PDGFRα联合VEGFR2靶向纳米超声造影剂早期诊断甲状腺癌及相关机制的研究

The incidence of thyroid cancer is increasing year by year. Early diagnosis of the malignant thyroid nodule is the key to the successful treatment. CEUS and elasticity imaging are the research hot recently. Currently sonovue is the only contrast agent which is approved for use, however, it's lacking of the target specificity on the lesion. So the diagnosis of early thyroid cancer rate is not high. The literature shows that it is related between VEGF and tumor angiogenesis. PDGF is involved in the progress of tumor fibrosis. Both showed high expression in thyroid cancer. Initially the research group used triblock copolymer mPEG-PLGA-PLL as the nanocarrier, and Cy3-siRNA was successfully wrapped up, and transfected successfully. Thus, this study intends to use the triblock copolymer mPEG-PLGA-PLL to wrap up both VEGFR2 and PDGFRα at the same time, in order to study the specificity of targeting and developing effects of this contrast agent on vitro and in nude mouse model of papillary thyroid carcinoma, and then to assess the diagnostic value of this new contrast agents for tumor angiogenesis and fibrosis. So the objective is to improve thyroid cancer ultrasound diagnosis rate and develop this new technology on nano molecules imaging of the early thyroid cancer ultrasound. In the meantime, via blocking PDGF on the expression in thyroid cancer cells, we could explore the mechanism of PDGF for the fibrosis progress of thyroid cancer, elaborate the mechanism that the hardness of malignant nodules are increased and provide strong theoretical basis for this new type of contrast agent on the early diagnosis of thyroid cancer.

甲状腺癌的发病率逐年上升，早期诊断是成功治疗的关键。超声造影及弹性成像是近期研究的热点，目前唯一批准使用的造影剂sonovue缺乏对病变部位的靶向特异性，对早期甲状腺癌诊断率不高。文献表明VEGF与肿瘤新生血管生成有关，PDGF参与肿瘤纤维化过程，两者在甲状腺癌中均呈高表达。课题组前期利用三嵌段共聚体mPEG-PLGA-PLL作为纳米载体，成功包裹Cy3－siRNA并转染成功，因此本课题设想利用三嵌段共聚体同时包裹VEGFR2和PDGFRα，构建具有双靶向功能的纳米级超声造影剂，考察此造影剂在体外及裸鼠甲状腺乳头状癌模型上的靶向特异性及显影效果，评估该造影剂对肿瘤新生血管及纤维化的诊断价值，提高甲状腺癌超声诊断率，实现早期甲状腺癌超声纳米分子成像新技术。同时通过阻断PDGF在甲状腺癌细胞中的表达，探讨PDGF在甲状腺癌纤维化中的发生机制，为新型造影剂诊断早期甲状腺癌提供有力的理论依据。

本课题主要建立了裸鼠甲状腺乳头状癌BCPAP细胞和下调PDGF的乳头状癌BCPAP细胞模型，成功配制了靶向VEGFR2,PDGFRα以及同时靶向VEGFR2+PDGFRα的超声造影剂，体外实验结果显示微泡和VEGFR2，PDGFRα以及VEGFR2+PDGFRα结合情况良好，通过对不同组的裸鼠肿瘤进行靶向超声造影以及超声弹性检查，并分析其超声造影特点，术后进行病理免疫组化和胶原纤维检测。结果发现1）正常BCPAP细胞组的四种超声造影剂在裸鼠体内均成功显影，以高增强或周边环状高增强为主要表现。2）在下调PDGF组的裸鼠肿瘤体内，靶向VEGFR2的超声造影剂正常显影，以高增强或环状增强为主，靶向PDGFRα的超声造影剂在瘤体内未见明显充填，靶向VEGFR2+PDGFRα的超声造影剂呈稀疏充填或低增强，说明下调PDGF成功，PDGF和甲状腺癌的肿瘤血管生成有关。3）裸鼠甲状腺乳头状癌的超声弹性数值随着肿瘤的生长，体积的增大而逐渐增高，PDGF和胶原纤维的含量也随之增长，说明PDGF、胶原纤维和肿瘤的弹性硬度密切相关，超声弹性可以评价肿瘤的纤维化情况。PDGF参与了甲状腺乳头状癌的发生发展，与肿瘤的血管生成和纤维化相关，为进一步抗纤维化和抗血管化的靶向PDGF的甲状腺癌治疗打下来的实验基础。