Natural compounds, such as resveratrol (Res), are currently used as adjuvants for anticancer therapies. To evaluate the effectiveness of Res for the treatment of ovarian cancer (OC), we screened the response of various OC cell lines to the combined treatment with cisplatin (CisPt) and Res. We identified A2780 cells as the most synergistically responding, thus optimal for further analysis. Because hypoxia is the hallmark of the solid tumor microenvironment, we compared the effects of Res alone and in combination with CisPt in hypoxia (pO2 = 1%) vs. normoxia (pO2 = 19%). Hypoxia caused an increase (43.2 vs. 5.0%) in apoptosis and necrosis (14.2 vs. 2.5%), reactive oxygen species production, pro-angiogenic HIF-1α (hypoxia-inducible factor-1α) and VEGF (vascular endothelial growth factor), cell migration, and downregulated the expression of ZO1 (zonula occludens-1) protein in comparison to normoxia. Res was not cytotoxic under hypoxia in contrast to normoxia. In normoxia, Res alone or CisPt+Res caused apoptosis via caspase-3 cleavage and BAX, while in hypoxia, it reduced the accumulation of A2780 cells in the G2/M phase. CisPt+Res increased levels of vimentin under normoxia and upregulated SNAI1 expression under hypoxia. Thus, various effects of Res or CisPt+Res on A2780 cells observed in normoxia are eliminated or diminished in hypoxia. These findings indicate the limitations in using Res as an adjuvant with CisPt therapy in OC.

Resistance to anti-cancer drugs is the main challenge in oncology. In pre-clinical studies, established cancer cell lines are primary tools in deciphering molecular mechanisms of this phenomenon. In this study, we proposed a new, transcriptome-focused approach, utilizing a model of isogenic cancer cell lines with gradually changing resistance. We analyzed trends in gene expression in the aim to find out a scaffold of resistance development process. The ovarian cancer cell line A2780 was treated with stepwise increased concentrations of paclitaxel (PTX) to generate a series of drug resistant sublines. To monitor transcriptome changes we submitted them to mRNA-sequencing, followed by the identification of differentially expressed genes (DEGs), principal component analysis (PCA), and hierarchical clustering. Functional interactions of proteins, encoded by DEGs, were analyzed by building protein-protein interaction (PPI) networks. We obtained human ovarian cancer cell lines with gradually developed resistance to PTX and collateral sensitivity to cisplatin (CDDP) (inverse resistance). In their transcriptomes, we identified two groups of DEGs: (1) With fluctuations in expression in the course of resistance acquiring; and (2) with a consistently changed expression at each stage of resistance development, constituting a scaffold of the process. In the scaffold PPI network, the cell cycle regulator—polo-like kinase 2 (PLK2); proteins belonging to the tumor necrosis factor (TNF) ligand and receptor family, as well as to the ephrin receptor family were found, and moreover, proteins linked to osteo- and chondrogenesis and the nervous system development. Our cellular model of drug resistance allowed for keeping track of trends in gene expression and studying this phenomenon as a process of evolution, reflected by global transcriptome remodeling. This approach enabled us to explore novel candidate genes and surmise that abrogation of the osteomimic phenotype in ovarian cancer cells might occur during the development of inverse resistance between PTX and CDDP.

The role of germline mutations in BRCA1 and BRCA2 genes in the risk of the development of ovarian cancer is clinically well established. BRCA1/2 testing seems to have increasing role in clinical management in patients with advanced ovarian cancer who require treatment with poly(ADP-ribose) polymerase inhibitors.

Recombinant human erythropoietin (rhEPO) is used in breast and ovarian cancer patients to alleviate cancer- and chemotherapy-related anemia. Some clinical trials have reported that rhEPO may adversely impact survival and increase the risk of thrombovascular events in patients with breast cancer but not with ovarian cancer. The latter may potentially benefit the most from rhEPO treatment due to the nephrotoxic and myelosuppresive effects of standard platinum-based chemotherapy used in ovarian cancer disease. However, over the last decade the preclinical data have revealed that EPO is not only the principal growth factor and the hormone which regulates erythropoiesis, but also a cytokine with a pleiotropic activity which also can affect cancer cells. EPO can stimulate survival, ability to form metastases and drug resistance not only in continuous breast- and ovarian cancer cell lines but also in breast cancer stem-like cells. EPO receptor (EPOR) can also be constitutively active in both these cancers and, in breast cancer cells, may act in an interaction with estrogen receptor (ER) and epidermal growth factor receptor-2 (HER-2). EPOR, by an EPO-independent mechanism, promotes proliferation of breast cancer cells in cooperation with estrogen receptor, resulting in decreased effectiveness of tamoxifen treatment. In another interaction, as a result of the molecular antagonism between EPOR and HER2, rhEPO protects breast cancer cells against trastuzumab. Both clinical and preclinical evidence strongly suggest the urgent need to reevaluate the traditional use of rhEPO in the oncology setting.

We reviewed the literature on the relationship between the Fanconi anemia pathway (FA) and response to chemotherapy in patients with ovarian cancer. Despite continuous developments in medicine, ovarian cancer remains a challenge for both, physicians and researchers seeking ways to achieve better results of chemotherapy combined with other targeted therapies. Clinically relevant resistance to chemotherapy is a major problem in treating ovarian cancer. Researchers continue to investigate mechanisms responsible for drug resistance in order to develop better therapeutic methods against ovarian cancer. Among the resistance mechanisms, defects in DNA repair, including the FA pathway may be important in increasing the sensitivity of ovarian cancer cells to chemotherapy agents at the clinical level. A growing number of data has shown that disruption of the FA genes may be a useful predictor of OC sensitivity to chemotherapy agents whose activity is based on DNA crosslinking mechanisms.

Recombinant human erythropoietin (rhEPO) has been used clinically to alleviate cancer- and chemotherapy-related anemia. However, recent clinical trials have reported that rhEPO also may adversely impact disease progression and survival. The expression of functional EPO receptors (EPOR) has been demonstrated in many human cancer cells where, at least in vitro, rhEPO can stimulate cell growth and survival and may induce resistance to selected therapies. Responses to rhEPO measured by alterations in tumor cell growth or survival, activation of signaling pathways or modulation of sensitivity to anticancer agents are variable. Both methodological and inherent biological issues underlie the differential cell responses, including reported difficulties in EPOR protein detection, potential involvement of EPOR isoforms or of cytoplasmic EPOR, possible differential structure and/or binding affinities of hematopoietic versus non-hematopoietic cell EPOR, possible aberrant regulation of EPOR activity, and a functional EPO/EPOR autocrine/paracrine loop. The modulation by rhEPO of tumor cell response to anticancer agents is coincident with modulation of multiple signaling pathways, BCL-2 family proteins, caspases and NFkB. The molecular interplay of pro-survival and pro-death signals, triggered by EPO and/or by anticancer agents, is multifactorial and tightly coordinated. Expression microarray analysis may prove critical for deciphering this potentially novel network and its broad spectrum of genes and proteins.

The identification of erythropoietin receptors (EpoR) on cancer cells has caused concern, since it implies the possibility that treatment of cancer patients with erythropoietin (Epo) and related agents with demonstrable antiapoptotic activity could enhance cancer growth and progression. However, the function and even the validity of the identification of these receptors have been called into question. We now report the characterization of EpoR and Epo expression by 4 human ovarian cancer cell lines: A2780, CaOV, SKOV and OVCAR‐3. Using semiquantitative RT‐PCR, restriction digestion of the PCR products and DNA sequence analysis, we determined that each of the lines expresses the EpoR and Epo at the mRNA level. A2780 cells were the highest expressers of both genes. We demonstrated EpoR protein both by western blotting and by immunofluorescence and biologically active Epo protein by quantitative in vitro bioassay. The EpoR on A2780 cells was shown to be functional, since Epo stimulation resulted in phosphorylation of Erk1/2, an important EpoR mitogenic signaling intermediate. None of the cell lines exhibited a growth response in culture to exogenous Epo. However, addition of a neutralizing anti‐Epo antibody to A2780 cells resulted in partial growth inhibition that was reversed by the addition of excess Epo, providing evidence for an autocrine/paracrine mechanism of growth enhancement in these cells. © 2007 Wiley‐Liss, Inc.

Leptin concentrations in blood serum are significantly elevated in women suffering from endometrial carcinoma or from advanced breast carcinoma. Controversial results on the relations in other cancers of genital organs (ovary, uterine cervix, vulva) and in endometriosis require some further studies.

BACKGROUNDThe reverse transcription-PCR tyrosinase assay (TYR test) cannot reliably detect malignant melanoma (MM) cells in blood as the cells often circulate at low concentrations. We evaluated the prognostic value of multiple TYR testing, the prognostic significance of individual positive TYR test results (TYR+) in asymptomatic melanoma patients, and whether statistical analysis could help in the interpretation of results of a test that measures phenomena that typically occur below its detection threshold.METHODSMM patients in stages I-IV (n = 150) underwent multiple testing with the TYR test during the course of their disease. TYR testing was performed as described by Smith et al. (Lancet 1991;38:1227-9). Statistical analyses were performed with the logistic function and t-test procedures.RESULTSThe relationship between MM stage and the frequency of TYR+ was statistically significant (P = 0.011). Higher frequency of TYR+ in clinically asymptomatic patients after complete resection of the primary tumor was associated with an increased risk of recurrence of MM (prognostic sensitivity, 62%; specificity, 78%).CONCLUSIONSA single positive TYR test provides a warning for disease relapse, suggesting that multiple TYR testing might provide more reliable predictions of disease progression. Multiple testing and statistical analysis using a logistic function might allow for the interpretation of apparently inconsistent results of tests for very rare cells.

Istotn1 rolê w postêpowaniu z chorym na nowotwór odgrywa coraz lepsza diagnostyka, która przekroczy3a granice wykrywalnoœci klasycznych metod oscyluj1ce wokó3 miliarda komórek nowotworowych (tab. 1.). Zasadnicz1 zmianê stanowi3o wprowadzenie w 1984 r. przez Kary Mullisa 3añcuchowej reakcji polimerazy (PCR) [1], pozwalaj1cej wykryæ pojedyncze kopie genu w badanym materiale, a nastêpnie wsparcie tej metody reakcj1 odwrotnej transkrypcji (RT), co pozwoli3o wykryæ pojedyncze kopie transkryptu takiego genu, a wiêc badaæ geny, które rzeczywiœcie ulegaj1 ekspresji w badanych komórkach, a nie tylko takie, które w nich s1. Z kolei zrozumienie, ¿e choroby nowotworowe s1 nabytymi chorobami genetycznymi, a wiêc chorobami, które s1 spowodowane mutacjami okreœlonych genów, a tak¿e s1 zwi1zane z wtórnie zaburzon1 ekspresj1 wielu innych genów zwróci3o uwagê na mo¿liwoœæ wykorzystania tych zmian genetycznych jako znaczników chorób nowotworowych, czyli ich swoistych wizytówek. To wszystko spowoDiagnostyka nowotworów coraz bardziej przesuwa siê w kierunku wykrywania choroby resztkowej, czyli niedostêpnej wykryciu i badaniu za pomoc1 klasycznych metod. Wprowadzenie techniki RT-PCR pozwoli3o wykrywaæ pojedyncze komórki nowotworowe w badanej próbce krwi lub tkanki i obni¿yæ poziom wykrywalnoœci do ok. 10 w ca3ej objêtoœci krwi obwodowej. Jednak¿e uwolnienie a¿ 1 tys. komórek do krwi dla wielu nowotworów mo¿e byæ zjawiskiem bardzo rzadkim i do jego oceny potrzebne s1 czulsze metody. Sprawa ta sta3a siê szczególnie kontrowersyjna w przypadku czerniaka z3oœliwego, gdzie wyniki badañ molekularnych wydawa3y siê nie korelowaæ z zaawansowaniem choroby. Nasze wczeœniejsze badania pos3u¿y3y do weryfikacji hipotezy, ¿e dzieje siê tak, dlatego ¿e liczba kr1¿1cych we krwi komórek czerniaka znajduje siê i zmienia w miarê progresji choroby, stale pozostaj1c znacznie poni¿ej dolnej granicy wykrywalnoœci. W chorobie niezaawansowanej przeciêtnie jedno na piêæ badañ jest dodatnie (ok. 200 kr1¿1cych komórek), a w zaawansowanej œrednio jedno na trzy badania jest dodatnie (ok. 333 kr1¿1cych komórek). Zosta3o to stwierdzone dziêki wielokrotnemu powtarzaniu testów u ka¿dego pacjenta oraz zastosowania odpowiednich analiz statystycznych. W ten sposób powsta3a najczulsza obecnie dostêpna metoda badania choroby resztkowej, pozwalaj1ca wykrywaæ nawet mniej ni¿ 100 kr1¿1cych we krwi komórek nowotworowych, któr1 autorzy nazwali MR-RT-PCR. Dla dalszej jej weryfikacji powinno siê j1 obecnie spróbowaæ wykorzystaæ w innych schorzeniach nowotworowych, takich jak przewlek3a bia3aczka szpikowa, ch3oniak p3aszcza czy rak prostaty. Powinno to zaowocowaæ technologi1 wykrywania bardzo rzadkich komórek, wystêpuj1cych rzadziej ni¿ w pojedynczej próbce badanego materia3u, a tak¿e dostarczyæ nowych informacji na temat biologii wielu nowotworów.

Leptyna, kodowana przez gen ob [1] jest polipeptydem o wybitnie plejotropowym dzia3aniu: hormonem, wytwarzanym g3ównie przez tkankê t3uszczow1, bior1cym udzia3 w regulacji masy cia3a [2] i stymuluj1cym wytwarzanie estrogenu [3] oraz cytokin1, bior1c1 udzia3 w hemopoezie [4, 5], angiogenezie [6], modulacji wytwarzania cytokin przez limfocyty T pomocnicze [7], regulacji cyklu komórkowego i byæ mo¿e onkogenezie.

Szenajch, J.; Jasinski, B.; Kozak, A.; Kulik, J.; Chomicka, M.; Struzyna, J.; Nowecki, Z.; Rutkowski, P.; Ruka, W.; Kupsc, W.; Siedlecki, J.; Wiktor-Jedrzejczak, W. Author Information

Reverse transcription and polymerase chain reaction (RT/PCR) with primers specific for tyrosinase allow for a new method of early detection of individual melanoma cells in peripheral blood. Using this test the effect of chemo-and chemoimmunotherapy on the spread of early micrometastatic cancer cells has been evaluated. No significant correlations have been found between RT/PCR results on the one hand and stage of disease, a kind of the therapy protocol used and usage of the therapy as an adjuvant or palliative on the other hand. Thus, although the RT/PCR test for detection of circulating individual melanoma cells might help in identification of minimal residual disease in some patients, it has no application for routine staging of more advanced disease and in monitoring the response to therapy.

Using reverse transcription and polymerase chain reaction (PCR) with primers specific for tyrosinase the individual melanoma cells were detected in peripheral blood of patients in different stages of disease, after excision of primary lesion and prior and after chemotherapy. No relation between stage of disease (including situations with overt generalized spread of melanoma) and probability of positive PCR reaction detecting transcript for tyrosinase gene was found. Many patients in III and IV stages were negative for prolonged periods. Therefore, this method cannot be used for monitoring of all patients, because many of them are negative prior as well as after chemotherapy. With regard to the effects of therapy, the patients differed one to another and although some persons positive prior treatment became negative thereafter, a similar number of initially negative patients became positive after treatment.

Macrophages have been suggested to play a major role in the immune response to cancer. They have also been suggested to stimulate the formation of tumor stroma and to promote tumor vascularization. The availability of the op/op mouse, which has no endogenous colony‐stimulating factor 1 (CSF‐1) and which possesses a profound macrophage deficiency, provides a new model to verify these notions. Subcutaneous growth of transplantable Lewis lung cancer (LLC) is markedly impaired in the op/op mice compared with normal littermates. Treatment of tumor‐bearing op/op mice with human recombinant CSF‐1 corrects this impairment. Histological analysis of tumors grown in op/op and normal mice revealed marked differences. Tumors grown in op/op mice display a decreased mitotic index and pronounced necrosis, particularly hemorrhagic. Moreover, particularly in the op/op tumors, peculiar sinusoid‐like abortive vessels (not filled with blood) have been observed. These tumors, in contrast to tumors grown in normal mice, are almost deprived of regular arteries and veins. In contrast to tumors grown in normal mice, they exhibit almost no Sirius red‐stained collagenous fibers and Gomori silver‐stained‐reticular fibers. Our data suggest that the CSF‐1‐dependent macrophage subpopulation missing in op/op mice plays a primary role in supporting tumor stroma formation and tumor vascularization in murine LLC tumors. © 1996 Wiley‐Liss, Inc.