[Show abstract][Hide abstract] ABSTRACT: Angiogenesis is the phenomenon by which new blood vessels are created from preexisting ones. But this natural process is also involved, in a chaotic way, in tumor development. Many molecules have shown particular efficiency in inhibiting this phenomenon, hopefully leading to either: (i) a reorganization of the neovessels allowing a better tumor uptake of cytotoxic molecules (as chemotherapy) or (ii) a deprivation of the tumor vascular network with the view to starve it. However, characterizing the anti-angiogenic effects of a molecule remains difficult, mainly because the proposed physical modeling approaches have barely been confronted to in vivo data, which are not directly available. This paper presents an original approach to characterize and analyze the anti-angiogenic responses in cancerology that allows biologists to account for spatial and dynamical dimensions of the problem. The proposed solution relies on the association of a specific biological in vivo protocol using skinfold chambers, image processing and dynamic system identification. An empirical model structure of the anti-angiogenic effect of a tested molecule is selected according to experimental data. Finally the model is identified and its parameters are used to characterize and compare responses of the tested molecule.
Biomedical Signal Processing and Control 07/2015; 20:52 - 60. DOI:10.1016/j.bspc.2015.04.008 · 1.42 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The general strategy developed aims to favor the vascular effect of photodynamic therapy by targeting tumor vasculature. Since angiogenic endothelial cells represent an interesting target to potentiate this vascular effect, we previously described the conjugation of a photosensitizer to a peptide targeting neuropilins (NRPs) over-expressed specially in tumor angiogenic vessels and we recently characterized the mechanism of photosensitization-induced thrombogenic events. Nevertheless, in glioma-bearing nude mice, we demonstrated that the peptide moiety was degraded to various rates according to time after intravenous administration. In this study, new peptidases-resistant pseudopeptides were tested, demonstrating a molecular affinity for NRP-1 and NRP-2 recombinant chimeric proteins and devoid of affinity for VEGF receptor type 1 (Flt-1). To argue the involvement of NRP-1, MDA-MB-231 breast cancer cells were used, strongly over-expressing NRP-1 receptor. We evidenced a statistically significant decrease of the different peptides-conjugated photosensitizers uptake after RNA interference-mediated silencing of NRP-1. Peptides-conjugated photosensitizers allowed a selective accumulation into cells. In mice, no degradation was observed in plasma in vivo 4h after intravenous injection by MALDI-TOF mass spectrometry. This study draws attention to this potential problem with peptides, especially in the case of targeting strategies, and provides useful information for the future design of more stable molecules.
[Show abstract][Hide abstract] ABSTRACT: Photodynamic therapy (PDT) is based on the interaction of a photosensitizing (PS) agent, light, and oxygen. Few new PS agents are being developed to the in vivo stage, partly because of the difficulty in finding the right treatment conditions. Response surface methodology, an empirical modeling approach based on data resulting from a set of designed experiments, was suggested as a rational solution with which to select in vivo PDT conditions by using a new peptide-conjugated PS targeting agent, neuropilin-1.
A Doehlert experimental design was selected to model effects and interactions of the PS dose, fluence, and fluence rate on the growth of U87 human malignant glioma cell xenografts in nude mice, using a fixed drug-light interval. All experimental results were computed by Nemrod-W software and Matlab.
Intrinsic diameter growth rate, a tumor growth parameter independent of the initial volume of the tumor, was selected as the response variable and was compared to tumor growth delay and relative tumor volumes. With only 13 experimental conditions tested, an optimal PDT condition was selected (PS agent dose, 2.80 mg/kg; fluence, 120 J/cm(2); fluence rate, 85 mW/cm(2)). Treatment of glioma-bearing mice with the peptide-conjugated PS agent, followed by the optimized PDT condition showed a statistically significant improvement in delaying tumor growth compared with animals who received the PDT with the nonconjugated PS agent.
Response surface methodology appears to be a useful experimental approach for rapid testing of different treatment conditions and determination of optimal values of PDT factors for any PS agent.
International journal of radiation oncology, biology, physics 08/2009; 75(1):244-52. DOI:10.1016/j.ijrobp.2009.04.004 · 4.26 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The strategy developed aims to favor the vascular effect of photodynamic therapy (PDT) by targeting tumor vasculature. This approach is considered by coupling a photosensitizer (PS) to an heptapeptide targeting neuropilin-1 (NRP-1). We previously demonstrated that this new conjugated PS, which binds to recombinant NRP-1 protein, was a much more potent PS compared to the non-conjugated PS in human umbilical vein endothelial cells (HUVEC) expressing NRP-1, due to the coupling of the peptide moiety. To argue the involvement of NRP-1 in the conjugated PS cellular uptake, MDA-MB-231 breast cancer cells were used, strongly over-expressing NRP-1 receptor, and we evidenced a significant decrease of the conjugated PS uptake after RNA interference-mediated silencing of NRP-1. In mice xenografted ectopically with U87 human malignant glioma cells, we demonstrated that only the conjugated PS allowed a selective accumulation in endothelial cells lining tumor vessels. Vascular endothelial growth factor (VEGF) plasma and tumor levels could not prevent the recognition of the conjugate by NRP-1. The vascular effect induced by the conjugated PS, was characterized by a reduction in tumor blood flow around 50% during PDT. In vivo, the photodynamic efficiency with the conjugated PS induced a statistically significant tumor growth delay compared to the non-coupled PS. The peptide-conjugated chlorin-type PS uptake involves NRP-1 and this targeting strategy favors the vascular effect of PDT in vivo.
Journal of photochemistry and photobiology. B, Biology 06/2009; 96(2):101-8. DOI:10.1016/j.jphotobiol.2009.04.008 · 2.96 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Destruction of the neovasculature is essential for efficient tumor eradication by photodynamic therapy (PDT). The PDT anti-vascular effect can be promoted by developing addressed photosensitizers localized preferentially to the tumor vascular compartment. A new photosensitizer conjugated to an heptapeptide [H-Ala-Thr-Trp-Leu-Pro-Pro-Arg-OH (ATWLPPR)] targeting neuropilin-1, a Vascular Endothelial Growth Factor (VEGF) co-receptor, has been synthesized. It was administered intravenously for an easier access to endothelial cells lining the vasculature in human malignant glioma-bearing nude mice. Plasma pharmacokinetic parameters were derived from plasma concentration-time data using a non-compartmental analysis and validated a relatively rapid elimination from the blood compartment with an elimination rate constant of 0.062 h(-1) and a biological half-life of 11.0 h. The photosensitizer was mainly concentrated in organs such as liver, spleen and kidneys, which are rich in reticuloendothelial cells. In these organs, the elimination profiles of the photosensitizer were comparable, with half-lives as short as 12.2, 15.1 and 19.7 h, respectively. The peptidic moiety of the conjugated photosensitizer was degraded to various rates depending on the organ considered, most of the degradation process occurred in organs of the reticuloendothelial system. A metabolic product resulting from the enzymatic cleavage of the peptide bond between Ala and Thr was detected in plasma at all the examined time points from 2 h post-injection. The conjugated photosensitizer accumulated rapidly and at high levels in the tumor, with 2.3% of injected dose per gram of tumor tissue at 1 h after injection. Taking into account the aspecific uptake of the degradation product, the tumor levels of total photoactivable compounds might exhibit an interesting photodynamic activity. On the contrary, levels of total photoactivable compounds remained low in the skin. This study provides essential information for the choice of the time interval not to exceed to activate the photosensitizer.
[Show abstract][Hide abstract] ABSTRACT: Destruction of the neovasculature is essential for efficient tumour eradication by photodynamic therapy (PDT). Since the over-expression of Vascular Endothelial Growth Factor (VEGF) receptors is correlated with tumour angiogenesis and growth, we conjugated a photosensitiser (5-4-carboxyphenyl)-10,15,20-triphenyl-chlorin, TPC) via a spacer (6-aminohexanoic acid, Ahx) to a neuropilin-1 (NRP-1) specific homing heptapeptide (ATWLPPR) targeting tumour vasculature (Tirand et al. 2006). The intratumoural localisation of the photosensitiser, its stability and its metabolic profile have been studied on a model of nude mice xenografted with U87 human malignant glioma cells (Tirand et al. 2007). By fluorescence microscopy, we evidenced a selective accumulation of the conjugated photosensitiser in the endothelial cells bordering the tumour vessels compared with unconjugated photosensitiser. TPC-Ahx-ATWLPPR accumulated at high levels in the tumour and tissues of the reticuloendothelial system, particularly by liver and spleen. TPC-Ahx-ATWLPPR was stable in vitro in plasma for at least 24 h at 37°C. In vivo, the peptide moiety was progressively degraded from 2h post injection, resulting in the formation of a metabolic product, TPC-Ahx-A (Tirand, et al. 2007). In order to improve the heptapeptide stability towards proteases (Adessi and Soto 2002) and to avoid any non-selective accumulation of the metabolic product, this photosensitiser has been coupled to pseudopeptides on solid support. We have, for the first time, studied the affinity of these pseudopeptides towards the different VEGF165 (isoform 165 of VEGF) receptors (NRP-1, NRP-2, Flt-1 and KDR) by competition experiments. .
[Show abstract][Hide abstract] ABSTRACT: Because angiogenic endothelial cells of the tumor vasculature represent an interesting target to potentiate the antivascular effect of photodynamic therapy, we recently described the conjugation of a photosensitizer [5-(4-carboxyphenyl)-10,15,20-triphenylchlorin (TPC)], via a spacer [6-aminohexanoic acid (Ahx)], to a vascular endothelial growth factor receptor-specific heptapeptide [H-Ala-Thr-Trp-Leu-Pro-Pro-Arg-OH (ATWLPPR)] and showed that TPC-Ahx-ATWLPPR binds to neuropilin-1. Because peptides often display low stability in biological fluids, we examined the in vivo and in vitro stability of this conjugate by high-performance liquid chromatography and matrix-assisted laser desorption ionization/time of flight mass spectrometry. TPC-Ahx-ATWLPPR was stable in vitro in human and mouse plasma for at least 24 h at 37 degrees C but, following i.v. injection in glioma-bearing nude mice, was degraded in vivo to various rates, depending on the organ considered. TPC-Ahx-A was identified as the main metabolic product, and biodistribution studies suggested that its appearance in plasma mainly resulted from the degradation of the peptidic moiety into organs of the reticuloendothelial system. According to in vitro cell culture experiments, TPC-Ahx-ATWLPPR was also significantly degraded after incorporation in human umbilical vein endothelial cells (HUVEC), mainly into TPC-Ahx-A and to a lesser extent into TPC-Ahx-AT and TPC-Ahx-ATWLPP. TPC-Ahx-ATWLPPR mostly localized into lysosomes, and when HUVEC were treated with the lysosomal enzymes' inhibitor ammonium chloride, this resulted in a significant decrease of the peptide degradation. This study provides essential information for the choice of the time of activation of the photosensitizer (drug-light interval) not to be exceeded and for the future design of more stable molecules.
Drug Metabolism and Disposition 06/2007; 35(5):806-13. DOI:10.1124/dmd.106.013763 · 3.25 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: La thérapie photodynamique (PDT) est une méthode proposée pour le traitement de certains cancers. Cette thérapie est basée sur une activation par la lumière de médicaments dits photo?activables appelés photosensibilisateurs. Les cellules endothéliales tumorales angiogéniques sont des cibles intéressantes pour promouvoir l'effet anti-vasculaire de la PDT. Nous avons récemment décrit (Schneider et al., 2006 ; Tirand et al., 2006) le couplage d'un photosensibilisateur de type chlorine (chlorine 5-(4-carboxyphényl)-10,15,20-triphényl, TPC), via un bras espaceur (6-acide aminohexanoïque, Ahx), à un heptapeptide (ATWLPPR), ciblant neuropiline-1 (NRP-1), un co-récepteur du VEGF (Vascular Endothelial Growth Factor) surexprimé dans les cellules endothéliales angiogéniques. TPC-Ahx-ATWLPPR s'est révélé stable dans du plasma in vitro à 37°C pendant 48h. In vivo, une dégradation peptidique apparaît progressivement dès 2h p.i. aboutissant principalement à la formation du composé TPC-Ahx-A (Tirand et al., 2007). Ce produit de dégradation a été mis en évidence majoritairement dans les organes du système réticulo-endothélial. Afin de stabiliser l'heptapeptide vis-à-vis des peptidases tissulaires (Adessi and Soto, 2002) et d'éviter toute accumulation non sélective du produit de dégradation, ce photosensibilisateur a été couplé à des pseudopeptides (aTWLPPR, rpplwta et Ay[CH2NH]TWLPPR) sur support solide. Nous avons, dans un premier temps, étudié l'affinité de ces pseudopeptides vis-à-vis des différents récepteurs au VEGF165 (NRP-1, NRP-2, Flt-1 et KDR) par compétition en utilisant du VEGF165 biotinylé. Les concentrations efficaces en peptides pour lesquelles la fixation du VEGF165 a été déplacée de 50 % (EC50) ont été comparées. Les récepteurs NRP-1 et NRP-2 sont reconnus par aTWLPPR (EC50=13µM et 7,6 µM) et Ay[CH2NH]TWLPPR (EC50=22µM et 8,9 µM) sans modification d'affinité par comparaison à ATWLPPR (EC50=13,2µM et 7,4 µM) alors que le peptide rétro-inverso n'est pas affin vis-à-vis de ces récepteurs. Ces peptides seront ensuite couplés à la TPC afin d'évaluer leur affinité après couplage à la molécule photoactivable et surtout, leur stabilité in vivo.