Study of the pO2-Sensitivity of the Dendrimeric and Free Forms of Pd-meso-tetra(4-carboxyphenyl)porphyrin, Incorporated or not in Chitosan-Based Nanoparticles

The concentration of oxygen and its rate of consumption are important factors in certain medical treatments, such as radiotherapy and photodynamic therapy (PDT). Measuring the tissue concentration of oxygen or its partial pressure (pO2) can be achieved by taking advantage of the oxygen-dependent luminescence lifetime of certain molecules, including metallo-porphyrin derivatives, due to the oxygen-dependent quenching of their triplet state. Unfortunately, most of these porphyrin derivatives are phototoxic due to the O21? produced in the pO2 measurement procedure. The aim of this work was to characterize new nanoparticle oxygen sensors, where the palladium-porhyrin molecule (Pd-meso-tetra(4-carboxyphenyl)porphyrin) or its dendrimer form, is incorporated into an oxygen permeable matrix of chitosan-based colloidal particles. It was hypothesized that the reactive O21? produced during the pO2 measurement would react inside the particle thus reducing its toxicity for the surrounding tissue, whereas the 3? ground state of O2, that is to be measured, would diffuse freely in the peptide. We observed that the incorporation of the porphyrin in the nanoparticles resulted in a reduction of the phosphorescence lifetime sensitivity to pO2 by about one order of magnitude. Our studies of these new sensors indicate that the oxygen concentration can be measured in aqueous solutions with a precision of ±20% for oxygen concentrations ranging between 0% and 25%

The chicken chorioallantoic membrane model in biology, medicine and bioengineering

The chicken chorioallantoic membrane (CAM) is a simple, highly vascularized extraembryonic membrane, which performs multiple functions during embryonic development, including but not restricted to gas exchange. Over the last two decades, interest in the CAM as a robust experimental platform to study blood vessels has been shared by specialists working in bioengineering, development, morphology, biochemistry, transplant biology, cancer research and drug development. The tissue composition and accessibility of the CAM for experimental manipulation, makes it an attractive preclinical in vivo model for drug screening and/or for studies of vascular growth. In this article we provide a detailed review of the use of the CAM to study vascular biology and response of blood vessels to a variety of agonists. We also present distinct cultivation protocols discussing their advantages and limitations and provide a summarized update on the use of the CAM in vascular imaging, drug delivery, pharmacokinetics and toxicology

Vascular regrowth following photodynamic therapy in the chicken embryo chorioallantoic membrane

Photodynamic therapy (PDT) induces damage to the endothelium, which can lead to increased vascular permeability and, under intensive PDT conditions, even to platelet aggregation, vasoconstriction, and blood flow stasis. Eventually, ischemia, hypoxia, and inflammation can occur, resulting in angiogenesis. We studied the sequence of the vascular events after Visudyne®-PDT in the chicken chorioallantoic membrane (CAM) at day 11 of development. Using epi-fluorescence microscopy, we monitored the regrowth of capillaries in the PDT treated area. Immediately after irradiation, the treatment resulted in blood flow arrest. And 24h post PDT, sprouting of new blood vessels was observed at the edge of the PDT zone. Neovessels looping out from the edge of the PDT zone gave rise to specialized endothelial tip structures guiding the vessels towards the center of the treated area. At 48h almost all of the treated area was repopulated with functional but morphologically altered vasculature. These observations also showed reperfusion of some of the vessels that had been closed by the PDT treatment. CAM samples were immunohistochemically stained for Ki-67 showing proliferation of endothelial cells in the PDT area. Also, several markers of immature and angiogenic blood vessels, such as αVβ3-integrin, vimentin and galectin-1, were found to be enhanced in the PDT area, while the endothelial maturation marker intercellular adhesion molecule (ICAM)-1 was found to be suppressed. These results demonstrate that the new vascular bed is formed by both neo-angiogenesis and reperfusion of existing vessels. Both the quantitative real-time RT-PCR profile and the response to pharmacological treatment with Avastin®, an inhibitor of angiogenesis, suggest that angiogenesis occurs after PDT. The observed molecular profiling results and the kinetics of gene regulation may enable optimizing combination therapies involving PDT for treatment of cancer and other disease

Thermoresponsive fluorinated small-molecule drugs: a new concept for efficient localized chemotherapy

Hyperthermia is currently being explored as an adjuvant treatment to conventional therapies with chemotherapeutic agents based on thermoresponsive macromolecules. Although the concept of hyperthermia has existed for many years it has yet to become routinely used in the clinical management of cancer. The development of small thermoresponsive molecules could help to change this paradigm. Temperaturesensitive compounds have recently been developed by covalently modifying drug and drug-like molecules with thermomorphic perfluorinated appendages. Lead thermoresponsive compounds have been validated in a pre-clinical model, displaying high tumor growth inhibition, with strong synergies observed between hyperthermia and the thermomorphic compounds

Thermoresponsive organometallic arene ruthenium complexes for tumour targeting

Application of mild hyperthermia can increase the cytotoxicity of anticancer drugs in tumour cells. In this report, we describe low molecular weight thermoactive ruthenium-based drugs with fluorous chains that are selectively triggered by mild hyperthermia. The organometallic complexes were prepared, characterized, and evaluated for their in vitro cytotoxicity against a panel of human cancer cell lines and non-cancerous immortalized cells. The compounds show considerable chemo-thermal selectivity towards cancer cells (ca. 5 mu M versus >500 mu M for healthy cells) for the compound with the longest fluorous chain

In vivo evaluation of small-molecule thermoresponsive anticancer drugs potentiated by hyperthermia

Hyperthermia used as an adjuvant with chemotherapy is highly promising in the treatment of certain cancers. Currently, the small molecule drugs used in combination with hyperthermia were not designed for this application. Herein, we report the evaluation of a chlorambucil and a ruthenium compound modified with a long fluorous chain, which exhibit thermoresponsive activity in colorectal adenocarcinoma xenografts in athymic mice in combination with mild hyperthermia (42 degrees C). Intraperitoneal injection of the derivatives followed by local hyperthermia showed a synergistic tumor growth reduction by 79% and 90% for the chlorambucil and ruthenium-based derivatives, respectively, with the latter exhibiting a higher synergy in combination with hyperthermia compared to the monotherapies. Histological analysis shows that both derivatives in combination with hyperthermia significantly decrease the number of proliferating tumor cells

Optimization and regeneration kinetics of lymphatic-specific photodynamic therapy in the mouse dermis

Lymphatic vessels transport fluid, antigens, and immune cells to the lymph nodes to orchestrate adaptive immunity and maintain peripheral tolerance. Lymphangiogenesis has been associated with inflammation, cancer metastasis, autoimmunity, tolerance and transplant rejection, and thus, targeted lymphatic ablation is a potential therapeutic strategy for treating or preventing such events. Here we define conditions that lead to specific and local closure of the lymphatic vasculature using photodynamic therapy (PDT). Lymphatic-specific PDT was performed by irradiation of the photosensitizer verteporfin that effectively accumulates within collecting lymphatic vessels after local intradermal injection. We found that anti-lymphatic PDT induced necrosis of endothelial cells and pericytes, which preceded the functional occlusion of lymphatic collectors. This was specific to lymphatic vessels at low verteporfin dose, while higher doses also affected local blood vessels. In contrast, light dose (fluence) did not affect blood vessel perfusion, but did affect regeneration time of occluded lymphatic vessels. Lymphatic vessels eventually regenerated by recanalization of blocked collectors, with a characteristic hyperplasia of peri-lymphatic smooth muscle cells. The restoration of lymphatic function occurred with minimal remodeling of non-lymphatic tissue. Thus, anti-lymphatic PDT allows control of lymphatic ablation and regeneration by alteration of light fluence and photosensitizer dose

Low-dose angiostatic tyrosine kinase inhibitors improve photodynamic therapy for cancer: lack of vascular normalization

Photodynamic therapy (PDT) is an effective clinical treatment for a number of different cancers. PDT can induce hypoxia and inflammation, pro-angiogenic side effects, which may counteract its angio-occlusive mechanism. The combination of PDT with anti-angiogenic drugs offers a possibility for improved anti-tumour outcome. We used two tumour models to test the effects of the clinically approved angiostatic tyrosine kinase inhibitors sunitinib, sorafenib and axitinib in combination with PDT, and compared these results with the effects of bevacizumab, the anti-VEGF antibody, for the improvement of PDT. Best results were obtained from the combination of PDT and low-dose axitinib or sorafenib. Molecular analysis by PCR revealed that PDT in combination with axitinib suppressed VEGFR-2 expression in tumour vasculature. Treatment with bevacizumab, although effective as monotherapy, did not improve PDT outcome. In order to test for tumour vessel normalization effects, axitinib was also applied prior to PDT. The absence of improved PDT outcome in these experiments, as well as the lack of increased oxygenation in axitinib-treated tumours, suggests that vascular normalization did not occur. The current data imply that there is a future for certain anti-angiogenic agents to further improve the efficacy of photodynamic anti-cancer therapy

In vivo anti-tumor activity of the organometallic ruthenium(II)-arene complex [Ru(eta(6)-p-cymene)-Cl-2(pta)] (RAPTA-C) in human ovarian and colorectal carcinomas

Based on the clinical success of platinum-based anti-cancer drugs such as cisplatin, carboplatin and oxaliplatin, a variety of other metal-based anti-cancer compounds are being investigated. In particular, a number of ruthenium-based compounds have been identified which exhibit unique biochemical properties and reduced toxicity profiles compared to the clinically used platinum-based drugs. We have developed a series of organometallic ruthenium(II)-arene complexes that were shown to exert anti-metastatic activity with relatively minor activity on primary tumor growth. Here, we show that the prototype compound, [Ru(eta(6)-p-cymene)Cl-2(pta)], where pta = 1,3,5-triaza-7-phosphaadamantane (RAPTA-C), reduces the growth of primary tumors in preclinical models for ovarian and colorectal carcinomas. When administered daily at relatively low doses (0.2 mg kg(-1)), RAPTA-C was shown to significantly reduce the growth of the A2780 ovarian carcinoma transplanted onto the chicken chorioallantoic membrane model. Similar activity was observed in LS174T colorectal carcinoma in athymic mice, albeit at a higher dose. In both models, a clear inhibition of microvessel density was observed, confirming the previously discovered anti-angiogenic mechanism of RAPTA-C. Biodistribution studies with radiolabeled (Ru-103) RAPTA-C indicate that the compound is rapidly cleared from the organs and the bloodstream through excretion by the kidneys. As such, RAPTA-C is a promising compound for translation to clinical evaluation