Synthesis And Antitumor Activity Of Camptothecin Prodrugs

Many small molecular chemotherapy drugs,such as camptothecin,etc.,have drawbacks in poor water solubility,dissatisfactory biodistribution,leading to ungratified antitumor effects after administration,and serious side effects,thereby limiting their clinical use.Prodrugs and nanotechnology are important ways to improve the efficacy of hydrophobic drugs(such as camptothecin,etc.)and reduce side effects.Prodrugs,which are precursor molecules that can be transformed into parent drugs in the body,are widely used to solve the problems of hydrophobicity,stability,or pharmacokinetics of parent drugs.Therefore,this thesis synthesized y-glutamyl transpeptidase(GGT)-responsive CPT and 7-ethyl-10-hydroxy camptothecin(SN38)prodrugs and albumin-binding CPT prodrug,aiming at improve the water solubility and the anticancer efficacy of CPT or SN38.The first part of this thesis includes Chapters 2 and 3,exploring GGT-responsive CPT prodrugs and SN38 prodrugs.Based on that GGT is overexpression on the tumor blood vessels and the surface of the cell membrane and specifically recognizes and cleaves the structure containing ?-glutamyl bond,our group previously designed the GGT-responsive charge reversal polymer-CPT conjugate,PBEAGA-CPT,which showed strong tumor tissue penetration ability,and achieved excellent antitumor effects.We further found that its antitumor activity was affected by the drug loading content(DLC).Thus,in Chapter 2,we synthesized three conjugates PBEAGA18-CPT5(DLC.20%),PBEAGA18-CPT2(DLC,9%),and PBEAGA23(DLC,0%)with the different CPT contents using radical polymerization by adjusting the CPT-monomer feed ratio.We compared these conjugates in terms of charge reversal performance under GGT catalysis,cellular uptake,tumor penetration ability in tumor spherical,and anticancer efficacy in subcutaneous HepG2 tumors.We found that the conjugate with the highest hydrophobic conjugate,PBEAGA18-CPT5,possessed excellent ability of GGT-sensitivity,cellular uptake in GGT overexpression tumor cells,tumor spheroids penetration,along with enhanced tumor inhibition in subcutaneous HepG2 tumor model.However,the highest hydrophilic conjugate,PBEAGA23 was not GGT-responsive.Molecular dynamics simulations showed that PBEAGA18-CPT5 underwent a transition from a relatively open structure to a compact one when interacted with GGT,which facilitated the close contact of ?-carbon of the glutamate moiety to the side chain oxygen of Thr381.However,when the more hydrophilic PBEAGA18-CPT2 or PBEAGA23 interacted with GGT,the molecular chains were maintained flexible,so they could not get close to the GGT catalytic center.In addition,polymers with different phenyl methacrylate(PMA)contents were prepared by copolymerization of GGT-responsive monomers and phenyl methacrylate,and further proved that the hydrophobicity of the polymers affected the interaction between the polymers and the GGT enzyme and the charge reversal ability,which in turn affected the endocytosis of the polymer and tumor penetration.Inspired by this,in Chapter 3,we designed and synthesized a GGT-responsive y-glutamyl-modified SN38 prodrug.G-SN38 prodrug was stable in PBS solution and had a strong sensitivity to GGT in vitro to release SN38.So,G-SN38 was more cytotoxic to GGT-overexpression tumor cells than normal cells.In the mice model,G-SN38 showed better efficacy and safety than CPT-11 for 4T1 subcutaneous tumors and 4T1 lung metastases.GGT-responsive prodrug G-SN38 was sensitive to GGT,but like other small molecular drugs,its rapid clearance limits its efficacy.Covalently binding the drug to albumin can slow down the drug's clearance rate and prolong its blood circulation time,but the linkage needs to be designed to cleave the carrier in the tumor microenvironment.2-Acetophenone boronic acid(2-APBA)can form iminoboronate bond with the lysine residue ?-amino group of the protein,which can quickly dissociate at the tumor acidity or in the presence of GSH.Thus,in Chapter 4,CPT and CCM were functionalized with 2-APBA to prepare prodrugs(CPT-SS-APBA and CCM-APBA).The prodrugs efficiently and reversibly bound to proteins to obtain protein/prodrug conjugates,which further self-assembled into BSA/CPT-SS-APBA and BSA/CCM-APBA nanoparticles,having long-circulation times.In vivo,BSA/CPT-SS-APBA nanoparticles showed enhanced anticancer efficacy compared with clinically used irinotecan or sorafenib in subcutaneous 4T1 or HepG2 tumor models.