DIFFERENT FORMULATIONS OF MICROALGAL CRUDE EXTRACTS AND SILVER NANOPARTICLES HAVING CYTOTOXIC EFFECTS ON BREAST CANCER CELLS

Technical description of this Malaysian patent

FIELD OF INVENTION

The present inventiongenerally relates to treating cancer especially breast cancer. More particularly, the present invention relates to different formulations having cytotoxic effects on breast cancer cells without klling the normal healthy cells, and with potent anti-oxidant and anti-microbial activities.

BACKGROUND OF INVENTION

Cancer is a group of diseases involving abnormal cell growth with the potential to invade or spread to other parts of the body. Possible symptoms may include such as a lump, abnormal bleeding, weight loss, and so on. Cancer starts when a single cell becomes mutated and proliferates abnormally.Additional mutation followed by selection of more rapidly growing cells within the population results in cancer cell progression and malignancy. Destroying the normal cellular homeostasis, the cancerous cells show variety of biochemical and physiological changes and may become resistant to conventional chemotherapeutics during transformation process (Majumder et al, 2015). There are various cancer treatments including chemotherapy, hormone therapy, surgery, radiation, immune therapy, and targeted therapy (ACS, 2015). However, the conventional treatment such as chemo- and radiation therapy has limitations due to unexpected drug- associated side effects, lack of specificity of low drug concentrations at the tumor target site, and the development of chemoresistance (Singh et al, 2012; Fanciullino et al, 2013).

For example, US Patent Publication US4261989A discloses Geldanamycin derivatives and antitumor drug. The Geldanamycin derivatives having the formula () can be produced by reacting Geldanamycin with an amine having the formula #STR8# wherein R7 and R8 respectively represent hydrogen atom or a saturated alkyl group having at least two carbon atoms; an unsaturated alkyl group, a substituted lower alkyl group a cycloalkyl group or an aralkyl group and both R7 and R8 cannot be hydrogen atom and R7 and R8 can bonded to form one alkylene group. As the anti-tumor drugs may have side effects causing normal cellular death, thefocus is to develop natural based drugs, with minimal effect on normal healthy cells whilst treating cancer. For example, Chinese Patent Publication CN1130088Adiscloses pure traditional Chinese medicine anti-cancer drug. The drug contains 45 kinds of medicines such as oldenlandiaalgida, camptotheca seed, Holboellia, barbed skullcap, nux-vomica seed, Rhizomaparidis, portulaca, fevervine, Jingangteng, Fructuscnidi, etc. The said drug adopts herbs from nature, and has no side-effect and safety, short course and quick action, and not only can be used for curing cancer, but also for preventing cancer. There is another revolutionary approach for developing anti-cancer drugs in combination with nanoparticles. The co-application of natural compounds with nanomedicineis a promising strategy to combat cancer. Nanomaterials could revolutionize cancer diagnosis and therapy (Gul e Saba& Audullah, 2015; Supraja et al, 2016) and the encapsulation of therapeutic agents with nanoparticles have been used as drug delivery systems (Abdullah et al, 2014). The use of metallic nanoparticles and medically relevant nanoparticles such as silver nanoparticles (AgNPs) have shown different degrees of in vitro cytotoxicity with the ability to target through passive or active targeting of particular diseased cells or tumor tissues (Wicki et al, 2015). To overcome the limitations of conventional chemotherapy, the challenges are to develop new nanoparticles in single platform-based strategies and to address the physiological barriers, limited carrying capacity, enhanced permeability and retention (EPR) effect, variability of nanoparticles, and the regulatory and manufacturing issues (Wicki et al, 2015). AgNPs have gained special interest over gold and copper NPs because of their surface plasmon resonance (SPR) energy which is located away from the inter band transition energy. Various applications have been reported for AgNPs including in catalysis, optoelectronics, detection and diagnostic, anti-microbials and therapeutics. The AgNPs can be exploited in medical and pharmaceutical application due to their low toxicity to human cells and high thermal stability (El-Sheekh& El-Kassas, 2014). The biosynthesis of AgNPs through green routes confer major benefits over chemical synthetic route of other metallic-based anti-cancer agents (Chaudhari et al, 2012; Caroling et al, 2013; Yazdi et al, 2015; Jaffat et al. 2017). For instance, US Patent Publication US8603499B2 discloses cancer cell targeting using nanoparticles. The nanoparticle includes docetaxel; adiblock copolymer of poly(ethylene glycol) and polylactic acid; and a prostate specific membrane antigen ligand conjugated to a diblock copolymer of poly(ethylene glycol) and polylactic acid. Another exemplary US Patent Publication US20100112077A1 discloses combination therapy methods of treating proliferative diseases (such as cancer) comprising a first therapy comprising administering to an individual an effective amount of a taxane in a nanoparticle composition, and a second therapy which may include, for example, radiation, surgery, administration of chemotherapeutic agents (such as an anti-VEGF antibody), or combinations thereof. Also provided are methods of administering to an individual a drug taxane in a nanoparticle composition based on a metronomic dosing regime. However, such nanoparticles are costly and require more effort. Antioxidants are capable of neutralizing free radicals prior to their detrimental physiological effect. However, synthetic antioxidants such as butylatedhydroxytoluene and butylatedhydroxyanisole have recently been reported to have adverse side effects in human health. The search for effective natural antioxidants in food, cosmetic and therapeutic industry is fast emerging as a promising alternative for synthetic antioxidants with regards to low cost, high biocompatibility with dietary intake and zero or reduced side effects (Chaudhuri et al, 2014). Most natural antioxidants are currently derived from plant sources and marine organisms are widely used in the life sciences as a source of compounds with diverse structural forfns and biological activities (Jo et al, 2012). Microalgae are microorganisms that have different morphological, physiological, and genetic traits that confer the ability to produce different biologically active metabolites and are therefore increasingly seen as the viable alternative source of biocompounds. Microalgae are rich in proteins, carbohydrates, minerals, and diverse functional pigments. Their proteins are good health food or animal feed, their carbohydrates are useful as stabilizers and emulsifiers in foods, and their bioactive materials have potential medicinal values. Their pigments could be used as natural food dyes and contain many beneficial minerals. Microalgae are therefore useful in functional foods (Jo et al, 2012; Abdullah et al., 2016; 2017) and can developed as an environmentally friendly and economically viable source of compounds of interest, via optimized controlled culture and integrated bioprocess engineering (Abdullah et al, 2016; 2017). Microalgae have the ability to promote health and reduce the risk of the development of degenerative diseases (de Morais et al, 2015). Bioactive compounds from microalgae can be obtained directly from primary metabolism such as proteins, fatty acids, vitamins, and pigments, or can be synthesized from secondary metabolism. These compounds exhibit antifungal, antiviral, antialgal, antienzymatic, or antibiotic actions (Volk, 2008; Abdullah et al, 2017). Many of these compounds (cyanovirin, oleic acid, linolenic acid, palmitoleic acid, vitamin E, B12, β-carotene, phycocyanin, lutein, and zeaxanthin) have antimicrobial, antioxidant, and ani-inflammatory capacities, with the potential for the reduction and prevention of diseases (Smee et al, 2008; Harun et al, 2010). In most microalgae, the bioactive compounds are accumulated in the biomass and in some cases, these metabolites are excreted into the medium, known as exometabolites. Due to their phototrophic nature, microalgae are subjected to intense oxidative stress generated by reactive oxygen species (ROS) produced during photosynthes To prevent cellular damage by these ROS, microalgae possess antioxidant defence mechanisms such as antixidant enzymes (e.g. superoxide dismutase or catalase) and low molecular weight antioxidants (LMWA) which include carotenoids, glutathione, vitamins (ascorbate and tocopherols), and phenolics (Goiris et al, 2012; Adulah et al, 2017).However, in some cases, the ICso values of the crude extracts may be much higher than the 20-30 ug/ml level stipulated for any compounds to be considered as cytotoxic. Therefore, there exists a need for developing a formulation having natural based compounds in combination with nanoparticles such that the formulations may be developed through low cost and greener method.

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