p53 restoration kills primitive leukemia cells in vivo and increases survival of leukemic mice

Loss of p53 function is a common feature of human cancers and it is required for differentiated tumor cell maintenance; however, it is not known whether sustained inactivation of the p53 pathway is needed for cancer stem cell persistence. Chronic myeloid leukemia (CML) is caused by a chromosome translocation that generates the BCRABL oncogene encoding a constitutively active protein tyrosine kinase. The disease originates in a hematopoietic stem cell and is maintained by leukemic stem cells (LSCs). Treatment with specific tyrosine kinase inhibitors does not eliminate LSCs because they do not depend on the oncogene for survival. We have combined a switchable p53 knock-in mouse model, p53^KI/KI, with the well-characterized Sca1-BCRABLp210 CML transgenic model, to show that transient restoration of p53 slows disease progression and significantly extends the survival of leukemic animals, being the mechanism responsible for this effect, apoptotic death of primitive leukemia cells. In agreement with these in vivo findings, in vitro assays show that restoring p53 reduces hematopoietic colony formation by cells of leukemic animals. These results suggest that reestablishing p53 function may be a therapeutic strategy for the eradication of leukemic stem cells and to prevent disease progression.     

Knockout of LASP1 in CXCR4 expressing CML cells promotes cell persistence,proliferation and TKI resistance

Chronic myeloid leukaemia (CML) is a clonal myeloproliferative stem cell disorder characterized by the constitutively active BCR-ABL tyrosine kinase. The LIM and SH3 domain protein 1 (LASP1) has recently been identified as a novel BCR-ABL substrate and is associated with proliferation, migration, tumorigenesis and chemoresistance in several cancers. Furthermore, LASP1 was shown to bind to the chemokine receptor 4 (CXCR4), thought to be involved in mechanisms of relapse. In order to identify potential LASP1-mediated pathways and related factors that may help to further eradicate minimal residual disease (MRD), the effect of LASP1 on processes involved in progression and maintenance of CML was investigated. The present data indicate that not only overexpression of CXCR4, but also knockout of LASP1 contributes to proliferation, reduced apoptosis and migration as well as increased adhesive potential of K562 CML cells. Furthermore, LASP1 depletion in K562 CML cells leads to decreased cytokine release and reduced NK cell-mediated cytotoxicity towards CML cells. Taken together, these results indicate that in CML, reduced levels of LASP1 alone and in combination with high CXCR4 expression may contribute to TKI resistance.     

Diverse glandular pathologies coexist with high-grade squamous intraepithelial lesion in cyto-histological review of atypical glandular cells on ThinPrep specimens

Objective: To identify in cytology, high‐grade squamous intraepithelial lesions with endocervical glandular extension in cases previously diagnosed as atypical glandular cells (AGC), analyse possible reasons for the diagnostic pitfall and document the frequency of glandular pathology coexisting with high‐grade cervical intraepithelial lesion in histology. Methods: Thirty‐nine ThinPrep® cervical smear (Pap) tests reported as AGC of undetermined significance and showing high‐grade lesions on histology [cervical intraepithelial neoplasia (CIN) 2 or 3, endometrial or extrauterine adenocarcinoma] were reviewed retrospectively to identify the cases of high‐grade squamous intraepithelial lesion with endocervical glandular extension, using the Bethesda 2001 system. Cyto‐histological correlation was performed. Results: A high frequency of diverse glandular pathologies coexisted with high‐grade cervical intraepithelial lesions on histology. This included endocervical glandular extension in 63%, benign glandular pathology in 33% and pre‐neoplastic or malignant glandular pathology (endocervical glandular dysplasia, adenocarcinoma in situ and metastatic breast carcinoma) in 17% cases. On cytology, the sensitivity was 40%, specificity was 80% and positive predictive value was 86% for endocervical gland extension in high‐grade squamous intraepithelial lesions. Conclusions: Special efforts to recognize endocervical glandular extension in high‐grade squamous intraepithelial lesions and glandular neoplasia coexisting with squamous intraepithelial lesions from the heterogeneous category of AGC can contribute to increasing the diagnostic accuracy. The identification of endocervical glandular extension on cervical cytology would alert the gynaecologist to perform a thorough assessment of the endocervix during colposcopy. This could also help to decide on the need to perform deeper conization rather than loop electrosurgical excision procedure to ensure negative margins when colposcopic biopsy shows CIN 2 or 3.     

Gene therapy legislation in The Netherlands

Several regulatory organisations are involved in the assessment of clinical gene therapy trials involving genetically modified organisms (GMOs) in The Netherlands. Medical, ethical and scientific aspects are, for instance, evaluated by the Central Committee on Research Involving Human Subjects (CCMO). The Ministry of Housing, Spatial Planning and the Environment (VROM) is the competent authority for the environmental risk assessment according to the deliberate release Directive 2001/18/EC. A Gene Therapy Office has been established in order to streamline the different national review processes and to enable the official procedures to be completed as quickly as possible. Although the Gene Therapy Office improved the application process at the national level, there is a difference of opinion between the EU member states with respect to the EU Directive according to which gene therapy trials are assessed, that urges for harmonisation. This review summarises the gene therapy legislation in The Netherlands and in particular The Netherlands rationale to follow Directive 2001/18/EC for the environmental risk assessment.     

Life cycle assessment of a multi-material car component

Background, Aims and Scope In recent years, the automotive industry has been experiencing an increasing concern with environmental requirements. A particular focus is being given to light-weighting of cars, to reducing fuel consumption and to the use of different recycling materials. Consequently, decisions on product design and development must involve economic and technological as well as environmental considerations. In adequate conditions, the LCA methodology enables one to assist an effective integration of the environmental considerations in the decision-making process [1]. In this paper, a multi-material car component which is part of the current automotive brake system, has been modified by its original manufacturer. Such a modification included the use of a new multi-material injection moulding process and the consumption of recyclable materials. The new and the current component were comparatively assessed throughout their life cycles in order to evaluate their respective environmental impacts and, thus, to verify if the new component offers a lower environmental load. The results described in this paper are part of the outcome of a broader research project involving industrial companies, university, technological centres and research institutes based in Portugal, Spain and Germany. Main Features The car component under focus has four subcomponents whose base materials consist of steel and plastic. The LCA methodology is used to evaluate two scenarios describing the new car component, on the one hand, and the reference scenario, which consists of the existing car component, on the other. The former results from the selection of new subcomponents materials, aiming to use a new production process together with a recycling strategy. Results and Discussion The inventory analysis shows a lower energy consumption in the alternative scenario (4.2 MJ) compared to the reference scenario (6.1 MJ). Most of that energy is still non-renewable, relating in particular to crude consumption in the car use phase and in the production phase (transports and plastics production). The life cycle inventory analysis indicates also that the alternative scenario has lower air emissions of CO₂, CO, NO_x, SO_x, NM VOC and PM10, as well as lower solid wastes and water emissions of oils and BOD5. Otherwise, the water emissions of undissolved substances and COD are higher for the alternative scenario. Most of the energy consumed and the air pollutants inventoried occur as a consequence of the use phase. Otherwise, for most of the life cycle water emissions inventoried and solid wastes, the production phase is the major contributor. The impact assessment, performed with the CML method, allows one to conclude that the alternative scenario exhibits lower results in all the impact categories. Both scenarios have similar environmental profiles, being: (i) the use phase, the major contributor for the abiotic depletion, global warming, photochemical oxidation, acidification and eutrophication; and (ii) the production phase, the main contributor for ozone depletion, human toxicity, fresh water aquatic ecotoxicity, marine aquatic ecotoxicity and terrestrial ecotoxicity. The sensitivity analysis, with respect to the fuel consumption reduction value, the impact assessment method and the final disposal scenario, performed in this study allows one to confirm, as a main conclusion, that the alternative scenario is environmentally preferable to the reference scenario. Conclusion The results obtained through the application of the LCA methodology enable one to conclude that the alternative component has a lower environmental load than the reference component. Recommendations and Perspectives Considering that the time required for the inventory data collection is a critical issue in LCA practise, the insights provided by this particular case study are likely to be useful to product developers in the car component manufacturing industry, particularly to brake system manufacturers supporting the environmental design within the sector.     

Co‐encapsulation of anti‐BCR‐ABL siRNA and imatinib mesylate in transferrin receptor‐targeted sterically stabilized liposomes for chronic myeloid leukemia treatment

Chronic myeloid leukemia (CML) is triggered by the BCR‐ABL oncogene. Imatinib is the first‐line treatment of CML; however imatinib resistance and intolerance have been detected in many patients. Therefore, new therapeutic approaches are required. The present work aimed at the development and application of transferrin receptor (TrfR) targeted liposomes co‐encapsulating anti‐BCR‐ABL siRNA and imatinib at different molar ratios. The encapsulation yields and drug loading of each molecule was evaluated. Anti‐leukemia activity of the developed formulations co‐encapsulating siRNA and imatinib and of the combination of Trf‐liposomes carrying siRNA and free imatinib under two different treatment schedules of pre‐sensitization was assessed. The results obtained demonstrate that the presence of imatinib significantly decreases the encapsulation yields of siRNA, whereas imatinib encapsulation yields are increased by the presence of siRNA. Cytotoxicity assays demonstrate that the formulations co‐encapsulating siRNA and imatinib promote a 3.84‐fold reduction on the imatinib IC₅₀ (from 3.49 to 0.91 µM), whereas a 8.71‐fold reduction was observed for the pre‐sensitization protocols (from 42.7 to 4.9 nM). It was also observed that the formulations with higher siRNA to imatinib molar ratios promote higher cell toxicity. Thus, the present work describes a novel triple targeting strategy with one single system: cellular targeting (through the targeting ligand, transferrin) and molecular targeting at the BCR‐ABL mRNA and Bcr‐Abl protein level. Biotechnol. Bioeng. 2010;107: 884–893. © 2010 Wiley Periodicals, Inc.     

Geothermal bryophyte habitats in the South Sandwich Islands,maritime Antarctic

Question: How does geothermal activity influence terrestrial plant colonization, species composition and community development in the Antarctic? Location: South Sandwich Islands, maritime Antarctic. Methods: Bryophytes were documented during a biological survey of the archipelago in January and February 1997. Particular attention was given to sites under current or recent influence of geothermal activity. Temperature profiles obtained across defined areas of activity on several islands were linked with the presence of specific bryophytes. Results: Greatest bryophyte richness was associated with geothermally influenced ground. Of 35 moss and nine liverwort species recorded, only four mosses were never associated with heated ground, while eight of the liverworts and 50% of the mosses were found only on actively or recently heated ground. Some species occur in unheated sites elsewhere in the maritime Antarctic, but were absent from such habitats on the South Sandwich Islands. Several species occurred in distinct zones around fumaroles. Maximum temperatures recorded within the upper 0.5 cm of the vegetation surface were 40 ‐ 47 °C, with only Campylopus introflexus tolerating such temperatures. Maximum temperatures 2.5 or 5 cm below the vegetation surface of this moss reached 75 °C. Other bryophytes regularly present in zoned vegetation included the mosses Dicranella hookeri, Sanionia georgico‐uncinata, Pohlia nutans and Notoligotrichum trichodon, and the liverworts Cryptochila grandiflora and Marchantia berteroana. Surface temperatures of 25 ‐ 35 °C and subsurface temperatures of 50 ‐ 60 °C were recorded in these species. Conclusions: These exceptional plant communities illustrate the transport of viable propagules into the Antarctic. Individually ephemeral in nature, the longer term existence of geothermal habitats on islands along the Scotia Arc may have provided refugia during periods of glacial expansion, facilitating subsequent recolonization of Antarctic terrestrial habitats.     

The antioxidant properties of serum albumin

Free radicals are a normal component of cellular oxygen metabolism in mammals. However, free radical-associated damage is an important factor in many pathological processes. Glycation and oxidative damage cause protein modifications, frequently observed in numerous diseases. Albumin represents a very abundant and important circulating antioxidant. This review brings together recent insights on albumin antioxidant properties. First, it focuses on the different activities of albumin concerning protein antioxidation. In particular, we describe the role of albumin in ligand binding and free radical-trapping activities. In addition, physiological and pathological situations that modify the antioxidant properties of albumin are reported.     

Activation of cytotoxic T lymphocytes against CML28-bearing tumors by dendritic cells transduced with a recombinant adeno-associated virus encoding the CML28 gene

Induction of anti-tumor immune responses by dendritic cells (DCs) transduced with a recombinant adeno-associated virus type 2 (rAAV2) encoding tumor antigens is considered a promising approach for cancer vaccine development. CML28, a novel antigen with the properties of cancer/testis (CT) antigens, is an attractive target for antigen-specific immunotherapy. Here we investigated the feasibility of inducing CML28-specific cytotoxic T lymphocyte (CTL) responses using DCs transduced with the rAAV2 vectors containing the CML28 gene (rAAV/CML28). Using an adenovirus-free packaging system, rAAV/CML28 was generated. The transduction efficiency of rAAV/CML28 in DCs increased in a multiplicity of infection (MOI)-dependent manner. The rAAV/CML28 transduction did not impair DC maturation, but even enhanced the CD80 expression. The rAAV/CML28-transduced DCs induced CML28-specific CTLs which exhibited a MHC class I-mediated antigen-specific lytic activity against CML28-bearing tumor cell lines (HepG2 and MCF-7) as well as the primary leukemia blasts. These findings suggest that rAAV/CML28-transduced DCs vaccine may serve as a feasible approach for the treatment of CML28-associated cancers.