Studies On The Preparation And Antitumor Properties Of Hyaluronic Acid-Alendronate-Methotrexate Self-assembled Nanoparticles

Methotrexate is an anti-folate antitumor drug,which inhibits dihydrofolate reductase and blocks tumor cell DNA synthesis to achieve anti-tumor effect,and plays an important role in the treatment of various malignant tumors(especially bone tumors).However,the clinical use of MTX is limited because of its low selectivity and high toxic side effects.In order to increase the accumulation of methotrexate in tumor tissues and reduce the toxicity caused by non-specific uptake of nonnal tissues,this study designed a nano drug delivery system with hyaluronic acid(HA)loaded methotrexate(MTX)and alendronate sodium(ALN)for the treatment of bone tumors in combination with the current research focus of targeted drug delivery.The advantage of this nano drug delivery system is that HA is a receptor for CD44 which is highly expressed on the surface of tumor cells,and thus has the function of targeting tumor cells;ALN,as a bone-targeting ligand,has high affinity with hydroxyapatite,which is the main mineral component of bone,and can be selectively accumulated in bone tumors;In addition,drugs can form into self-assembled nanoparticles through intermolecular hydrophilic/hydrophobic interactions,due to their unique size effects,nanoparticles are be easily retained at the targeted site through high permeability and long retention(EPR)effect.This drug delivery system was designed to provide an idea for achieving targeted therapy of bone tumors by MTXIn this study,the hydrophilic macromolecular compound HA was firstly linked to the ALN under the action of a high-efficiency catalyst by an amide bond,which was then linked to the hydrophobic small molecule drug MTX via an ester bond.Thus formed the HA-ALN-MTX conjugate.HA-ALN-MTX nanoparticles were prepared according to the hydrophilic/hydrophobic effect of the drug.And their structure was verified by NMR and FT-IR The crystal structure was determined by XRD.The particle size was examined by DLS.The apparent morphology of nanaoparticles was studied by TEM and AFM,and the cytotoxicity effect in vitro was studied.The research content mainly includes1.Preparation of HA-ALN conjugate.Human normal cells(HUVEC)were cultured to investigate the cytocompatibility of the conjugate.The results conformed that,the HA was successfully coupled with ALN and its cell compatibility was good.This part is important for the following self-assembled nanoparticles preparation.2.Preparation of HA-MTX self-assembling nanoparticles.The-CH2OH of HA and the-COOH of MTX are covalently linked to form an ester bond to form a HA-MTX conjugate.Compared with free MTX,HA-MTX nanoparticles have enhanced proliferation inhibition effects on lung cancer cells(A549)and osteosarcoma cells(MG 63),and the toxic side effects on human umbilical vein endothelial cells(HUVEC)are reduced.3.Preparation of HA-ALN-MTX self-assembling nanoparticles.The HA-ALN and MTX were linked by an ester bond,and it confirmed that the HA-ALNwas successfully coupled with MTX.The prepared nanoparticles have a uniform nano-size distribution and good stability.In vitro cytotoxicity studies showed that HA-ALN-MTX nanoparticles had higher proliferation inhibition effects on tumor cells A549 and MG 63 than free MTX and HA-MTX nanoparticles,and were toxic to normal cells HUVEC.The side effects are lower.In summary,the HA-loaded MTX and ALN nano-drag delivery system designed by this study can be taken up by tumor cells,selectively inhibit the proliferation of tumor cells,and reduce the toxic side effects of normal cells,which is expected to improve the therapeutic effect of MTX.It is applied to targeted therapy of bone tumors.