Precisely Tuned Acid-activatable Nanoparticles For Dual-mode Imaging-guided Synergetic Cancer Therapy

Cancer has become the leading cause of death worldwide.Traditional treatment to tumors includes surgery,chemotherapy,radiotherapy and so on.However,surgery means the possibility of recurrence,chemotherapy may cause overall side effect while radiotherapy is limited by accumulated dosage of radiation.Researchers ceased to find new treatments to tumors.Photodynamic therapy?PDT?is a noninvasive tumor therapy,which produce reactive oxygen species?ROS?to kill tumor cells through the interactions between laser with suitable wavelength,photosensitizer and oxygen.However,becausephotosensitizerhaspoorselectivitytowardstumors,photosensitizer could be activated by laser or light even when it didn't reach tumor site,resulting the damage to normal tissue.Porphyrins are used as photosensitizers for PDT because it has distinctive biological activity and affinity to cancer cells.Tetrakis?4-carboxyphenyl?porphyrin?TCPP?has short photodynamic period,needs longer wavelength of laser to be activated so that the laser could penetrate deeper tissues.It also generates more singlet oxygen and more selectively towards tumors.However,with the strong inter attraction between aromatic macrocyclic of porphyrins,the molecules could self aggregate,which leads to the low efficacy of photodynamic therapy.Especially in the acidic environment of tumor,the carboxyl groups of porphyrins could cause the aggregation more easily.On account of this,we put some amine with large stereospecific blockade onto the aromatic macrocyclic to reduce thep-pinteractions.With some amine,the pK_a of porphyrin could be altered,which means they could be dissolved in the acidic environment of tumors,promoting the photodynamic efficacy.The main work of the dissertation was concentrated on the synthesis of amine modified TCPP molecules with a series of pK_a and acid-triggered micelles.Then,GPLGLAG peptide and polyethyleneglycol?PEG?chain were attached to the molecule to form an amphiphilic polymer which could self-assemble to form nanoparticles with better biocompatibility.The multifunctional nanoparticles could be used as dual-mode imaging guided synergetic therapy micelles.Chapter 1.IntroductionIn this chapter,we firstly illustrated the meaning of micro-environment of tumor tissue especially the acidic milieu in tumor detection and treatment.Then,we introduced the mechanism,photodynamic therapy and the classification of photosensitizers.Then,the application of nanomaterials were introduced in the photodynamic therapy.At last,the aim and the meaning of our work were expressed.Chapter 2.Synthesis and Characterization of Acid-triggered TCPP moleculesIn this chapter,we used amine to modify the four carboxyl group of TCPP molecules and obtained different acid-responsive molecules with a series of pK_a.Because of its inter attraction between aromatic macrocyclic and four carboxyl groups,especially in tumor micro-environment,TCPP are prone to be aggregated in acidic environment,so the treatment efficacy may be decreased because of the aggregation in tumor milieu.So we used amine to take place of the four carboxyl group,then the molecules would give out strong fluorescence at acidic environment but aggregate at base environment.Different lengths of hydrophobia chains of amine could tune the pK_a of the molecules,so the molecules could be applied in the imaging of tumor environment or organelles.At last,we had molecules with a series of aimed pK_a:The pK_a of N,N-dimethylethylenediamine-TCPP is 7.5,the pK_a of N,N-diisopropyl-ethylenediamine-TCPP is 6.8,the pK_a of N,N-dibutylethylenediamine is 5.7,the pK_a of 2-diisopropylaminoethanol is4.9.The fluorescent responses are 8 times,7 times,11times and 50 times respectively.Their ROS generation towards different pH had the same pK_a as the fluorescent response towards pH.So it provided possibility of tumor fluorescent imaging and PDT.Chapter 3.Synthesis and Characterization of Acid and MMP-2-responsive NanoparticlesBesides acidic environment,matrix metalloproteinase-2?MMP-2?is also overexpressed in tumor tissue.In this chapter,we modified the TCPP molecules with the GPLGLAG modified polyethylene glycol?PEG?chain and amine to obtain amphiphilic polymers.The amphiphilic polymers could self assemble to be nanoparticles.We firstly connected the GPLGLAG peptide with hydrophilic PEG chain.Then the TCPP was synthesized onto the GPLGLAG to obtain amphiphilic polymers.They could self assemble to be nanoparticles with a diameter of 30 to 40nm or so by nanoprecipitation method.The modification of GPLGLAG could concentrate nanoparticles on the tumor cells where MMP-2 were overexpressed,which means the photosensitizer and the loading drug could be better concentrated.We chose N,N-diisopropylethylenediamine?DPA?and N,N-dibutylethylenediamine?DBA?to modify TCPP molecule,so that the nanoparticles would have the pK_a of 6.8and 5.7 respectively.We could see from the characterization of TEM and DLS that,when pH>pK_a,the nanoparticles were homogeneous spheres,when pH<pK_a,the nanoparticles were dissolved to be amorphous polymers.We also proved the nanoparticles could response to MMP-2.The pK_a value of two nanoparticles was within the pH of tumor environment and lysosome,so they could give out fluoresence in these positions.We could use them to detect tumors.Chapter 4.Dual-mode Imaging and Synergetic Therapy of the Acid-responsive nanoparticlesThis chapter mainly functionalize the acid-triggered nanoparticles,making it into a kind of nanomaterial which could achieve dual-mode imaging?fluorescent and magnetic resonance imaging?and synergetic therapy.Firstly,we could coordinate Mn²⁺into the aromatic macrocycle of TCPP,so that the magnetic signal could also be acid-responsive.As a result,we had successfully synthesized Mn-DBA-TGP nanoparticles whose magnetic response was over 3 times and fluorescent response was over 130 times.Thus,the nanoparticles could be used as magnetic resonance imaging?MRI?reagent.At last,we encapsulated some hydrophobia DOX into the core of the nanoparticles.We could calculate that the DOX loading efficiency was 7wt%by the standard curve of fluorescence.With such prepared nanoparticles,we did the cytotoxicity test in vitro,imaging experiment and pharmacodynamic test in vivo,proving the effective use of the nanoparticles to image tumors and treat cancer.