Dual turn-on fluorescence signal-based controlled release system for real-time monitoring of drug release dynamics in living cells and tumor tissues

Abstract

Controlled release systems with capabilities for direct and real-time monitoring of the release and dynamics of drugs in living systems are of great value for cancer chemotherapy. Herein, we describe a novel dual turn-on fluorescence signal-based controlled release system (CDox), in which the chemotherapy drug doxorubicin (Dox) and the fluorescent dye (CH) are conjugated by a hydrazone moiety, a pH-responsive cleavable linker. CDox itself shows nearly no fluorescence as the fluorescence of CH and Dox is essentially quenched by the C=N isomerization and N-N free rotation. However, when activated under acidic conditions, CDox could be hydrolyzed to afford Dox and CH, resulting in dual turn-on signals with emission peaks at 595 nm and 488 nm, respectively. Notably, CDox exhibits a desirable controlled release feature as the hydrolysis rate is limited by the steric hindrance effect from both the Dox and CH moieties. Cytotoxicity assays indicate that CDox shows much lower cytotoxicity relative to Dox, and displays higher cell inhibition rate to cancer than normal cells. With the aid of the dual turn-on fluorescence at different wavelengths, the drug release dynamics of CDox in living HepG2 and 4T-1 cells was monitored in double channels in a real-time fashion. Importantly, two-photon fluorescence imaging of CDox in living tumor tissues was also successfully performed by high-definition 3D imaging. We expect that the unique controlled release system illustrated herein could provide a powerful means to investigate modes of action of drugs, which is critical for development of much more robust and effective chemotherapy drugs.

A Facile, One-Pot, Surfactant-Free Nanoprecipitation Method for the Preparation of Nanogels from Polyglycerol–Drug Conjugates that Can Be Freely Assembled for Combination Therapy Applications

“…In fact, the quenching efficiency was even up to 86% (calculated with Equation (1)). This is in the range of similar systems reported in the literature [ 43 ]. Quenching of DOX fluorescence has proven useful for monitoring pH-labile bond cleavage in real time in cells.…”

“…The observation that DOX-EMCH can be quenched by itself has been reported in the literature and appears to be due to C=N isomerization and N–N free rotation of the hydrazone linker. When activated under acidic conditions, hydrolyzation of the hydrazone bond afforded free DOX and therefore an increase of the fluorescence [ 43 ]. To confirm our interpretation of the fluorescence spectra as quenching and release, we synthesized a NG built from a PG-DOX conjugate which was not cleavable.…”

Recent advances in construction of small molecule-based fluorophore-drug conjugates

“…Hydrazone is a well-known linkage capable of creating acid-labile response. It is stable at physiological pH 7.4 but is slowly hydrolyzed in the acidic tumor environment and degrades much more quickly after entering cells due to exposure to late endosomes and lysosomes (pH 6.5–5.5) [ 61 , 62 ]. Vendrell and co-workers [ 62 ] developed an acid-sensitive conjugate 11 ( Fig.…”

Recent advances in drug release monitoring

“…DT-diaphorase, an enzyme overexpressed in some malignant tumors, can be exploited to catalyze the reduction of quinine propionic acid moiety incorporated into prodrug, which could be utilized for concomitant drug release imaging and cancer chemotherapy [43,44]. In order to monitor drug release more accurately and understand pharmacological mechanisms better, Kong et al [45] developed a dual turn-on fluorescence based controlled release system (CDox), in which doxorubicin (Dox) and fluorescent dye (CH) are conjugated by a pH-responsive cleavable hydrazone linker. The C=N isomerization and N-N free rotation efficiently quenched the fluorescence of CH and Dox in the CDox complex.…”