Sirtuin 6 (SIRT6), a member of sirtuin family (SIRT1-7), regulates a variety of cellular processes involved in aging, metabolism, and cancer. Dysregulation of SIRT6 is widely observed in different breast cancer subtypes; however, the role and function of SIRT6 in cancer development remain largely unexplored. The aim of this study was to identify novel compounds targeting SIRT6 which may provide a new approach in development of anti-cancer therapy for breast cancer. Virtual screening was utilized to discover potential compounds targeting SIRT6 for in vitro screening. In addition, novel 1,4-dihydropyridine derivatives were synthetized and further subjected for the screening. The impact of the compounds on the deacetylation activity of SIRT6 was determined with HPLC method. The anti-cancer activities were screened for a panel of breast cancer cells. A set of 1,4-dihydropyridine derivatives was identified as SIRT6 inhibitors. A SIRT6 activating compound, (2,4-dihydroxy-phenyl)-2-oxoethyl 2-(3-methyl-4-oxo-2-phenyl-4H-chromen-8-yl)acetate (later called as 4H-chromen), was discovered and it provided 30-40-fold maximal activation. 4H-chromen was proposed to bind similarly to quercetin and place to previously reported SIRT6 activator sites. 4H-chromen was investigated in various breast cancer cells, and it decreased cell proliferation in all cells as well as arrested cell cycle in triple negative cells. Overall, this study describes a highly potent SIRT6 activator and new inhibitors that represent a novel tool to study the mechanism of SIRT6 function.

• Klíčová slova
• Activator, Breast cancer, Inhibitor, Sirtuin 6, Virtual screening,
• MeSH
• antitumorózní látky chemie   farmakologie   terapeutické užití   MeSH
• časná detekce nádoru metody   MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• nádory prsu farmakoterapie   metabolismus   MeSH
• sekundární struktura proteinů MeSH
• simulace molekulového dockingu metody   MeSH
• sirtuiny antagonisté a inhibitory   chemie   metabolismus   MeSH
• terciární struktura proteinů MeSH
• viabilita buněk účinky léků   fyziologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antitumorózní látky MeSH
• SIRT6 protein, human MeSH Prohlížeč
• sirtuiny MeSH

Spheroid cultures of cancer cells reproduce the spatial dimension-induced in vivo tumor traits more effectively than the conventional two-dimensional cell cultures. With growing interest in spheroids for high-throughput screening (HTS) assays, there is an increasing demand for cost-effective miniaturization of reproducible spheroids in microtiter plates (MPs). However, well-to-well variability in spheroid size, shape, and growth is a frequently encountered problem with almost every culture method that has prevented the transfer of spheroids to the HTS platform. This variability partly arises due to increased susceptibility of MPs to edge effects and evaporation-induced changes in the growth of spheroids. In this study, we examined the effect of evaporation on the reproducibility of spheroids of tumor and nontumor cell lines in 384-well plates, and show that culture conditions that prevent evaporation-induced medium loss result in the formation of uniform spheroids across the plate. Additionally, we also present a few technical improvements to increase the scalability of the liquid-overlay spheroid culturing technique in MPs, together with a simple software routine for the quantification of spheroid size. We believe that these cost-effective improvements will aid in further improvement of spheroid cultures for HTS drug discovery.

• Klíčová slova
• edge effect, evaporation, liquid-overlay culture, reproducibility, spheroids,
• MeSH
• antitumorózní látky izolace a purifikace   terapeutické užití   MeSH
• buněčné kultury metody   MeSH
• buněčné sféroidy účinky léků   MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• objevování léků metody   MeSH
• rychlé screeningové testy metody   MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antitumorózní látky MeSH

AIMS: To develop an in vitro tool for assessing the efficacy and toxicity of anticancer drugs using mixed culture containing both tumor and non-tumor cells. Such in vitro tool should have high application potential in drug-screening and personalized cancer care. METHODS: Fibroblasts were spiked as non-tumor cells into tumor cells of an established line. The mixed culture was treated with a test drug at various concentrations. After the treatment, DNA was prepared directly from the survived adhesive cells in the wells of the 96-well plates using a simple and inexpensive method, and subjected to digital PCR for measuring relative copy numbers of a target gene NF1 to that of a reference gene RPP30. The NF1 gene is known to be heterozygously deleted in these tumor cells while the RPP30 gene has two copies in both tumor and non-tumor cells. Using the NF1/ RPP30 ratios resulting from the dual digital PCR assay, the proportions of tumor cells were calculated for each drug concentration. RESULTS: Digital PCR confirmed that the tumor cells have only one copy of the NF1 gene while the non-tumor fibroblasts have two copies. By contrast, both types of cells have two copies of the reference gene RPP30. Using the ratio of the two genes, we successfully calculated the proportion of tumor cells which decreased as the dose of the test drug increased up to a certain concentration, indicating that the drug is more effective for the tumor cells than for the non-tumor cells in this dose-range. At the highest dose, we observed a slight increase in the proportion of tumor cells, likely reflecting the toxic effect of the drug on both tumor and non-tumor cells. CONCLUSION: This pilot study demonstrated the feasibility of a genetic- and cell-based tool for testing efficacy and toxicity of anticancer drugs in vitro. The promising results suggest that additional efforts are merited, for further development since such a tool will likely have high application potential (1) in drug discovery where it enables simultaneously assessing therapeutic effect on target cells and toxic effect on non-target cells, and (2) in personalized adjuvant chemotherapy where multiple drugs can be tested in primary cultures derived from surgically removed tumor.

• Klíčová slova
• drug discovery; drug specificity; digital PCR; primary culture; mixed culture; chemotherapy; drug selection; mutation; copy number variation,
• MeSH
• antitumorózní látky farmakologie   toxicita   MeSH
• autoantigeny účinky léků   genetika   MeSH
• DNA nádorová účinky léků   genetika   MeSH
• fibroblasty účinky léků   MeSH
• geny neurofibromatózy 1 účinky léků   MeSH
• inhibitory enzymů farmakologie   MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• mutace účinky léků   genetika   MeSH
• nádorové buněčné linie MeSH
• objevování léků metody   MeSH
• pilotní projekty MeSH
• polymerázová řetězová reakce metody   MeSH
• pyrimidiny farmakologie   MeSH
• ribonukleasa P účinky léků   genetika   MeSH
• studie proveditelnosti MeSH
• techniky in vitro MeSH
• tyrosinkinasy antagonisté a inhibitory   MeSH
• variabilita počtu kopií segmentů DNA účinky léků   genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• Názvy látek
• antitumorózní látky MeSH
• autoantigeny MeSH
• DNA nádorová MeSH
• inhibitory enzymů MeSH
• nilotinib MeSH Prohlížeč
• pyrimidiny MeSH
• ribonukleasa P MeSH
• RPP30 protein, human MeSH Prohlížeč
• tyrosinkinasy MeSH

Treatment of benign meningiomas remains a challenge, especially when they involve the skull-base or when surgery and radiation fail. Moreover, a recent in vitro MTT (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide) study testing hydroxyurea, temozolomide and other targeting agents failed to identify drugs effective in their treatment; therefore the search for further more effective agents continues. We performed a thorough review of in vitro investigations, animal studies and human clinical trials and endeavoured to integrate our results of MTT assay into current concepts of chemotherapy in benign meningiomas. Our results demonstrated that other chemotherapeutics with various mechanisms of action have the potential to be incorporated into second line therapy. Our study shows for the first time that chemosensitivity/resistance may be associated with histopathological variants of benign meningiomas.

• Klíčová slova
• Benign meningioma, Chemotherapy, MTT assay,
• MeSH
• antitumorózní látky terapeutické užití   MeSH
• chemorezistence účinky léků   MeSH
• dospělí MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé středního věku MeSH
• lidé MeSH
• meningeální nádory farmakoterapie   patologie   MeSH
• meningeom farmakoterapie   patologie   MeSH
• nádorové buňky kultivované MeSH
• následné studie MeSH
• proliferace buněk účinky léků   MeSH
• senioři MeSH
• techniky in vitro MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antitumorózní látky MeSH

The alarming rate of failure of clinical trials is a major hurdle in cancer therapy that partly results from the inadequate use of in vitro tumor models for the screening of promising hits and leads in preclinical studies. 2D cultures of cancer cell lines that are primarily used for drug screening do not adequately recapitulate tumor microenvironment (TME) complexities compared with 3D cancer cell cultures and tumor-derived primary cell cultures. In this review, we focus on the potential use of in vitro tumor models that reproduce in vivo tumor complexities for effective drug selection in the preclinical stages of drug development.

• MeSH
• alternativy testů na zvířatech MeSH
• antitumorózní látky farmakologie   MeSH
• buněčné kultury MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• mezibuněčná komunikace účinky léků   MeSH
• nádorové buněčné linie MeSH
• nádorové mikroprostředí účinky léků   MeSH
• nádory farmakoterapie   metabolismus   patologie   patofyziologie   MeSH
• objevování léků metody   MeSH
• reprodukovatelnost výsledků MeSH
• rychlé screeningové testy MeSH
• signální transdukce účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• antitumorózní látky MeSH

The platinum(II) dichlorido and oxalato complexes of the general formula cis-[PtCl(2)(nHaza)(2)] (1-3) [Pt(ox)(nHaza)(2)] (4-6) involving 7-azaindole halogeno-derivatives (nHaza) were prepared and thoroughly characterized. A single-crystal X-ray analysis of cis-[PtCl(2)(3ClHaza)(2)]·DMF (1·DMF; 3ClHaza symbolizes 3-chloro-7-azaindole) revealed a distorted square-planar arrangement with both the 3ClHaza molecules coordinated through their N7 atoms in a cis fashion. In vitro cytotoxicity of the complexes was evaluated by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay against the HOS (osteosarcoma), MCF7 (breast adenocarcinoma) and LNCaP (prostate adenocarcinoma) human cancer cell lines. The dichlorido complexes 1-3 (IC(50)=3.8, 3.9, and 2.5 μM, respectively) showed significantly higher in vitro anticancer effect against HOS as compared with cisplatin, whose IC(50)=37.7 μM. The biological effect of cisplatin against MCF7 (IC(50)=24.5 μM) and LNCaP (IC(50)=3.8 μM) was also exceeded by 1-3 (except for 2 against LNCaP), but the difference can be classified as significant only in the case of 1 (IC(50)=3.4 μM) and 3 (IC(50)=2.0 μM) against MCF7. The molecular pharmacological studies (RNA synthesis by T7 RNA polymerase in vitro) proved that 1-3 bind to DNA in a similar manner as cisplatin, since the RNA synthesis products of 1-3 and cisplatin showed a similar sequence profile of major bands.

• MeSH
• antitumorózní látky * chemická syntéza   chemie   farmakologie   MeSH
• cisplatina * analogy a deriváty   chemická syntéza   chemie   farmakologie   MeSH
• cytotoxiny * chemická syntéza   chemie   farmakologie   MeSH
• indoly chemie   MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• nádory farmakoterapie   metabolismus   MeSH
• RNA nádorová biosyntéza   MeSH
• RNA biosyntéza   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 7-azaindole dimer MeSH Prohlížeč
• antitumorózní látky * MeSH
• cisplatina * MeSH
• cytotoxiny * MeSH
• indoly MeSH
• RNA nádorová MeSH
• RNA MeSH

Certain hope is entertained in the prediction of chemosensitivity in vitro/ ex vivo for the purpose of selecting the most effective treatment of malignant diseases with minimal patient loading. The possible choice of an effective substance based on the results of a simple ex vivo test would increase the success of the treatment in case of standard chemotherapy failure or in the treatment of primary chemoresistant tumor. MTT test seems to be an easy process for the prediction of chemosensitivity of isolated malignant cells ex vivo, however each method represents a simple tool, which can provide false results if incorrectly preformed. Numerous limitations significantly reduce the successful evaluation and constituent aspects of the methodic press to further reflections about the proper application of the test.

• MeSH
• chemorezistence MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• nádorové buňky kultivované MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• přehledy MeSH

ATP-binding drug efflux transporters decrease intracellular concentrations of cytotoxic drugs, causing multidrug resistance in cancer. In this study, we examined possible interactions of ABCG2 transporter with platinum cytotoxic drugs. We demonstrate here an interference of platinum drugs with enhanced green fluorescence protein (EGFP) in the cellular models, where EGFP was employed as a reporter gene. Cytotoxicity of cisplatin (CIP), carboplatin (CAP) and oxaliplatin (OXP) was significantly lowered in MDCKII cells transfected with ABCG2 transporter and EGFP reporter. The IC(50) values in MDCKII-ABCG2 were 25.7, 164 and 165 microM for CIP, CAP and OXP, respectively, whereas IC(50) for the same cytostatics in MDCKII cells were as follows: 15.4, 133 and 50.3 microM. Addition of fumitremorgin C (FTC), a potent ABCG2 inhibitor, significantly suppressed the resistance of MDCKII-ABCG2 to OXP, suggesting that OXP interacts with ABCG2. However, FTC did not change the sensitivity of the cells to CIP and CAP. We assume that EGFP rather than ABCG2 causes the diminished toxicity of the platinum cytostatics in the transfected cells. This hypothesis was confirmed in human Hep2 cells expressing EGFP: using MTT test, IC(50) of 30.0, 247 and 27.9 microM were obtained for CIP, CAP and OXP, respectively, while 12.3, 106 and 20.5 microM were observed in the parent Hep2 cells. Employing neutral red cytotoxicity assay, similar data were obtained (IC(50) 7.73, 685 and 112 microM for CIP, CAP, and OXP, respectively, in the Hep2-EGFP cells and 1.65, 79.4 and 24.5 microM in the parent Hep2 cells). Caspase-3/7 assay revealed lower susceptibility of EGFP expressing Hep2 cells to apoptosis induced by CIP when compared to the parent cell line. We therefore conclude that EGFP in transfected cells interferes with cytotoxicity of platinum drugs by hindering the drug induced apoptosis and could cause misinterpretation of results obtained in cytotoxicity studies.

• MeSH
• ABC transportér z rodiny G, člen 2 MeSH
• ABC transportéry metabolismus   MeSH
• antitumorózní látky aplikace a dávkování   farmakologie   toxicita   MeSH
• apoptóza účinky léků   MeSH
• biologické modely MeSH
• chemorezistence MeSH
• cisplatina aplikace a dávkování   farmakologie   toxicita   MeSH
• inhibiční koncentrace 50 MeSH
• karboplatina aplikace a dávkování   farmakologie   toxicita   MeSH
• kultivované buňky MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• nádorové proteiny metabolismus   MeSH
• organoplatinové sloučeniny aplikace a dávkování   farmakologie   toxicita   MeSH
• oxaliplatin MeSH
• psi MeSH
• transfekce metody   MeSH
• zelené fluorescenční proteiny farmakologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• psi MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ABC transportér z rodiny G, člen 2 MeSH
• ABC transportéry MeSH
• ABCG2 protein, human MeSH Prohlížeč
• antitumorózní látky MeSH
• cisplatina MeSH
• enhanced green fluorescent protein MeSH Prohlížeč
• karboplatina MeSH
• nádorové proteiny MeSH
• organoplatinové sloučeniny MeSH
• oxaliplatin MeSH
• zelené fluorescenční proteiny MeSH

Determination of chemosensitivity/chemoresistance is becoming increasingly important for individualization of breast cancer chemotherapy. We developed a simple non-destructive test of cellular activity (NTCA) for assessment of the cytopathic effect of antitumour drugs in vitro. Contrary to routinely used methods (e.g. MTT), besides the comparative evaluation of metabolic activity using pH (given by the medium colour), the NTCA enables the simultaneous assessment of proliferation and morphology of cultured cells (phase-contrast microscopy) at any time during the incubation with cytostatics. Moreover, the regenerative potential of the cells can be examined by cell recovery and growth after drug removal. We provide evidence for the relevance of NTCA in chemosensitivity testing of primary breast cancer cells and breast cancer cell lines for cisplatin, gemcitabine and tamoxifen. NTCA represents a simple addition to the chemosensitivity assessment and could also serve for rapid screening of new antitumour strategies.

• MeSH
• antitumorózní látky farmakologie   MeSH
• buněčné linie MeSH
• buňky - růstové procesy účinky léků   MeSH
• cisplatina aplikace a dávkování   MeSH
• deoxycytidin analogy a deriváty   farmakologie   MeSH
• gemcitabin MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• mléčné žlázy lidské cytologie   účinky léků   MeSH
• nádorové buněčné linie MeSH
• nádory prsu farmakoterapie   metabolismus   patologie   MeSH
• protokoly antitumorózní kombinované chemoterapie farmakologie   MeSH
• receptory pro estrogeny biosyntéza   MeSH
• reprodukovatelnost výsledků MeSH
• tamoxifen aplikace a dávkování   MeSH
• tetrazoliové soli MeSH
• thiazoly MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antitumorózní látky MeSH
• cisplatina MeSH
• deoxycytidin MeSH
• gemcitabin MeSH
• receptory pro estrogeny MeSH
• tamoxifen MeSH
• tetrazoliové soli MeSH
• thiazoly MeSH
• thiazolyl blue MeSH Prohlížeč

A novel anti-tumor platinum(IV) complex, coded as LA-12, with a bulky adamantylamine ligand displaying oral activity was prepared and its oral activity was evaluated. The murine ADJ/PC6 plasmacytoma and human A2780 ovarian carcinoma tumor model were used to evaluate the in vivo anti-tumor activity of a single dose and also of repeated doses with comparison to the activity of cisplatin and of the platinum(IV) complex satraplatin. The acute toxicity of LA-12 in mice is relatively low (maximum tolerated dose 1000 mg/kg), and the effective dose is comparable to that of cisplatin and higher than that of satraplatin. The therapeutic index derived from this is very high (250). In the human tumor model, two repeated dose schedule regimens were evaluated. LA-12 exerted a significantly higher anti-tumor activity than other substances, i.e. cisplatin and satraplatin, in repeated doses on the murine ADJ/PC6 plasmacytoma tumor model. The dailyx5 repeated dose regimen was selected for further evaluation.

• MeSH
• adenokarcinom farmakoterapie   MeSH
• amantadin aplikace a dávkování   analogy a deriváty   MeSH
• antitumorózní látky aplikace a dávkování   MeSH
• aplikace orální MeSH
• inbrední kmeny myší MeSH
• léky antitumorózní - screeningové testy metody   MeSH
• lidé MeSH
• maximální tolerovaná dávka MeSH
• modely nemocí na zvířatech MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• nádorové buňky kultivované MeSH
• nádory vaječníků farmakoterapie   MeSH
• organoplatinové sloučeniny aplikace a dávkování   MeSH
• plazmocytom farmakoterapie   MeSH
• xenogenní modely - testy antitumorózní aktivity MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• amantadin MeSH
• antitumorózní látky MeSH
• bis(acetato)(1-adamantylamine)amminedichloroplatinum(IV) MeSH Prohlížeč
• organoplatinové sloučeniny MeSH
• satraplatin MeSH Prohlížeč