الفهرس 
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Abstract
This study was designed to find out the promising drug delivery system for the co-delivery of the potent drug pemetrexed (PMT) and the herbal drug honokiol (HK) for the treatment of different kinds of cancers. Naturally -derived nanogels (NGs) were tailored and fabricated of lactoferrin(Lf) and carboxymethylcellulose via a green self-assembly technique. Such naturally derived targeted NGs were proposed for the incorporation of the two drugs in the same drug delivery system. Therefore, different methods and approaches were designed for loading the NGs with the two drugs.
The first concern of this study was the optimization of Lf /CMC nanogels. Herein, the NGs fabrication was based on heat-induced complexation between protein and polysaccharides.
Different factors including mass ratio of lactoferrin and CMC, pH and reaction temperature were studied to optimize the physical and chemical properties of the NGs. These factors were found to have significant effects on the preparation of nanogels At that point, it could be concluded that the optimal conditions which aided in the fabrication of the proper and customizable blank (Lf-CMC) NGs with particle size 190.7±2.1nm, PDI of 0.376±0.003 and Z-potential of -32±0.8 were as follows :1.Lf and CMC in mass ratio of 4:1 were used to form the Lf: CMC NGs2.pH was adjusted to pH 7.5 for Lf to be positively charged.3.Temperature used for denaturation was 70°C for 45 minBeside the physical cross linking utilized for green self -assembly of NGs, chemical cross linking using cocktail of EDC.HCL/ K.Oxyma (molar ratio of EDC to carboxyl group of the CMC was set to be 1:1), was used in our fabricated NGs as they provided superior sustainability during drug release, customizable mean particle size and stability compared to physically self-assembled NGs.
Particle size of the cross-linked Lf-CMC NGs was found to be 123 ±0.09 nm with PDI of 0.29±0.002 and Z-potential of -28.9±0.7Concerning the loading of the first drugPMT, it was conjugated to Lf polymer which offers specific targeting to tissues or cells.
The conjugation of PMT to Lf was performed by carbodiimide reaction where an amide bond was formed betweenPMT and Lf in the presence of EDC.HCL/K.Oxyma as a cross linker.
The carboxylic group of PMT was activated by using EDC.HCl/K salt of Oxyma to form an intermediate active ester capable of covalent attachment to the amino groups of Lf.
Several conjugation ratios between Lf and PMT were studiedto obtain the most stable conjugate. The most stable PMT-Lf conjugate was obtained with conjugation efficiency of 80 % and PMT drug content of 10.7 %.
< PMT-Lf conjugate was characterized by using H1-NMR and MALDI-TOF to prove the successful synthesis of the amide linkage.
With regard to the seconddrug (HK) loading, the use of cyclodextrins molecules represents another approach to increase drug solubility. Therefore, the inclusion complex of honokiol (HK) with hydroxypropyl-β-cyclodextrin (HPβCD) was prepared to improve the solubility and 103 stability of HK.
<The molar ratio of honokiol to HPβCD was 1:2. Three different approaches of addition weretested to get the optimized NGformulations. The physical loaded HK-HP-β-CDinclusion complex in cross-linked F4CPLformulation showed the most promising particle size of 193.4±2.02nm with PDI of 0.32±0.001 and Z-potential of -34.5±0.7.
<In vitro release study showed preferable sustained release profiles of the most promising F4CPL when compared toF4CEand F4UENGs and to the free drugHKat two different pHs of 7.4 and 5.5. Our investigations showed that the HK released from F4CPLNGs was a pH dependent. At pH 7.4, F4CPL showed the slowest release during the first 8 h of about 22.22mg% followed by slower release with about 34 mg% of total HK released for the remaining 24 h. At pH 5.5, the HK released fromF4CPLNGs showed the slowest release during the first 8 h of about 25.11mg% followed by slower release with about 35.04 mg% of total HK released for the 24h at when compared to either of their uncross-linked counterparts (F4UE) or (F4CE).
The formulation F4CPLshowed good criteria in all the accessed points. The formulation was physically stable for three months with no significant changes in their size and zeta potential.
As it showed particle size of 208 nm after three months of storage, compared to freshly prepared NGs (193.5 nm).
Furthermore, F4CPLNGs showed good in vitroserum stability without any aggregation observed in serum up to 6 h and accepted hemo-compatibility.
Biological screening to the fabricated HK-PMT-LF/CMC F4CPLNGs was also achieved.
In vitrocytotoxicity study which was performed on three different cell lines (MDA-MB-231, A549 and HepG2 cell lines), showedhigher cytotoxic effect, highest DRI and the ideal CI 0.17 in MDA-MB-231 cell line when compared to other tested cell lines (A549 and HepG2 cell lines) and the free drugs.
In vitrocellular uptake showed higher cellular uptake efficiency of F1CNGs byabout 1.7 folds compared to F1UNGs and to the free dye (Rhodamine) which proved the added benefit of using chemical cross linking after following the green approach in NGs synthesis