Identification of novel small molecules that inhibit STAT3-dependent transcription and function

Activation of Signal Transducer and Activator of Transcription 3 (STAT3) has been linked to several processes that are critical for oncogenic transformation, cancer progression, cancer cell proliferation, survival, drug resistance and metastasis. Inhibition of STAT3 signaling has shown a striking ability to inhibit cancer cell growth and therefore, STAT3 has become a promising target for anti-cancer drug development. The aim of this study was to identify novel inhibitors of STAT-dependent gene transcription. A cellular reporter-based system for monitoring STAT3 transcriptional activity was developed which was suitable for high-throughput screening (Z’ = 0,8). This system was used to screen a library of 28,000 compounds (the ENAMINE Drug-Like Diversity Set). Following counter-screenings and toxicity studies, we identified four hit compounds that were subjected to detailed biological characterization. Of the four hits, KI16 stood out as the most promising compound, inhibiting STAT3 phosphorylation and transcriptional activity in response to IL6 stimulation. In silico docking studies showed that KI16 had favorable interactions with the STAT3 SH2 domain, however, no inhibitory activity could be observed in the STAT3 fluorescence polarization assay. KI16 inhibited cell viability preferentially in STAT3-dependent cell lines. Taken together, using a targeted, cell-based approach, novel inhibitors of STAT-driven transcriptional activity were discovered which are interesting leads to pursue further for the development of anti-cancer therapeutic agents

Loss of functional pRB is not a ubiquitous feature of B-cell malignancies

Human cancers frequently sustain genetic mutations that alter the function of their G1 cell cycle control check point. These include changes to the retinoblastoma gene and to the genes that regulate its phosphorylation, such as the cyclin-dependent kinase inhibitor p16(INK4a). Altered expression of retinoblastoma protein (pRb) is associated with non-Hodgkin's lymphoma, particularly centroblastic and Burkitt's lymphomas. pRb is expressed in normal B-cells and its regulatory phosphorylation pathway is activated in response to a variety of stimuli. Since human B-lymphoma-derived cell lines are often used as in vitro model systems to analyse the downstream effects of signal transduction, we examined the functional status of pRb in a panel of human B-cell lines. We identified eleven cell lines which express the hyperphosphorylated forms of pRb. Furthermore, we suggest that the pRb protein appears to be functional in these cell lines

Особенности клеточных реакций в легких во время обострения бронхиальной астмы

The cell population in BALF of asthmatic patients and 9 subjects free of lung pathology was studied. The percentages of cells in BALF of asthmatics were: alveolar macrophages (AM) — 85,2+1,5 % C — 92,4+1,3 %, lymphocytes — 6,1 ±0,9 C — 5,8±1,2 %, neutrophils — 2,9+0,8 % C — 0,8+0,3 %, eosinophils — 5,8±0,8 % C —0,5±0,1%, mast cells — 0,19+0,1 % C — 0. The significant increase of absolute and relative number of eosinophils was revealed in BALF of asthmatic patients, the number of neutrophils didn’t correlate with eosinophil level.Acute stage of bronchial asthma is accompanied with AM destruction increase and functional activation of the rest AM population. Proportions of BALF cells in asthmatic patients reflect the character and degree of protective and inflammmatory reactions in lung. Acute bronchial astma is accompanied with bronchiolitis and alveolitis mainly of eosinophilic type.

RBSP3 (HYA22) is a tumor suppressor gene implicated in major epithelial malignancies

Chromosome 3p21.3 region is frequently (>90%) deleted in lung and other major human carcinomas. We subdivided 3p21.3 into LUCA and AP20 subregions and discovered frequent homozygous deletions (10-18%) in both subregions. This finding strongly implies that they harbor multiple tumor suppressor genes involved in the origin and/or development of major epithelial cancers. In this study, we performed an initial analysis of RBSP3/HYA22, a candidate tumor suppressor genes located in the AP20 region. Two sequence splice variants of RBSP3/HYA22 (A and B) were identified, and we provide evidence for their tumor suppressor function. By sequence analysis RBSP3/HYA22 belongs to a gene family of small C-terminal domain phosphatases that may control the RNA polymerase II transcription machinery. Expression of the gene was drastically (>20-fold) decreased in 11 of 12 analyzed carcinoma cell lines and in three of eight tumor biopsies. We report missense and nonsense mutations in tumors where RBSP3/HYA22 was expressed, growth suppression with regulated transgenes in culture, suppression of tumor formation in severe combined immunodeficient mice, and dephosphorylation of ppRB by RBSP3/HYA22, presumably leading to a block of the cell cycle at the G(1)/S boundary

Interferon α Induces Nucleus-independent Apoptosis by Activating Extracellular Signal-regulated Kinase 1/2 and c-Jun NH2-Terminal Kinase Downstream of Phosphatidylinositol 3-Kinase and Mammalian Target of Rapamycin

Interferon (IFN)α induces apoptosis via Bak and Bax and the mitochondrial pathway. Here, we investigated the role of known IFNα-induced signaling cascades upstream of Bak activation. By pharmacological and genetic inhibition of the kinases protein kinase C (PKC)δ, extracellular signal-regulated kinase (ERK), and c-Jun NH2-terminal kinase (JNK) in U266-1984 and RHEK-1 cells, we could demonstrate that all three enzymes are critical for the apoptosis-associated mitochondrial events and apoptotic cell death induced by IFNα, at a step downstream of phosphatidylinositol 3-kinase (PI3K) and mammalian target of rapamycin (mTOR). Furthermore, the activation of JNK was found to occur in a PKCδ/ERK-dependent manner. Inhibition of these kinases did not affect the canonical IFNα-stimulated Janus tyrosine kinase-signal transducer and activator of transcription signaling or expression of IFN-responsive genes. Therefore, enucleated cells (cytoplasts) were examined for IFNα-induced apoptosis, to test directly whether this process depends on gene transcription. Cytoplasts were found to undergo apoptosis after IFNα treatment, as analyzed by several apoptosis markers by using flow cytometry, live cell imaging, and biochemical analysis of flow-sorted cytoplasts. Furthermore, inhibition of mTOR, ERK, and JNK blocked IFNα-induced apoptosis in cytoplasts. In conclusion, IFNα-induced apoptosis requires activation of ERK1/2, PKCδ, and JNK downstream of PI3K and mTOR, and it can occur in a nucleus-independent manner, thus demonstrating for the first time that IFNα induces apoptosis in the absence of de novo transcription