Synthesis and Investigation of a Radioiodinated F3 Peptide Analog as a SPECT Tumor Imaging Radioligand

A radioiodinated derivative of the tumor-homing F3 peptide, (N-(2-{3-[125I]Iodobenzoyl}aminoethyl)maleimide-F3Cys peptide, [125I]IBMF3 was developed for investigation as a SPECT tumor imaging radioligand. For this purpose, we custom synthesized a modified F3 peptide analog (F3Cys) incorporating a C-terminal cysteine residue for site-specific attachment of a radioiodinated maleimide conjugating group. Initial proof-of-concept Fluorescence studies conducted with AlexaFluor 532 C5 maleimide-labeled F3Cys showed distinct membrane and nuclear localization of F3Cys in MDA-MB-435 cells. Additionally, F3Cys conjugated with NIR fluorochrome AlexaFluor 647 C2 maleimide demonstrated high tumor specific uptake in melanoma cancer MDA-MB-435 and lung cancer A549 xenografts in nude mice whereas a similarly labeled control peptide did not show any tumor uptake. These results were also confirmed by ex vivo tissue analysis. No-carrier-added [125I]IBMF3 was synthesized by a radioiododestannylation approach in 73% overall radiochemical yield. In vitro cell uptake studies conducted with [125I]IBMF3 displayed a 5-fold increase in its cell uptake at 4 h when compared to controls. SPECT imaging studies with [125I]IBMF3 in tumor bearing nude mice showed clear visualization of MDA-MB-435 xenografts on systemic administration. These studies demonstrate a potential utility of F3 peptide-based radioligands for tumor imaging with PET or SPECT techniques

The synthetic peptide P111-136 derived from the C-terminal domain of heparin affin regulatory peptide inhibits tumour growth of prostate cancer PC-3 cells

<p>Abstract</p> <p>Background</p> <p>Heparin affin regulatory peptide (HARP), also called pleiotrophin, is a heparin-binding, secreted factor that is overexpressed in several tumours and associated to tumour growth, angiogenesis and metastasis. The C-terminus part of HARP composed of amino acids 111 to 136 is particularly involved in its biological activities and we previously established that a synthetic peptide composed of the same amino acids (P111-136) was capable of inhibiting the biological activities of HARP. Here we evaluate the ability of P111-136 to inhibit <it>in vitro </it>and <it>in vivo </it>the growth of a human tumour cell line PC-3 which possess an HARP autocrine loop.</p> <p>Methods</p> <p>A total lysate of PC-3 cells was incubated with biotinylated P111-136 and pulled down for the presence of the HARP receptors in Western blot. <it>In vitro</it>, the P111-136 effect on HARP autocrine loop in PC-3 cells was determined by colony formation in soft agar. <it>In vivo</it>, PC-3 cells were inoculated in the flank of athymic nude mice. Animals were treated with P111-136 (5 mg/kg/day) for 25 days. Tumour volume was evaluated during the treatment. After the animal sacrifice, the tumour apoptosis and associated angiogenesis were evaluated by immunohistochemistry. <it>In vivo </it>anti-angiogenic effect was confirmed using a mouse Matrigel™ plug assay.</p> <p>Results</p> <p>Using pull down experiments, we identified the HARP receptors RPTPβ/ζ, ALK and nucleolin as P111-136 binding proteins. <it>In vitro</it>, P111-136 inhibits dose-dependently PC-3 cell colony formation. Treatment with P111-136 inhibits significantly the PC-3 tumour growth in the xenograft model as well as tumour angiogenesis. The angiostatic effect of P111-136 on HARP was also confirmed using an <it>in vivo </it>Matrigel™ plug assay in mice</p> <p>Conclusions</p> <p>Our results demonstrate that P111-136 strongly inhibits the mitogenic effect of HARP on <it>in vitro </it>and <it>in vivo </it>growth of PC-3 cells. This inhibition could be linked to a direct or indirect binding of this peptide to the HARP receptors (ALK, RPTPβ/ζ, nucleolin). <it>In vivo</it>, the P111-136 treatment significantly inhibits both the PC-3 tumour growth and the associated angiogenesis. Thus, P111-136 may be considered as an interesting pharmacological tool to interfere with tumour growth that has now to be evaluated in other cancer types.</p

Met-Independent Hepatocyte Growth Factor-mediated regulation of cell adhesion in human prostate cancer cells

BACKGROUND: Prostate cancer cells communicate reciprocally with the stromal cells surrounding them, inside the prostate, and after metastasis, within the bone. Each tissue secretes factors for interpretation by the other. One stromally-derived factor, Hepatocyte Growth Factor (HGF), was found twenty years ago to regulate invasion and growth of carcinoma cells. Working with the LNCaP prostate cancer progression model, we found that these cells could respond to HGF stimulation, even in the absence of Met, the only known HGF receptor. The new HGF binding partner we find on the cell surface may help to clarify conflicts in the past literature about Met expression and HGF response in cancer cells. METHODS: We searched for Met or any HGF binding partner on the cells of the PC3 and LNCaP prostate cancer cell models, using HGF immobilized on agarose beads. By using mass spectrometry analyses and sequencing we have identified nucleolin protein as a novel HGF binding partner. Antibodies against nucleolin (or HGF) were able to ameliorate the stimulatory effects of HGF on met-negative prostate cancer cells. Western blots, RT-PCR, and immunohistochemistry were used to assess nucleolin levels during prostate cancer progression in both LNCaP and PC3 models. RESULTS: We have identified HGF as a major signaling component of prostate stromal-conditioned media (SCM) and have implicated the protein nucleolin in HGF signal reception by the LNCaP model prostate cancer cells. Antibodies that silence either HGF (in SCM) or nucleolin (on the cell surfaces) eliminate the adhesion-stimulatory effects of the SCM. Likewise, addition of purified HGF to control media mimics the action of SCM. C4-2, an LNCaP lineage-derived, androgen-independent human prostate cancer cell line, responds to HGF in a concentration-dependent manner by increasing its adhesion and reducing its migration on laminin substratum. These HGF effects are not due to shifts in the expression levels of laminin-binding integrins, nor can they be linked to expression of the known HGF receptor Met, as neither LNCaP nor clonally-derived C4-2 sub-line contain any detectable Met protein. Even in the absence of Met, small GTPases are activated, linking HGF stimulation to membrane protrusion and integrin activation. Membrane-localized nucelolin levels increase during cancer progression, as modeled by both the PC3 and LNCaP prostate cancer progression cell lines. CONCLUSION: We propose that cell surface localized nucleolin protein may function in these cells as a novel HGF receptor. Membrane localized nucleolin binds heparin-bound growth factors (including HGF) and appears upregulated during prostate cancer progression. Antibodies against nucleolin are able to ameliorate the stimulatory effects of HGF on met-negative prostate cancer cells. HGF-nucleolin interactions could be partially responsible for the complexity of HGF responses and met expression reported in the literature

Forêt : relever les défis du changement climatique en France métropolitaine - Avant-propos

Forests: meeting the challenges of climate change in metropolitan France - ForewordForêt : relever les défis du changement climatique en France métropolitaine - Avant-propo

Interactions between under- and over-storeys: Consequences to design silvicultures adapted to climate changes

International audienceForests comprise several interacting strata of vegetation; overstorey trees are most obvious but sub-canopy strata, collectively termed understorey or undergrowth, are typically present including herbaceous plants, shrubs, seedlings and saplings, and suppressed under- and midstorey trees. For decades only adult trees have been considered in forests and most often, only crop trees. However the understorey also plays a fundamental role in ecosystem functioning and health. Various silvicultural systems are currently designed or experimented in relation with climate changes, and in particular a reduction of soil water availability together with an increase of scorching temperatures in many regions of the temperate area. Modifying tree species composition and density of the forest ecosystem will have consequences on the understorey composition and functioning, which in turn will interact with the overstorey. These interactions will modify the whole ecosystem functioning in terms of biodiversity, tree regeneration, wild fauna habitats, pest and diseases, etc. Therefore it is compulsory to account for the interplays between the different strata in forest to better design silvicultural operations relative to climate changes. Examples of such interactions will be given and consequences for designing experimental systems and new silvicultures will be discussed

Interactions between under- and over-storeys: Consequences for designing silvicultural systems adapted to climate changes

International audienceForests comprise several interacting strata of vegetation; overstorey trees are most obvious but sub-canopy strata, collectively termed understorey or undergrowth, are typically present including herbaceous plants, shrubs, seedlings and saplings, and suppressed under- and midstorey trees. For decades only adult trees have been considered in forests and most often, only crop trees. However the understorey also plays a fundamental role in ecosystem functioning and health. Various silvicultural systems are currently designed or experimented in relation with climate changes, and in particular a reduction of soil water availability together with an increase of scorching temperatures in many regions of the temperate area. Modifying tree species composition and density of the forest ecosystem will have consequences on the understorey composition and functioning, which in turn will interact with the overstorey. These interactions will modify the whole ecosystem functioning in terms of biodiversity, tree regeneration, wild fauna habitats, pest and diseases, etc. Therefore it is compulsory to account for the interplays between the different strata in forest to better design silvicultural operations relative to climate changes. Examples of such interactions will be given and consequences for designing experimental systems and new silvicultures will be discussed

Mixed stand growth models - The sessile oak / Scots pine

Mixed forests currently account for 50% of the French forest cover. Nonetheless, there are very few management aid tools in France relating to these stands. Growth model describe the development of stands over time and are able to simulate forestry practices – in this respect, they are very useful tools for planning operations and harvesting, or as aids in producing silvicultural guidelines. The purpose of this article is to present a specific issue raised by these models: the choice of the level of detail for a growth model. This choice can indeed have significant implications for the use of the model. So as to choose the level of detail, we propose building and comparing a series of simplified models. This method is applied to the growth of sessile oak / Scots pine stands in the context of a partnership between the French Forestry Board (ONF) and Cemagref. We sampled 101 oaks and 116 pines distributed in four experimental plots in the Orleans state forest for the purpose of studying radial increment. With this data, we built a first model – a distance-dependent tree model. With this model, we were able to specify the interaction between and within species. One finding was that pines surrounded by pines are at a disadvantage compared to those surrounded by oak. The model was integrated in a module of the Capsis forest simulation system. Starting from an actual stand, we conducted simulations that enabled us to quantify the production of the mixture and furthermore to provide information or input as to the future development of that combination. We also showed the relevance of the distant-dependent tree model for testing marking strategies on the scale of the individual tree. This work is a first step in building a family of models for sessile oak / Scots pine stands at various levels of detail. In the longer term, by comparing the various models in the family, we hope to identify the appropriate scale for one or several management issues. We will then be able to supplement this model by bringing regeneration into the equation so as to develop a comprehensive management aid tool for these mixtures.Les peuplements mélangés couvrent actuellement 50 % de la surface forestière française. Pour autant, il existe en France très peu d’outils d’aide à la gestion de ces peuplements. Les modèles de croissance décrivent l’évolution d’un peuplement au cours du temps et sont capables de simuler des interventions sylvicoles : ils sont, à ce titre, des outils très intéressants pour la planification des travaux et des récoltes ou pour l’aide à la production de guides sylvicoles. L’objectif de cet article est de présenter une question spécifique posée par ces modèles : le choix du niveau de détail d’un modèle de croissance. En effet, ce choix peut avoir des implications importantes par rapport à l’utilisation du modèle. Pour choisir ce niveau de détail, nous proposons de construire et de comparer une série de modèles simplifiés. Cette méthode est appliquée à la croissance des peuplements mélangés Chêne sessile – Pin sylvestre dans le cadre d’un partenariat entre l’ONF et le Cemagref. Pour étudier la croissance radiale, nous avons effectué des sondages dans 101 chênes et 116 pins répartis dans 4 dispositifs de la forêt domaniale d’Orléans. Nous avons utilisé ces données pour construire un premier modèle : un modèle arbre dépendant des distances. Ce modèle nous a permis de préciser les interactions intra- et interspécifiques. Il ressort notamment que les pins entourés de pins sont plus défavorisés que les pins entourés de chênes. Le modèle a été intégré dans un module de la plateforme de simulation forestière Capsis. À partir d’un peuplement réel, nous avons réalisé des simulations qui nous ont permis de quantifier la production du mélange mais aussi d’apporter des éléments de réponses ou de réflexion sur le devenir du mélange. Nous avons également illustré l’intérêt du modèle arbre dépendant des distances pour tester des stratégies de martelage à l’échelle de l’arbre. Ce travail est un premier pas vers la construction d’une famille de modèles des peuplements Chêne sessile - Pin sylvestre à différents niveaux de détail. À terme, la comparaison des différents modèles de cette famille nous permettra d’identifier l’échelle adéquate pour une ou plusieurs problématiques de gestion. Nous pourrons ensuite compléter ce modèle pour prendre en compte notamment la régénération et développer un outil complet d’aide à la gestion de ces peuplements mélangés