Design Of Tertiary-amine-oxide-based Active Transcytosis Nanomedicines For Cancer Therapy

Liposomes and dendrimers have been widely explored for anticancer drug delivery.However,their clinical therapeutic outcomes are unsatisfactory due to the insufficient tumor accumulation and penetration efficiencies.In our previous studies,the active transcytosis nanomedicines were found to effectively target and penetrate throughout the solid tumors,exhibiting potent therapeutic efficacies in preclinical tumor models.To this end,this thesis proposed the strategy to fabricate efficient active transcytosis liposomes and dendrimers via TAO modification.The obtained TAO-containing nanocarriers greatly enhanced the accumulation and penetration of the payloads in tumor tissues,exhibiting potent antitumor efficacies in preclinical tumor models.1.In the first part of this thesis,we developed surface functionalized TAO liposomes(TAOLs)capable of effective active transcytosis(chapter 2).Sixteen TAO lipids were first synthesized through combinatorial chemistry involving four fatty acids and four tertiary amine oxides.A library of 96 TAOLs was prepared using these TAO lipids and loaded with a chemotherapeutic nanodrug(SC).The SC-loaded TAOLs(TAOLSCs)with optimal particle size and stability were obtained at a mass ratio of N,N-dimethyl(ethyl)-amino-N-oxide-containing lipids,cholesterol and TPGS at 4:1:1.The active transporting efficiency of TAOLSCs was measured by transwell models,whereas palmitoyl-or stearyl-based ones exhibited the highest trans-endothelial transport rates.Finally,the stearyl-based and N,N-dimethyl(ethyl)-amino-N-oxidecontaining liposomes(ODM(E)LSC)were chosen for further evaluation due to the high overall yield,optimal size,high stability,and excellent active transporting capability.2.In the second part of this thesis,we elucidated the mechanism of ODM(E)LSCinduced transcytosis,and evaluated their drug delivery efficiencies as well as the antitumor activities(chapter 3).The internalization behaviors,subcellular distributions,endocytosis behaviors,intercellular transports and in vitro tumor penetration behaviors of ODM(E)LSC were evaluated,whereas ODM(E)LSC were found to transport transcellularly,penetrate deep into multicellular tumor spheroids(MTSs)and induce potent cytotoxicity via the micropinocytosis-associated rapid cellular uptake and endoplasmic reticulum(ER)/Golgi-involved exocytosis.In vivo pharmacokinetic,biodistribution and tumor penetration studies showed that ODM(E)LSC could undergo long blood circulation via RBC-hitchhiking,effectively accumulate in tumor sites,rapidly extravasate from tumor blood vessels and penetrate deep into tumor tissues via transcytosis.These effects enabled ODM(E)LSC to deliver chemotherapeutic drugs to the whole tumor tissue,reducing the growth of hepatocellular carcinoma HepG2 tumors and exhibiting superior antitumor activity compared to traditional liposomal medicine.3.In the third part of this thesis,we synthesized a TAO-modified dendrimer conjugated with triptolide(TPL)(ODMP-T)because TPL was unable to be loaded into the above liposomes.Then,we investigated its capacities in triggering transcytosismediated transcellular transport,boosting ER stress(ERS)-associated apoptosis of tumor cells,optimizing drug delivery process,enhancing antitumor efficacy and reducing systemic side effect(chapter 4).The ODMP-T was synthesized by covalent conjugation of TPL to polyamidiamine dendrimer through the hydrolytic ester bond,followed by surface modification with TAO moieties.In vitro studies demonstrated that ODMP-T could effectively transport across tumor vascular endothelial cells,transport intercellularly between pancreatic cancer cells,and infiltrate into MTSs via transcytosis.Besides,the ER-targeting ODMP-T greatly promoted TPL-induced unfolding protein stress response and TPL-mediated GRP78 downregulation,resulting in lethal ERS in pancreatic cancer cell lines as well as MTSs.In vivo studies revealed that ODMP-T could effectively accumulate and infiltrate throughout the whole tumor tissue.As a result,ODMP-T could not only reduce the growth of BxPC3 and patient-derived subcutaneous xenografted tumors,but also prevent the progression of BxPC3 orthotopic xenografted tumors,exhibiting superior tumor inhibition rate than TPL or ethylene glycol methyl-modified contrast.Meanwhile,ODMP-T substantially avoided the TPL-related hepatorenal toxicity,showing reliable biosafety.