Strategies For Highly Efficient Cancer Therapy And Their Applications Based On Multifunctional Photodynamic Nanomaterials

Cancer is one of the dominant causes for human deaths.The improvement of therapeutic efficacy and inhibition of tumor metastasis and recurrence have remained as the main challenges in clinical cancer therapy.With the development of nanotechnology and precision medicine,explore strategies for efficient cancer therapy and develop safe and efficient nanomedicine with precise targeting are desperately desired.The therapeutic outcome of nanomedicine depends on the efficiency of the biological hierarchical delivery and the toxicity to active sites.The low efficiency of drug payload,intratumor accumulation,tumor penetration,cellular internalization,subcellular targeting and drug resistance have severely limited the therapeutic efficiency of nanomedicines.Even only one limitation exists,the treatment of nanomedicine would be ineffective or noneffective.Cancer treatment by photodynamic therapeutics have been studied in our group for many years and possesses several advantages:1)instantly photosensitive response for precise controlled release;2)the controllability of generation and concentration of cytotoxic ROS;3)accurate subcellular targeting;4)surface modification with excellent biocompatibility.Thus,we utilize PDT as therapeutic model,designed multi-functional UCNs-PSs nanomaterials and proposed different strategies for highly efficient therapy to overcome the above bottlenecks.The main works are as follow:(1)Synergetic targeting strategy.UCNs-GOQD nanohybrids were prepared with triggering new photosensitizer GOQD to release ROS by the emissions of UCNs after NIR absorption.Considering the short action radius of cytotoxic ROS(<150 nm)and that mitochondrion is the efficient target of PDT,we functionalized the nanohybrids with tumor/cell targeting agent folic acid(FA)and mitochondrial targeting agent triphenylphosphine(TPP)to evaluate the synergetic efficacy in intratumor accumulation,cellular internalization and mitochondrial localization.In the in-vitro experiments,compared to the nanoparticles modified with single target agent FA or TPP,the intracellular uptake of FA/TPP modified nanoparticles improved 20%and 40%respectively and the mitochondrial targeting efficiency improved 46%and 50%respectively,thus showed enhanced damage to mitochondria and Hela cells.In the in-vivo therapeutic model,the intratumor accumulation of nanoparticles with FA/TPP modification improved 40%compared to modification of FA and 75%compared to modification of TPP;meanwhile the mitochondrial localization of FA/TPP functionalized nanomaterials improved 16%and 10%respectively,thus showing more significant inhibition to the tumor growth.These results indicated that the intratumor accumulation,intracellular internalization,subcellular localization and tumor therapeutic outcome of nanomedicines could be enhanced by synergetic targeting.(2)Cell delivery strategy.UCNs@Pp IX nano-conjugations were synthesized with modification of biocompatible oli GOQDmer poly-L-lysine(PLL)and tumor/cell targeting agent FA to study the interaction between tumor cells and the tumor penetration effects.In the in-vitro experiments,PLL/FA functionalized UCNs@Pp IX nanoparticles could be efficiently delivered between different cell layers in transwell system and the intracellular uptake was 1.51 times of3-aminopropyltriethoxysilane(APTS)modified UCNs while the exocytosis does was 1.8 times;by labeling of Rab proteins,the colocalization of PLL/FA modified nanoparticle to recycling proteins Rab11 could reach 55%while the colocalization of APTS modified UCNs was only 32%;indicating the efficient transcytosis-mediated cell delivery effects of PLL/FA functionalized nanoparticles between Hela cells.In the in-vivo tumor models,the significantly enhanced tumor penetration of PLL/FA modified nanoparticles were observed and the cellular internalization dose in tumor tissues reached 62.8%,which was2.6 times to APTS modified one,thus the tumor was eradicated.When administrated with inhibitors,the endocytosis and exocytosis of PLL/FA modified nanoparticles were significantly inhibited and the colocalization of nanoparticles to recycling proteins Rab11 was decreased,resulting in the inhibited tumor penetration,decreased cellular internalization,tumor residual and tumor recurrence.By analyzing the results of tumor penetration and intracellular endocytosis-exocytosis dose,the positive dependence was found,indicating the tumor penetration and therapeutic outcome could be enhanced by cell delivery of nanomedicine in tumor tissues.(3)Drug resistance circumvention strategy.The UCNs@Pp IX nanoparticles were used to study the therapeutic effects on heterogenous and drug resistant glioblastoma cells and the corresponding tumor models.In cell experiments,the tolerance of glioblastoma cells LN229,U87,T98G to the clinical first-line medicine temozolomide(TMZ)were strongly different and the 50%inhibition concentration(IC₅₀)were 194?M,485?M and 1442?M respectively.Meanwhile,with the long time incubation of TMZ under a low concentration,the resistant cell strains LN229/TMZ and U87/TMZ were developed with IC₅₀improving to 606?M and 980?M respectively.When being treated by PDT nanoparticles,all of the cells could be efficiently killed with the comparable IC₅₀at 70-90 ug/ml.In the in-vivo treatment models,the therapeutic outcome of TMZ to different tumor models were significantly different while these tumors were efficiently inhibited and eradicated by PDT nano-conjugations.In the study of gene regulation by Q-PCR analysis,multiple drug resistance pathways were activated by TMZ treatment while except for the down-regulation of antiapoptotic proteins and the up-regulation of the apoptotic proteins,no drug resistant pathway was significantly regulated by nano-PDT treatment.It indicated that nano-PDT treatment could initiate cell apoptotic cascade via its specific toxicity mechanism and circumvent the activation of drug resistance pathway,thus achieve the highly efficient cancer therapy.Based on the above results,we designed and prepared the multi-functionalized nanomedicine UCNs@Pp IX-PLL-FA.The photosensitizers Pp IX were conjugated to PLL with the improved drug loading efficacy as25wt%-43wt%,which was far higher than the reported 14 wt%.In the in-vivo experiments,the PDT nano-conjugations could efficiently accumulate to tumor,penetrated into deep tumor tissues and efficiently internalized into tumor cells.Additionally,by anti-CD133 labeling,the PDT nano-conjugations could efficiently kill cancer stem-like Hela cells and non-stem-like Hela cells.With the integration of the above three strategies and the high loading efficiency,the PDT nano-conjugations successfully achieved eradication of Hela tumor models without tumor recurrence in 40 days under the injection dose of 5.6 mg/kg(i.p.),which is very low and safe compared to the reported the injection dose(>20mg/kg)and the safe injection dose of UCNs(<10 mg/kg),providing a potential possibility of application and clinical translation of the safe and efficient photodynamic nanomedicines.