Construction Of Novel Choline Phosphate Liposomes And Their Use As Nanocarriers In Tumor Therapy

Liposomes play an important role as a nanocarrier in tumor therapy.Due to their biological composition and structure mimicking cell membranes,they possess good biocompatibility and can simultaneously deliver hydrophilic and hydrophobic drugs,making them widely used in areas such as tumor imaging,chemical therapy,gene therapy,and immunotherapy.Currently,more than a dozen liposomal drugs including paclitaxel and doxorubicin have been approved by the FDA for clinical treatment.Doxorubicin HCl liposome injection produced in China also obtained clinical permission in 2019.However,the traditional phosphatidylcholine(PC)liposomes cannot be modified structurally,which seriously hinders their development as nanomedicine carriers in personalized therapy.Generally,the functionalization can be achieved by incorporating phosphatidylethanolamine(PE)with biologically orthogonal functional groups.But the biocompatibility decreases due to the loss of zwitterionic structure of PE after modification.In addition,PC derivatives,such as sulfo betaine phosphatidylcholine(SB)and carboxyl betaine phosphatidylcholine(CB),which have similar zwitterionic characteristics,have also been applied in nano medicine and tissue engineering.Due to the rigidness of the carbon and sulfur electronic structures,SB and CB lack further development in terms of biologically orthogonal functionalization.Therefore,it is urgent to design a new zwitterionic lipid to solve this dilemma.In this paper,we have designed a novel choline phosphate(CP)lipid based on the charge orientation heterogeneity of the PC head group,and the CP liposomes(CP-lipo)prepared from it can not only retain the good biocompatibility and drug delivery capability of traditional liposomes,but also utilize biologically orthogonal functionalization to endow it with various functions,thereby improving the in vivo transport efficiency and tumor treatment effect.The specific results are as follows:(1)We have designed and synthesized a novel zwitterionic CP lipid(CP-lip),with charge oriented translocation at the PC head group,which overcomes the limitations of traditional zwitterionic lipids.Due to its opposite charge orientation to PC,CP-lip can maintain the natural zwitterionic characteristics,including good bioprotection and biocompatibility,and promote cellular uptake through supramolecular interactions with cell membranes.Based on the modifiability of CP-lip,we introduce folic acid to prepare CP liposomes with active targeting function(tCP-lipo),and demonstrate the feasibility of using them as drug carriers.(2)During the early exploration of CP-lip physical and chemical properties,we discovered that the melting temperature(Tm)of CP-lip was around 43 □.Thus,we designed a novel photosensitive Lip-cRGDfk/ICG/Dox formulation for synergistic therapy of breast cancer.This formulation simultaneously loads the chemotherapeutic drug Dox and the photothermal conversion agent Indocyanine Green(ICG)into choline phospholipid liposomes.Upon irradiation with 808 nm near-infrared laser,ICG is activated,causing an increase in temperature that induces thermal ablation of cancer cells.Furthermore,the molecular conformation of the lipids changes when the temperature rises to Tm,leading to the breakdown of the liposome structure and triggering the release of Dox to attack cancer cells.In vitro experiments showed that the Lip-cRGDfk/ICG/Dox formulation exhibited good breast cancer cell targeting and spatiotemporal control of photodynamic attack against cancer cells.In the mouse tumor suppression experiment,the tumor almost disappeared after an 18-day treatment cycle.It can serve as an effective platform for non-invasive and spatiotemporally controlled activation of tumor cell toxicity in precise breast cancer treatment.(3)At present,the transfection efficiency of cationic liposomes and Hyperbranched polyethylene iMine(hPEI)containing genes is high,but their biological toxicity is relatively large.We constructed a highly stable and shielded interlocking protective system CP-lipo/(PEI/DNA),using CP-lip,hPEI,and DNA.It not only retains the high transfection efficiency of traditional hPEI,but also greatly reduces the overall toxicity.Based on a series of in vitro and in vivo experiments,this protective system can safely deliver gene to the tumor site after circulation in the blood and kill cancer cells through loaded gene transfection,effectively suppressing tumor growth.The interlocking protective system built by CP-lip and PEI provides an efficient and safe platform for targeted gene delivery in vivo.