Analysis in epithelial ovarian cancer identifies KANSL1 as a biomarker and target gene for immune response and HDAC inhibition.

OBJECTIVE: There is an immunoreactive subtype of ovarian cancer with a favorable prognosis, but the majority of ovarian cancers have limited immune reactivity. The reason for this is poorly understood. This study aimed to approach this question by identifying prognostically relevant genes whose prognostic mRNA expression levels correlated with a genomic event. METHODS: Expression microarray and 5-year survival data on 170 ovarian tumors and aCGH data on 45 ovarian cancer cell lines were used to identify amplified/deleted genes associated with prognosis. Three immune-response genes were identified mapping to epigenetically modified chromosome 6p21.3. Genes were searched for roles in epigenetic modification, identifying KANSL1. Genome-wide association studies were searched to identify genetic variants in KANSL1 associated with altered immune profile. Sensitivity to HDAC inhibition in cell lines with KANSL1 amplification/rearrangement was studied. RESULTS: Expression of 196 genes was statistically significantly associated with survival, and expression levels correlated with copy number variations for 82 of them. Among these, 3 immune-response genes (HCP5, PSMB8, PSMB9) clustered together at epigenetically modified chromosome 6p21.3 and their expression was inversely correlated to epigenetic modification gene KANSL1. KANSL1 is amplified/rearranged in ovarian cancer, associated with lymphocyte profile, a biomarker for response to HDAC inhibition, and may drive expression of immune-response genes. CONCLUSION: This study identifies 82 genes with prognostic relevance and genomic alteration in ovarian cancer. Among these, immune-response genes have correlated expression which is associated with 5-year survival. KANSL1 may be a master gene altering immune-response gene expression at 6p21.3 and drive response to HDAC inhibitors. Future research should investigate KANSL1 and determine whether targeting it alters the immune profile of ovarian cancer and improves survival, HDAC inhibition, and/or immunotherapy response.

Lack of Benefit and Potential Harm of Induction Therapy in Simultaneous Liver-Kidney Transplants.

The number of simultaneous liver-kidney transplantations (SLKTs) and use of induction therapy for SLKT have increased recently, without much published evidence, especially in the context of maintenance immunosuppression containing tacrolimus (TAC) and mycophenolic acid (MPA). We queried the Organ Procurement and Transplant Network registry for SLKT recipients maintained on TAC/MPA at discharge in the United States for 2002-2016. The cohort was divided into 3 groups on the basis of induction type: rabbit antithymocyte globulin (r-ATG; n = 831), interleukin 2 receptor antagonist (IL2RA; n = 1558), and no induction (n = 2333). Primary outcomes were posttransplant all-cause mortality and acute rejection rates in kidney and liver allografts at 12 months. Survival rates were analyzed by the Kaplan-Meier method. A propensity score analysis was used to control potential selection bias. Multivariate inverse probability weighted Cox proportional hazard and logistic regression models were used to estimate the hazard ratios (HRs) and odds ratios. Among SLKT recipients, survival estimates at 3 years were lower for recipients receiving r-ATG (P = 0.05). Compared with no induction, the multivariate analyses showed an increased mortality risk with r-ATG (HR, 1.29; 95% confidence interval [CI], 1.10-1.52; P = 0.002) and no difference in acute liver or kidney rejection rates at 12 months across all induction categories. No difference in outcomes was noted with IL2RA induction over the no induction category. In conclusion, there appears to be no survival benefit nor reduction in rejection rates for SLKT recipients who receive induction therapy, and r-ATG appears to increase mortality risk compared with no induction.

Proteasome ubiquitin receptor PSMD4 is an amplification target in breast cancer and may predict sensitivity to PARPi.

Poly (ADP-ribose) polymerase 1 (PARP1) is an enzyme involved in DNA repair under investigation as a chemotherapeutic target. Current randomized phase three trials of PARPi in metastatic breast cancer are limited to patients with documented BRCA1/2 mutations and no biomarker of PARPi beyond BRCA status is available. In an effort to identify novel biomarkers for PARP inhibition, we created a cell line (HCC1187/TALRES) resistant to the PARP1 inhibitor talazoparib. Herein we show by array-CGH that HCC1187/TALRES has a selective loss of the proteasome ubiquitin receptor PSMD4 amplicon resulting in significant down-regulation of PSMD4. Conversely, we find that breast cancer cell lines that have copy number gain or amplification for PSMD4 are significantly more sensitive to talazoparib. Functional studies reveal that knock-down of PSMD4 in amplified breast cancer cells and loss of the PSMD4 amplicon result in knock-down of PARP1 protein. We show that PSMD4 is amplified and overexpressed in breast cancer and its overexpression correlates with poor survival. Knock-down of PSMD4 results in a significant decrease in cell growth. We provide evidence that PSMD4 is a proteasomal amplification target in breast cancer that PSMD4 amplification confers sensitivity to PARP inhibition, and that PSMD4 amplification is lost in the process of acquiring resistance to PARPi. Finally, this study shows not only that PSMD4 copy number correlates with PARPi sensitivity, but also, that it may be a better predictor of sensitivity to PARPi than BRCA1/2 mutation.

ADRM1-amplified metastasis gene in gastric cancer.

The proteasome ubiquitin receptor ADRM1 has been shown to be a driver for 20q13.3 amplification in epithelial cancers including ovarian and colon cancer. We performed array-CGH on 16 gastric cancer cell lines and found 20q13.3 to be amplified in 19% with the minimal amplified region in gastric cancer cell line AGS spanning a 1 Mb region including ADRM1. Expression microarray analysis shows overexpression of only two genes in the minimal region, ADRM1 and OSBPL2. While RNAi knockdown of both ADRM1 and OSBPL2 led to a slight reduction in growth, only ADRM1 RNAi knockdown led to a significant reduction in migration and growth in soft-agar. Treatment of AGS cells with the ADRM1 inhibitor RA190 resulted in proteasome inhibition, but RNAi knockdown of ADRM1 did not. However, RNAi knockdown of ADRM1 led to a significant reduction in specific proteins including MNAT1, HRS, and EGFR. We hypothesize that ADRM1 may act in ADRM1-amplified gastric cancer to alter protein levels of specific oncogenes resulting in an increase in metastatic potential. Selective inhibition of ADRM1 independent of proteasome inhibition may result in a targeted therapy for ADRM1-amplified gastric cancer. In vivo models are now warranted to validate these findings. © 2015 Wiley Periodicals, Inc.

In vitro activity of the mTOR inhibitor everolimus, in a large panel of breast cancer cell lines and analysis for predictors of response.

Everolimus (RAD001, Afinitor(®)) is an oral, selective mTOR inhibitor recently approved by the US-FDA in combination with exemestane for treatment of hormone receptor positive advanced breast cancer. To date, no molecular predictors of response to everolimus in breast cancer have been identified. We hypothesized predictive markers could be identified using preclinical models. Using a molecularly characterized panel of human breast cancer and immortalized breast epithelial cell lines, we determined sensitivity to everolimus alone or in combination with ER- or HER2- targeted therapy. Gene expression microarrays and comparative genomic hybridization were performed on the cell lines to identify predictors of response to everolimus. Among 13 everolimus-sensitive cell lines, 10/13(77 %) were luminal, while in 26 resistant cell lines, 16/26(62 %) were non-luminal, and 10/26(38 %) were luminal. Only 3/24 non-luminal lines were sensitive, two of which were HER2+. Everolimus enhanced the anti-proliferative effect of both tamoxifen (TAM) and fulvestrant (FUL) in ER+ breast cancer cell lines, as well as trastuzumab in HER2+ cell lines. Everolimus + FUL but not everolimus + TAM reversed acquired resistance to TAM. Everolimus inhibited mTOR in tested cell lines by decreasing S6 phosphorylation, mediating its anti-proliferative effect by G0/G1 cell cycle arrest and induction of apoptosis. Chromosomal amplifications of AURKA (p value = 0.04) and HER2 (p value = 0.03) were each associated with increased sensitivity to everolimus. Transcript expression microarrays identified GSK3A, PIK3R3, KLF8, and MAPK10 among the genes overexpressed in sensitive luminal lines, while PGP, RPL38, GPT, and GFAP were among the genes overexpressed in resistant luminal cell lines. These preclinical in vitro data provide further support for continued clinical development of everolimus in luminal (ER+ or HER2+) breast cancer in combination with targeted therapies. We identified several potential molecular markers associated with response to everolimus that will require validation in clinical material.

POSTN/TGFBI-associated stromal signature predicts poor prognosis in serous epithelial ovarian cancer.

OBJECTIVE: To identify molecular prognosticators and therapeutic targets for high-grade serous epithelial ovarian cancers (EOCs) using genetic analyses driven by biologic features of EOC pathogenesis. METHODS: Ovarian tissue samples (n = 172; 122 serous EOCs, 30 other EOCs, 20 normal/benign) collected prospectively from sequential patients undergoing gynecologic surgery were analyzed using RNA expression microarrays. Samples were classified based on expression of genes with potential relevance in ovarian cancer. Gene sets were defined using Rosetta Similarity Search Tool (ROAST) and analysis of variance (ANOVA). Gene copy number variations were identified by array comparative genomic hybridization. RESULTS: No distinct subgroups of EOC could be identified by unsupervised clustering, however, analyses based on genes correlated with periostin (POSTN) and estrogen receptor-alpha (ESR1) yielded distinct subgroups. When 95 high-grade serous EOCs were grouped by genes based on ANOVA comparing ESR1/WT1 and POSTN/TGFBI samples, overall survival (OS) was significantly shorter for 43 patients with tumors expressing genes associated with POSTN/TGFBI compared to 52 patients with tumors expressing genes associated with ESR1/WT1 (median 30 versus 49 months, respectively; P = 0.022). Several targets with therapeutic potential were identified within each subgroup. BRCA germline mutations were more frequent in the ESR1/WT1 subgroup. Proliferation-associated genes and TP53 status (mutated or wild-type) did not correlate with survival. Findings were validated using independent ovarian cancer datasets. CONCLUSIONS: Two distinct molecular subgroups of high-grade serous EOCs based on POSTN/TGFBI and ESR1/WT1 expressions were identified with significantly different OS. Specific differentially expressed genes between these subgroups provide potential prognostic and therapeutic targets.

AMG 900, pan-Aurora kinase inhibitor, preferentially inhibits the proliferation of breast cancer cell lines with dysfunctional p53.

Aurora kinases play important roles in cell division and are frequently overexpressed in human cancer. AMG 900 is a novel pan-Aurora kinase inhibitor currently being tested in Phase I clinical trials. We aimed to evaluate the in vitro activity of AMG 900 in a panel of 44 human breast cancer and immortalized cell lines and identify predictors of response. AMG 900 inhibited proliferation at low nanomolar concentrations in all cell lines tested. Response was further classified based on the induction of lethality. 25 cell lines were classified as highly sensitive (lethality at 10 nM of AMG 900 >10 %), 19 cell lines as less sensitive to AMG 900 (lethality at 10 nM of AMG 900 <10 %). Traditional molecular subtypes of breast cancer did not predict for this differential response. There was a weak association between AURKA amplification and response to AMG 900 (response ratio = 2.53, p = 0.09). mRNA expression levels of AURKA, AURKB, and AURKC and baseline protein levels of Aurora kinases A and B did not significantly associate with response. Cell lines with TP53 loss of function mutations (RR = 1.86, p = 0.004) and low baseline p21 protein levels (RR = 2.28, p = 0.0004) were far more likely to be classified as highly sensitive to AMG 900. AMG 900 induced p53 and p21 protein expression in cell lines with wt TP53. AMG 900 caused the accumulation of cells with >4 N DNA content in a majority of cell lines independently of sensitivity and p53 status. AMG 900 induced more pronounced apoptosis in highly sensitive p53-dysfunctional cell lines. We have found that AMG 900 is highly active in breast cancer cell lines and that TP53 loss of function mutations as well as low baseline expression of p21 protein predict strongly for increased sensitivity to this compound in vitro.

Amplification Target ADRM1: Role as an Oncogene and Therapeutic Target for Ovarian Cancer.

Approximately 25,000 ovarian cancers are diagnosed in the U.S. annually, and 75% are in the advanced stage and largely incurable. There is critical need for early detection tools and novel treatments. Proteasomal ubiquitin receptor ADRM1 is a protein that is encoded by the ADRM1 gene. Recently, we showed that among 20q13-amplified genes in ovarian cancer, ADRM1 overexpression was the most highly correlated with amplification and was significantly upregulated with respect to stage, recurrence, and metastasis. Its overexpression correlated significantly with shorter time to recurrence and overall survival. Array-CGH and microarray expression of ovarian cancer cell lines provided evidence consistent with primary tumor data that ADRM1 is a 20q13 amplification target. Herein, we confirm the ADRM1 amplicon in a second ovarian cancer cohort and define a minimally amplified region of 262 KB encompassing seven genes. Additionally, using RNAi knock-down of ADRM1 in naturally amplified cell line OAW42 and overexpression of ADRM1 via transfection in ES2, we show that (1) ADRM1 overexpression increases proliferation, migration, and growth in soft agar, and (2) knock-down of ADRM1 results in apoptosis. Proteomic analysis of cells with ADRM1 knock-down reveals dysregulation of proteins including CDK-activating kinase assembly factor MAT1. Taken together, the results indicate that amplified ADRM1 is involved in cell proliferation, migration and survival in ovarian cancer cells, supporting a role as an oncogene and novel therapeutic target for ovarian cancer.

Community science designed ribosomes with beneficial phenotypes.

Functional design of ribosomes with mutant ribosomal RNA (rRNA) can expand opportunities for understanding molecular translation, building cells from the bottom-up, and engineering ribosomes with altered capabilities. However, such efforts are hampered by cell viability constraints, an enormous combinatorial sequence space, and limitations on large-scale, 3D design of RNA structures and functions. To address these challenges, we develop an integrated community science and experimental screening approach for rational design of ribosomes. This approach couples Eterna, an online video game that crowdsources RNA sequence design to community scientists in the form of puzzles, with in vitro ribosome synthesis, assembly, and translation in multiple design-build-test-learn cycles. We apply our framework to discover mutant rRNA sequences that improve protein synthesis in vitro and cell growth in vivo, relative to wild type ribosomes, under diverse environmental conditions. This work provides insights into rRNA sequence-function relationships and has implications for synthetic biology.

Knockdown of ovarian cancer amplification target ADRM1 leads to downregulation of GIPC1 and upregulation of RECK.

Approximately 25,000 ovarian cancers are diagnosed in the United States annually, and 75% of cases are in the advanced stage when they are largely incurable. There is a critical need for improved early detection tools and development of novel treatments. Recently, we showed that among 20q13-amplified genes in ovarian cancer, ADRM1 overexpression was the most highly correlated with amplification and was significantly upregulated with respect to stage, recurrence, and metastasis. In addition, overexpression of ADRM1 correlated significantly with shorter time to recurrence and overall survival. Herein, array-CGH and microarray expression of ovarian cancer cell lines provides evidence consistent with the primary tumor data that ADRM1 is a 20q13 amplification target. Knockdown of ADRM1 in amplified ovarian cell-line OAW42 results in downregulation of growth factor GIPC1 and upregulation of tumor-suppressor RECK RNA and protein. In our dataset of 141 ovarian primary tumors, ADRM1 overexpression significantly correlates with GIPC1 overexpression. In addition, there is a significant anticorrelation between ADRM1 overexpression and RECK expression. Further research is necessary to determine whether targeting knockdown of ADRM1 in 20q13-amplified ovarian cancers results in growth inhibition and tumor suppression via downstream targets GIPC1 and RECK.

Evaluation of well-differentiated/de-differentiated liposarcomas by high-resolution oligonucleotide array-based comparative genomic hybridization.

Well-differentiated/de-differentiated liposarcomas (WDLS/DDLS) encompass an intriguing disease model in which a temporal intersection occurs between the malignant transformation of mesenchymal cells and the process of adipogenesis. Deciphering the molecular events that trigger and are characteristic of the intersection of these oncogenic and normal processes is critical to affect the often morbid and lethal consequences of malignant tumors of fat. High-resolution genome-wide oligonucleotide array-based comparative genomic hybridization (aCGH) with matched gene expression analyses was performed on seven lipomas, one hibernoma, and 38 WD and DDLS to define and compare the genomic events associated with these tumors. WD and DDLS had complex karyotypes. On average, WDLS had 11.1 and DDLS had 22.7 chromosomal copy number aberrations. All of the liposarcomas had 12q13-q15 amplifications with varying peaks at CDK4 (12q14.1), HMGA2 (12q14.3), and MDM2 (12q15); 24% of the DDLS and no WDLS had 1p32.2 (JUN) amplifications; 33% WDLS and 35% DDLS had 1q24.3 amplifications involving DNM3 and miR-214/miR-199a2; 24% of the liposarcomas had 6q23-q24 amplifications (including MAP3K5). Amplifications in GLI1 (12q13.3), JUN, and MAP3K5 (6q23.3) were mutually exclusive and occurred predominately in the DDLS. 6q amplifications occurred primarily in retroperitoneal tumors and females represented the majority of those patients who developed fatty tumors prior to the age of 50 years old. This detailed genetic mapping provides insight into the heterogeneity of WD and DDLS and the chromosomal and genetic abnormalities that are present in and distinguish these mesenchymal malignancies.

Periostin promotes ovarian cancer angiogenesis and metastasis.

OBJECTIVE: Perostin (PN) has been found to be overexpressed in a variety of human malignancies including ovarian cancer. In the present study, we investigated PN expression status in a large cohort of ovarian tumors with the focus on biological influence of PN related on ovarian tumor angiogenesis and metastasis. METHODS: PN expression was determined by cDNA microarray, PN northern blot and PN IHC tissue array analyses. Exogenous PN expression in ovarian cancer cells OVCAR-3 and OV2008 were achieved through retroviral transfection and confirmed by PN western blot and ELISA. The effects of exogenous PN expression on tumor angiogenesis and metastatic growth were accessed in orthotopic mouse models. The in vitro cell adhesion, migration and invasion assays were performed to investigate the potential mechanisms involved in PN's in vivo effects. RESULTS: PN was frequently overexpressed in ovarian tumors. Higher PN levels significantly correlated with clinical late stages (III/IV) and cancer recurrence. PN was produced by engineered PN-overexpressing cells at levels comparable to that of A2780 cells, an ovarian carcinoma cell line with endogenous PN expression. PN overexpression did not change cell growth rates in vitro; however it significantly promoted intraperitoneal tumor metastatic growth in immunodeficient mice, which was associated with increased tumor angiogenesis and decreased tumor cell apoptosis. In vitro purified PN promoted cell adhesion, migration, and invasion of both human umbilical endothelial cells (HUVECs) and/or ovarian cancer cells. CONCLUSIONS: Our data indicate PN plays a critical role in both ovarian tumor angiogenesis and metastasis. Thus PN may represent a clinically effective new target for therapy of ovarian cancer.

Quality and access - Early experience of implementing a virtual stereotactic chart round across a national network.

INTRODUCTION: Stereotactic radiation therapy is a highly specialised technique which requires careful and structured implementation. As part of a national stereotactic programme implementation, protocols were developed and a national stereotactic chart round was formed, which strongly recommended attendance and presentation of all cases before treatment. Herein, we describe our experiences launching a national chart round and its importance in a stereotactic programme. METHOD: Stereotactic chart rounds were held via videoconference between July 2018 and July 2019. Data collected included attendances, patient-related information including, diagnosis, clinical background, treatment intent, prescribed dose and fractionation and technical approach. Consensus recommendations regarding changes to treatment approaches were also recorded. RESULTS: For the 12 months recorded, there were 1126 attendances, from 144 individual attendees, across 21 locations. In total, 285 cases (237 new cases, and 48 re-presentations) were presented by 27 radiation oncologists (ROs) from 13 different locations. From the cases presented, 65 changes were recommended from 53 patients (22.3%), including 27 (11.4%) changes to contours, 18 (7.6%) changes to dose prescription/fractionation, 9 (3.8%) changes to plan dosimetry, 1 (0.4%) changes to treatment technique and 10 (4.2%) recommendations for which stereotactic radiation therapy was not advised. A significant inverse relationship was found between frequency of recommended changes and the individual RO's stereotactic case load (P < 0.002). CONCLUSION: The implementation of a national stereotactic chart held via videoconference has ensured national protocol compliance across the network of locations. Furthermore, the chart rounds have allowed the entire multidisciplinary team to be provided with mentorship and guidance. Increasing number of cases presented was associated with lower rates of recommended changes highlighting the impact of experience and the need for continued mentorship.

Controlling the Structures, Flexibility, Conductivity Stability of Three-Dimensional Conductive Networks of Silver Nanoparticles/Carbon-Based Nanomaterials with Nanodispersion and their Application in Wearable Electronic Sensors.

This research has successfully synthesized highly flexible and conductive nanohybrid electrode films. Nanodispersion and stabilization of silver nanoparticles (AgNPs) were achieved via non-covalent adsorption and with an organic polymeric dispersant and inorganic carbon-based nanomaterials-nano-carbon black (CB), carbon nanotubes (CNT), and graphene oxide (GO). The new polymeric dispersant-polyisobutylene-b-poly(oxyethylene)-b-polyisobutylene (PIB-POE-PIB) triblock copolymer-could stabilize AgNPs. Simultaneously, this stabilization was conducted through the addition of mixed organic/inorganic dispersants based on zero- (0D), one- (1D), and two-dimensional (2D) nanomaterials, namely CB, CNT, and GO. Furthermore, the dispersion solution was evenly coated/mixed onto polymeric substrates, and the products were heated. As a result, highly conductive thin-film materials (with a surface electrical resistance of approximately 10-2 Ω/sq) were eventually acquired. The results indicated that 2D carbon-based nanomaterials (GO) could stabilize AgNPs more effectively during their reductNion and, hence, generate particles with the smallest sizes, as the COO- functional groups of GO are evenly distributed. The optimal AgNPs/PIB-POE-PIB/GO ratio was 20:20:1. Furthermore, the flexible electrode layers were successfully manufactured and applied in wearable electronic sensors to generate electrocardiograms (ECGs). ECGs were, thereafter, successfully obtained.

Comprehensive analysis of 20q13 genes in ovarian cancer identifies ADRM1 as amplification target.

Approximately 25,000 ovarian cancers are diagnosed in the US annually, and 75% of cases are in the advanced stage when they are largely incurable. There is a critical need for improved early detection tools and development of novel treatments. Because chromosome band 20q13 is a commonly DNA amplified region in ovarian cancer and increase in 20q13 copy number may be an early event, we examined the DNA amplification and RNA expression pattern of 239 microarray probes mapping to this region with the goal of identifying gene(s) associated with ovarian cancer. Using Agilent expression microarray analysis and FISH to tumor tissue arrays, we narrowed the candidates to 19 genes that were consistently overexpressed in a subset of tumors amplified for both ZNF217 and TPD54, although, interestingly the candidates do not include these two amplified genes. Unsupervised clustering of 225 ovarian samples with respect to RNA expression of these 19 genes allowed identification of a 20q-amplified subset of 51 (23%) tumors and this subset was significantly correlated with poor outcome. Of the 19 candidate genes in this subset, ADRM1 overexpression was the most highly correlated with amplification, was amplified in a higher percentage of tumors than ZNF217 and TPD54, and was significantly upregulated with respect to stage, recurrence and metastasis. In addition, overexpression of ADRM1 correlates significantly with shorter time to recurrence and overall survival. Functional analysis is now warranted to determine whether ADRM1 is a target for early screening and/or therapy for ovarian cancer.

Principles for Predicting RNA Secondary Structure Design Difficulty.

Designing RNAs that form specific secondary structures is enabling better understanding and control of living systems through RNA-guided silencing, genome editing and protein organization. Little is known, however, about which RNA secondary structures might be tractable for downstream sequence design, increasing the time and expense of design efforts due to inefficient secondary structure choices. Here, we present insights into specific structural features that increase the difficulty of finding sequences that fold into a target RNA secondary structure, summarizing the design efforts of tens of thousands of human participants and three automated algorithms (RNAInverse, INFO-RNA and RNA-SSD) in the Eterna massive open laboratory. Subsequent tests through three independent RNA design algorithms (NUPACK, DSS-Opt and MODENA) confirmed the hypothesized importance of several features in determining design difficulty, including sequence length, mean stem length, symmetry and specific difficult-to-design motifs such as zigzags. Based on these results, we have compiled an Eterna100 benchmark of 100 secondary structure design challenges that span a large range in design difficulty to help test future efforts. Our in silico results suggest new routes for improving computational RNA design methods and for extending these insights to assess "designability" of single RNA structures, as well as of switches for in vitro and in vivo applications.

Enzyme-catalysed nitrate reduction-themes and variations as revealed by protein film voltammetry.

Protein film voltammetry has been used to define the catalytic performance of two nitrate reductases: the respiratory nitrate reductase, NarGH, from Paracoccus pantotrophus and the assimilatory nitrate reductase, NarB, from Synechococcus sp. PCC 7942. NarGH and NarB present distinct "fingerprints" of catalytic activity when viewed in this way. Potentials that provide insufficient driving force for significant rates of nitrate reduction by NarB result in appreciable rates of nitrate reduction by NarGH. However, both enzymes display complex modulations in their rate of substrate reduction when viewed across the electrochemical potential domain.

PD-0332991, a potent and selective inhibitor of cyclin-dependent kinase 4/6, demonstrates inhibition of proliferation in renal cell carcinoma at nanomolar concentrations and molecular markers predict for sensitivity.

BACKGROUND: PD-0332991 is an inhibitor of cyclin-dependent kinases (CDK) 4 and 6, and was evaluated to determine its anti-proliferative effects in 25 renal cell carcinoma (RCC) cell lines. MATERIALS AND METHODS: Half-maximal inhibitory concentrations (IC50) of PD-0332991 were determined with cell line proliferation assays, as were its effects on the cell cycle, apoptosis, and retinoblastoma (RB) phosphorylation. Molecular markers for response prediction, including p16, p15, cyclin D1 (CCND1), cyclin E1 (CCNE1), E2F transcription factor 1 (E2F1), RB, CDK4 and CDK6, were studied using array comparative genomic hybridization (CGH) and gene expression. RESULTS: IC50 values for PD-0332991 ranged from 25.0 nM to 700 nM, and the agent demonstrated G0/G1 cell-cycle arrest, induction of late apoptosis, and blockade of RB phosphorylation. Through genotype and expression data p16, p15 and E2F1 were identified as having significant association between loss and sensitivity to PD-0332991: p16 (p=0.021), p15 (p=0.047), and E2F1 (p=0.041). CONCLUSION: PD-0332991 has antiproliferative activity in RCC cell lines, and molecular markers predict for sensitivity to this agent.

Activity of the fibroblast growth factor receptor inhibitors dovitinib (TKI258) and NVP-BGJ398 in human endometrial cancer cells.

The recent identification of activating fibroblast growth factor receptor 2 (FGFR2) mutations in endometrial cancer has generated an opportunity for a novel target-based therapy. Here, we explore the therapeutic potential of 2 FGFR inhibitors, the multikinase inhibitor dovitinib (TKI258) and the more selective FGFR inhibitor NVP-BGJ398 for the treatment of endometrial cancer. We examined the effects of both inhibitors on tumor cell growth, FGFR2 signaling, cell cycle, and apoptosis using a panel of 20 molecularly characterized human endometrial cancer cell lines. Anchorage-independent growth was studied using soft agar assays. In vivo studies were conducted using endometrial cancer xenograft models. Cell lines with activating FGFR2 mutations (S252W, N550K) were more sensitive to dovitinib or NVP-BGJ398 when compared with their FGFR2 wild-type counterparts (P = 0.073 and P = 0.021, respectively). Both agents inhibited FGFR2 signaling, induced cell-cycle arrest, and significantly increased apoptosis in FGFR2-mutant lines. In vitro, dovitinib and NVP-BGJ398 were both potent at inhibiting cell growth of FGFR2-mutant endometrial cancer cells, but the activity of dovitinib was less restricted to FGFR2-mutant lines when compared with NVP-BGJ398. In vivo, dovitinib and NVP-BGJ398 significantly inhibited the growth of FGFR2-mutated endometrial cancer xenograft models. In addition, dovitinib showed significant antitumor activity in FGFR2 wild-type endometrial cancer xenograft models including complete tumor regressions in a long-term in vivo study. Dovitinib and NVP-BGJ398 warrant further clinical evaluation in patients with FGFR2-mutated endometrial cancer. Dovitinib may have antitumor activity in endometrial cancer beyond FGFR2-mutated cases and may permit greater flexibility in patient selection.