Cost-effectiveness of genome sequencing for diagnosing patients with undiagnosed rare genetic diseases.

PURPOSE: To estimate the cost-effectiveness of genome sequencing (GS) for diagnosing critically ill infants and noncritically ill pediatric patients (children) with suspected rare genetic diseases from a United States health sector perspective. METHODS: A decision-analytic model was developed to simulate the diagnostic trajectory of patients. Parameter estimates were derived from a targeted literature review and meta-analysis. The model simulated clinical and economic outcomes associated with 3 diagnostic pathways: (1) standard diagnostic care, (2) GS, and (3) standard diagnostic care followed by GS. RESULTS: For children, costs of GS ($7284) were similar to that of standard care ($7355) and lower than that of standard care followed by GS pathways ($12,030). In critically ill infants, when cost estimates were based on the length of stay in the neonatal intensive care unit, the lowest cost pathway was GS ($209,472). When only diagnostic test costs were included, the cost per diagnosis was $17,940 for standard, $17,019 for GS, and $20,255 for standard care followed by GS. CONCLUSION: The results of this economic model suggest that GS may be cost neutral or possibly cost saving as a first line diagnostic tool for children and critically ill infants.

The Effect of Temperature and Moisture on Colonization of Apple Fruit and Branches by Botryosphaeria dothidea.

Botryosphaeria dothidea causes severe disease of apple trees in China. The process of conidium germination, colonization, and infection of apple fruit and branches was examined on 'Fuji' apple and the effect of temperature, surface wetness and relative humidity (RH), and host surface washates on these processes was studied in controlled environments. Initial germ tube development and hyphal growth resulted in the colonization of the host surface without forming an infection structure. Hyphae expanded radially across the host surface and, after entering lenticels, developed into a dense mycelium mass or differentiated pseudoparenchyma. Hyphae from the bottom of the pseudoparenchyma either directly penetrated the lenticel surface intercellularly through the cell layer, or formed an undifferentiated hypha that invaded the lenticel through cracks formed during the lenticel development. Conidial germination and hyphal colonization occurred at 10 to 40°C, with an optimum of approximately 28°C. Conidial germination required an RH > 95% or surface wetness but, for hyphal colonization, an RH > 90% was sufficient. Conidia germinated and formed germ tubes within 1 h under optimum conditions. However, the pathogen required a longer period at RH > 90% or surface wetness for hyphae to colonize and form pseudoparenchyma or dense mycelia on the host surface. Hyphal colonization is a crucial stage for infection of apple tissues by B. dothidea.

Progression of Symptoms Caused by Botryosphaeria dothidea on Apple Branches.

Until recently, the causal agent of Botryosphaeria canker was assumed to differ from that causing ring rot on fruit and warts on branches on apple trees in China and East Asia. However, recent research documented that Botryosphaeria dothidea caused both disease symptoms on apple. Inoculations with strains isolated from cankers and warts on branches were conducted to investigate symptom progression caused by B. dothidea and conditions inducing the two symptom types. The results confirmed that both cankers and warts are caused by B. dothidea. Warts are the results of hyperplasia and suberization of bark tissues induced by fungal infection, whereas cankers result from the rapid growth of hyphae from inside warts, lenticels, or wounds. Resistance to B. dothidea exists in living apple branches. When a living branch is infected via lenticels, the pathogen induces proliferation and suberization of cortical cells that restricts the growth and expansion of the hyphae, leading to warts. However, under certain stress conditions such as drought, the hyphae inside host tissues expand rapidly and kill cortical cells, leading to canker development. Host resistance may recover during active growth periods, which suppresses or even stops rapid expansion of the hyphae, leading to the intermediate symptom of canker warts. Abiotic factors, such as drought or high temperature in early spring, can result in rapid extension of colonized hyphae in branches and conversion of warts to cankers. Preventing this transition can be an important measure in managing Botryosphaeria canker on apple.

Effect of Rainfall and Temperature on Perithecium Production of Botryosphaeria dothidea on Cankered Apple Branches.

Botryosphaeria dothidea is a fungal pathogen causing canker, dieback, and fruit rot of apple trees worldwide. Ascospores are an important source of inoculum of Botryosphaeria canker in China. Experiments were conducted under both controlled and natural conditions to study perithecium formation in relation to environmental conditions. Perithecia of B. dothidea were detected on cankered lesions throughout the apple growing season except in July and in some years including August under natural conditions. On newly formed canker lesions, the first perithecium was detected as early as August, about 1 week after rainfall. Perithecia matured successively, lasting from early August to June of the next year, with a peak in late September or early October. Temperature and rainfall are two key environmental factors affecting perithecium formation. Under controlled conditions, perithecia were produced only on cankered shoots incubated at test temperatures of 20 and 25°C and wetted by >3 days of simulated rainfall per week. The number of perithecia produced on canker lesions increased with the increase in rainfall duration. Perithecia were formed on canker shoots exposed to rainfall only in June, July, and August but not in September. Rainfall of >3 days per week can be used to predict the initial formation of perithecia in the main apple production areas in China to assist disease management.

Excision of uracil from DNA by hSMUG1 includes strand incision and processing.

Uracil arises in DNA by hydrolytic deamination of cytosine (C) and by erroneous incorporation of deoxyuridine monophosphate opposite adenine, where the former event is devastating by generation of C → thymine transitions. The base excision repair (BER) pathway replaces uracil by the correct base. In human cells two uracil-DNA glycosylases (UDGs) initiate BER by excising uracil from DNA; one is hSMUG1 (human single-strand-selective mono-functional UDG). We report that repair initiation by hSMUG1 involves strand incision at the uracil site resulting in a 3'-α,ß-unsaturated aldehyde designated uracil-DNA incision product (UIP), and a 5'-phosphate. UIP is removed from the 3'-end by human apurinic/apyrimidinic (AP) endonuclease 1 preparing for single-nucleotide insertion. hSMUG1 also incises DNA or processes UIP to a 3'-phosphate designated uracil-DNA processing product (UPP). UIP and UPP were indirectly identified and quantified by polyacrylamide gel electrophoresis and chemically characterised by matrix-assisted laser desorption/ionisation time-of-flight mass-spectrometric analysis of DNA from enzyme reactions using 18O- or 16O-water. The formation of UIP accords with an elimination (E2) reaction where deprotonation of C2' occurs via the formation of a C1' enolate intermediate. A three-phase kinetic model explains rapid uracil excision in phase 1, slow unspecific enzyme adsorption/desorption to DNA in phase 2 and enzyme-dependent AP site incision in phase 3.

Food Microstructure and Fat Content Affect Growth Morphology, Growth Kinetics, and Preferred Phase for Cell Growth of Listeria monocytogenes in Fish-Based Model Systems.

Food microstructure significantly affects microbial growth dynamics, but knowledge concerning the exact influencing mechanisms at a microscopic scale is limited. The food microstructural influence on Listeria monocytogenes (green fluorescent protein strain) growth at 10°C in fish-based food model systems was investigated by confocal laser scanning microscopy. The model systems had different microstructures, i.e., liquid, xanthan (high-viscosity liquid), aqueous gel, and emulsion and gelled emulsion systems varying in fat content. Bacteria grew as single cells, small aggregates, and microcolonies of different sizes (based on colony radii [size I, 1.5 to 5.0 µm; size II, 5.0 to 10.0 µm; size III, 10.0 to 15.0 µm; and size IV, ≥15 µm]). In the liquid, small aggregates and size I microcolonies were predominantly present, while size II and III microcolonies were predominant in the xanthan and aqueous gel. Cells in the emulsions and gelled emulsions grew in the aqueous phase and on the fat-water interface. A microbial adhesion to solvent assay demonstrated limited bacterial nonpolar solvent affinities, implying that this behavior was probably not caused by cell surface hydrophobicity. In systems containing 1 and 5% fat, the largest cell volume was mainly represented by size I and II microcolonies, while at 10 and 20% fat a few size IV microcolonies comprised nearly the total cell volume. Microscopic results (concerning, e.g., growth morphology, microcolony size, intercolony distances, and the preferred phase for growth) were related to previously obtained macroscopic growth dynamics in the model systems for an L. monocytogenes strain cocktail, leading to more substantiated explanations for the influence of food microstructural aspects on lag phase duration and growth rate.IMPORTANCEListeria monocytogenes is one of the most hazardous foodborne pathogens due to the high fatality rate of the disease (i.e., listeriosis). In this study, the growth behavior of L. monocytogenes was investigated at a microscopic scale in food model systems that mimic processed fish products (e.g., fish paté and fish soup), and the results were related to macroscopic growth parameters. Many studies have previously focused on the food microstructural influence on microbial growth. The novelty of this work lies in (i) the microscopic investigation of products with a complex composition and/or structure using confocal laser scanning microscopy and (ii) the direct link to the macroscopic level. Growth behavior (i.e., concerning bacterial growth morphology and preferred phase for growth) was more complex than assumed in common macroscopic studies. Consequently, the effectiveness of industrial antimicrobial food preservation technologies (e.g., thermal processing) might be overestimated for certain products, which may have critical food safety implications.

Inoculation of drought-stressed strawberry with a mixed inoculum of two arbuscular mycorrhizal fungi: effects on population dynamics of fungal species in roots and consequential plant tolerance to water deficiency.

The effect of inoculation with two arbuscular mycorrhizal fungi (AMF) on growth and drought tolerance of cultivated strawberry (Fragaria × ananassa) was studied. Three treatments (a single treatment either of Funneliformis mosseae BEG25, Funneliformis geosporus BEG11 or a 50:50 mixed inoculation treatment of both species) were compared to uninoculated plants. Species-specific primers for qPCR quantification of F. geosporus and F. mosseae DNA were developed to quantify the relative abundance of each fungus in roots of strawberry under different conditions of water stress. Co-occupation of the same root by both species was shown to commonly occur, but their relative abundance varied with water stress (reduced irrigation of up to 40%). Greater root colonisation was observed microscopically under water stress, but this increased colonisation was often accompanied with decreased amounts of fungal DNA in the root. F. mosseae tended to become more abundant under water stress relative to F. geosporus. There was significant correlation in the fungal colonisation measurements from the microscopic and qPCR methods under some conditions, but the nature of this relationship varied greatly with AMF inoculum and abiotic conditions. Single-species inoculation treatments gave similar benefits to the host to the mixed inoculation treatment regardless of irrigation regime; here, amount of colonisation was of greater importance than functional diversity. The addition of AMF inocula to plants subjected to reduced irrigation restored plant growth to the same or higher values as the non-mycorrhizal, fully-watered plants. The water use efficiency of plants was greater under the regulated deficit irrigation (RDI) regime and in AMF-inoculated plants, but there were no significant differences between plants inoculated with the single or combined inoculum. This study demonstrated that the increase in plant growth was directly influenced by an increase in root colonisation by AMF when individual plants were examined.

Parkinson disease protein DJ-1 binds metals and protects against metal-induced cytotoxicity.

The progressive loss of motor control due to reduction of dopamine-producing neurons in the substantia nigra pars compacta and decreased striatal dopamine levels are the classically described features of Parkinson disease (PD). Neuronal damage also progresses to other regions of the brain, and additional non-motor dysfunctions are common. Accumulation of environmental toxins, such as pesticides and metals, are suggested risk factors for the development of typical late onset PD, although genetic factors seem to be substantial in early onset cases. Mutations of DJ-1 are known to cause a form of recessive early onset Parkinson disease, highlighting an important functional role for DJ-1 in early disease prevention. This study identifies human DJ-1 as a metal-binding protein able to evidently bind copper as well as toxic mercury ions in vitro. The study further characterizes the cytoprotective function of DJ-1 and PD-mutated variants of DJ-1 with respect to induced metal cytotoxicity. The results show that expression of DJ-1 enhances the cells' protective mechanisms against induced metal toxicity and that this protection is lost for DJ-1 PD mutations A104T and D149A. The study also shows that oxidation site-mutated DJ-1 C106A retains its ability to protect cells. We also show that concomitant addition of dopamine exposure sensitizes cells to metal-induced cytotoxicity. We also confirm that redox-active dopamine adducts enhance metal-catalyzed oxidation of intracellular proteins in vivo by use of live cell imaging of redox-sensitive S3roGFP. The study indicates that even a small genetic alteration can sensitize cells to metal-induced cell death, a finding that may revive the interest in exogenous factors in the etiology of PD.

Modeling the Adenoma and Serrated pathway to Colorectal CAncer (ASCCA).

Several colorectal cancer (CRC) screening models have been developed describing the progression of adenomas to CRC. Currently, there is increasing evidence that serrated lesions can also develop into CRC. It is not clear whether screening tests have the same test characteristics for serrated lesions as for adenomas, but lower sensitivities have been suggested. Models that ignore this type of colorectal lesions may provide overly optimistic predictions of the screen-induced reduction in CRC incidence. To address this issue, we have developed the Adenoma and Serrated pathway to Colorectal CAncer (ASCCA) model that includes the adenoma-carcinoma pathway and the serrated pathway to CRC as well as characteristics of colorectal lesions. The model structure and the calibration procedure are described in detail. Calibration resulted in 19 parameter sets for the adenoma-carcinoma pathway and 13 for the serrated pathway that match the age- and sex-specific adenoma and serrated lesion prevalence in the COlonoscopy versus COlonography Screening (COCOS) trial, Dutch CRC incidence and mortality rates, and a number of other intermediate outcomes concerning characteristics of colorectal lesions. As an example, we simulated outcomes for a biennial fecal immunochemical test screening program and a hypothetical one-time colonoscopy screening program. Inclusion of the serrated pathway influenced the predicted effectiveness of screening when serrated lesions are associated with lower screening test sensitivity or when they are not removed. To our knowledge, this is the first model that explicitly includes the serrated pathway and characteristics of colorectal lesions. It is suitable for the evaluation of the (cost)effectiveness of potential screening strategies for CRC.

Induction of apoptosis in hormone-resistant human prostate cancer PC3 cells by inactivated Sendai virus.

OBJECTIVE: Inactivated Sendai virus particle [hemagglutinating virus of Japan envelope (HVJ-E)] has a potential oncolytic effect due to its ability to induce apoptosis in tumor cells. However, the molecular mechanism of apoptosis induction in cancer cells mediated by HVJ-E has not been fully elucidated. This paper aims to investigate the underlying mechanism of apoptosis induction by HVJ-E in prostate cancer cells (PC3). METHODS: PC3 cells were treated with HVJ-E at various MOI, and then interferon-ß (IFN-ß) production, and the cell viability and apoptosis were detected by ELISA, MTT-based assay and flow cytometry, respectively. Next, the roles of Jak-Stat, MAPK and Akt pathways played in HVJ-E-induced apoptosis in PC3 cells were analyzed by immunoblot assay. To further evaluate the cytotoxic effect of HVJ-E on PC3 cells, HVJ-E was intratumorally injected into prostate cancers on BALB/c-nude mice, and the tumor volume was monitored for 36 days. RESULTS: HVJ-E induced IFN-ß production and activated Jak-Stat signaling pathway, which resulted in the activation of caspase-8, caspase-3, and PARP in PC3 prostate cancer cells post HVJ-E treatment. Furthermore, we observed for the first time that p38 and Jnk MAPKs in PC3 cells contributed to HVJ-E-induced apoptosis. In addition, intratumoral HVJ-E treatment displayed a direct inhibitory effect in an in vivo BALB/c nude mouse prostate cancer model. CONCLUSION: Our findings have provided novel insights into the underlying mechanisms by which HVJ-E induces apoptosis in tumor cells.

The Arabidopsis DJ-1a protein confers stress protection through cytosolic SOD activation.

Mutations in the DJ-1 gene (also known as PARK7) cause inherited Parkinson's disease, which is characterized by neuronal death. Although DJ-1 is thought to be an antioxidant protein, the underlying mechanism by which loss of DJ-1 function contributes to cell death is unclear. Human DJ-1 and its Arabidopsis thaliana homologue, AtDJ-1a, are evolutionarily conserved proteins, indicating a universal function. To gain further knowledge of the molecular features associated with DJ-1 dysfunction, we have characterized AtDJ-1a. We show that AtDJ-1a levels are responsive to stress treatment and that AtDJ-1a loss of function results in accelerated cell death in aging plants. By contrast, transgenic plants with elevated AtDJ-1a levels have increased protection against environmental stress conditions, such as strong light, H(2)O(2), methyl viologen and copper sulfate. We further identify superoxide dismutase 1 (SOD1) and glutathione peroxidase 2 (GPX2) as interaction partners of both AtDJ-1a and human DJ-1, and show that this interaction results in AtDJ-1a- and DJ-1-mediated cytosolic SOD1 activation in a copper-dependent fashion. Our data have highlighted a conserved molecular mechanism for DJ-1 and revealed a new protein player in the oxidative stress response of plants.

Inactivated Sendai virus suppresses murine melanoma growth by inducing host immune responses and down-regulating ß-catenin expression.

OBJECTIVE: This paper aims to investigate the anti-tumor mechanism of inactivated Sendai virus (Hemagglutinating virus of Japan envelope, HVJ-E) for murine melanoma (B16F10). METHODS: The murine dendritic cells (DCs) were treated with HVJ-E, and then the cytokines secreted from DCs and costimulation-related molecules on DCs were measured. Meanwhile, the expression of ß-catenin in HVJ-E treated murine melanoma cells was detected. In addition, HVJ-E was intratumorally injected into the melanoma on C57BL/6 mice, and the immune cells, CTL response and tumor volume were analyzed. RESULTS: HVJ-E injected into B16F10 melanoma obviously inhibited the growth of the tumor and prolonged the survival time of the tumor-bearing mice. Profiles of cytokines secreted by dendritic cells (DCs) after HVJ-E stimulation showed that the number of cytokines released was significantly higher than that elicited by PBS (1P<0.05). The co-stimulation-related molecules on DCs were comparable to those stimulated by LPS. Immunohistochemical examinations demonstrated the repression of ß-catenin in B16F10 melanoma cells after HVJ-E treatment. Meanwhile, real-time reverse transcription PCR revealed that HVJ-E induced a remarkable infiltration of CD11c positive cells, chemokine ligand 10 (CXCL10) molecules, interleukin-2 (IL-2) molecule, CD4(+) and CD8(+) T cells into HVJ-E injected tumors. Furthermore, the mRNA expression level of ß-catenin in the HVJ-E injected tumors was also down-regulated. In addition, B16F10-specific CTLs were induced significantly after HVJ-E was injected into the tumor-bearing mice. CONCLUSION: This is the first report to show the effective inhibition of melanoma tumors by HVJ-E alone and the mechanism through which it induces antitumor immune responses and regulates important signal pathways for melanoma invasion. Therefore, HVJ-E shows its prospect as a novel therapeutic for melanoma therapy.

Intrinsic Strand-Incision Activity of Human UNG: Implications for Nick Generation in Immunoglobulin Gene Diversification.

Uracil arises in cellular DNA by cytosine (C) deamination and erroneous replicative incorporation of deoxyuridine monophosphate opposite adenine. The former generates C → thymine transition mutations if uracil is not removed by uracil-DNA glycosylase (UDG) and replaced by C by the base excision repair (BER) pathway. The primary human UDG is hUNG. During immunoglobulin gene diversification in activated B cells, targeted cytosine deamination by activation-induced cytidine deaminase followed by uracil excision by hUNG is important for class switch recombination (CSR) and somatic hypermutation by providing the substrate for DNA double-strand breaks and mutagenesis, respectively. However, considerable uncertainty remains regarding the mechanisms leading to DNA incision following uracil excision: based on the general BER scheme, apurinic/apyrimidinic (AP) endonuclease (APE1 and/or APE2) is believed to generate the strand break by incising the AP site generated by hUNG. We report here that hUNG may incise the DNA backbone subsequent to uracil excision resulting in a 3´-α,ß-unsaturated aldehyde designated uracil-DNA incision product (UIP), and a 5´-phosphate. The formation of UIP accords with an elimination (E2) reaction where deprotonation of C2´ occurs via the formation of a C1´ enolate intermediate. UIP is removed from the 3´-end by hAPE1. This shows that the first two steps in uracil BER can be performed by hUNG, which might explain the significant residual CSR activity in cells deficient in APE1 and APE2.

Arabidopsis stromal 70-kDa heat shock proteins are essential for chloroplast development.

70 kDa heat shock proteins (Hsp70s) act as molecular chaperones involved in essential cellular processes such as protein folding and protein transport across membranes. They also play a role in the cell's response to a wide range of stress conditions. The Arabidopsis family of Hsp70s homologues includes two highly conserved proteins, cpHsc70-1 and cpHsc70-2 which are both imported into chloroplasts (Su and Li in Plant Physiol 146:1231-1241, 2008). Here, we demonstrate that YFP-fusion proteins of both cpHsc70-1 and cpHsc70-2 are predominantly stromal, though low levels were detected in the thylakoid membrane. Both genes are ubiquitously expressed at high levels in both seedlings and adult plants. We further show that both cpHsc70-1 and cpHsc70-2 harbour ATPase activity which is essential for Hsp70 chaperone activity. A previously described T-DNA insertion line for cpHsc70-1 (DeltacpHsc70-1) has variegated cotyledons, malformed leaves, growth retardation, impaired root growth and sensitivity to heat shock treatment. In addition, under stress conditions, this mutant also exhibits unusual sepals, and malformed flowers and sucrose concentrations as low as 1% significantly impair growth. cpHsc70-1/cpHsc70-2 double-mutants are lethal. However, we demonstrate through co-suppression and artificial microRNA (amiRNA) approaches that transgenic plants with severely reduced levels of both genes have a white and stunted phenotype. Interestingly, chloroplasts in these plants have an unusual morphology and contain few or no thylakoid membranes. Our data show that cpHsc70-1 and cpHsc70-2 are essential ATPases, have overlapping roles and are required for normal plastid structure.

[Significance of K-ras detection in colorectal cancer].

OBJECTIVE: To determine the mutation status of K-ras gene in colorectal cancer and analyze the associations between its mutation status and clinicopathological characteristics in colorectal cancer so as to select the patients likely to benefit from a targeted therapy. METHODS: A total of 208 colorectal cancer tissue samples were collected from September 2008 to February 2009. DNA was extracted with a genomic DNA miniprep kit. Then PCR was performed with the designed primers and the product directly sequenced by the Sanger method. Then the associations between K-ras mutation status and clinicopathological characteristics in colorectal cancer were analyzed. RESULTS: Of 208 cases, 91 cases of K-ras gene mutation were detected. The 12 or 13 codon had a mutation rate of 43.8%. There were no significant differences in gender, tumor location, histopathological grading and Duke's stage between the wild and mutated groups. CONCLUSION: Detection of K-ras gene status in colorectal cancer will help to select the patients likely to benefit from the monoclonal antibody therapy of targeting epidermal growth factor receptor.

Pathological significance of abnormal recepteur d'origine nantais and programmed death ligand 1 expression in colorectal cancer.

BACKGROUND: Programmed death ligand 1 (PD-L1) immunotherapy remains poorly efficacious in colorectal cancer (CRC). The recepteur d'origine nantais (RON) receptor tyrosine kinase plays an important role in regulating tumor immunity. AIM: To identify the patterns of RON and PD-L1 expression and explore their clinical significance in CRC. METHODS: Gene expression data from the Gene Expression Omnibus database (GEO; n = 290) and patients at the First Affiliated Hospital, Zhejiang University School of Medicine (FAHZUSM; n = 381) were analyzed to determine the prognostic value of RON and PD-L1 expression within the tumor microenvironment of CRC. HT29 cell line was treated with BMS-777607 to explore the relationship between RON activity and PD-L1 expression. Signaling pathways and protein expression perturbed by RON inhibition were evaluated by cellular immunofluorescence and Western blot. RESULTS: In the GEO patient cohort, cut-off values for RON and PD-L1 expression were determined to be 7.70 and 4.3, respectively. Stratification of patients based on these cutoffs demonstrated that high expression of RON and PD-L1 was associated with a poor prognosis. In the FAHZUSM cohort, rates of high expression of RON in tumor cells, high PD-L1 expression in tumor cells and tumor infiltrating monocytes, and both high RON and high PD-L1 expression in the tumor microenvironment were 121 (32%), 43 (11%), 91 (24%), and 51 (13.4%), respectively. High expression of RON was significantly correlated with high expression of PD-L1 in the tumor cell compartment (P < 0.001). High expression of RON and that of PD-L1 were independent prognostic factors for poorer overall survival. Concurrent high expression of both RON and PD-L1 in the tumor microenvironment was significantly associated with a poor prognosis. In vitro, BMS-777607 inhibited the phosphorylation of RON, inhibited PD-L1 expression, and attenuated activation of the ERK1/2 and AKT signaling pathways in CRC cells. CONCLUSION: RON, PD-L1, and their crosstalk are significant in predicting the prognostic value of CRC. Moreover, phosphorylation of RON upregulates PD-L1 expression, which provides a novel approach to immunotherapy in CRC.

RON and MET Co-overexpression Are Significant Pathological Characteristics of Poor Survival and Therapeutic Targets of Tyrosine Kinase Inhibitors in Triple-Negative Breast Cancer.

PURPOSE: Triple-negative breast cancer (TNBC) is highly malignant and has poor prognosis and a high mortality rate. The lack of effective therapy has spurred our investigation of new targets for treating this malignant cancer. Here, we identified RON (macrophage-stimulating 1 receptor) and MET (MET proto-oncogene, receptor tyrosine kinase) as a prognostic biomarker and therapeutic targets for potential TNBC treatment. MATERIALS AND METHODS: We analyzed RON and MET expression in 187 primary TNBC clinical samples with immunohistochemistry. We validated the targeted therapeutic effects of RON and MET in TNBC using three tyrosine kinase inhibitors (TKIs): BMS-777607, INCB28060, and tivantinib. The preclinical therapeutic efficacy of the TKIs was mainly estimated using a TNBC xenograft model. RESULTS: Patients with TNBC had widespread, abnormal expression of RON and MET. There was RON overexpression, MET overexpression, and RON and MET co-overexpression in 63 (33.7%), 63 (33.7%), and 43 cases (23.0%), respectively, which had poor prognosis and short survival. In vivo, the TKI targeting RON ant MET inhibited the activation of the downstream signaling molecules, inhibited TNBC cell migration and proliferation, and increased TNBC cell apoptosis; in the xenograft model, they significantly inhibited tumor growth and shrank tumor volumes. The TKI targeting RON and Met, such as BMS-777607 and tivantinib, yielded stronger anti-tumor effects than INCB28060. CONCLUSION: RON and MET co-overexpression can be significant pathological characteristics in TNBC for poor prognosis. TKIs targeting RON and MET have stronger drug development potential for treating TNBC.

Aberrant RON and MET Co-overexpression as Novel Prognostic Biomarkers of Shortened Patient Survival and Therapeutic Targets of Tyrosine Kinase Inhibitors in Pancreatic Cancer.

RON (recepteur d'origine nantais) and MET (hepatocyte growth factor receptor) are tyrosine kinase receptors. Various cancers have aberrant RON and MET expression and activation, which contribute to cancer cell proliferation, invasiveness, and metastasis. Here, we explored RON and MET expression in pancreatic cancer and their relationship with overall survival (OS) time, and evaluated their significance as therapeutic targets of tyrosine kinase inhibitors in pancreatic cancer. We enrolled 227 patients with pancreatic cancer in the study. RON and MET expression was analyzed by immunohistochemical staining. Four human pancreatic cancer cell lines expressing variable levels of RON or MET and four MET superfamily inhibitors (BMS777607, PHA665752, INCB28060, Tivantinib) were used. The effect of the four tyrosine kinase inhibitors on cell viability, migration, and apoptosis were determined using cell viability, scratch wound healing, and Caspase-Glo 3/7 assays. Cellular signaling was analyzed by immunoprecipitation and western blotting. The therapeutic efficacy of the tyrosine kinase inhibitors was determined with mouse xenograft pancreatic cancer models in vivo. There was wide aberrant RON and MET expression in the cancer tissues. In 227 pancreatic cancer samples, 33% had RON overexpression, 41% had MET overexpression, and 15.4% had RON and MET co-overexpression. RON and MET expression were highly correlated. RON and MET expression levels were significantly related to OS. Patients with RON and MET co-overexpression had poorer OS. BMS777607 and PHA665752 inhibited pancreatic cancer cell viability and migration, and promoted apoptosis by inhibiting RON and MET phosphorylation and further inhibiting the downstream signaling pathways in vitro. They also inhibited tumor growth and further inhibited phosphorylated (phosphor)-RON and phospho-MET expression in the mouse xenograft models in vivo effectively. INCB28060, which inhibits the MET signaling pathway alone, was not effective. RON and MET can be important indicators of prognosis in pancreatic cancer. Tyrosine kinase inhibitors targeting RON and MET in pancreatic cancer are a novel and potential approach for pancreatic cancer therapy.

Adult sacrococcygeal teratoma: a retrospective study over eight years at a single institution.

OBJECTIVE: To determine the clinical, imaging, and histological features, and surgical resection modalities and outcomes of adult sacrococcygeal teratoma (SCT). METHODS: Adult patients with histopathologically diagnosed SCT were enrolled in our hospital between August 2010 and August 2018. Each patient's characteristics and clinical information were reviewed. RESULTS: There were 8 patients in the study (2 males, 6 females) with a median age of 34 years (range, 18-67 years). The time to clinical symptoms was 14 d to 35 years, with a median time of 4 years. Six patients presented with symptoms of sacrococcygeal pain, and four with signs of sacrococcygeal mass and ulceration in the sacrococcygeal region. Six patients were evaluated using a combination of computed tomography (CT) and magnetic resonance imaging (MRI). All patients showed a presacral tumor with heterogeneous intensity on CT images. All patients underwent surgical treatment, including 6 parasacral, 1 transabdominal, and 1 combined anterior-posterior surgery cases. Seven patients were histopathologically diagnosed with benign mature SCT, and have shown no recurrence. One patient had malignant SCT, with recurrence at 84 months after surgery. After a second surgery, the patient had no recurrence within 6 months follow-up after re-resection. CONCLUSIONS: Our retrospective study demonstrated: (1) adult SCT is difficult to diagnose because of a lack of typical clinical symptoms and signs; (2) a combination of CT and MRI examination is beneficial for preoperative diagnosis; (3) the choice of surgical approach and surgical resection modality depends on the size, location, and components of the tumor, which can be defined from preoperative CT and MRI evaluation; (4) most adult SCTs are benign; the surgical outcome for the malignant SCT patient was good after complete resection. Even for the patient with recurrent malignant SCT, the surgical outcome was good after re-resection.

Iron-sulfur cluster biogenesis systems and their crosstalk.

The biogenesis of iron-sulfur clusters ([Fe-S]) plays a very important role in many essential functions of life. Several [Fe-S] biogenesis systems have been discovered, such as the NIF (nitrogen fixation), SUF (mobilisation of sulfur) and ISC (iron-sulfur cluster) systems in bacteria, and the ISC-like and CIA (cytosolic iron-sulfur protein assembly) systems in yeast. Experimental evidence has revealed that SUF and ISC in bacteria communicate with each other partly through IscR to coordinate the utilisation of iron and cysteine. The ISC-like system in yeast is localised to the mitochondria, while the ISC-dependent CIA system is localised to the cytosol; this suggests a possible role for the ISC mitochondrial export machinery in mediating crosstalk between the two systems. Based on genetic analysis, the model plant Arabidopsis thaliana contains three [Fe-S] biogenesis systems similar to SUF, ISC and CIA named AtSUF, AtISC and AtCIA. Possible communication between these three systems has been proposed.