Novel Photosensitizers Trigger Rapid Death of Malignant Human Cells and Rodent Tumor Transplants via Lipid Photodamage and Membrane Permeabilization

BACKGROUND: Apoptotic cascades may frequently be impaired in tumor cells; therefore, the approaches to circumvent these obstacles emerge as important therapeutic modalities. METHODOLOGY/PRINCIPAL FINDINGS: Our novel derivatives of chlorin e(6), that is, its amide (compound 2) and boronated amide (compound 5) evoked no dark toxicity and demonstrated a significantly higher photosensitizing efficacy than chlorin e(6) against transplanted aggressive tumors such as B16 melanoma and M-1 sarcoma. Compound 5 showed superior therapeutic potency. Illumination with red light of mammalian tumor cells loaded with 0.1 µM of 5 caused rapid (within the initial minutes) necrosis as determined by propidium iodide staining. The laser confocal microscopy-assisted analysis of cell death revealed the following order of events: prior to illumination, 5 accumulated in Golgi cysternae, endoplasmic reticulum and in some (but not all) lysosomes. In response to light, the reactive oxygen species burst was concomitant with the drop of mitochondrial transmembrane electric potential, the dramatic changes of mitochondrial shape and the loss of integrity of mitochondria and lysosomes. Within 3-4 min post illumination, the plasma membrane became permeable for propidium iodide. Compounds 2 and 5 were one order of magnitude more potent than chlorin e(6) in photodamage of artificial liposomes monitored in a dye release assay. The latter effect depended on the content of non-saturated lipids; in liposomes consisting of saturated lipids no photodamage was detectable. The increased therapeutic efficacy of 5 compared with 2 was attributed to a striking difference in the ability of these photosensitizers to permeate through hydrophobic membrane interior as evidenced by measurements of voltage jump-induced relaxation of transmembrane current on planar lipid bilayers. CONCLUSIONS/SIGNIFICANCE: The multimembrane photodestruction and cell necrosis induced by photoactivation of 2 and 5 are directly associated with membrane permeabilization caused by lipid photodamage

CONFOCAL MICROSCOPY STUDY OF BIOLOGICAL PECULIARITIES OF SCAFFOLD MADE FROM RECOMBINANT SPIDER SILK

We studied the viability and dynamic of cell distribution during long-term cultivation of broblasts 3T3 in spider silk spidroin 1-based scaffold. Laser scanning confocal microscopy is shown to have advantages for visualization of cells situated on the external and internal surfaces of scaffold. Fibroblasts maintain high proliferative ability and viability during long term cultivation. Spidroin 1-based scaffold are the perspective materials for bioengineering

Functional bread enreached with regional bioactive food additives

The scientific work presents materials of theoretical and experimental substantiation of the use of regional bioactive food additives, namely wild plants of the legume family (Fabaceae), permitted for use in the food industry: sweet clover (Melilotus officinalis), red meadow clover (Trifolium pratense), white acacia (Robinia pseudoacacia) in the production of functional foods [1]. Experimental studies of the safety indicators of food wild-growing plants (FWP) have been carried out. It was found that the studied FWP growing in ecologically clean areas of the North Ossetia-Alania are distinguished by a low mass fraction of toxic substances. Two fractions of cellulose were investigated: 1 - water-soluble (pectic acid, pectin, methylcellulose); 2 - water-insoluble (protopectin, cellulose, hemicellulose). The increased content of dietary fiber in WGP powders has been established, corresponding to the physiological norms of satisfying the daily requirement for FWP and, in this regard, indicating the expediency of using functional food in formulations for the prevention and treatment of diabetes mellitus, obesity, and atherosclerosis. The sorption capacity of WGP powders with respect to lead has been investigated. All studied powders of WGP modification products are characterized by high sorption capacity. The technologies of flour products with the addition of FWP powders have been developed. The results of clinical and preclinical studies of the developed functional bread show a corrective effect of glycemic blood parameters, as well as a decrease in the lead content in the blood of fed laboratory rat pups by almost two times, compared with the control

Protein-Mediated Carotenoid Delivery Suppresses the Photoinducible Oxidation of Lipofuscin in Retinal Pigment Epithelial Cells

Lipofuscin of retinal pigment epithelium (RPE) cells is a complex heterogeneous system of chromophores which accumulates as granules during the cell’s lifespan. Lipofuscin serves as a source of various cytotoxic effects linked with oxidative stress. Several age-related eye diseases such as macular degeneration of the retina, as well as some severe inherited eye pathologies, are accompanied by a significant increase in lipofuscin granule concentration. The accumulation of carotenoids in the RPE could provide an effective antioxidant protection against lipofuscin cytotoxic manifestations. Given the highly lipophilic nature of carotenoids, their targeted delivery to the vulnerable tissues can potentially be assisted by special proteins. In this study, we demonstrate how protein-mediated delivery of zeaxanthin using water-soluble Bombyx mori carotenoid-binding protein (BmCBP-ZEA) suppresses the photoinducible oxidative stress in RPE cells caused by irradiation of lipofuscin with intense white light. We implemented fluorescence lifetime imaging of the RPE cell culture ARPE-19 fed with lipofuscin granules and then irradiated by white light with and without the addition of BmCBP-ZEA. We demonstrate that after irradiation the mean fluorescence lifetime of lipofuscin significantly increases, while the presence of BmCBP-ZEA at 200 nM concentration suppresses the increase in the average lifetime of lipofuscin fluorescence, indicating an approx. 35% inhibition of the oxidative stress. This phenomenon serves as indirect yet important evidence of the efficiency of the protein-mediated carotenoid delivery into pigment epithelium cells

Amides of pyrrole- and thiophene-fused anthraquinone derivatives: A role of the heterocyclic core in antitumor properties

Heteroarene-fused anthraquinone derivatives represent a class of perspective anticancer drug candidates capable of targeting multiple vital processes including drug resistance. Taking advantage of previously demonstrated potential of amide derivatives of heteroarene-fused anthraquinones, we herein dissected the role of the heterocyclic core in antitumor properties. A new series of naphtho[2,3-f]indole-3- and anthra[2,3-b]thiophene-3-carboxamides was synthesized via coupling the respective acids with cyclic diamines. New compounds demonstrated a submicromolar antiproliferative potency close to doxorubicin (Dox) against five tumor cell lines of various tissue origin. In contrast to Dox, the new compounds were similarly cytotoxic for HCT116 colon carcinoma cells (wild type p53) and their isogenic p53 knockout counterparts. Modification of the heterocyclic core changed the targeting properties: the best-in-series naphtho[2,3-f]indole-3-carboxamide 8 formed more affine complexes with DNA duplex than furan and thiophene analogs, a property that can be translated into a stronger inhibition of topoisomerase 1 mediated DNA unwinding. At tolerable doses the water soluble derivative 8 significantly inhibited tumor growth (up to 79%) and increased the lifespan (153%) of mice bearing P388 lymphoma transplants. Together with better solubility for parenteral administration and well tolerance by animals of the indole derivative 8 indicates prospects for further search of new antitumor drug candidates among the heteroarene-fused anthraquinones.status: publishe

ИСПОЛЬЗОВАНИЕ МЕТОДА КОНФОКАЛЬНОЙ МИКРОСКОПИИ ДЛЯ ИЗУЧЕНИЯ БИОЛОГИЧЕСКИХ СВОЙСТВ МАТРИКСА ИЗ РЕКОМБИНАНТНОЙ ПАУТИНЫ

We studied the viability and dynamic of cell distribution during long-term cultivation of broblasts 3T3 in spider silk spidroin 1-based scaffold. Laser scanning confocal microscopy is shown to have advantages for visualization of cells situated on the external and internal surfaces of scaffold. Fibroblasts maintain high proliferative ability and viability during long term cultivation. Spidroin 1-based scaffold are the perspective materials for bioengineering. Изучена жизнеспособность и динамика распределения мышиных фибробластов 3T3 в матрице на ос- нове рекомбинантного белка каркасной нити паутины спидроина 1 при длительном культивировании. Показана возможность и преимущества использования конфокальной микроскопии как для визуализа- ции клеток, находящихся в глубоких слоях трехмерной матрицы, так и для наблюдения за изменением состояния культивируемых клеток. Популяция фибробластов сохраняла высокие показатели жизнеспо- собности и пролиферативной активности, а инфильтрация матрицы клеточной популяцией становилась однородной в течение длительного времени культивирования. Это позволяет рассматривать матрицы на основе спидроина 1 в качестве перспективного материала для создания биоинженерных конструкций.