Significant Growth Delay Research Articles

Agroforestry enables high efficiency of light capture, photosynthesis and dry matter production in a semi-arid climate

Agroforestry systems, which combine annual crops with trees, are used widely in semi-arid regions to reduce wind erosion and improve resource (e.g. water) use efficiency. Limited knowledge is available on optimizing such systems by the choice of crop species with specific physiological traits (i.e. C3 vs C4, N-fixing vs non-N-fixing). In this study we quantified the light interception and utilization efficiency of trees and crops in agroforestry systems comprising apricot trees and a C3 species (sweet potato), a C4 species (millet) or an N-fixing legume species (peanut), and used measurements in the sole stands as a reference. A significant delay in leaf growth was found in millet. Maximum LAI of millet was 17% higher in agroforestry then expected from sole crop LAI, taking into account the relative density of 2/3, while a 25% decrease in maximum LAI compared to expected was observed in peanut and sweet potato. The total light interception in agroforestry was 54% higher than in sole tree stands and 23% higher than in sole crops. The millet intercepted more light and produced more biomass in agroforestry than peanut and sweet potato. The LUE values of the crops in the mixed systems were higher than those of the sole crops, as was the photosynthetic efficiency of individual leaves, especially in plants in the border rows of the crop strips. High light capture in agroforestry made a greater contribution to productivity of understory crops than the increases in light use efficiency. We conclude that agroforestry systems with apricot trees and annual crops, especially millet, can improve light utilization in semi-arid climates and contribute to regional sustainability and adaptation to climate change.

Electrotransfer of Different Control Plasmids Elicits Different Antitumor Effectiveness in B16.F10 Melanoma.

Several studies have shown that different control plasmids may cause antitumor action in different murine tumor models after gene electrotransfer (GET). Due to the differences in GET protocols, plasmid vectors, and experimental models, the observed antitumor effects were incomparable. Therefore, the current study was conducted comparing antitumor effectiveness of three different control plasmids using the same GET parameters. We followed cytotoxicity in vitro and the antitumor effect in vivo after GET of control plasmids pControl, pENTR/U6 scr and pVAX1 in B16.F10 murine melanoma cells and tumors. Types of cell death and upregulation of selected cytosolic DNA sensors and cytokines were determined. GET of all three plasmids caused significant growth delay in melanoma tumors; nevertheless, the effect of pVAX1 was significantly greater than pControl. While DNA sensors in vivo were not upregulated significantly, cytokines IFN β and TNF α were upregulated after GET of pVAX1. In vitro, the mRNAs of some cytosolic DNA sensors were overexpressed after GET; however, with no significant difference among the three plasmids. In summary, although differences in antitumor effects were observed among control plasmids in vivo, no differences in cellular responses to plasmid GET were detected in tumor cells in vitro. Thus, the tumor microenvironment as well as some plasmid properties are most probably responsible for the antitumor effectiveness.

Overcoming Resistance to Cetuximab with Honokiol, A Small-Molecule Polyphenol.

Overexpression and activation of the EGFR have been linked to poor prognosis in several human cancers. Cetuximab is a mAb against EGFR that is used for the treatment in head and neck squamous cell carcinoma (HNSCC) and metastatic colorectal cancer. Unfortunately, most tumors have intrinsic or will acquire resistance to cetuximab during the course of therapy. Honokiol is a natural compound found in the bark and leaves of the Chinese Magnolia tree and is established to have several anticancer properties without appreciable toxicity. In this study, we hypothesized that combining cetuximab and honokiol treatments could overcome acquired resistance to cetuximab. We previously developed a model of acquired resistance to cetuximab in non-small cell lung cancer H226 cell line. Treatment of cetuximab-resistant clones with honokiol and cetuximab resulted in a robust antiproliferative response. Immunoblot analysis revealed the HER family and their signaling pathways were downregulated after combination treatment, most notably the proliferation (MAPK) and survival (AKT) pathways. In addition, we found a decrease in phosphorylation of DRP1 and reactive oxygen species after combination treatment in cetuximab-resistant clones, which may signify a change in mitochondrial function. Furthermore, we utilized cetuximab-resistant HNSCC patient-derived xenografts (PDX) to test the benefit of combinatorial treatment in vivo There was significant growth delay in PDX tumors after combination treatment with a subsequent downregulation of active MAPK, AKT, and DRP1 signaling as seen in vitro Collectively, these data suggest that honokiol is a promising natural compound in overcoming acquired resistance to cetuximab. Mol Cancer Ther; 17(1); 204-14. ©2017 AACR.

Qualitative improvement of rabbit burgers using Zingiber officinale Roscoe powder

The object of this study was to evaluate the effect of Zingiber officinale powder on physical-chemical traits, microbiological growth and sensory properties of rabbit burger. Raw burgers (only meat and meat added with 1 and 2% w/w ginger powder) were stored at 4°C for 1, 4 and 7 d and then cooked. Ginger modified the colour of both raw and cooked burgers, leading to more yellow hue and reducing lightness. Aspect of burgers were affected by ginger powder addition, leading to a noticeable difference between the samples. During storage time, the highest modifications were recorded for control samples, followed by burgers with added ginger. Sensory evaluation highlighted that ginger enhanced the juiciness of the burgers; moreover, burgers with ginger powder presented a significant delay in microbial growth. Ginger powder might be considered as a potential ingredient in rabbit meat products to increase their quality and extend their shelf-life.

Selenotriapine – An isostere of the most studied thiosemicarbazone with pronounced pro-apoptotic activity, low toxicity and ability to challenge phenotype reprogramming of 3-D mammary adenocarcinoma tumors

Triapine, the most studied α-N-heterocyclic thiosemicarbazone, revealed potent activity against advanced leukemia, but was ineffective against a variety of solid tumors. Moreover, methemoglobinemia, which is a side effect of triapine administration, may limits all clinical application. To enhance anticancer activity and reduce side effects, we applied an isosteric replacement of sulfur to selenium atom was performed by synthesis and characterization of selenium triapine analog, 3-aminopyridine-2-carboxaldehyde selenosemicarbazone (selenotriapine). Compared to triapine, selenotriapine revealed superior pro-apoptotic activity with activation of intrinsic apoptotic pathway in both human monocytic leukemia (THP-1) and mammary adenocarcinoma (MCF-7) cell lines. For MCF-7 2-D cultures, selenotriapine induced notable increase in mitochondrial superoxide radical generation and dissipation of mitochondrial transmembrane potential. A significant delay in growth of MCF-7 spheroids (3-D culture) was accompanied by phenotypic stem cell reprogramming (Oct-4 expression). Additionally, selenotriapine demonstrated a very low toxicity profile as compared to triapine, confirmed over alleviated extent of methemoglobin formation and higher IC50 value in brine shrimp cytotoxicity assay.

Influence of particle properties on iron flocculation

Abstract In this study, the importance of charge interactions during flocculation of Fe3+ in the presence of particles and anions/cations at various pH values was investigated. SiO2, (s) and ZnO(s) were dosed as particles to promote charge interactions and/or serve as a nucleus to accelerate floc formation. In the pH range 6–9, SiO2, (s) is negatively charged, while ZnO(s) carries a positive charge. Ca2+ and HPO42− were selected to investigate charge interactions in the water phase. A significant delay in floc growth due to charge repulsion between negatively charged iron species was observed at pHini 9. For positively charged species at pHini 6, a delay in floc growth was observed as well, but to a lesser degree. These effects could be neutralized by either dosing (positively charged) ZnO(s) or Ca2+ at pHini 9, or (negatively charged) SiO2, (s) at pHini 6. The addition of phosphate did not hinder floc growth at pHini 6. While phosphate completely inhibited floc growth at pHini 7–9 in the presence of negatively charged SiO2, (s), the presence of positively charged ZnO(s) partly neutralized the detrimental influence of phosphate on floc growth. Similarly, dosing Ca2+ partly neutralized the effect of phosphate.

First-Trimester Crown-Rump Length and Risk of Chromosomal Aberrations-A Systematic Review and Meta-analysis.

Lower than expected first-trimester crown-rump length (CRL) is a common sonographic finding, usually leading to reassessment of gestational age. The aim of this study was to perform a meta-analysis defining the risk of chromosomal aberrations in pregnancies with decreased first-trimester CRL. A search was conducted by a research librarian in 5 databases, with no time or language restrictions. Original researches examining the risk of chromosomal aberrations in pregnancies with low CRL were selected. The effect estimates were presented as odds ratios with 95% confidence intervals). Following screening of 3894 references, 12 articles were selected. Only 2 articles directly fulfilled the review goals, comparing the rates of chromosomal aberrations in small-CRL versus normal-CRL fetuses. Combined analysis of these 2 studies, encompassing a total of 403 fetuses with small CRL and 4047 control pregnancies, yielded an odds ratio of 5.54 (95% confidence interval, 1.2-26.1) for abnormal karyotype in small-CRL fetuses. Ten studies compared CRL in fetuses with specific chromosomal aberrations to pregnancies with normal karyotype, yielding a significant growth delay in fetuses with trisomy 18 (6 of 7 articles) and triploidy, but not in fetuses with trisomy 21. Overall quality of evidence was rated as "very low" using Grading of Recommendations Assessment, Development and Evaluation criteria. Our review results indicate that low first-trimester CRL might be associated with a significantly increased risk of chromosomal anomalies. Thus, invasive prenatal testing or cell-free DNA screening might be offered in such pregnancies, particularly if dating is certain.

Anti-tumor Activity of miniPEG-γ-Modified PNAs to Inhibit MicroRNA-210 for Cancer Therapy.

MicroRNAs (miRs) are frequently overexpressed in human cancers. In particular, miR-210 is induced in hypoxic cells and acts to orchestrate the adaptation of tumor cells to hypoxia. Silencing oncogenic miRs such as miR-210 may therefore offer a promising approach to anticancer therapy. We have developed a miR-210 inhibition strategy based on a new class of conformationally preorganized antisense γ peptide nucleic acids (γPNAs) that possess vastly superior RNA-binding affinity, improved solubility, and favorable biocompatibility. For cellular delivery, we encapsulated the γPNAs in poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs). Our results show that γPNAs targeting miR-210 cause significant delay in growth of a human tumor xenograft in mice compared to conventional PNAs. Further, histopathological analyses show considerable necrosis, fibrosis, and reduced cell proliferation in γPNA-treated tumors compared to controls. Overall, our work provides a chemical framework for a novel anti-miR therapeutic approach using γPNAs that should facilitate rational design of agents to potently inhibit oncogenic microRNAs.

Abstract 4599: Antibody targeting PDZ domain of TIP-1 induces proliferation arrest through AKT/mTOR signaling inhibition in lung cancer and glioblastoma

Abstract Antigens that are over-expressed in cancer in response to radiation are being used as novel targets. We showed tax interacting protein 1 (TIP-1) to be radiation-inducible that translocated to the surface of the cancer cell following irradiation. TIP-1, which consists of a single PDZ domain plays an important role in cell signaling, cancer development, and progression. TIP-1’s involvement in various survival pathways makes it an attractive target for anticancer therapeutics. We used antibodies specific to this PDZ domain to determine its role in cancer cell survival. We monitored proliferation in lung cancer (A549 and H460) and glioblastoma (D54 and U251) cells after 24, 48, 72 and 96h treatment with the anti-PDZ antibody. We observed a time-dependent proliferation arrest with anti-PDZ antibody treatment which was associated with increased apoptosis. The anti-PDZ antibody when combined with radiation (3Gy) led to reduced proliferation and colony formation. Anti-PDZ antibody had no effect on the proliferation of normal lung (MRC-5) and endothelial (HUVEC) cells. Cells treated with anti-PDZ antibody showed decreased levels of the phosphorylated forms of AKT, mTOR, and a downstream substrate of mTOR, 4EBP1. Anti-PDZ antibody treatment also led to an overall reduction in basal levels of AKT, mTOR, and 4EBP1. Further, we evaluated the effect of the anti-PDZ antibody on tumor growth in heterotopic mouse models of lung cancer (A549) and glioma (U251). We observed significant growth delay in mice treated with anti-PDZ antibody treatment when compared to mice treated with the isotype control. The combination of the anti-PDZ antibody with radiation showed an additive effect. Immunoblot analysis of tumor tissues also showed downregulation of phosphorylated and total levels of AKT, mTOR and 4EBP1 in the tumors treated with anti-PDZ antibody. Overall, our results suggest that TIP-1 is a promising therapeutic target for treatment of lung cancer and glioblastoma. Antibodies specific to the PDZ domain of TIP-1 enhance the efficacy of radiotherapy. The anti-PDZ antibodies need to be optimized further before translating it into the clinic. Citation Format: Vaishali Kapoor, David Dadey, Kelly Hoye, Andrea Collins, Dinesh Thotala, Dennis Hallahan. Antibody targeting PDZ domain of TIP-1 induces proliferation arrest through AKT/mTOR signaling inhibition in lung cancer and glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4599. doi:10.1158/1538-7445.AM2017-4599

MerTK as a therapeutic target in glioblastoma.

Glioma-associated macrophages and microglia (GAMs) are components of the glioblastoma (GBM) microenvironment that express MerTK, a receptor tyrosine kinase that triggers efferocytosis and can suppress innate immune responses. The aim of the study was to define MerTK as a therapeutic target using an orally bioavailable inhibitor, UNC2025. We examined MerTK expression in tumor cells and macrophages in matched patient GBM samples by double-label immunohistochemistry. UNC2025-induced MerTK inhibition was studied in vitro and in vivo. MerTK/CD68+ macrophages increased in recurrent tumors while MerTK/glial fibrillary acidic protein-positive tumor cells did not. Pharmacokinetic studies showed high tumor exposures of UNC2025 in a syngeneic orthotopic allograft mouse GBM model. The same model mice were randomized to receive vehicle, daily UNC2025, fractionated external beam radiotherapy (XRT), or UNC2025/XRT. Although median survival (21, 22, 35, and 35 days, respectively) was equivalent with or without UNC2025, bioluminescence imaging (BLI) showed significant growth delay with XRT/UNC2025 treatment and complete responses in 19%. The responders remained alive for 60 days and showed regression to 1%-10% of pretreatment BLI tumor burden; 5 of 6 were tumor free by histology. In contrast, only 2% of 98 GBM mice of the same model treated with XRT survived 50 days and none survived 60 days. UNC2025 also reduced CD206+ macrophages in mouse tumor samples. These results suggest that MerTK inhibition combined with XRT has a therapeutic effect in a subset of GBM. Further mechanistic studies are warranted.

Analysis of 31-year-old patient with SYNGAP1 gene defect points to importance of variants in broader splice regions and reveals developmental trajectory of SYNGAP1-associated phenotype: case report

BackgroundWhole exome sequencing is a powerful tool for the analysis of genetically heterogeneous conditions. The prioritization of variants identified often focuses on nonsense, frameshift and canonical splice site mutations, and highly deleterious missense variants, although other defects can also play a role. The definition of the phenotype range and course of rare genetic conditions requires long-term clinical follow-up of patients.Case presentationWe report an adult female patient with severe intellectual disability, severe speech delay, epilepsy, autistic features, aggressiveness, sleep problems, broad-based clumsy gait and constipation. Whole exome sequencing identified a de novo mutation in the SYNGAP1 gene. The variant was located in the broader splice donor region of intron 10 and replaced G by A at position +5 of the splice site. The variant was predicted in silico and shown experimentally to abolish the regular splice site and to activate a cryptic donor site within exon 10, causing frameshift and premature termination. The overall clinical picture of the patient corresponded well with the characteristic SYNGAP1-associated phenotype observed in previously reported patients. However, our patient was 31 years old which contrasted with most other published SYNGAP1 cases who were much younger. Our patient had a significant growth delay and microcephaly. Both features normalised later, although the head circumference stayed only slightly above the lower limit of the norm. The patient had a delayed puberty. Her cognitive and language performance remained at the level of a one-year-old child even in adulthood and showed a slow decline. Myopathic facial features and facial dysmorphism became more pronounced with age. Although the gait of the patient was unsteady in childhood, more severe gait problems developed in her teens. While the seizures remained well-controlled, her aggressive behaviour worsened with age and required extensive medication.ConclusionsThe finding in our patient underscores the notion that the interpretation of variants identified using whole exome sequencing should focus not only on variants in the canonical splice dinucleotides GT and AG, but also on broader splice regions. The long-term clinical follow-up of our patient contributes to the knowledge of the developmental trajectory in individuals with SYNGAP1 gene defects.

Spheroid-based 3D Cell Cultures Enable Personalized Therapy Testing and Drug Discovery in Head and Neck Cancer.

Chemo-radiation currently serves as first-line therapy of advanced and recurrent head and neck cancer, while new chemotherapy regimens are emerging. However, response rates to any treatment are difficult to predict and underlie broad variation. This study shows the development of a standardized, high-throughput in vitro assay to assess patients' individual response to therapy regimens as a future tool for personalized tumor therapy. Viability and proliferation analyses after chemo +/- radiation treatment of single spheroids (low adhesion plates/Hanging Drop (HD)) were generated from head and neck squamous cell carcinoma (HNSCC) cell lines and primary human cells from fresh tumor specimens. All cell lines showed reliable growth in all cell culture methods. The spheroids showed significant delay of growth and/or necrosis compared to control groups when exposed to current standard chemotherapeutic regimens. Single 3D spheroids ready for therapy susceptibility testing could be generated from actual tumor specimens after enzymatic and mechanical separation. In its current form, this single spheroid-based in vitro assay was able to test individual therapy susceptibility to current standard therapy regimens or, potentially, for testing new targeted drugs in HNSCC treatment. With recent discoveries regarding tumor heterogeneity and individual mutation status, a reliable assay is a prerequisite for personalized therapy in head and neck cancer.

The glycogen synthase kinase MoGsk1, regulated by Mps1 MAP kinase, is required for fungal development and pathogenicity in Magnaporthe oryzae

Magnaporthe oryzae, the causal agent of blast disease, is one of the most destructive plant pathogens, causing significant yield losses on staple crops such as rice and wheat. The fungus infects plants with a specialized cell called an appressorium, whose development is tightly regulated by MAPK signaling pathways following the activation of upstream sensors in response to environmental stimuli. Here, we show the expression of the Glycogen synthase kinase 3 (GSK3) MoGSK1 in M. oryzae is regulated by Mps1 MAP kinase, particularly under the stressed conditions. Thus, MoGSK1 is functionally characterized in this study. MoGsk1 is functionally homologues to the Saccharomyces cerevisiae GSK3 homolog MCK1. Gene replacement of MoGSK1 caused significant delay in mycelial growth, complete loss of conidiation and inability to penetrate the host surface by mycelia-formed appressorium-like structures, consequently resulting in loss of pathogenicity. However, the developmental and pathogenic defects of Δmogsk1 are recovered via the heterologous expression of Fusarium graminearum GSK3 homolog gene FGK3, whose coding products also shows the similar cytoplasmic localization as MoGsk1 does in M. oryzae. By contrast, overexpression of MoGSK1 produced deformed appressoria in M. oryzae. In summary, our results suggest that MoGsk1, as a highly conservative signal modulator, dictates growth, conidiation and pathogenicity of M. oryzae.

Mechanism of the Antitumor and Radiosensitizing Effects of a Manganese Porphyrin, MnHex-2-PyP.

Cationic manganese (Mn)-substituted N-pyridylporphyrin-based potent mimics of the family of superoxide dismutases (SODs) protect normal tissues from injury related to ionizing radiation (IR) by reducing levels of reactive oxygen and nitrogen species (ROS/RNS). Furthermore, Mn-porphyrins have demonstrated antitumor and radiosensitizing effects on cancer cells by promoting IR-induced tumor vasculature damage and apoptotic processes. In this study, we explored the underlying mechanisms of Mn-porphyrin-mediated tumor radiosensitization using murine mammary carcinoma 4T1 and melanoma B16 cells in vitro and in vivo. Combination treatment with MnTnHex-2-PyP and IR substantially reduced cell viability, clonogenic cell survival, and DNA damage repair and synergistically increased IR-induced apoptosis of 4T1 and B16 cells. MnTnHex-2-PyP in combination with IR caused a significant delay in growth of 4T1 and B16 xenograft tumors. MnTnHex-2-PyP dose-dependently enhanced IR-mediated production of H2O2-derived species, but not superoxide. Catalase overexpression reversed MnTnHex-2-PyP-enhanced ROS production and apoptosis. Demonstrated suppression of phosphorylation of several mitogen-activated protein (MAP) kinases and activation of NF-κB by MnTnHex-2-PyP/IR, which presumably inhibited activation of the antiapoptotic pathway, are in agreement with our other data on the apoptosis of cancer cells. Innovation and Conclusions: MnTnHex-2-PyP exerted a radiosensitizing effect on 4T1 and B16 tumor models in vitro and in vivo via pro-oxidative actions and therefore bears a large therapeutic potential. When combined with IR, it attenuated DNA damage repair and triggered a shift from prosurvival pathways to apoptotic cell death, likely due to increased ROS production and disturbed cellular redox balance, acting at the level of nuclear factor κB (NF-κB). Antioxid. Redox Signal. 27, 1067-1082.

Correlation of Ultrasound Shear Wave Elastography with Pathological Analysis in a Xenografic Tumour Model

The objective of this study was to evaluate the potential value of ultrasound (US) shear wave elastography (SWE) in assessing the relative change in elastic modulus in colorectal adenocarcinoma xenograft models in vivo and investigate any correlation with histological analysis. We sought to test whether non-invasive evaluation of tissue stiffness is indicative of pathological tumour changes and can be used to monitor therapeutic efficacy. US-SWE was performed in tumour xenografts in 15 NCr nude immunodeficient mice, which were treated with either the cytotoxic drug, Irinotecan, or saline as control. Ten tumours were imaged 48 hours post-treatment and five tumours were imaged for up to five times after treatment. All tumours were harvested for histological analysis and comparison with elasticity measurements. Elastic (Young's) modulus prior to treatment was correlated with tumour volume (r = 0.37, p = 0.008). Irinotecan administration caused significant delay in the tumour growth (p = 0.02) when compared to control, but no significant difference in elastic modulus was detected. Histological analysis revealed a significant correlation between tumour necrosis and elastic modulus (r = −0.73, p = 0.026). SWE measurement provided complimentary information to other imaging modalities and could indicate potential changes in the mechanical properties of tumours, which in turn could be related to the stages of tumour development.

Rapeseed-legume intercrops: plant growth and nitrogen balance in early stages of growth and development.

In this study we tested whether legumes can improve the growth and N and S nutrition of rapeseed in an intercropping system and compared the effect of mixtures on legume N-fixation and soil N-resources. Rapeseed was cultivated in low N conditions in monocrops using one (R) or two plants (RR) per pot and in mixtures with lupine, clover or vetch.The R monocrop was the most relevant control, intraspecific competition inducing a significant growth delay resulting in a significantly lower leaf number, in RR monocrop compared to R and the three mixtures considered. Plant biomass, and the N and S contents of rapeseed grown in mixtures were the same than those measured in R monocrop. Compared to the monocrop, the proportion of N derived from the atmosphere was increased by 34, 140 and 290% in lupine, clover and vetch, respectively when intercropped with rapeseed. In mixture with clover and lupine, the soil N pool at harvest was higher than in other treatments, while N export by crop was constant. Legumes suffered from competition for soil S resulting in a decrease of 40% in their S content compared to the monocrop. Compared to rapeseeds grown in R monocrop and in mixture with lupine and vetch, rapeseed mixed with clover showed significantly higher SPAD values in old leaves.In our conditions, mixing legumes with rapeseed is relevant to reduce N fertilization and improve nutrition and growth of rapeseed.

213 - Targeting the HER family with Pan-HER effectively overcomes resistance to cetuximab

Cetuximab, an antibody against the EGFR, has shown efficacy in treating head and neck squamous cell carcinoma (HNSCC), metastatic colorectal cancer, and non-small cell lung cancer (NSCLC). Despite the clinical success of cetuximab, many patients do not respond to cetuximab. Furthermore, virtually all patients who do initially respond become refractory, highlighting both intrinsic and acquired resistance to cetuximab as significant clinical problems. To understand mechanistically how cancerous cells acquire resistance, we previously developed models of acquired resistance using the H226 NSCLC and UM-SCC1 HNSCC cell lines. Cetuximab-resistant clones showed a robust upregulation and dependency on the HER family receptors EGFR, HER2, and HER3. Here, we examined pan-HER, a mixture of six antibodies targeting these receptors on cetuximab-resistant clones. In cells exhibiting acquired or intrinsic resistance to cetuximab, pan-HER treatment decreased all three receptors' protein levels and downstream activation of AKT and MAPK. This correlated with decreased cell proliferation in cetuximab-resistant clones. To determine whether pan-HER had a therapeutic benefit in vivo, we established de novo cetuximab-resistant mouse xenografts and treated resistant tumors with pan-HER. This regimen resulted in a superior growth delay of cetuximab-resistant xenografts compared with mice continued on cetuximab. Furthermore, intrinsically cetuximab-resistant HNSCC patient-derived xenograft tumors treated with pan-HER exhibited significant growth delay compared with vehicle/cetuximab controls. These results suggest that targeting multiple HER family receptors simultaneously with pan-HER is a promising treatment strategy for tumors displaying intrinsic or acquired resistance to cetuximab. Mol Cancer Ther; 15(9); 2175-86. ©2016 AACR.

Small molecule inhibition of polo-like kinase 1 by volasertib (BI 6727) causes significant melanoma growth delay and regression in vivo

The objective of this study was to determine the therapeutic potential of polo-like kinase 1 (Plk1) inhibition in melanoma, in vivo. Employing Vectra technology, we assessed the Plk1 expression profile in benign nevi, malignant (stages I–IV) and metastatic melanomas. We found a significant elevation of Plk1 immunostaining in melanoma tissues. Further, a second generation small molecule Plk1 inhibitor, BI 6727, resulted in reductions in growth, viability and clonogenic survival, as well as an increase in apoptosis of A375 and Hs 294T melanoma cells. BI 6727 treatment also resulted in a G2/M-as well as S-phase cell cycle arrest in melanoma cells. Importantly, BI 6727 (intravenous injection; 10 and 25 mg/kg body weight) treatment resulted in significant tumor growth delay and regression in vivo in A375-and Hs 294T-implanted xenografts in athymic nude mice. These anti-melanoma effects were accompanied with a decreased cellular proliferation (Ki-67 staining) and induction of apoptosis (caspase 3 activation). In addition, BI 6727 treatment caused a marked induction of p53 and p21 in vitro as well as in vivo. Overall, we suggest that Plk1 inhibition may be a useful approach as a monotherapy as well as in combination with other existing therapeutics, for melanoma management.

Effect of inhibitors of two isoprenoid biosynthetic pathways on physiological and biosynthetic characteristics of Dioscorea deltoidea cell suspension culture

The effect of phosmidomycin and mevinolin, which inhibit MEP and MVA isoprenoid biosynthetic pathways, respectively, on the growth (biomass accumulation, growth index, specific growth rate), physiological (respiration intensity and ratio between the cytochrome and cyanide-resistant respiration types), and biosynthetic (steroid glycoside biosynthesis) characteristics of the cell suspension culture of Dioscorea deltoidea Wall. has been studied. Both inhibitors decreased the growth index of a cell culture by 20–25%, but their influence on the cell growth dynamics was different. Mevinolin treatment reduced the maximum biomass accumulation by 20% as against the control but did not change the character of a growth curve. Phosmidomycin treatment caused a significant growth delay (a 6-day lag phase) followed by a short active growth period (μ = 0.29 days-1). Treatment of cells with inhibitors did not significantly influence on their total oxygen uptake rate, whose average value at different growth phases was equal to 100–200 mg О2/g of dry cell weight per hour, but cardinally changed characteristics of respiratory metabolism. The inhibitors increased the activity of a cyanide-resistant respiration and decreased the intensity of a cytochrome respiration; each inhibitor worked in its specific manner. In the case of mevinolin, the maximum level of cyanide-resistant respiration (70% of the total respiration intensity) was observed at initial growth phases; during the further cell culture growth, this value gradually reduced to 6–8%. Phosmidomycin treatment caused a reverse dynamics: at the initial growth phases, the contribution of cyanide-resistant respiration was 30%, whereas at the stationary and degradation phases it increased to 50–60%. The treatment of cells with phosmidomycin resulted in a double increase in the content of furostanol glycosides at the stationary growth phase, whereas the use of mevinolin-containing medium reduced the content of these compounds as against the control. Inhibitors also influenced on the ratio of individual glycosides, such as protodioscin, deltoside, and their 26-S-isomers. The obtained results confirm the hypothesis of a possible intermediate exchange between the plastid (MEP) and cytosolic (MVA) isoprenoid biosynthetic pathways; this exchange is directed mainly from the plastids to the cytosol.

Accumulation and developmental toxicity of hexabromocyclododecanes (HBCDs) on the marine copepod Tigriopus japonicus

The brominated flame retardants hexabromocyclododecanes (HBCDs) are ubiquitous environmental contaminants, widely distributed in aquatic systems including the marine environment and marine organisms. HBCDs are toxic to the development of both freshwater and marine fish. However, the impacts of HBCDs on marine invertebrates are not well known. In this study, the marine copepod, Tigriopus japonicus, was used to assess the bioaccumulation and developmental toxicity of technical HBCD (tHBCD) through water-borne exposure. The uptake rate constant of tHBCD by T. japonicus was high, which resulted in high bioaccumulation potential. The bioconcentration factors of tHBCD were 8.73 × 104 and 6.34 × 104 L kg−1 in T. japonicus, calculated using the kinetic and steady-state methods, respectively. Exposure of T. japonicus nauplii to tHBCD caused significant growth delay. The lowest-observable-effect-concentrations of tHBCD induced developmental delay were 30 and 8 μg L−1 for the F0 and F1 generations, respectively, which suggested that the F1 generation was more sensitive to tHBCD than the F0 generation and warranted multiple-generation toxicity tests for future studies. Furthermore, exposure of the adult copepods to tHBCD induced the transcription of oxidative stress response genes and apoptotic genes, e.g., SOD,CAT, GST, OGG1, P53 and Caspase-3. It was therefore speculated that tHBCD exposure induced the generation of reactive oxygen species in T. japonicus, which activated the oxidative stress defense genes and meanwhile resulted in oxidative DNA damage. The damaged DNA activated the transcription of p53 and triggered the caspase-mediated apoptosis pathway, which may be the reason for the tHBCD induced developmental delay in T. japonicus nauplii.