Iman Gomaa

Chlorophyll derivative mediated PDT versus methotrexate: An in vitro study using MCF-7 cells

BACKGROUND Breast cancer is the most common cause of cancer deaths among women worldwide. Although chemotherapy is a standard method for the treatment of breast cancer, the photodynamic therapy (PDT) is a recent promising modality for cancer diagnosis and treatment. Its major advantages over chemotherapy are better selectivity of tumour tissue destruction and lack of severe local and systemic complications. This work is directed towards evaluation of the efficacy of Photodynamic therapy using chlorophyll derivative (CHL) as a photosensitizer in treatment of breast cancer. It also aims at investigation of the genetic safety of chlorophyll mediated PDT in comparison to the conventional chemotherapy. METHODS Both methotrexate (MTX) and light activated chlorophyll derivative were used to target MCF-7 breast cancer cell line. Standard karyotyping and alkaline single cell microgel electrophoresis assay (Comet assay) were applied on normal human peripheral blood lymphocytes (HPL) in order to investigate the respective possible mutagenic and genotoxic side effects that might result from application of each therapeutic modality. RESULTS Results obtained from this study showed that 50% of MCF-7 tumour cell death (LC(50)) was reached by using a concentration of chlorophyll derivative that is 138 times lower than MTX. Moreover, chlorophyll derivative exerted no genetic side effects as compared to MTX that resulted into several types of chromosomal breakages. CONCLUSIONS Compared to MTX, light activated chlorophyll derivative proved to be a better candidate for breast cancer cell toxicity, referring to its higher efficacy at tumour cells killing, safety to normal cells and simple method of extraction.

Synthesis and cytotoxic activities of some pyrazoline derivatives bearing phenyl pyridazine core as new apoptosis inducers

The cyclization of chalcones 3a-3u with 3-hydrazinyl-6-phenylpyridazine 7 under basic condition led to the formation of new pyrazoline derivatives 8a-8u. All final compounds were characterized by spectral and elemental analyses. They were screened for their antiproliferative activities against A549 (lung), HepG-2 (liver), CaCo-2 (intestinal) and MCF-7 (breast) cancer cell lines. Some of the synthesized compounds exhibited promising antiproliferative activities especially compound 8k with IC50 values of 8.33, 1.67 and 10 μM against HepG-2, MCF-7 and CaCo-2 cancer cell lines, respectively. Moreover, their antiproliferative activity was due to apoptosis rather than necrosis induction except compound 8h which exhibited equal apoptotic and necrotic properties. Compound 8k showed 5 fold increase in caspase-3 activity indicating that the apoptosis proceeds via caspase-3 activation.

Liposomal delivery of ferrous chlorophyllin: A novel third generation photosensitizer for in vitro PDT of melanoma

BACKGROUND Cutaneous melanoma (CM) has substantially increased among Caucasian populations in the past few decades. This increased the number of CM deaths throughout the world. Pigmentation of melanoma reduces the efficacy of photodynamic therapy (PDT). Third generation photosensitizers (PSs) are characterized by improved targeting to the diseased tissue and reduced systemic side effects. This study is directed towards synthesis and characterization of liposomes encapsulating sodium ferrous chlorophyllin (Fe-CHL) and assessing its efficacy as a PS in PDT of melanoma. METHODS Phenylthiourea (PTU) was used as a melanin synthesis inhibitor. PDT has been applied on de-pigmented melanoma cells using liposomes-encapsulated Fe-CHL. Cell death mechanisms after PDT were evaluated. RESULTS Treatment of melanoma cells with 200μM of PTU for 48h provided 49.9% melanin inhibition without significant cytotoxicity. Transmission electron microscope (TEM) results proved an increase in the cellular uptake of liposomes by increasing incubation period from 6 to 24h via endocytosis with preferential accumulation in the mitochondria and the nucleus. Following de-pigmentation, PDT was applied resulting in LC50 of 18.20 and 1.77μM after 24 and 48h incubation with liposomes-encapsulated Fe-CHL respectively and exposure to 56.2J/cm2 monochromatic red laser of wavelength of 652nm. Mechanism of cell death of Fe-CHL mediated PDT was found to be a combination of both apoptosis and necrosis. CONCLUSIONS Liposomes could be efficiently employed as a potential sustained release delivery system in the Fe-CHL-mediated PDT of de-pigmented melanoma.

Targeted penetration of MCF-7 cells using iron-oxide nanoparticles in vitro

We achieve selective penetration of MCF-7 breast cancer cells using iron-oxide nanoparticles without causing a permanent damage to the membrane and without any effect on the cell morphology. The nanoparticles are controllably pulled towards the cells under the influence of the magnetic field gradients. First, the nanoparticles are fabricated and their magnetic dipole moment is characterized to be 7.8×10-8 A.m2, at magnetic field of 60 mT and mass of 1.80×10-9 kg. This characterization is done by measuring the magnetic force exerted on their dipole moment under the influence of controlled magnetic field gradient. Second, a magnetic-based control system is designed and used to achieve selective targeting of the cells under microscopic guidance. We find that the magnetic control achieves immediate uptake of nanoparticles in the MCF-7 cells without incubation for relatively long time, using magnetic force less than 51 nN. In addition, a microforce sensing probe is used to characterize the impendence of the cells to limit the exerted magnetic force during penetration and to avoid causing damage to the membrane. We find that a single cell can overcome penetration force in excess of 13.3 μN.

Dermal delivery of Fe-chlorophyllin via ultradeformable nanovesicles for photodynamic therapy in melanoma animal model

ABSTRACT Melanoma is resistant to chemotherapeutics with poor prognosis and high potential of metastasis. Photodynamic therapy (PDT) represents a localized therapeutic modality, as cytotoxicity occurs when light activates photosensitizer (PS) at the tumour site. The aim of this study is dermal delivery of a high molecular weight hydrophilic photosensitizer (PS), ferrous chlorophyllin (Fe‐CHL) via transethosomes for treatment of melanoma by PDT. Transethosomes were made of phosphatidyl choline, edge activator and 20%w/v Ethanol. They were evaluated for mean size, zeta potential, entrapment efficiency, ex‐vivo permeation, localization in skin layers by transmission electron microscope (TEM), and finally, evaluated in melanoma animal model. Transethosomes of different mean vesicle size were evaluated for their skin retention and permeation through mice skin. TE of ˜500nm (E3) being ultradeformable showed deep localization in skin confirmed by ex‐vivo and TEM micrographs without permeation of PS to recipient compartment due to its size. The proposed study offers successful treatment of resistant melanoma by PDT, where complete tumour regression of small tumours occurred after single PDT, while large tumours after double PDT without recurrence for 8months. This indicates the efficiency of nanovesicles in PS delivery and the efficiency of Fe‐CHL in production of reactive oxygen species.

In vivo suppression of solid Ehrlich cancer via chlorophyllin derivative mediated PDT: an albino mouse tumour model

In this study, copper chlorophyllin was used as a photosensitizer for photodynamic therapy (PDT) in Ehrlich tumour mouse model. Six groups of animals comprising 5 animals per group were subcutaneously transplanted with 1x106 Ehrlich tumour cells. A single dose of 200 μg/gm body weight chlorophylin derivative was administered by intravenous (IV) or intratumoral (IT) routes. Mice were exposed to monochromatic red laser of 630 nm for 1 h, and tumour regression was followed up for three consecutive months post treatment. Several Biochemical, histological and molecular tests were performed in order to evaluate the efficacy and safety of the applied treatment. An interest has been also directed towards investigating the molecular mechanisms underlying chlorophyllin derivative mediated PDT. PDT-treated animals via either the IV or IT routes showed significant decrease in tumour size 72 h post-treatment. Tumours at the IV-PDT group disappeared totally within a week with no recurrence over three months follow up. In the IT-PDT, the decrease in tumour size at the first week was interrupted by a slight increase; however never reached the original size. Histological examination of tumour tissues of the IV-PDT group at 24 h post treatment demonstrated restoring the normal muscle tissue architecture, and the biochemical assays indicated normal liver functions. The immunohistochemical analysis of caspase-3, and the quantitative PCR results of caspases-8 and 9 proved the presence of extrinsic apoptotic pathway after cholorphyllin derivative-mediated PDT. In conclusion IV-PDT strategy proved better cure rate than the IT-PDT, with no recurrence over three months of follow up.