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Purpose: Cell authentication is a necessary procedure to avoid scientific data from cell culture experiments with cross-contamination or false classification. A genetic fingerprint pattern of a specimen by short tandem repeats (STR) is self-evident. Due to high amount of chromosomal rearrangements, known in epithelia ovary cancer cells and the instable STR pattern described in other tumour entities like leukaemia, this study explores the suitability of STR profiling for primary cultured epithelial ovary cancer cells.Methods: STR profiles of epithelial ovary cancers of 16 patients were compared with corresponding blood and corresponding primary cell cultures. The primary cell cultures of epithelial ovary tumours were passaged up to 28 times. In between, cultures were cryo conserved and recultured again, two to five times per patient.Results: In two cases, the STR pattern of tumour lost alleles (1/16 and 3/16) in comparison of corresponding STR-pattern from blood. In comparison to blood, cell culture of a third case, lost four alleles (4/16) accompanied with morphologic changes after 14th passage. It is equal after cryo conservation of the seventh passage from the same patient. The only changes in STR profiles we recognized are losses of alleles. Remaining STR markers allow authentication.Conclusions: Very likely, the allelic drop-outs beyond passage 14 assume complex genetic losses of heterozygosis resulting in changed growth behaviour of cells. All other STR-profiles of remaining 15 patients analysed in this study are stable over all passages and freeze-thaw processes. Thus, ovary cancer cell cultures in research should be authenticated by STR-profile in general. [ABSTRACT FROM AUTHOR]

The human hepatic cell line LX-2 has been described as tool to study mechanisms of hepatic fibrogenesis and the testing of antifibrotic compounds. It was originally generated by immortalisation with the Simian Vacuolating Virus 40 (SV40) transforming (T) antigen and subsequent propagation in low serum conditions. Although this immortalized line is used in an increasing number of studies, detailed genetic characterisation has been lacking. We here have performed genetic characterisation of the LX-2 cell line and established a single-locus short tandem repeat (STR) profile for the cell line and characterized the LX-2 karyotype by several cytogenetic and molecular cytogenetic techniques. Spectral karyotyping (SKY) revealed a complex karyotype with a set of aberrations consistently present in the metaphases analyses which might serve as cytogenetic markers. In addition, various subclonal and single cell aberrations were detected. Our study provides criteria for genetic authentication of LX-2 and offers insights into the genotype changes which might underlie part of its phenotypic features. [ABSTRACT FROM AUTHOR]

Purpose: The treatment of ovarian tumors is carried out with platinum medicine which can lead to incompatibilities or resistances. Thus, it is of great interest to check new medicine suitability for its application. AZD1152 is an Aurora kinase inhibitor predominantly works against Aurora kinase B involved in the chromosome segregation. Cells become polyploidy and reduce the proliferation by this impairment. To investigate whether AZD1152, may play a role in the treatment of ovarian carcinoma we serving it to the cisplatinum-resistant cell line SKOV3 alone and in combination with platinum. Methods: We look at the proliferation, the ploidy, the phases of cell cycle and the apoptosis activity of the cells. Results and conclusion: We could show that the combination of both medicines in the preclinical experiment produces a working advantage. [ABSTRACT FROM AUTHOR]

Although ovarian cancer is a rare disease, it constitutes the fifth leading cause of cancer death among women. It is of major importance to develop new therapeutic strategies to improve survival. Combining P8-D6, a novel dual topoisomerase inhibitor with exceptional anti-tumoral properties in ovarian cancer and compounds in preclinical research, and olaparib, a PARP inhibitor targeting DNA damage repair, is a promising approach. P8-D6 induces DNA damage that can be repaired by base excision repair or homologous recombination in which PARP plays a major role. This study analyzed benefits of combining P8-D6 and olaparib treatment in 2D and 3D cultures with ovarian cancer cells. Measurement of viability, cytotoxicity and caspase activity were used to assess therapy efficacy and to calculate the combination index (CI). Further DNA damage was quantified using the biomarkers RAD51 and γH2A.X. The combinational treatment led to an increased caspase activity and reduced viability. CI values partially show synergisms in combinations at 100 nM and 500 nM P8-D6. More DNA damage accumulated, and spheroids lost their membrane integrity due to the combinational treatment. While maintaining the same therapy efficacy as single-drug therapy, doses of P8-D6 and olaparib can be reduced in combinational treatments. Synergisms can be seen in some tested combinations. In summary, the combination therapy indicates benefits and acts synergistic at 100 nM and 500 nM P8-D6. [ABSTRACT FROM AUTHOR]

Purpose: Endometrial cancer (EC) therapy is characterized by the heterogeneity of EC subtypes resulting in unclear clinical behavior as well as in unsatisfactory treatment options. The available biomarkers, such as cellular tumor antigen p53 (TP53), phosphatidylinositol 3,4,5-trisphosphate 3-phosphatase and dual-specificity protein phosphatase (PTEN), and phosphatidylinositol 4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) genes alone might not be sufficient, and thus, new predictive and prognostic biomarkers are urgently required. The biomolecule class of microRNA represents a group of endogenously expressed regulatory factors primarily involved in control of pivotal cancer-related mechanisms including cell cycle, proliferation, apoptosis, and metastasis. Here, we review the current state of science regarding microRNA functionality in EC progression. [ABSTRACT FROM AUTHOR]

Ovarian cancer (OC) cells with homologous recombination deficiency (HRD) accumulate genomic scars (LST, TAI, and LOH) over a value of 42 in sum. PARP inhibitors can treat OC with HRD. The detection of HRD can be done directly by imaging these genomic scars, or indirectly by detecting mutations in the genes involved in HR. We show that HRD detection is also possible using high-resolution aCGH. A total of 30 OCs were analyzed retrospectively with high-resolution arrays as a test set and 19 OCs prospectively as a validation set. Mutation analysis was performed by HBOC TruRisk V2 panel to detect HR-relevant mutations. CNVs were clustered with respect to the involved HR genes versus the OC cases. In prospective validation, the HRD status determined by aCGH was compared with external HRD assessments. Two BRCA mutation carriers did not have HRD. OC could approximately differentiate into two groups with characteristic CNV patterns with different survival rates. Mutation frequencies have a linear regression on the HRD score. Mutations in individual HR-relevant genes do not always indicate HRD. This may depend on the mutation frequency in tumor cells. The aCGH shows the genomic scars of an HRD inexpensively and directly. [ABSTRACT FROM AUTHOR]

Ovarian cancer (OvCa) is the gynaecological disorder with the poorest prognosis due to the fast development of chemoresistance. We sought to connect chemoresistance and cancer cell‐derived extracellular vesicles (EV). The mechanisms of how chemoresistance is sustained by EV remained elusive. One potentially contributing factor is A Disintegrin and Metalloprotease 17 (ADAM17)—itself being able to promote chemoresistance and inducing tumour cell proliferation and survival via the Epidermal Growth Factor Receptor (EGFR) pathway by shedding several of its ligands including Amphiregulin (AREG). We now demonstrate that upon chemotherapeutic treatment, proteolytically active ADAM17 is released in association with EV from OvCa cells. In terms of function, we show that patient‐derived EV induce AREG shedding and restore chemoresistance in ADAM17‐deficient cells. Confirming that ADAM17‐containing EV transmit chemoresistance in OvCa, we propose that ADAM17 levels (also on EV) might serve as an indicator for tumour progression and the chemosensitivity status of a given patient. [ABSTRACT FROM AUTHOR]

Purpose: The dysregulation of cell cycle kinases plays a crucial role in carcinogenesis and the expression of various kinases has been attributed to aggressive tumor growth and an unfavourable prognosis in oncological patients. We, therefore, aimed to evaluate the expression of Ki67 among five additional cell cycle kinases in a collective of mammary and ovarian tumor specimens and to find a correlation with clinicopathological parameters.Methods: 76 mammary and 93 ovarian benign and malignant tumor samples were immunohistochemically stained and evaluated for the expression of Aurora A and B, Repp86, CDK1 and 2 (only breast specimens) and Ki67. The expression patterns of these cell cycle kinases were matched with retrospectively collected clinicopathological parameters.Results: All examined cell cycle kinases accurately discriminated benign from malignant breast and ovarian tissues. In breast cancer, Aurora A and B-, Repp86-, CDK2- and Ki67-expression was inversely associated with ER expression. No correlation with the HER2-status was found in our collective. Importantly, we found a significant correlation between the expression of Aurora A and CDK1 and axillary lymph node metastasis in breast cancer. Furthermore, a shortened disease free survival (DFS) upon expression of Aurora B and CDK2 was shown in breast cancer patients. None of the cell cycle kinases was associated with predictive or prognostic factors in epithelial ovarian cancer.Conclusion: The prognostic value of the expression of Ki67 is overtrumped by alternative cell cycle kinases when it comes to prediction of axillary tumor spread and a shortened DFS, which might allow a further risk stratification in breast cancer patients. [ABSTRACT FROM AUTHOR]

Simple Summary: Ovarian cancer is usually diagnosed in an advanced stage of the disease and is associated with a poor prognosis. Therefore, it is important to find reliable biomarkers that allow early diagnosis. In this study, we detected elevated Nectin-4 and HB-EGF serum levels in ovarian cancer patients which correlate with prognostically favorable parameters, such as early FIGO stage and successful tumor debulking. In addition, Nectin-4 was elevated in patients with low blood levels of Ca-125. This suggests that Nectin-4 and Ca-125 provide complementary diagnostic information. Therefore, Nectin-4 and HB-EGF appear to be promising blood-based biomarkers for the diagnosis of early stages of ovarian cancer. Ovarian cancer is the third most common gynecological malignancy and has the highest mortality rate. Owing to unspecific symptoms, ovarian cancer is not detected until an advanced stage in about two-thirds of cases. Therefore, it is crucial to establish reliable biomarkers for the early stages to improve the patients' prognosis. The aim of this study is to investigate whether the ADAM17 substrates Nectin-4, Heparin-binding EGF-like growth factor (HB-EGF) and Amphiregulin (AREG) could function as potential tumor markers for ovarian cancer. In this study a set of 231 sera consisting of 131 ovarian cancer patients and 100 healthy age-matched controls were assembled. Nectin-4, HB-EGF and AREG levels of preoperatively collected sera were determined by enzyme-linked immunosorbent assay (ELISA). Our analysis revealed that Nectin-4 and HB-EGF were significantly increased compared to the age-matched control group (p < 0.0001, p = 0.016). Strikingly, significantly higher Nectin-4 and HB-EGF levels were detected in early-stage FIGO I/II (p <0.001; p = 0.025) compared to healthy controls. Eighty-four percent (16/19) of patients with low Ca-125 levels showed increased Nectin-4 levels. Our study proposes Nectin-4 and HB-EGF as promising blood-based biomarkers for the detection of early stages of ovarian cancer patients that would not have been detected by Ca-125. [ABSTRACT FROM AUTHOR]

Objective: To characterize a complex chromosome rearrangement previously detected by G-banding in peripheral blood lymphocytes as 46,X, inv(X)(p11;q2?), inv(4)(q?),ins(8)(q?) in a patient with primary amenorrhea. Design: Case report. Setting: University faculty of medicine and hospital. Patient(s): A 16-year-old girl with primary amenorrhea. Intervention(s): Microdissection of chromosomes labeled by fluorescence in situ hybridization and by reverse painting. Main Outcome Measure(s): Use of commercial whole-chromosome painting probes for the detection of the aberrant chromosomes 4, 8, and X. Fluorescence probes of the isolated derivative chromosomes are self-generated. Result(s): The use of whole-chromosome painting probes allowed reliable identification of all chromosomes involved in the complex chromosome rearrangements. The DNA of those chromosomes was amplified and fluorescence labeled by polymerase chain reaction using degenerated oligonucleotide primers. These probes revealed breakpoints of the complex chromosome rearrangement by hybridization on normal and original chromosomes in 4q31.1, 8q24.1, Xp22.1, Xp11.4, and Xq24. Conclusion(s): We report on an Indian patient who has premature ovarian failure with primary amenorrhea as well as a hormone level increased for LH and FSH but decreased for TSH. She has a balanced complex translocation with three breakpoints in the X chromosome that were located by fluorescence in situ hybridization by chromosome microdissection, but no breakpoints localized in the critical regions for premature ovarian failure on the X chromosome. The breakpoint in Xq24 may be associated with the amenorrhea. [Copyright &y& Elsevier]

Purpose: Preimplantation genetic screening wants to improve artificial reproductive technologies, primarily by raising the rates of pregnancy, implantation and birth. We investigated if embryos derived from oocytes detected euploid for five chromosomes implant better than those which were biopsied but where the genetic detection failed. They were nevertheless transferred, thus serving as a sham control. Method: From 2004 to 2008 we performed 104 cycles of PGS with laser biopsy of the first polar body and FISH with five chromosomes. It was offered to all patients with eight or more oocytes, free of charge. The average female age was 36 years. If no euploid oocytes were available, not detected oocytes were transferred. Result: In 104 cycles 99 embryo transfers (95 %) were performed, resulting in 28 pregnancies (27 %), 20 births (71 %) and 8 miscarriages (29 %). The implantation rate in the euploid group was 19 vs. 13 % in the not detected group (n.s.). This trend was the same independent of age and embryo morphology. Conclusion: The pregnancy rate does not differ significantly from the national average. The trend in better implantation rates of euploid oocytes justifies a continuation of studies in this matter. [ABSTRACT FROM AUTHOR]

Purpose: For better selection of oocytes and embryos, preimplantation genetic screening (PGS) was introduced. As from the beginning of IVF, morphology was used as selection criteria; we investigated the combination of both. If there was a correlation between phenotype and genotype, invasive PGS might be replaced. Method: Therefore, 104 cycles with PGS were done by biopsy of the first polar body and FISH with five chromosomes. Morphology of the oocyte was recorded digitally and noted for 12 categories in 4-13 values; evaluation of the chromosomes was noted for five chromosomes in five values. Morphology and genetics were correlated to each other. Result: Correlations between morphology and genetics for day 0 were found: oocytes with an irregular or dark zona are less probable to have a normal chromosome 13 (80 vs. 53 %, p = 0.001). A medium amount of detritus in the perivitelline space makes it more probable to have a normal chromosome 18 (94 vs. 78 %, p = 0.001). A halo in the cytoplasm makes it less probable to be euploid for chromosome 22 (56 vs. 75 %, p = 0.018). For day 1, pattern '1, 2, 3 and fine' in the pronuclei makes it more probable to be euploid for chromosome 22 (78 vs. 63 %, p = 0.002). Conclusion: There are correlations between the oocyte genome and its morphology also on day 0. These correlations are not sufficient to replace PGS. [ABSTRACT FROM AUTHOR]

Background Complex small supernumerary marker chromosomes (sSMC) constitute one of the smallest subgroups of sSMC in general. Complex sSMC consist of chromosomal material derived from more than one chromosome; the best known representative of this group is the derivative chromosome 22 {der(22)t(11;22)} or Emanuel syndrome. In 2008 we speculated that complex sSMC could be part of an underestimated entity. Results Here, the overall yet reported 412 complex sSMC are summarized. They constitute 8.4% of all yet in detail characterized sSMC cases. The majority of the complex sSMC is contributed by patients suffering from Emanuel syndrome (82%). Besides there are a der(22)t(8;22)(q24.1;q11.1) and a der(13)t(13;18)(q11;p11.21) or der(21)t(18;21)(p11.21;q11.1) = der(13 or 21)t(13 or 21;18) syndrome. The latter two represent another 2.6% and 2.2% of the complex sSMC-cases, respectively. The large majority of complex sSMC has a centric minute shape and derives from an acrocentric chromosome. Nonetheless, complex sSMC can involve material from each chromosomal origin. Most complex sSMC are inherited form a balanced translocation in one parent and are non-mosaic. Interestingly, there are hot spots for the chromosomal breakpoints involved. Conclusions Complex sSMC need to be considered in diagnostics, especially in non-mosaic, centric minute shaped sSMC. As yet three complex-sSMC-associated syndromes are identified. As recurrent breakpoints in the complex sSMC were characterized, it is to be expected that more syndromes are identified in this subgroup of sSMC. Overall, complex sSMC emphasize once more the importance of detailed cytogenetic analyses, especially in patients with idiopathic mental retardation. [ABSTRACT FROM AUTHOR]

Simple Summary: Despite significant advancement in therapeutic strategies, breast cancers remain the most prevalent type of cancer in terms of incidence and mortality worldwide. Therefore, new alternative therapies have become an urgent clinical need. The present study shows a significantly enhanced cytotoxicity and apoptosis of a treatment with the dual topoisomerase inhibitor P8-D6 compared to standard cytostatics. These effects were visible in various breast cancer cell lines and primary patient cells in 2D monolayer and 3D spheroids. In summary, P8-D6 seems to be a potent chemotherapeutic agent for various breast cancer cell treatments. Breast cancer constitutes the leading cause of cancer deaths among females. However, numerous shortcomings, including low bioavailability, resistance and significant side effects, are responsible for insufficient treatment. The ultimate goal, therefore, is to improve the success rates and, thus, the range available treatment options for breast cancer. Consequently, the identification, development and evaluation of potential novel drugs such as P8-D6 with seminal antitumor capacities have a high clinical need. P8-D6 effectively induces apoptosis by acting as a dual topoisomerase I/II inhibitor. This study provides an overview of the effectiveness of P8-D6 in breast cancer with both 2D monolayers and 3D spheroids compared to standard therapeutic agents. For this drug effectiveness review, cell lines and ex vivo primary cells were used and cytotoxicity, apoptosis rates and membrane integrity were examined. This study provides evidence for a significant P8-D6-induced increase in apoptosis and cytotoxicity in breast cancer cells compared to the efficacy of standard therapeutic drugs. To sum up, P8-D6 is a fast and powerful inductor of apoptosis and might become a new and suitable therapeutic option for breast cancer in the future. [ABSTRACT FROM AUTHOR]

Simple Summary: Ovarian cancer has the highest lethality among gynecological tumors. Therefore, it is essential to find reliable biomarkers to improve early detection. This is the first report describing ADAM17 detection in serum and ascites fluid of ovarian cancer patients. A high ADAM17 concentration in serum at primary diagnosis is associated with early FIGO stages and predicts complete resection of the tumor mass. In addition, ADAM17 and CA-125 complement each other, especially in the diagnosis of early stages. In summary, ADAM17 appears to be a promising screening marker for detecting early-stage ovarian cancer. Ovarian cancer has the highest mortality rate among gynecological tumors. This is based on late diagnosis and the lack of early symptoms. To improve early detection, it is essential to find reliable biomarkers. The metalloprotease ADAM17 could be a potential marker, as it is highly expressed in many solid tumors, including ovarian and breast cancer. The aim of this work is to evaluate the relevance of ADAM17 as a potential diagnostic blood-based biomarker in ovarian cancer. Ovarian cancer cell lines IGROV-1 and A2780, as well as primary patient-derived tumor cells obtained from tumor tissue and ascitic fluid, were cultured to analyze ADAM17 abundance in the culture supernatant. In a translational approach, a cohort of 117 well-characterized ovarian cancer patients was assembled and ADAM17 levels in serum and corresponding ascitic fluid were determined at primary diagnosis. ADAM17 was quantified by enzyme-linked immunosorbent assay (ELISA). In the present study, ADAM17 was detected in the culture supernatant of ovarian cancer cell lines and primary cells. In addition, ADAM17 was found in serum and ascites of ovarian cancer patients. ADAM17 level was significantly increased in ovarian cancer patients compared to an age-matched control group (p < 0.0001). Importantly early FIGO I/II stages, which would not have been detected by CA-125, were associated with higher ADAM17 concentrations (p = 0.007). This is the first study proposing ADAM17 as a serum tumor marker in the setting of a gynecological tumor disease. Usage of ADAM17 in combination with CA-125 and other markers could help detect early stages of ovarian cancer. [ABSTRACT FROM AUTHOR]

HP1 proteins are thought to be modulators of chromatin organization in all mammals, yet their exact physiological function remains unknown. In a first attempt to elucidate the function of these proteins in vivo, we disrupted the murine Cbx1 gene, which encodes the HP1-β isotype, and show that the Cbx1[sup -/-]-null mutation leads to perinatal lethality. The newborn mice succumbed to acute respiratory failure, whose likely cause is the defective development of neuromuscular junctions within the endplate of the diaphragm. We also observe aberrant cerebral cortex development in Cbx1[sup -/-] mutant brains, which have reduced proliferation of neuronal precursors, widespread cell death, and edema. In vitro cultures of neurospheres from Cbx1[sup -/-] mutant brains reveal a dramatic genomic instability. Our results demonstrate that HP1 proteins are not functionally redundant and that they are likely to regulate lineage-specific changes in heterochromatin organization. [ABSTRACT FROM AUTHOR]

Several investigations have supposed that tumor suppressor genes might be located on human chromosome 8. We used microcell-mediated transfer of chromosome 8 into MDA-MB-231 breast cancer cells and generated independent hybrids with strongly reduced tumorigenic potential. Loss of the transferred chromosome results in reappearance of the malignant phenotype. Expression analysis identified a set of 109 genes (CT8-ps) differentially expressed in microcell hybrids as compared to the tumorigenic MDA-MB-231 and rerevertant cells. Of these, 44.9%are differentially expressed in human breast tumors. The expression pattern of CT8-ps was associated with prognostic factors such as tumor size and grading as well as loss of heterozygosity at the short arm of chromosome 8. We identified CT8-ps networks suggesting that these genes act cooperatively to cause reversion of tumorigenicity in MDA-MB-231 cells. Our findings provide a conceptual basis and experimental system to identify and evaluate genes and gene networks involved in the development and/or progression of breast cancer.Oncogene (2005) 24, 869-879. doi:10.1038/sj.onc.1208260 Published online 6 December 2004 [ABSTRACT FROM AUTHOR]

Simple Summary: Ovarian cancer (OvCa) treatment is still a challenge, mainly due to acquired resistance mechanisms during the course of chemotherapy. Here, we show the enhanced cytotoxicity of the combined treatment with the ADAM17 inhibitor GW280264X and cisplatin in comparison with cisplatin monotherapy. This effect was visible in five of five ovarian cancer cell lines grown as a monolayer and two of three tested cell lines in three-dimensional tumor spheroids. Tumor spheroids derived from primary tumor and ascites cells were sensitized to cisplatin treatment by GW280264X. In summary, the combination of ADAM17 inhibition with conventional chemotherapy seems to be a promising strategy to overcome chemotherapy resistance in OvCa. Chemotherapy resistance is a major challenge in ovarian cancer (OvCa). Thus, novel treatment combinations are highly warranted. However, many promising drug candidates tested in two-dimensional (2D) cell culture have not proved successful in the clinic. For this reason, we analyzed our drug combination not only in monolayers but also in three-dimensional (3D) tumor spheroids. One potential therapeutic target for OvCa is A disintegrin and metalloprotease 17 (ADAM17). ADAM17 can be activated by chemotherapeutics, which leads to enhanced tumor growth due to concomitant substrate cleavage. Therefore, blocking ADAM17 during chemotherapy may overcome resistance. Here, we tested the effect of the ADAM17 inhibitor GW280264X in combination with cisplatin on ovarian cancer cells in 2D and 3D. In 2D, the effect on five cell lines was analyzed with two readouts. Three of these cell lines formed dense aggregates or spheroids (HEY, SKOV-3, and OVCAR-8) in 3D and the treatment effect was analyzed with a multicontent readout (cytotoxicity, viability, and caspase3/7 activation). We tested the combined therapy on tumor spheroids derived from primary patient cells. In 2D, we found a significant reduction in the half minimal (50%) inhibitory concentration (IC50) value of the combined treatment (GW280264X plus cisplatin) in comparison with cisplatin monotherapy in all five cell lines with both 2D readout assays (viability and caspase activation). In contrast, the combined treatment only showed an IC50 reduction in HEY and OVCAR-8 3D tumor spheroid models using caspase3/7 activity or CelltoxTM Green as the readout. Finally, we found an improved effect of GW280264X with cisplatin in tumor spheroids derived from patient samples. In summary, we demonstrate that ADAM17 inhibition is a promising treatment strategy in ovarian cancer. [ABSTRACT FROM AUTHOR]