Chelators as Antineuroblastomas Agents.

Neuroblastoma represents 8-10 % of all malignant tumors in childhood and is responsible for 15 % of cancer deaths in the pediatric population. Aggressive neuroblastomas are often resistant to chemotherapy. Canonically, neuroblastomas can be classified according to the MYCN (N-myc proto-oncogene protein) gene amplification, a common marker of tumor aggressiveness and poor prognosis. It has been found that certain compounds with chelating properties may show anticancer activity, but there is little evidence for the effect of chelators on neuroblastoma. The effect of new chelators characterized by the same functional group, designated as HLZ (1-hydrazino phthalazine), on proliferation (WST-1 and methylene blue assay), cell cycle (flow cytometry), apoptosis (proliferation assay after use of specific pharmacological inhibitors and western blot analysis) and ROS production (fluorometric assay based on dichlorofluorescein diacetate metabolism) was studied in three neuroblastoma cell lines with different levels of MYCN amplification. The molecules were effective only on MYCN-non-amplified cells in which they arrested the cell cycle in the G0/G1 phase. We investigated the mechanism of action and identified the activation of cell signaling that involves protein kinase C.

Pilot study of gene mutations associated with Lynch syndrome in Slovak patients with breast cancer.

BACKGROUND: Lynch syndrome (LS) is an autosomal dominant inherited disorder which causes an increased risk of cancer, especially colorectal and endometrial carcinomas. Recent studies have shown an association between LS and breast cancer as well. The aim of our study is to highlight the possible presence of mutations in genes associated with LS in patients with breast cancer and the need to include the examination of Lynch-associated genes in patients with a family history of breast cancer as well as in patients with recurrent breast cancer, as well as with the occurrence of other Lynch-associated cancer. MATERIALS AND METHODS: We analyzed tumor tissue samples from 78 patients with primary breast cancer. Our samples were tested with a gene panel associated with the risk of developing breast cancer, while in our study we focused primarily on the occurrence of mutations in mismatch-repair genes. DNA isolated from tumor tissue was sequenced using next generation sequencing (NGS) and analyzed using the Ingenuity Variant Analysis tool. To confirm the germline mutation, we examined the patient's blood sample using NGS sequencing. RESULTS: As a result of our analysis, we managed to identify a mutation in the PMS2 gene in one patient's breast tumor tissue. The presence of this mutation indicates that the resulting cancer may be a consequence of LS. As for pathogenicity, this was probably a pathogenic variant, as we detected deletions in the exon region, which led to frameshift mutation. Moreover, we also identified single-nucleotide pathogenic variants in the TP53 and PIK3CA genes. To definitively establish the diagnosis of LS in the patient, we examined a blood sample, where we also identified a mutation of the PMS2 gene. CONCLUSION: LS is underdiagnosed in many Lynch-associated cancers. However, in the case of a familial occurrence of breast cancer and other Lynch-associated genes, it is important to think about a possible diagnosis of LS and, if the patient meets the diagnostic criteria, to carry out a genetic examination of Lynch-associated genes.

PTEN mutations as predictive marker for the high-grade endometrial cancer development in slovak women.

Endometrial carcinoma (ECa) is one of the most common neoplasia of the female genital tract. The phosphatase and tensin (PTEN) homolog is the most frequently mutated tumor suppressor gene in endometrial carcinoma. PTEN encodes a phosphatase, a key regulatory enzyme involved in a signal transduction pathway that regulates cell growth, migration and apoptosis. The study evaluates an association between the morphological appearance of endometrial hyperplasia and ECa, and the presence of PTEN variations, PTEN protein´s level and intracellular localization. A total of 67 archived formalin-fixed and paraffin-embedded human biopsy tissue specimens with normal proliferative and secretory endometrium, endometrial hyperplasia without atypia and endometrial atypical hyperplasia, endometrioid the grade G1 and G3 and serous subtype of ECa were evaluated by sequencing for the presence of mutations in coding regions of PTEN gene of endometrial epithelial cells. The PTEN gene expression and intercellular localization of PTEN protein were evaluated immunohistochemically by immunoreactive score (IRS). PTEN mutation spectrum in endometrial carcinoma was identified for Slovak population. 28 non-silent mutations were identified in PTEN, twelve of them were novel, not annotated in Catalogue of Somatic Mutations in Cancer. Higher frequency of PTEN mutations was observed in serous carcinoma compared to global average. No correlation was observed between samples´ IRS, PTEN cellular localization and identified mutations. PTEN sequencing can be beneficial for patients considering prognosis of disease and sensitivity to treatment.

Importance of the genetics in the diagnostics of hydatidiform mole.

OBJECTIVE: To summarize the possibilities of the genetic analysis of hydatidiform moles and point out its perspectives in the diagnostics of this disease. DESIGN: Review. SETTING: Institute of Medical Biology, Genetics and Clinical Genetics, Faculty of Medicine, Comenius University in Bratislava, Slovak Republic. METHODS: Analysis of published literature data from the internet databases PubMed, ScienceDirect, Scopus and printed literature from the period 1963-2019. RESULTS: This review refers on karyotyping, flow cytometry, FISH (Fluorescent in Situ Hybridization), VNTR-RFLP analysis (Variable Number of Tandem Repeats-Restriction Fragment Length Polymorphism), VNTR-PCR analysis (Variable Number of Tandem Repeats-Polymerase Chain Reaction) and STR (Short Tandem Repeat) genotyping of hydatidiform moles. The article summarizes possible application of these methods in the differential diagnostics of molar pregnancy (partial and complete hydatidiform moles) and nonmolar hydropic abortions. CONCLUSION: Genetic analyses offer precise identification of types of molar pregnancies when histopathological diagnosis is not clear during early stages of pathology.

Schiff base Cu(II) complexes as inhibitors of proteasome in human cancer cells.

BACKGROUND: It has been demonstrated that proteasome inhibitors might be potential anticancer drugs. The copper complexes can be used as specific proteasome inhibitors in tumor cells able to induce apoptosis by the ubiquitin-proteasome pathway. The goal of our study was to test the cytotoxic and proteasome inhibitory effects of five Schiff base Cu(II) complexes - [Cu2(sal-D,L-glu)2(isoquinoline)2] . 2C2H5OH (1), [Cu(sal-5-met-L-glu)(H2O)].H2O (2), [Cu(ethanol)2(imidazole)4][Cu2(sal-D,L-glu)2(imidazole)2] (3), [Cu(sal-D,L-glu)(2-methylimidazole)] (4) on human lung carcinoma cells A549, cervix carcinoma cells HeLa and glioblastoma cells U-118MG. MATERIAL AND METHODS: For the cytotoxic analysis we used MTT test and for monitoring the proteasome inhibition western blot analysis. RESULTS: We have observed different cytotoxic effects of tested complexes on human cancer cells depending on the ligand present in their structure. Cu(II) complexes 4 and 5 were the most effective against A549 cells; all complexes were cytotoxic against HeLa cells and the complex 4 was the most effective against U-118MG. Moreover, we have detected the inhibition of the proteasome activity in human cancer cells A549 by Cu(II) complexes 1, 2 and 4 at IC50 concentration. CONCLUSION: Results of our study suggest that isoquinoline- and imidazole-based copper complexes could be used as inhibitors of the proteasome system in cancer cells A549 (Tab. 1, Fig. 1, Ref. 26).

ERVW-1 gene polymorphisms related to preeclampsia.

OBJECTIVES: Identification of genetic association between the gene ERVW-1 and preeclampsia. BACKGROUND: Preeclampsia is a multifactorial disease affecting women during pregnancy and it is one of the main causes of perinatal and maternal morbidity and mortality. The pathophysiology of preeclampsia is very complex and several aspects of the disease have not been elucidated yet. Abnormal placentation frequently occurs during severe preeclampsia. Protein syncytin 1, a product of the ERVW-1 gene, plays a crucial role in the syncytiotrophoblast differentiation and optimal placentation. The syncytin 1 expression is disturbed during preeclampsia. The main focus of this study was the analysis of the ERVW-1 regulatory regions and identification of DNA polymorphisms associated with preeclamptic cases in Slovak population. METHODS: Regulatory region of gene ERVW-1 was analyzed by sequencing to identify genetic variants. RESULTS: We identified four DNA variants, namely rs4727276, rs148592540, rs569899772 and rs555416193, in samples of Slovak population. CONCLUSION: No relation between polymorphisms and preeclampsia was observed, indicating that further investigations with a larger sampling are still required. However, our work represents new original approach in genetic differential diagnosis of preeclampsia with possible useful findings in the future (Tab. 3, Fig. 1, Ref. 34).

Heme oxygenase is not involved in the anti-proliferative effects of statins on pancreatic cancer cells.

BACKGROUND: Pancreatic cancer is recognized as one of the most fatal tumors due to its aggressiveness and resistance to therapy. Statins were previously shown to inhibit the proliferation of cancer cells via various signaling pathways. In healthy tissues, statins activate the heme oxygenase pathway, nevertheless the role of heme oxygenase in pancreatic cancer is still controversial. The aim of this study was to evaluate, whether anti-proliferative effects of statins in pancreatic cancer cells are mediated via the heme oxygenase pathway. METHODS: In vitro effects of various statins and hemin, a heme oxygenase inducer, on cell proliferation were evaluated in PA-TU-8902, MiaPaCa-2 and BxPC-3 human pancreatic cancer cell lines. The effect of statins on heme oxygenase activity was assessed and heme oxygenase-silenced cells were used for pancreatic cancer cell proliferation studies. Cell death rate and reactive oxygen species production were measured in PA-TU-8902 cells, followed by evaluation of the effect of cerivastatin on GFP-K-Ras trafficking and expression of markers of invasiveness, osteopontin (SPP1) and SOX2. RESULTS: While simvastatin and cerivastatin displayed major anti-proliferative properties in all cell lines tested, pravastatin did not affect the cell growth at all. Strong anti-proliferative effect was observed also for hemin. Co-treatment of cerivastatin and hemin increased anti-proliferative potential of these agents, via increased production of reactive oxygen species and cell death compared to individual treatment. Heme oxygenase silencing did not prevent pancreatic cancer cells from the tumor-suppressive effect of cerivastatin or hemin. Cerivastatin, but not pravastatin, protected Ras protein from trafficking to the cell membrane and significantly reduced expressions of SPP1 (p < 0.05) and SOX2 (p < 0.01). CONCLUSIONS: Anti-proliferative effects of statins and hemin on human pancreatic cancer cell lines do not seem to be related to the heme oxygenase pathway. While hemin triggers reactive oxygen species-induced cell death, cerivastatin targets Ras protein trafficking and affects markers of invasiveness.

PTEN sequence analysis in endometrial hyperplasia and endometrial carcinoma in Slovak women.

Phosphatase and tensin homolog (PTEN) is a protein that acts as a tumor suppressor by dephosphorylating the lipid second messenger phosphatidylinositol 3,4,5-trisphosphate. Loss of PTEN function has been implicated in the pathogenesis of a number of different tumors, particularly endometrial carcinoma (ECa). ECa is the most common neoplasia of the female genital tract. Our study evaluates an association between the morphological appearance of endometrial hyperplasia and endometrial carcinoma and the degree of PTEN alterations. A total of 45 endometrial biopsies from Slovak women were included in present study. Formalin-fixed and paraffin-embedded tissue samples with simple hyperplasia (3), complex hyperplasia (5), atypical complex hyperplasia (7), endometrioid carcinomas G1 (20) and G3 (5), and serous carcinoma (5) were evaluated for the presence of mutations in coding regions of PTEN gene, the most frequently mutated tumor suppressor gene in endometrial carcinoma. 75% of the detected mutations were clustered in exons 5 and 8. Out of the 39 mutations detected in 24 cases, 20 were frameshifts and 19 were nonsense, missense, or silent mutations. Some specimens harboured more than one mutation. The results of current study on Slovak women were compared to a previous study performed on Polish population. The two sets of results were similar.