Anti-non Small Cell Lung Cancer Activity And Mechanism Of A New Type Of Polyoxometalate

The morbidity of non-small cell lung cancer?NSCLC?is increasing yearly,the age of onset is decreasing,the mortality is high in recent years,threaten human's health seriously.There are 18.1million new cases of cancer and 9.6million death cases of cancer in the world wide which according to statistics by the American cancer society on September 2018.Lung cancer is the most common cancer?11.6%in whole?in human,and it's also the leading cause of cancer death?18.4%of total cancer death?.Non-small cell lung cancer accounts for 85%of lung cancer.China is one of the country which have high incidence of lung cancer.It is a burning question to improve the effectiveness of lung cancer therapy.Surgery,radiotherapy,interventional therapy,chemotherapy,target therapy and immunotherapy are current therapies for NSCLC.There are many limiting factors in surgical treatment,and the injury to the body is great.Radiotherapy has poor selectivity,it would damage the normal lung cells during the therapy,and cause high incidence of radiation pneumonitis.Thought medication has many side effects,it is also one of the most importent therapy of NSCLC.The drug treatment of tumors mainly includes three categories:the first one is immunotherapy,it developed rapidly recent years.But when is the best time to star the therapy and how to select the beneficiaries are difficult problems.The second one is targeted drug,it has obvious curative effect and low damage to other normal cells,but the biggest problem is the drug resistance three to six months after treatment.Doctors should to change the drug to solve drug resistance,but all of the target drugs are very expensive,it's a huge financial burden for patients.The third one is cytotoxic agent,that is typical chemotherapy drug,it would damage the normal cell during therapy for it has poor selectivity.Severe adverse drug reactions even lead to the failure of chemotherapy termination in many patients.Therefore,the research and development of anti-tumor drugs is still the focus of research.It is urgent to develop effective and cost-effective drugs against NSCLC.Polyoxometalate is a kind of polymetallic oxygen cluster compound.It has the characteristics of spatial structure diversity,skeletal structural stability,easy to modify,and strong electron acceptability,high redox properties.Most of the study of polyoxometalate focous on invitro tests,dates from 1970s,however,there is no systematic study on the effect and mechanism of polymetalate anti-NSCLC therapy.The objective of this study is to design synthesis new polyoxometalate on the basic of preliminary study.First,character the structure of it.Then research the antitumor activity and toxicity in vitro,acute toxicity studies,and in vitro?in vivo mechanism of anti-NSCLC from cell and protein levels at last.The main study contents include the following five parts:Part I Synthesis characterization and stability of two polyoxometalates?VMOP-31 and AsMOP-8?Two polyoxometalates VMOP-31 and AsMOP-8 were synthesized by conventional synthesis method in this part.Infrared spectroscopy,UV-vis spectroscopy,single crystal diffraction NMR analysis,and thermogravimetric analysis were used to characterize the two polyoxometalates.The results showed that the structure of VMOP-31 was nano-molecular cage with octahedral configuration,and the AsMOP-8 wasPart II Screen antitumor activities and toxicity of polyoxometalates in vitroMTT was used to screen the antitumor activity of three polyoxometalates in this part.SMMC-7721,MCF-7,A549 and HL-60 four cell lines were chosen as module cells to study the antitumor activities of polyoxometalates,and normal lung cell BEAS-2B was also chosen to research the cytotoxicity at the same time.The results showed that VMOP-31 and AsMOP-8 expressed better antitumor activity.The IC₅₀ of the two polyoxometalates to BEAS-2B cells is higher than the IC50 to A549 cells.That means the two polyoxometalates have better anti-NSCLC activity and lower cytotoxicity to normal lung cells.Finally,the anti-NSCLC activity and cytotoxicity to BEAS-2B were compared with positive drugs cisplatin and arsenic trioxide.The results showed that the two drugs were better than the two positive drugs.Part III Acute toxicity study of AsMOP-8Acute toxicity of AsMOP-8 was studied by intraperitoneal injection in ICR mice,LD₅₀ was 35mg/kg,95%confidential interval was 90.16¹20.50mg/kg.That would lay a foundation for the selection of anti-tumor dosage in vivo.Part IV Study of anti-NSCLC activity and toxicity of AsMOP-8 in vitroMice with Lewis lung cancer cells were chosen as study model in this part,the groups were divided according to LD50 which got from acute toxicity test,normal saline was chosen to be negative control group,cisplatin was chosen to be positive control group.The anti-tumor effect of polyoxometallate and its effect on each system were studied.The result showed that AsMOP-8 had better antitumor activities than the positive and negative control groups,had no significant effects on organs of mice in vivo.Part V Study of anti-NSCLC mechanism of AsMOP-8 in vitro and in vivoFirst,we use ultraviolet spectroscopy to study the interaction of AsMOP-8 with biomolecule ctDNA and BSA;next the morphological changes of A549 cells and the changes of chromatin in cell nucleus were observed macroscopically;then the influences of polyoxometalate to cell cycle,apoptosis,cell membrane potential,intracellular ROS levels and intracellular calcium concentration were researched by flow cytometry;western blot was used to find the apoptosis pathway at last.Mice with Lewis lung cancer cells were chosen as study model in vivo experiment,the groups were divided according to LD₅₀ which got from acute toxicity test,normal saline was chosen to be negative control group,cisplatin was chosen to be positive control group,study the effect of polyoxometalate to immune system by observe the pathological changes of thymus and spleen,calculate thymus index and spleen index;observe the apoptosis of tumor cells by HE staining of tumor sections;western blot was used to find the apoptosis pathway.The results showed that it could combined with ctDNA and BSA;induce A549 cells apoptosis and chromatin fragmentation;induce cell cycle arrested in G2/M phase;A549 cells apoptosis,reduce cell memberane potential,increase intracellular ROS level,and increase intracellular calcium concentration in vitro.It could active the immune system of mice and inducing the apoptosis of tumor cell in vivo.The apoptosis pathway is Caspase depended and Caspase independent mitochondrial pathway both in vitro and in vivo.Finally,a series of polyoxometalates were synthesized by self-designed synthesis in this study,and AsMOP-8 was one of the effective polyoxometalates in anti-NSCLC screened from the whole.Then the activity and mechanism of AsMOP-8 on anti-NSCLC were studied systematically in vitro and in vivo.The results showed that it had a clear anti-NSCLC effect in vitro and in vivo,the mechanism was induced tumor cell apoptosis,and the apoptosis pathway was also founded.Part results of the research have been granted the national invention patent.The study provides a scientific basis for the development of new polyoxometalate drugs to anti-tumor with independent intellectual property rights.