Synthesis,Biological Activity And Imaging Application Of Quinoxaline/phenothiazine Oxazine Conjugated Compounds

As the number of cancer patients continues to increase,cancer diagnosis and treatment have received more and more attention.Currently,cancer treatment includes traditional treatment methods,such as surgical resection and chemotherapy,as well as new treatment methods,such as immunotherapy and phototherapy.However,different cancer treatment methods have their own advantages and disadvantages.Therefore,a combination therapy model that organically combines multiple cancer treatment methods has been developed.Through this combined treatment model,different treatment methods can complement each other and improve efficacy.Among them,the combination of chemotherapy and light therapy is one of the most popular combination therapies in recent years.In addition,on the basis of combination therapy,multifunctional cancer diagnosis and treatment reagents have been developed,which can understand the lesion range and drug distribution through imaging,and then realize precise treatment of image-guided therapy.Among these diagnostic and therapeutic reagents,those with fluorescence-photoacoustic dual-mode imaging capabilities can improve the temporal and spatial resolution and accuracy of tumor imaging,which has become a hot spot in current research.At present,this clinical treatment reagent is mainly through the application of nanomaterials,which are composed of imaging reagents,therapeutic drugs and other components.However,they have many disadvantages,such as complex composition,unclear metabolic mechanism,high liver and kidney toxicity and so on.In response to the above problems,in this thesis,Oxazine 1,a symmetric oxazine dyes with anti-cancer potential,are used as the basis to develop a series of new multifunctional organic small molecule compounds.At the same time,the performance and"structure-activity"relationship of these multifunctional organic small molecule compounds have been studied in detail,and some of them have been used in the coordinated treatment of chemotherapy and phototherapy under the guidance of fluorescence-photoacoustic dual-mode imaging.Based on the above,the following three tasks are mainly carried out:（1）Oxazine 1 has potential chemotherapy effect.However,their absorption wavelength is only about 650 nm,which limits their application prospects in phototherapy.In the previous work of our research group,the quinoxaline-oxazine conjugated compound DQF-683 with an absorption wavelength of about 680 nm has been synthesized by introducing quinoxaline into the oxazine structure.In this chapter,DQF-683 was studied in detail,including its optical,photothermal conversion properties and biological activity.The experimental results show that,under the irradiation of 660 nm laser,the photothermal conversion efficiency of DQF-683（about 38.9%）is significantly improved than that of Oxazine 1（about 30.5%）.In addition,the cell viability test results showed that DQF-683(IC₅₀ is 1.051±0.0253μM)has a higher chemotherapeutic effect than the positive control doxorubicin(IC₅₀ is 10.44±0.9405μM).At the same time,further research results show that DQF-683 can enter the mitochondria and endoplasmic reticulum,thereby inhibiting the proliferation and migration of tumor cells.In this chapter,DQF-683 was used in in vivo treatment experiments,and realized fluorescence-photoacoustic dual-mode imaging-guided tumor treatment,effectively inhibiting tumor growth through the synergistic effect of photothermal therapy and chemotherapy.（2）In order to further improve the imaging performance of DQF-683 and enhance its therapeutic effect,the thiophene-oxazine conjugated compound NCN2 was synthesized through the quinoxaline moiety in DQF-683 being substituted by thiophene.Compared with DQF-683,the maximum absorption wavelength of NCN2 is red-shifted by about 20 nm,and it has a strong absorption at 808 nm(λ_ex=705 nm,λ_em=840 nm).Under the irradiation of808 nm laser,compared with DQF-683,the light energy absorbed by NCN2 is further released through non-radiative transition,and the photothermal conversion efficiency of it is about 44.8%.The cell viability test results show that NCN2 still has superior chemotherapy effect(IC₅₀ is 6.518±0.3079μM),and it is higher than the positive control group doxorubicin(IC₅₀ is 10.44±0.9405μM).Furthermore,in vitro optical performance test results show that NCN2 also has a good fluorescence-photoacoustic dual mode Imaging potential.In summary,NCN2 is a multifunctional organic molecule with the potential of fluorescence-photoacoustic dual-mode imaging and chemotherapy-photothermal combined treatment,and it has potential biological application prospects.（3）In view of the lack of tumor selectivity in traditional chemotherapy and photothermal therapy,the structure of NCN2 was optimized,and a series of p H-activated oxazine dye NHNX was designed and synthesized.Through in vitro spectroscopy tests,the oxazine dye NHN3(λ_ex=618 nm,λ_em=747 nm,p K_a=5.40),which is more compatible with the fluorescence-photoacoustic imaging wavelength,was screened for the next step.Subsequently,the oxazine dye NHNQ-PEG2K with good water solubility was synthesized,which was further modified with polyethylene glycol through NHN3.The maximum absorption of NHNQ-PEG2K under neutral conditions is at 630 nm.And as the p H decreases,its maximum absorption wavelength red-shifts to 755 nm.Meanwhile,the p K_a of NHNQ-PEG2K increased to 5.78,which was closer to the weakly acidic microenvironment of the tumor.In in vitro experiments,NHNQ-PEG2K successfully achieved fluorescence and ratiometic photoacoustic imaging.In addition,further light-to-heat conversion experiments and cytotoxicity experiments show that NHNQ-PEG2K maintains effective light-to-heat conversion ability（light-to-heat conversion efficiency of 45.46%）under weak acid conditions,and it still maintains high cytotoxicity.Therefore,NHNQ-PEG2K is expected to become a activatable small molecule diagnosis and treatment reagent in tumor with fluorescence-photoacoustic dual-mode imaging and chemotherapy-photothermal combined treatment capabilities.