Design And Synthesis Of Novel Near Infra-red Fluorescent Small Molecular Probes For Tumor Theranostic Application

Background:It is well known that tumor is still one of the most important diseases threatening human life and health.In recent years,with the increase of tumor incidence and mortality year by year,it is urgent to develop new materials and technologies for tumor diagnosis and treatment.Actually,optical probes can provide new possibilities for early diagnosis and targeted therapy of malignant tumors because of high sensitivity,high selectivity,easy operation and real-time monitoring.Compared with visible light,the near-infrared fluorescent probe has deeper tissue penetration and lower spontaneous fluorescence interference.And combining the physiological differences between tumor microenvironment and normal tissue,the development of effective and safe tumor-targeted near-infrared fluorescent diagnostic and therapeutic agents is expected to improve the efficacy of in vivo tumor diagnosis and treatment.With the development of optical physics,molecular biology and computational analysis,the integration of optical diagnosis and therapeutic technology will move towards high sensitivity and spatial-temporal resolution.It will promote the integration of cancer diagnosis and treatment,and eventually change the current status of cancer theranostics.Objective:This paper was focused on the critical scientific problem of accurate diagnosis and efficient treatment of malignant tumors,and construction of a serious of novel small molecule fluorescent probes realizing the specific,sensitive and non-invasive detection of tumor-associated enzymes or tumor tissues.Moreover,the emission wavelength of the probe is advanced from near-infrared I region to near-infrared II region to achieve the sensitive imaging from superficial tumor to deep tumor,and the leap from single modal to multimodal imaging as well as tumor theranostics.Methods:In the first part,we report a new ALP specific NIR fluorescent probe QMTP with AIE characteristics by conjugating a hydrophobic AIE fluorophore quinolone-malononitrile to a hydrophilic phosphate-modified phenol moiety.When QMTP enters the tumor tissue and is specifically dephosphorylated by the ALP due to overexpressed ALP in most tumor cells,it will activate the fluorescence signal of the probe and produce strong near-infrared emission,thus realizing the highly sensitive detection and imaging of ALP activity in vitro and in vivo.In the second part,we report an activatable NIR fluorescence/ratiometric PA bimodal probe Indol-Glu by incorporating the NIR hemicyanine dye to y-glutamyl group which can be specifically recognized and hydrolyzed by GGT for in vivo dual-modal imaging and image-guided surgery of tumors.Indol-Glu will release strong near-infrared fluorescence remarkably and light the tumor tissue selectively due to specific cleavage by GGT within tumor region.Meanwhile,hydrophobic hemiocyanine residues of Indol-Glu tend to self-assemble into uniform nanoparticles resulting in prolonged retention and amplified imaging signals,which facilitates the image-guided surgical resection.Because the specific cleavage of probes by GGT shifted the absorbance of the probe to the NIR region,we can accurately and noninvasively assess the differences of in the expressed level of GGT enzymes in different tumor models by using enzyme-activated NIR fluorescence/ratiometric PA properties,which provides an effective means for visualizing the invasion and metastasis of malignant tumors.In the third part,we report a novel second near-infrared(NIR-?)fluorescent probe QT-RGD constructed with a NIR-? emissive organic fluorophore and two cyclic-(arginine-glycine-aspartic)(cRGD)peptides for in vivo multimodal NIR-?/PA/SPECT imaging and PTT of tumors.The cRGD can specifically bind to the tumor biomarker ?v?3 integrin receptor and effectively promote the accumulation of the probe at the tumor site.Therefore,when the probe targets the tumor site,taking advantage of the strong absorption and emission characteristics of the probe in the NIR-? rerion as well as the radioisotope 125I labeling,we can readily carry out the multimodal imaging and the theranostics of deep tumors in vivo.Results:In the first part,the probe QMTP showed specific and sensitive response to ALP enzymes in vitro.Taking advantage of the activated near-infrared fluorescence signal,the probes enable to specific detection and imaging of ALP activity both in vitro and in vivo.Because the emission wavelength of the probe was significantly red-shifted to the near-infrared region together with AIE characteristics,the probe had deeper tissue penetration and lower fluorescent background.By establishing the drug-induced liver injury model in zebrafish,QMTP was demonstrated to be able to sensitively image the liver injury in zebrafish.Meanwhile,direct serum fluorescence detection further demonstrated that the probe could distinguish the mice bearing tumor from normal mice.In the second part,the probe Indol-Glu can specifically respond to GGT enzymes both in vitro and in vivo with the activated NIR fluorescence and ratiometric PA signals allowing for non-invasive and quantitative monitoring of GGT levels,which can be used to assess the differences of enzyme expression levels in different tumor models.Due to the enzyme-activated NIR fluorescence and enzyme-mediated self-assembly functions resulting in long retention and amplified fluorescence signals in tumor tissues,Indol-Glu has been successfully applied for precise localization of peritoneal tumors in mice and fluorescence-guided tumor surgical resection.In the thrid part,the hydrophilic polypeptide cRGD of the probe QT-RGD can not only improve the water solubility of the probe,but also improve the tumor targeting ability through ?v?3 integrin-mediated endocytosis.The pharmacokinetic results showed that the elimination half-life t1/2? of QT-RGD was 2.41 h,which is favorable to the fast clearance through renal/hepatobiliary excretion from body.Notably,the 125I-labeled QT-RGD exhibited strong absorption and emission in NIR-? region,which makes it highly useful for multimodal NIR-?/PA/SPECT imaging and imaging-guided PTT applications.Conclusion:In this paper,by using tumor-associated protease and membrane receptor protein as targets in tumor microenvironment,we designed and synthesized two kinds of activated near-infrared ? region small-molecule fluorescent probes QMTP and Indol-Glu,and a near-infrared ? region small-molecule fluorescent probe QT-RGD with diagnostic and therapeutic integration function.We have achieved specific,sensitive and non-invasive detection and imaging of tumor-associated enzymes or tumors,and we also established an advanced strategy of multi-modal imaging and theranostics of tumors in vivo.