الفهرس 
IntroductionOnly 14 pages are availabe for public view
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Abstract
Skin disorders in the human population vary from cancer to non-cancerous diseases.
Non-cancerous skin disorders include a broad spectrum ofinfectious, inflammatory, and autoimmune diseases.
Skin cancer is more common in people with immunosuppressive diseases or with diseases managed by chronic immunosuppressive therapy.
Skin cancers are by far the most common malignancy among humans.
Theincidence of skin cancer increases essentially with age, apparently reflecting the long latency between carcinogen exposure and cancer establishment.
The three common forms of skin cancer are squamous cell carcinomas (SCC) and basal cell carcinomas (BCC), (also known as nonmelanocytic skin cancer, NMSC) and cutaneous malignant melanoma(MM)).
Skin squamous cell carcinoma is a malignancy arising from epithelial keratinocyte in which the causative factors are correlated with ultraviolet radiation, chemicals exposure, immunosuppressive drug application, trauma, scar and some precancerous diseases.
Nanoparticles (NPs) exhibit interesting features such as small sizes, large surface-to -volume ratios, self-assembly capability and biocompatibility.
Consequently, theyhave the potential to increase the selectivity and efficacy of chemical, physical and biological approaches to cancer cell death while reducing non-malignant cell toxicity.
Nanoparticles can deliver higher concentrations of drugs to target areas in the skin and provide targeted treatments when modified with cell-specific ligands.
Drug loaded charged NPs also accumulate in hair follicles and thus promote the penetration of drug molecules through the shallow layers of the stratum corneum.
Natural polymers have outstanding properties for a wide variety of clinical applications, including biodegradability, non-antigenicity, durability, high nutritional value, plentiful renewable sources, strong tailoring capacity and remarkable binding capacity of different drugs.
Lactoferrin (Lf) is an iron-binding non-hem glycoprotein present in the transferrin family of proteins.
One of the characteristics of Lf structure is its positive charge contained on the surface.
This enables Lf to bind on the negatively charged surface of different immune cells which is thought to cause various pathways leading to cellular 95responses such as activation, differentiation, and proliferation.
Hyaluronic acid (HA) or Hyaluronan is a natural linear polysaccharide formed by repeating units of D-glucoronic acid and N-acetyl-D-glucosamine disaccharide, this bio macromolecule is one of the major constituents of the skin and can be found in extracellular tissues of various parts of the body.
Hydrogels made from HA have the potential to degrade intosafe products and have been commonly used in various biomedical applications, such as tissue regeneration, drug delivery, gene therapy, diagnostics, etc.
Some hydrogels regulate the release of the drugs in response to stimuli of the environment such as pH, temperature and enzyme.
In this study, we propose for the first time up to our knowledge, the co-delivery of two herbal drugs, which are resveratrol (RSV) & honokiol (HK) through the conjugation using a linker to form a polymeric nanoconjugate.
There was a synergistic effect of the formulated drug nanoconjugate that effectively boost skin permeation with a homogenous distribution through all skin layers especially to squamous layer.
Lactoferrin-succ-resveratrol conjugate (Lf-succ-RSV) was firstly prepared, followed by preparation of honokiol-lactoferrin-resveratrol conjugate (HK-succ-Lf-succ-RSV) and incorporatingthe conjugate in hydrogel.
The two hydrophobic drugs are conjugated vialinker (succinic anhydride)throughester bonds.
The viscosity and spread ability of HA/HK-succ-Lf-succ-RSV hydrogelsat two different concentration of HA were determined to obtain the most applicable hydrogel. Different physicochemical characterizations were done.Lf-succ-RSV and HK-succ-Lf-succ-RSVconjugates exhibited particle size of 163.5±12.4 nmand188.3 ± 12.3 nmrespectively.
Zeta potentialwas also measured forLf-succ-RSVand HK-succ-Lf-succ-RSVto ensure the physical stability of the formulations upon storage for three months.
The results indicated thatthe prepared nanoconjugates were physically stable as manifested by their particle size and zeta potential.
< No significant changes in these parameters were observedbefore and after storage.
<Fourier Transform Infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC) were performed on Lf, RSV, HK, HK-succ-Lf-succ-RSV for the characterization of possible interactions between RSV and HK to the Lf through the linker by formation the ester bond.
In addition, Nuclear Magnetic Resonance (1H-NMR) was done to confirm the formation of HK-succ-Lf-succ-RSV nanoconjugate, 96whereas mass spectrometry (MALDI TOF-MS) was done to calculate the exact mass of Lf and Lf-succ-RSV nanoconjugate.
<HPLC method was developedand validated to quantify RSV and HK in HK-succ-Lf-succ-RSV.
The RSV and HK showed conjugation efficiency of52%and 32.5%respectively.
<n vitro release of RSV and HK from HK-succ-Lf-succ-RSV nanoconjugate and hydrogel was performed.
<Release of drugs was tested in both PBS (pH7.4)at physiological condition and (pH 5.5)to mimic acidic skin condition.
< In-vitrorelease profiles of free RSV and free HK were recorded and compared to their release profiles from the HK-succ-Lf-succ-RSV nanoconjugate and hydrogel.
Reduced and sustained release profiles of the drugs from the nanoconjugate were observed incomparison to the free drugs as the cleavage of the ester bond needed esterase enzymes that are found in sufficient amount in the skin.Additionally,In vitrocytotoxicity assay was performed on A431 cell line.
It showed synergistic action between RSV and HK.
< It also provided a proof that the nanoconjugate improved the free drugs combination potency demonstrating an IC50value with 8-foldreduction in comparison to the IC50of the free combined drug solution.
<In-vitro cellular uptake study and flow cytometry study were done on A431 cell line.
<Such experiments demonstrated the efficient intracellular delivery of the nanoconjugate.
<Moreover,the permeation and retention offree RSV, free HK, HK-succ-Lf-succ-RSV nanoconjugate, HA/HK-succ-Lf-succ-RSV hydrogelacross human skin was investigatedto determine the percentage of drugs permeated and retained in stratum corneum, epidermis and dermis.
Additionally, the dermal distribution offluorescently labelled Lfaloneand HK-succ-Lf-succ-RSVnanoconjugate within the human skin was visualizedby CLSM.
<Finally, it can be concluded that HK-succ-Lf-succ-RSV nanoconjugate showedoptimum size, narrow PDI, appropriate zeta potential, high and excellent conjugation efficiency of RSV and HK with a pH dependent release and the final hydrogel that incorporated the nanoconjugate had promising viscosity and spreadability when topically applied on the skin.
< The dual loaded nanoconjugate embedded in biomimetic hyaluronate gel offersa promising approach for the local management of cutaneous cancer via natural based ingredints.