Synthesis, Nanosizing And In Vitro Drug Release Of Two Different Anti-HIV Polymeric Prodrug Between Chitosan And Nucleoside

A novel approach to improve the antiviral efficacy of nucleoside reverse transcriptase inhibitors (NRTIs) and reduce their side effects was developed by constructing a nanosized NRTI monophosphate-polymer conjugate using d4T as a model NRTI. Firstly, a novel chitosan-O-isopropyl-5'-O-d4T monophosphate conjugate with a phosphoramidate linkage was efficiently synthesized through Atherton-Todd reaction under mild conditions and analyzed by NMR. The anti-HIV activity and cytotoxicity of the polymeric conjugate were evaluated in MT4 cell line. Then the conjugate nanoparticles were prepared by the process of ionotropic gelation between TPP and chitosan-d4T conjugate to improve their delivery to viral reservoirs, and their physicochemical properties were characterized by transmission electron microscopy (TEM), dynamic light scattering (DLS) techniques and X-ray diffraction (XRD). In vitro drug release studies in pH 1.1 and pH 7.4 suggested that both chitosan-d4T conjugate and its nanoparticles prefer to release d4T 5'-(O-isopropyl) monophosphate than free d4T for prolonged periods, which resulted in the enhancement of anti-HIV selectivity of the polymeric conjugate relative to free d4T due to bypassing the metabolic bottleneck of monophosphorylation. Additionally, the crosslinked conjugate nanoparticles can prevent the coupled drug from leaking out of the nanoparticles before entering the target viral reservoirs and provide a mild sustained release of d4T 5'-(O-isopropyl) monophosphate without the burst release.For comparison, the conventional chemical conjugation approaches have been accomplished to form the polymer-drug conjugates, the formation of amide linkages between drug and chitosan by direct coupling reactions using carbodiimides as zero lengths cross-linkers to form amide linkage between chitosan and 5'-O-succinylstavudine. Further more, the chitosan-5'-O-succinylstavudine conjugate nanoparticles were prepared and characterized. In vitro anti-HIV activity study showed chitosan-5'-O-succinylstavudine conjugate has a much lower HIV inhibition activity than chitosan-O-isopropyl-5'-O-d4T monophosphate conjugate. The results suggested that the novel kind of chitosan-O-isopropyl-5'-O-d4T monophosphate conjugate nano-prodrugs may be used as a targeting and sustained polymeric prodrugs for improving therapy efficacy and reducing side effects in antiretroviral treatment.In the end, aim to enhance the solubility in water, we regioselectivly synthesize 3,6-O-disulfated chitosan-O-isopropyl-5'-O-d4T monophosphate conjugate. The obtained 3,6-O-sulfated chitosan-d4T conjugate manifested an increased solubility in aqueous phase and considerable anti-HIV activity.