancers, which might suggest the expression of alternative isoforms of ERα (other than classical ERα of 66 kDa). In the current work, we have investigated the gene dosage in 402 primary breast cancer patients as well as the expression of ERα isoforms-ERα66 and ERα36-on mRNA and protein levels. The obtained results were correlated with clinicopathological data of the patients. Results showed that increased gene dosage is not related to gene amplification measured by fluorescent in situ hybridization (FISH), but it correlates with the decreased expression of isoform ( = 0.01). Interestingly, the short ER isoform was expressed in samples with increased gene dosage, suggesting that genomic aberration might influence the expression of that particular isoform. Similarly to increased gene dosage, high expression was linked with the decreased disease-free survival of the patients ( = 0.05), which was independent of the status of the classical level in breast tumors.
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http://dx.doi.org/10.3390/ijms20081881DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6514554PMC
April 2019

Spectrum of Epithelial-Mesenchymal Transition Phenotypes in Circulating Tumour Cells from Early Breast Cancer Patients.

Cancers (Basel) 2019 Jan 9;11(1). Epub 2019 Jan 9.

Department of Medical Biotechnology, Intercollegiate Faculty of Biotechnology of the University of Gdansk and Medical University of Gdansk, 80-211 Gdańsk, Poland.

Circulating tumour cells (CTCs) can provide valuable prognostic information in a number of epithelial cancers. However, their detection is hampered due to their molecular heterogeneity, which can be induced by the epithelial-mesenchymal transition (EMT) process. Therefore, current knowledge about CTCs from clinical samples is often limited due to an inability to isolate wide spectrum of CTCs phenotypes. In the current work, we aimed at isolation and molecular characterization of CTCs with different EMT status in order to establish their clinical significance in early breast cancer patients. We have obtained CTCs-enriched blood fraction from 83 breast cancer patients in which we have tested the expression of epithelial, mesenchymal and general breast cancer CTCs markers (), cancer stem cell markers (, , , , ) and cluster formation gene (plakoglobin). We have shown that in the CTCs-positive patients, epithelial, epithelial-mesenchymal and mesenchymal CTCs markers were detected at a similar rate (in 28%, 24% and 24%, respectively). Mesenchymal CTCs were characterized by the most aggressive phenotype (significantly higher expression of , , , , < 0.05 for all), presence of lymph node metastases ( = ), larger tumour size () and 7.33 higher risk of death in the multivariate analysis (95% CI 1.06⁻50.41, ). Epithelial-mesenchymal subtype, believed to correspond to highly plastic and aggressive state, did not show significant impact on survival. Gene expression profile of samples with epithelial-mesenchymal CTCs group resembled pure epithelial or pure mesenchymal phenotypes, possibly underlining degree of EMT activation in particular patient's sample. Molecular profiling of CTCs EMT phenotype provides more detailed and clinically informative results, proving the role of EMT in malignant cancer progression in early breast cancer.
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http://dx.doi.org/10.3390/cancers11010059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6356662PMC
January 2019

The NnaR orphan response regulator is essential for the utilization of nitrate and nitrite as sole nitrogen sources in mycobacteria.

Sci Rep 2018 12 3;8(1):17552. Epub 2018 Dec 3.

Institute for Medical Biology, Polish Academy of Sciences, Łódź, Poland.

Nitrogen is an essential component of biological molecules and an indispensable microelement required for the growth of cells. Nitrogen metabolism of Mycobacterium smegmatis is regulated by a number of transcription factors, with the glnR gene product playing a major role. Under nitrogen-depletion conditions, GlnR controls the expression of many genes involved in nitrogen assimilation, including the msmeg_0432 gene encoding NnaR, the homologue of a nitrite/nitrate transport regulator from Streptomyces coelicolor. In the present study, the role of NnaR in the nitrogen metabolism of M. smegmatis was evaluated. The ∆glnR and ∆nnaR mutant strains were generated and cultured under nitrogen-depletion conditions. Total RNA profiling was used to investigate the potential role of NnaR in the GlnR regulon under nitrogen-depletion and in nitrogen-rich media. We found that disruption of MSMEG_0432 affected the expression of genes involved in nitrite/nitrate uptake, and its removal rendered mycobacteria unable to assimilate nitrogen from those sources, leading to cell death. RNA-Seq results were validated using quantitative real-time polymerase chain reaction (qRT-PCR) and electrophoretic mobility shift assays (EMSAs). The ability of mutants to grow on various nitrogen sources was evaluated using the BIOLOG Phenotype screening platform and confirmed on minimal Sauton's medium containing various sources of nitrogen. The ∆glnR mutant was not able to convert nitrates to nitrites. Interestingly, NnaR required active GlnR to prevent nitrogen starvation, and both proteins cooperated in the regulation of gene expression associated with nitrate/nitrite assimilation. The ∆nnaR mutant was able to convert nitrates to nitrites, but it could not assimilate the products of this conversion. Importantly, NnaR was the key regulator of the expression of the truncated haemoglobin trHbN, which is required to improve the survival of bacteria under nitrosative stress.
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http://dx.doi.org/10.1038/s41598-018-35844-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6277429PMC
December 2018

Characterization of Mutations Conferring Resistance to Rifampin in Mycobacterium tuberculosis Clinical Strains.

Antimicrob Agents Chemother 2018 10 24;62(10). Epub 2018 Sep 24.

Institute of Medical Biology, Polish Academy of Sciences, Łódź, Poland

Resistance of to rifampin (RMP), mediated by mutations in the gene coding for the beta-subunit of RNA polymerase, poses a serious threat to the efficacy of clinical management and, thus, control programs for tuberculosis (TB). The contribution of many individual mutations to the development and level of RMP resistance remains elusive. In this study, the incidence of mutations throughout the gene among 115 clinical isolates, both resistant and susceptible to RMP, was determined. Of the newly discovered mutations, the role of three substitutions in the causation of RMP resistance was empirically tested. The results from mutagenesis experiments were combined with the assessment of the prevalence of mutations, and their reciprocal co-occurrences, across global populations. Twenty-two different types of mutations in the gene were identified and distributed among 58 (89.2%) RMP-resistant strains. The MICs of RMP were within the range of 40 to 800 mg/liter, with MIC and MIC values of 400 and 800 mg/liter, respectively. None of the mutations (Gln429His, Met434Ile, and Arg827Cys) inspected for their role in the development of RMP resistance produced an RMP-resistant phenotype in isogenic H37Rv strain-derived mutants. These mutations are supposed to compensate for fitness impairment incurred by other mutations directly associated with drug resistance.
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http://dx.doi.org/10.1128/AAC.01093-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6153850PMC
October 2018

Aggressive Phenotype of Cells Disseminated via Hematogenous and Lymphatic Route in Breast Cancer Patients.

Transl Oncol 2018 Jun 16;11(3):722-731. Epub 2018 Apr 16.

Intercollegiate Faculty of Biotechnology, University of Gdansk and Medical University of Gdansk, Poland. Electronic address:

Intratumoral heterogeneity of breast cancer remains a major challenge in successful treatment. Failure of cancer therapies can also be accredited to inability to systemically eradicate cancer stem cells (CSCs). Recent evidence points to the role of epithelial-mesenchymal transition (EMT) in expanding the pool of tumor cells with CSCs features. Thus, we assessed expression level as well as heterogeneity of CSCs markers in primary tumors (PT), lymph node metastasis (LNM), and circulating tumor cells (CTCs)-enriched blood fractions in order to correlate them with signs of EMT activation as well as clinicopathological data of breast cancer patients. Level of CSCs markers (ALDH1, CD44, CD133, OCT-4, NANOG) and EMT markers was quantified in PT (N=107), LNM (N=56), and CTCs-enriched blood fractions (N=85). Heterogeneity of CSCs markers expression within each PT and LNM was assessed by calculating Gini Index. Percentage of ALDH1-positive cells was elevated in PT in comparison to LNM (P = .005). However, heterogeneity of the four CSCs markers: ALDH1 (P = .019), CD133 (P = .009), OCT-4 (P = .027), and CD44 (P < .001) was decreased in LNM. Samples classified as mesenchymal (post-EMT) showed elevated expression of CSCs markers (OCT-4 and CD44 in PT; OCT-4 in LNM; ALDH1, OCT-4, NANOG, CD44 in CTCs). Patients with mesenchymal-like CTCs had worse prognosis than patients with epithelial-like or no CTCs (P = .0025). CSCs markers are enriched in PT, LNM, and CTCs with mesenchymal features, but their heterogeneity is decreased in metastatic lymph nodes. Mesenchymal CTCs phenotype correlates with poor prognosis of the patients.
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http://dx.doi.org/10.1016/j.tranon.2018.03.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6056759PMC
June 2018

MiR-192 and miR-662 enhance chemoresistance and invasiveness of squamous cell lung carcinoma.

Lung Cancer 2018 04 6;118:111-118. Epub 2018 Feb 6.

Laboratory of Cell Biology, Department of Medical Biotechnology, Intercollegiate Faculty of Biotechnology of University of Gdańsk and Medical University of Gdańsk, ul. Dębinki 1, 80-211 Gdańsk, Poland. Electronic address:

Objectives: Overexpression of miR-192, miR-192* and miR-662 was previously found to correlate with poor prognosis of early-stage squamous cell lung cancer (SCC) patients. In this study, we investigated the relevance of these miRNAs to cancer cell biology and chemoresistance.

Materials And Methods: MiRNA expression profile was analysed in 10 non-small cell lung cancer (NSCLC) cell lines using RT-qPCR. H520 and H1703 cells were transfected with miRNA inhibitors (anti-miR-192, -192* and -662) for functional studies. Chemoresistance to cisplatin and etoposide was evaluated using MTT colorimetric assay. H520 cells were subjected to 3D soft-agar colony formation assay and H1703 cells to wound healing assay. Whole transcriptome analysis was used to assess the effect of miR-192 and miR-662 inhibition on gene expression.

Results: SCC cell lines, H520 and H1703, differed in miRNA expression and phenotypic features. MiR-192 and miR-662 inhibition decreased clonogenicity and motility of SCC cells. MiR-192 and miR-662 inhibition sensitized SCC cells to etoposide but not to cisplatin. Whole transcriptome analysis revealed genes regulated by miR-192 and miR-662 in SCC, relevant to maintaining chemoresistance, invasiveness, epithelial-mesenchymal transition (EMT) and immune evasion.

Conclusions: We showed for the first time that miR-192 and miR-662 have functional role in SCC cells. Our findings suggest that targeting these miRNAs may impact both chemoresistance and invasiveness of SCC, and add to the evidence linking these aspects of tumour biology. Overexpression of miR-192 and miR-662 might be useful as a marker of resistance to etoposide.
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http://dx.doi.org/10.1016/j.lungcan.2018.02.002DOI Listing
April 2018

Molecular typing of Mycobacterium kansasii using pulsed-field gel electrophoresis and a newly designed variable-number tandem repeat analysis.

Sci Rep 2018 03 13;8(1):4462. Epub 2018 Mar 13.

Department of Applied Microbiology, Institute of Microbiology, Faculty of Biology, University of Warsaw, Warsaw, Poland.

Molecular epidemiological studies of Mycobacterium kansasii are hampered by the lack of highly-discriminatory genotyping modalities. The purpose of this study was to design a new, high-resolution fingerprinting method for M. kansasii. Complete genome sequence of the M. kansasii ATCC 12478 reference strain was searched for satellite-like repetitive DNA elements comprising tandem repeats. A total of 24 variable-number tandem repeat (VNTR) loci were identified with potential discriminatory capacity. Of these, 17 were used to study polymorphism among 67 M. kansasii strains representing six subtypes (I-VI). The results of VNTR typing were compared with those of pulsed-field gel electrophoresis (PFGE) with AsnI digestion. Six VNTRs i.e. (VNTR 1, 2, 8, 14, 20 and 23) allow to differentiate analyzed strains with the same discriminatory capacities as use of a 17-loci panel. VNTR typing and PFGE in conjunction revealed 45 distinct patterns, including 11 clusters with 33 isolates and 34 unique patterns. The Hunter-Gaston's discriminatory index was 0.95 and 0.66 for PFGE and VNTR typing respectively, and 0.97 for the two methods combined. In conclusion, this study delivers a new typing scheme, based on VNTR polymorphism, and recommends it as a first-line test prior to PFGE analysis in a two-step typing strategy for M. kansasii.
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http://dx.doi.org/10.1038/s41598-018-21562-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5849605PMC
March 2018

Sensitive Detection of Single-Cell Secreted HO by Integrating a Microfluidic Droplet Sensor and Au Nanoclusters.

Anal Chem 2018 04 14;90(7):4478-4484. Epub 2018 Mar 14.

Institute of Functional Nano and Soft Materials (FUNSOM), Jiangsu Key Laboratory for Carbon-Based Functional Materials and Devices , Soochow University , 199 Ren Ai Road, Suzhou Industrial Park , Suzhou , Jiangsu 215123 , China.

As an important signaling molecule, hydrogen peroxide (HO) secreted externally by the cells influences cell migration, immunity generation, and cellular communications. Herein, we have developed a microfluidic approach with droplets in combination with Au nanoclusters for the sensitive detection of HO secreted by a single cell. Isolated in the ultrasmall volume (4.2 nL) of a microdroplet, single-cell secreted HO can initiate dramatic fluorescence changes of horseradish peroxidase-Au nanoclusters. We have demonstrated an ultrahigh sensitivity (200-400 attomole HO directly measured from a single cell) with good specificity. It offers a useful research tool to study the cell-to-cell differences of HO secretion at the single-cell level.
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http://dx.doi.org/10.1021/acs.analchem.7b04798DOI Listing
April 2018

RSK1 promotes murine breast cancer growth and metastasis.

Folia Histochem Cytobiol 2018 2;56(1):11-20. Epub 2018 Mar 2.

Department of Molecular Enzymology, Intercollegiate Faculty of Biotechnology, University of Gdansk and Medical University of Gdansk, Poland.

Introduction: Triple-negative breast cancer (TNBC), representing over 15% of all breast cancers, has a poorer prognosis than other subtypes. There is no effective targeted treatment available for the TNBC sufferers. Ribosomal S6 kinases (RSKs) have been previously proposed as drug targets for TNBC based on observations that 85% of these tumors express activated RSKs.

Materials And Methods: Herein we examined an involvement of RSK1 (p90 ribosomal S6 kinase 1) in a regulation of TNBC growth and metastatic spread in an animal model, which closely imitates human disease. Mice were inoculated into mammary fat pad with 4T1 cells or their RSK1-depleted variant. We examined tumor growth and formation of pulmonary metastasis. Boyden chamber, wound healing and soft agarose assays were performed to evaluate cells invasion, migration and anchorage-independent growth.

Results: We found that RSK1 promoted tumor growth and metastasis in vivo. After 35 days all animals inoculated with control cells developed tumors while in the group injected with RSK1-negative cells, there were 75% tumor-bearing mice. Average tumor mass was estimated as 1.16 g and 0.37 g for RSK1-positive vs. -negative samples, respectively (p < 0.0001). Quantification of the macroscopic pulmonary metastases indicated that mice with RSK1-negative tumors developed approximately 85% less metastatic foci on the lung surface (p < 0.001). This has been supported by in vitro data presenting that RSK1 promoted anchorage-independent cell growth and migration. Moreover, RSK1 knock-down corresponded with decreased expression of cell cycle regulating proteins, i.e. cyclin D3, CDK6 and CDK4.

Conclusions: We provide evidence that RSK1 supports tumor growth and metastatic spread in vivo as well as in vitro migration and survival in non-adherent conditions. Further studies of RSK1 involvement in TNBC progression may substantiate our findings, laying the foundations for development of anti-RSK1-based therapeutic strategies in the management of patients with TNBC.
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http://dx.doi.org/10.5603/FHC.a2018.0001DOI Listing
November 2018

PdtaS Deficiency Affects Resistance of Mycobacteria to Ribosome Targeting Antibiotics.

Front Microbiol 2017 3;8:2145. Epub 2017 Nov 3.

Institute for Medical Biology, Polish Academy of Sciences, Łódź, Poland.

Two-component regulatory systems (TCSSs) are key regulatory elements responsible for the adaptation of bacteria to environmental stresses. A classical TCSS is typically comprised of a sensory histidine kinase and a corresponding response regulator. Here, we used homologous recombination to construct a mutant defective in the synthesis of cytosolic histidine kinase PdtaS (Msmeg_1918). The resulting Δ mutant strain was tested in the Phenotype Microarray screening system, which allowed us to identify aminoglycoside antibiotic sensitivity, tetracyclines antibiotic resistance as well as membrane transport and respiration, as the main processes affected by removal of . The antibiotic sensitivity profiles were confirmed by survival assessment and complementation studies. To gain insight into the molecular mechanisms responsible for the observed phenotype, we compared ribosomal RNA and protein profiles of the mutant and wild-type strains. We carried out Northern blotting and qRT-PCR to compare rRNA levels and analyzed ribosome sedimentation patterns of the wild-type and mutant strains on sucrose gradients. Isolated ribosomes were further used to estimate relative abundance of individual proteins in the ribosomal subunits using label free mass spectrometry analysis. Additionally, the Δ mutant revealed lower activity of the respiratory chain as measured by the rate of TTC (triphenyltetrazolium chloride) reduction, while at the same time showing only insignificant changes in the uptake of aminoglycosides. We postulate that deficiency of PdtaS affects the oxidative respiration rates and ribosomal composition causing relevant changes to intrinsic resistance or susceptibility to antibiotics targeting ribosomes, which are commonly used to treat mycobacterial infections.
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http://dx.doi.org/10.3389/fmicb.2017.02145DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5676007PMC
November 2017

The Landscape of Circulating Tumor Cell Research in the Context of Epithelial-Mesenchymal Transition.

Pathobiology 2017 14;84(5):264-283. Epub 2017 Oct 14.

Department of Medical Biotechnology, Medical University of Gdansk, Gdansk, Poland.

The metastatic spread of cancer accounts for the vast majority of cancer-related deaths. It is mediated by tumor cells circulating in blood (called circulating tumor cells, CTCs), which escaped from their established niches. CTCs give a unique opportunity to look into the metastatic cascade and to study the molecular processes supporting the spread of tumor cells throughout the body. As current therapies are not sufficiently effective in treating metastatic disease, it is important to determine cellular and molecular features of cancer cells that "seed" new tumors in distant organs at early stages. In this review we focus on the role of the epithelial-mesenchymal transition program in providing a survival advantage to metastasizing tumor cells, especially CTCs, and put it in the context of clinical findings.
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http://dx.doi.org/10.1159/000477812DOI Listing
July 2018

Keratin 7 expression in lymph node metastases but not in the primary tumour correlates with distant metastases and poor prognosis in colon carcinoma.

Pol J Pathol 2016;67(3):228-234

Department of Pathomorphology, Medical University of Gdańsk, Gdańsk, Poland.

Colorectal carcinoma (CRC) is one of the leading causes of cancer-related deaths worldwide. Alterations in keratin expression, including keratin 7 (K7), are frequent findings in multiple cancers, and they constitute a prognostic factor. The aim of our study was to evaluate the prognostic significance of K7 in the primary tumour and lymph node metastases in two separate cohorts of patients: the first one with lymph node involvement (LN+, 129 cases) and the second one free of LN metastases (LN-, 85 cases). Keratin 7 expression in CRC was analysed on tissue microarrays with immunohistochemistry and evaluated using the h-score. In the LN+ group K7 positivity was identified in 7/129 (5.4%) of primary tumours (PT) and lymph node metastases (LNM); concordance between them was 94% ( 0.396). Keratin 7 was expressed in 8/85 cases (9.4%) in the LN- group. K7 expression in LNM of the LN+ cohort correlated with shorter overall survival (OS) (p = 0.047) and presence of distant metastases at diagnosis (p = 0.005). Expression of K7 in the primary tumour in both cohorts did not correlate with survival. We conclude that the status of K7 expression in metastatic lymph nodes from CRC is a poor prognostic factor.
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http://dx.doi.org/10.5114/pjp.2016.63774DOI Listing
June 2017

Fibroblast growth factor signalling induces loss of progesterone receptor in breast cancer cells.

Oncotarget 2016 Dec;7(52):86011-86025

Department of Molecular Enzymology, Intercollegiate Faculty of Biotechnology, University of Gdansk and Medical University of Gdansk, Poland.

We have recently demonstrated that, fibroblast growth factor 2 (FGFR2), signalling via ribosomal S6 kinase 2 (RSK2), promotes progression of breast cancer (BCa). Loss of progesterone receptor (PR), whose activity in BCa cells can be stimulated by growth factor receptors (GFRs), is associated with poor patient outcome. Here we showed that FGF7/FGFR2 triggered phosphorylation of PR at Ser294, PR ubiquitination and subsequent receptor`s degradation via the 26S proteasome pathway in BCa cells. We further demonstrated that RSK2 mediated FGF7/FGFR2-induced PR downregulation. In addition, a strong synergistic effect of FGF7 and progesterone (Pg), reflected in the enhanced anchorage-independent growth and cell migration, was observed. Analysis of clinical material demonstrated that expression of PR inversely correlated with activated RSK (RSK-P) (p = 0.016). Patients with RSK-P(+)/PR(-) tumours had 3.629-fold higher risk of recurrence (p = 0.002), when compared with the rest of the cohort. Moreover, RSK-P(+)/PR(-) phenotype was shown as an independent prognostic factor (p = 0.006). These results indicate that the FGF7/FGFR2-RSK2 axis promotes PR turnover and activity, which may sensitize BCa cells to stromal stimuli and contribute to the progression toward steroid hormone negative BCa.
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http://dx.doi.org/10.18632/oncotarget.13322DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5349893PMC
December 2016

Interactions between FGFR2 and RSK2-implications for breast cancer prognosis.

Tumour Biol 2016 Oct 30;37(10):13721-13731. Epub 2016 Jul 30.

Department of Molecular Enzymology, Intercollegiate Faculty of Biotechnology, University of Gdańsk and Medical University of Gdańsk, 80-210, Gdansk, Poland.

We have previously demonstrated that fibroblast growth factor receptor 2 (FGFR2) activates ribosomal s6 kinase 2 (RSK2) in mammary epithelial cells and that this pathway promotes in vitro cell growth and migration. Potential clinical significance of FGFR2 and RSK2 association has never been investigated. Herein, we have undertaken an evaluation of a possible relationship between FGFR2/RSK2 interdependence and disease outcome in breast cancer (BCa) patients. The clinical analysis was complemented by an in vitro investigation of an involvement of RSK2 in the regulation of FGFR2 function. Primary tumour samples from 152 stage I-III BCa patients were examined for FGFR2 and RSK2 gene and protein expression. FGFR2 showed a positive correlation with RSK2 at both protein (p = 0.003) and messenger RNA (mRNA) (p = 0.001) levels. Lack of both FGFR2 and activated RSK (RSK-P) significantly correlated with better disease-free survival (DFS) (p = 0.01). Patients with tumours displaying immunoreactivity for either or both FGFR2 and RSK-P had 4.89-fold higher risk of recurrence when compared to the FGFR2/RSK-P-negative subgroup. FGFR2-RSK2 interactions were verified by co-immunoprecipitation and internalization assays in HB2 mammary epithelial cell line (characterized by high endogenous FGFR2 and RSK2 expression). In vitro analyses revealed that FGFR2 and RSK2 formed an indirect complex and that activated RSK exerted a significant impact on fibroblast growth factor 2 (FGF2)-triggered internalization of FGFR2. Our results suggest that the FGFR2-RSK2 signalling pathway is involved in pathophysiology of BCa and evaluation of FGFR2/RSK-P expression may be useful in disease prognostication.
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http://dx.doi.org/10.1007/s13277-016-5266-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5097089PMC
October 2016

Rak błony śluzowej trzonu macicy u chorych po przebytej chorobie nowotworowej - aspekty kliniczne i molekularne.

Ginekol Pol 2016 ;87(2):88-93

Katedra i Klinika Ginekologii, Ginekologii Onkologicznej i Endokrynologii Ginekologicznej Gdańskiego Uniwersytetu Medycznego, Kliniczna 1a, 80-402 Gdańsk, Polska.

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http://dx.doi.org/10.17772/gp/60559DOI Listing
July 2018

Methodological and Clinical Aspects of the Molecular Epidemiology of Mycobacterium tuberculosis and Other Mycobacteria.

Clin Microbiol Rev 2016 Apr;29(2):239-90

Mycobacterium Genetics and Physiology Unit, Institute of Medical Biology, Polish Academy of Sciences, Lódź, Poland.

Molecular typing has revolutionized epidemiological studies of infectious diseases, including those of a mycobacterial etiology. With the advent of fingerprinting techniques, many traditional concepts regarding transmission, infectivity, or pathogenicity of mycobacterial bacilli have been revisited, and their conventional interpretations have been challenged. Since the mid-1990s, when the first typing methods were introduced, a plethora of other modalities have been proposed. So-called molecular epidemiology has become an essential subdiscipline of modern mycobacteriology. It serves as a resource for understanding the key issues in the epidemiology of tuberculosis and other mycobacterial diseases. Among these issues are disclosing sources of infection, quantifying recent transmission, identifying transmission links, discerning reinfection from relapse, tracking the geographic distribution and clonal expansion of specific strains, and exploring the genetic mechanisms underlying specific phenotypic traits, including virulence, organ tropism, transmissibility, or drug resistance. Since genotyping continues to unravel the biology of mycobacteria, it offers enormous promise in the fight against and prevention of the diseases caused by these pathogens. In this review, molecular typing methods for Mycobacterium tuberculosis and nontuberculous mycobacteria elaborated over the last 2 decades are summarized. The relevance of these methods to the epidemiological investigation, diagnosis, evolution, and control of mycobacterial diseases is discussed.
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http://dx.doi.org/10.1128/CMR.00055-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4786889PMC
April 2016

Immunohistochemical characterisation of molecular subtypes in endometrial cancer.

Int J Clin Exp Med 2015 15;8(11):21981-90. Epub 2015 Nov 15.

Department of Medical Biotechnology, Intercollegiate Faculty of Biotechnology, University of Gdańsk and Medical University of Gdańsk Dębinki 1, 80-211 Gdańsk, Poland.

Four molecular subtypes have lately been established in endometrial cancer basing on estrogen receptor (ER), progesterone receptor (PR) and HER2 status: ER+/PR+/HER2+, ER+/PR+/HER2-, ER-/PR-/HER2+ and ER-/PR-/HER2-. The subtypes have shown diversity in terms of prognosis, clinicopathological and molecular characteristics, with ER+/PR+/HER2- and ER-/PR-/HER2+ group exhibiting exceptionally benign and aggressive behavior, respectively. We have further characterized the subtypes in the context of pathways known to drive endometrial carcinogenesis: phosphatidylinositol 3-kinase (PI3K)-AKT pathway (ERBB/PI3K pathway), TP53 system, and the mismatch repair (MMR) mechanism. Analysis of tumor heterogeneity was also included. ER+/PR+/HER2+ was characterized by active ERBB/PI3K pathway occurring in 58% of cases. Subtype ER-/PR-/HER2+ was characterized by the most frequent TP53 mutations (83% of cases). Triple negative phenotype utterly lacked active ERBB/PI3K pathway. Analyzed major pathways rarely correlated with clinicopathologial data but mutated TP53 and retained MMR did correlate with shorter overall survival (both P<0.01). The presence of tumor heterogeneity was most frequent in ER-/PR-/HER2+ subtype (53% of all cases). The presented results further emphasize that the molecular subtype distinction, along with MMR and TP53 status, could be a useful diagnostic tool in guiding individualized therapy.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4724016PMC
February 2016

HOTAIR in relation to epithelial-mesenchymal transition and cancer stem cells in molecular subtypes of endometrial cancer.

Int J Biol Markers 2016 Jul 30;31(3):e245-51. Epub 2016 Jul 30.

Department of Medical Biotechnology, Intercollegiate Faculty of Biotechnology, University of Gdańsk and Medical University of Gdańsk, Gdańsk - Poland.

Background: Endometrial cancer (EC) is a hormone-related disease, showing highly diverse features of ER/PR/HER2 status-based molecular subtypes. Long noncoding RNA (lncRNA) HOX antisense intergenic RNA (HOTAIR) has recently emerged as a key molecule in many cancers, triggering epithelial-mesenchymal transition (EMT)-mediated cancer stem cell (CSC) formation, but little is known about its significance in EC. Thus, we aimed to investigate the clinical significance of HOTAIR itself in different molecular subtypes of EC and possible links between HOTAIR, EMT and CSC-related markers.

Methods: The study group included 156 consecutive, stage I-IV EC patients treated between 2000 and 2010. ER, PR and HER2 protein expression were examined by immunohistochemistry (IHC) on tissue microarrays. RT-qPCR was used to analyze the expression levels of HOTAIR, EMT-related genes - SNAIL and SLUG - and the CSCs marker CD133.

Results: Molecular subtypes, defined as ER/PR+HER2+, ER/PR+HER2-, ER-PR-HER2+ and ER-PR-HER2-, occurred in 40.2%, 52.3%, 4.7% and 1.9% of cases, respectively. The expression of HOTAIR did not differ between the subtypes, but high HOTAIR expression correlated with shorter overall survival (p = 0.04) in the entire group. The expression levels of HOTAIR, SNAIL, SLUG and CD133 were similar in defined EC molecular subtypes.

Conclusions: Our data do not confirm the role of HOTAIR in EMT-mediated CSC formation in EC. Neither does the diversity of EC molecular subtypes influence these processes. But HOTAIR expression could serve as an independent prognostic factor in EC. The clinical importance of the above discoveries requires further studies.
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http://dx.doi.org/10.5301/jbm.5000187DOI Listing
July 2016

Epidemiological Analysis of Mycobacterium tuberculosis Strains Isolated from Patients of Small Communities Living in the South-East of Poland.

Pol J Microbiol 2015 ;64(3):289-93

The diversity of Mycobacterium tuberculosis clinical isolates, collected from a single hospital, was analyzed by ligation-mediated PCR techniques: FLiP and FLAP, and hybridization technique, IS6110-RFLP. The isolated strains were divided in terms of location (3 towns of Podkarpackie voivodeship differing in population size) and relationship (8 members of 4 families, each represented by 2 patients). Within each family identical DNA profiles, as well as drug resistance patterns were identified indicating a great chance of transmission of strains within the same family. Identical, or very similar patterns were also shared by strains isolated from unrelated patients living in a very small town (1 200 inhabitants) or hospitalized in the same place and time.
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February 2016