放射性药物对疾病早期诊断与精准治疗具有重要意义。当前我国放射性诊疗药物的发展水平与欧美发达国家仍存在一定差距，远不能满足临床需求。本论文围绕临床急需的新型放射性药物进行研发及临床转化研究：（1）前列腺特异性膜抗原（Prostate specific membrane antigen, PSMA）靶向放射性治疗药物可以精准有效治疗前列腺癌等恶性肿瘤，是当前国际核医学领域的研究重点，本论文设计研发新型^125/131I标记的PSMA靶向化合物，以期成为具有临床转化潜力的PSMA靶向放射性治疗药物。（2）¹⁸F核素是当前PET显像药物的首选核素，18F药物的发展远不能满足临床需求，由于半衰期短（109.8 min），药物即时制备的关键技术亦成其临床转化瓶颈，甚至导致许多性质优良的探针“胎死腹中”。针对国际上新一代Aβ分子探针[¹⁸F]D3FSP临床制备关键技术——快速纯化方法进行研究，建立简单、快速、高效、低成本的探针制备新工艺，推动[18F]D3FSP在国内外的临床转化和应用。具体研究内容如下：
一、新型125/131I-PSMA肿瘤治疗药物的研究
由于放射性碘同位素（¹²³I/¹²⁴I/¹²⁵I/¹³¹I）的多样性，其标记药物既可用于SPECT显像（¹²³I/¹³¹I）、PET显像（¹²⁴I），也可用于肿瘤靶向治疗（多个¹³¹I药物已应用于临床）及肿瘤植入治疗（¹²⁵I籽源），成为核医学重要的诊疗核素。本论文选择国内自主生产且规模化供应的¹³¹I作为治疗核素，在PSMA特异性靶向结合基团谷氨酸-脲-赖氨酸结构基础上，首次引入谷氨酸和酪氨酸以探究¹³¹I标记的PSMA靶向化合物的构效关系，共设计合成54个化合物，其中23个化合物未见文献报道： 9个含碘PSMA靶向目标化合物、7个放射性碘标记前体、7个标记中间体。鉴于¹²⁵I核素半衰期长，与¹³¹I生物化学性质相似，论文采用¹²⁵I代替¹³¹I进行临床前初步药物评价与筛选。
体外竞争结合实验和放射自显影实验结果表明，9个新合成目标化合物均具有良好的PSMA靶向亲和性和特异性。考虑对碘苯环结构体内脱碘现象较轻，因此选取7个化合物进行“两步法”放射性碘标记，所得¹²⁵I标记物放射化学纯度均大于95%（除[¹²⁵I]39放射化学纯度约为90%），比活度约为2200 Ci/mmol。体外细胞实验结果表明，在¹²⁵I标记物中引入酪氨酸可以提高肿瘤细胞内化摄取率。
荷瘤裸鼠体内分布实验结果表明，含有酪氨酸的125I标记物在肝肠组织中摄取较高，在肾脏、PC3-PIP肿瘤中摄取低；而引入1 - 2个谷氨酸基团后，脂溶性较低的¹²⁵I标记物主要通过泌尿系统代谢排出体外，在肾脏和PC3-PIP肿瘤中摄取明显升高，说明在连接基团中引入不同数量的谷氨酸、酪氨酸，会改变化合物的PSMA靶向亲和性及药代动力学性质。其中，[125I]43在血液和肝脏中的摄取明显低于国际上报道的第一代PSMA靶向治疗药物[^125/131I]MIP-1095，有望减轻[^125/131I]MIP-1095在临床试验阶段出现的血液毒性，值得进一步深入研究。
二、Aβ分子探针[18F]D3FSP的制备新工艺研究
[¹⁸F]D3FSP可早期诊断阿尔茨海默病（AD），在美国已进入Ⅰ/Ⅱ期临床试验阶段。[¹⁸F]D3FSP原制备工艺中采用传统HPLC纯化方法，耗时长（90 - 100 min），产率较低（25 - 30%，衰变校正）。本论文通过优化[¹⁸F]D3FSP的亲核氟化标记条件及固相萃取（SPE）纯化方法，简化探针制备流程、提高其放射化学产率。
为探究放射化学合成条件及SPE纯化方法对制剂中化学杂质含量的影响，以及化学杂质的Aβ靶向亲和性，论文共合成10种化合物，包括：1种“两步法”标记前体化合物、3种标记中间体、3种标记反应中产生的非放射性化学杂质。实验得出[¹⁸F]D3FSP的最佳亲核氟化标记条件是：以DMSO为氟化反应溶剂、130 ℃ 下反应10 min，再加入1 mL 3 N HCl溶液、100 ℃ 下反应5 min进行脱保护反应，得到最终产物[¹⁸F]D3FSP。
体外竞争结合实验表明，D3FSP（62）（IC₅₀ = 9.8 ± 0.5 nM）与AV-45（IC₅₀ = 8.6 ± 0.5 nM）具有相似的Aβ靶向亲和性，OH-衍生物（63）（IC₅₀ ​​​​​​​= 19.5 ± 0.5 nM）和Cl-衍生物（65）（IC₅₀ = 18.6 ± 3.9 nM）也显示一定的靶向结合，说明制剂中化学杂质会影响[¹⁸F]D3FSP与脑内Aβ斑块结合，进而影响显像结果。因此，论文通过优化SPE纯化方法，降低制剂中化学杂质含量：使用不同体积（2 - 10 mL）、不同浓度（体积分数10 - 100%）的乙腈/水或乙醇/水溶液淋洗7种不同类型的固相萃取小柱，定量分析纯化效果，确定SPE最佳纯化条件：使用6 mL 30%乙腈/水溶液淋洗Sep-Pak^® light tC18小柱洗脱产物中含量最大的OH-衍生物（63），最后用2 mL 50%乙醇/生理盐水溶液将目标产物[18F]D3FSP淋洗至产品瓶中。[¹⁸F]D3FSP制备新工艺耗时50 min，放射化学产率为44.4 ± 5.7%（n = 10，衰变校正），放射化学纯度﹥95%，制剂中化学量为21 ± 5 μg/批次（低于FDA规定同类型分子探针的化学量标准：< 50 μg/单人制剂 ），有望进行自动化制备和临床转化研究。
综上所述，本论文首次在^125/131I标记的PSMA靶向药物中引入不同数量的谷氨酸或酪氨酸基团，筛选出的候选化合物[^125/131I]43具有良好的PSMA靶向亲和性和药代动力学性质，表现出精准治疗前列腺癌等恶性肿瘤的临床转化潜力；研究了制剂中化学杂质对[¹⁸F]D3FSP显像的影响，利用SPE纯化方法取代[¹⁸F]D3FSP原制备工艺中复杂、耗时的HPLC纯化方法，建立了简单、快速、高效、低成本的探针制备新工艺，加速[¹⁸F]D3FSP在国内外的临床转化与应用，为核医学领域放射性药物的临床推广提供思路和依据。

Radiopharmaceuticals is of great significance for early diagnosis and precision medicine of diseases, while there is still a certain disparity between the research of radiopharmaceuticals in China and that of in Europe and the North America. This dissertation work focuse on the development and translation of urgently needed radiopharmaceuticals in clinic, including: (1) The radioligand therapy, targeting prostate specific membrane antigen (PSMA), can treat prostate cancer and other malignant tumors accurately and effectively, which is the research focus in the field of nuclear medicine. Great efforts were made to design and develop novel ^125/131I-labeled PSMA compounds, in order to obtain potential radioligands for clinical translation for tumor treatment. (2) ¹⁸F is the preferred raidonuclide of PET imaging agents, while the development of ¹⁸F-labeled agents is far from meeting the clinical needs. Due to short half-life (109.8 min) of 18F, the rapid preparation process of 18F-labeled agents has become the bottleneck of clinical translation, even lead to many promising agents “stillborn”. A rapid purification, the key preparation technologies of the novel β-amyloid (Aβ) imaging agent [¹⁸F]D3FSP, was studied. A simple, rapid, efficient and low-cost preparation process was established for conributing to accelerate the clinical translation and application of [¹⁸F]D3FSP in China.
1. Development of novel ^125/131I-labeled PSMA-targeted ligands for radiotherapy
Since the diversity of medical radiouiodine isotopes (¹²³I/¹²⁴I/¹²⁵I/¹³¹I), radioiodinated ligands can be used not only for SPECT imaging (¹²³I/¹³¹I) and PET imaging (¹²⁴I), but also for targeted radio-therapy ⁽¹³¹I) and implantation therapy (¹²⁵I) of tumors. Radioiodine has become important theranostic nuclide in nuclear medicine. Based to the key PSMA-targeted motif glutamic-urea-lysine, we selected domestically produced and large-scale supplied ¹³¹I as radiotherapy nuclide, explore the structure-activity relationship of ¹³¹I-labeled PSMA ligands by adding glutamate acid or subtracting tyrosine for the first time. A total of fifity-four compounds were designed and synthesized, of which twenty-three compounds have not been reported, including nine PSMA-targeted compounds, seven radioiodine-labeled precursors and seven radioiodine-labeled intermediates. Taking advantage of the long half-life and similar biochemical properties into consideration, ¹²⁵I was used as a suragate instead of ¹³¹I for preliminary preclinical evaluations.

In vitro competitive binding assay and in vitro autoradiography study showed that nine iodinated compounds displayed high binding affinities and specificity towards PSMA. Due to the relatively low deiodination of p-iodobenzene, seven ¹²⁵I-labeled ligands were selected for “two-step” radioiodine labeling, and obtained in high radiochemical purity of ﹥ 95% (except for 90% of [¹²⁵I]39) and specific activity of ~ 2200 Ci/mmol. In vitro cell uptake studies demonstrated that the introduction of tyrosine could improve the internalization of ¹²⁵I-labeled ligands in tumor cells.
Biodistribution studies in mice bearing tumors showed that ¹²⁵I-labeled ligands containing tyrosine exhibited highe accumulations in the liver and intestine, and low uptake in kidney and PC3-PIP tumor. Introducing one or two glutamic acid group(s), ¹²⁵I-labeled ligands with reduced lipophilicity were excreted through urinary system, resulting in increased accumulations in kidney and PC3-PIP tumor. It indicated that the linker modification of adding glutamate acid or subtracting tyrosine has a significant impact on the binding affinity and pharmacokinetic properties of ¹²⁵I-labeled ligands. Among them, [¹²⁵I]43 showing significantly lower uptakes in the liver and blood than [125I]MIP-1095, which is expected to reduce the blood toxicity of [^125/131I]MIP-1095 in clinical trials. [^125/131I]43 is worthy of further study.
2. Study of new preparation process of Aβ-targeted PET imaging agent [¹⁸F]D3FSP
[¹⁸F]D3FSP is the new generation of Aβ-targeted agent for early diagnosis of Alzheimer's disease (AD), which is in phase I/II clinical trials in US and clinical translation in China. Since [¹⁸F]D3FSP was originally achieved within a long time (90 - 100 min) and low radiochemical yield (25 - 30%, decay-corrected) with the traditional semi-preparative liquid chromatography (HPLC) purification, we herein aimed to simplify the preparation process and improve the radiochemical yield by optimization of nucleophilic fluorination reaction conditions and solid phase extraction (SPE) purification.
In order to study the amount and the Aβ-targeted affinity of chemical impurities, ten compounds were synthesized, including one "two-step" labeled precursor, three labeled intermediates, and three nonradioactive chemical impurities produced in the nucleophilic fluorination reaction. Results of the optimization studies suggested that optimal fluorination condition was accomplished by heating in DMSO at 130 ℃ for 10 min, followed by the deprotection step, which was carried out at 100 ℃ for 5 min, using 1 mL HCl solution (3 N) as acid of hydrolysis.
In vitro competitive binding assay showed that D3FSP (62) (IC₅₀ = 9.8 ± 0.5 nM) displayed similar Aβ-targeted affinity as AV-45 (IC₅₀ = 8.6 ± 0.5 nM), meanwhile OH-derivative (63) (IC₅₀ = 19.5 ± 0.5 nM) and Cl-derivative (65) (IC₅₀ = 18.6 ± 3.9 nM) showed a slightly lower binding abilities, indicating that the high amount of chemical impurities will inhibit the binding of [¹⁸F]D3FSP towards Aβ plaques in the brain. Therefore, the SPE purification method was optimized herein to reduce the amount of chemical impurities in the [¹⁸F]D3FSP product. The separation of the OH-pseudo carrier (63) and the desired product [18F]D3FSP was analyzed quantitatively through using different concentrations (10 - 100%) and volumes (2 - 10 mL) of ethanol/water or acetonitrile/water to wash seven types of solid-phase extraction (SPE) cartridges. The optimal SPE purification method of [¹⁸F]D3FSP was determined as the following: washing Sep-Pak? light tC18 cartridge with 6 mL 30%acetonitrile/water for removing OH-pseudo carrier (63), followed by eluting the desired [¹⁸F]D3FSP as product with 2 mL 50% ethanol/saline.Using the optimized radiosynthesis and SPE purification method, [¹⁸F]D3FSP was successfully prepared in radiochemical yield of 44.4 ± 5.7% (n = 10, decay-corrected) and radiochemical purity of ﹥ 95% in 50 min. Total mass of chemical in the per batch of [¹⁸F]D3FSP product was 21 ± 5 μg (lower than 50 μg/dose of [¹⁸F]AV-45 required by FDA). The modified method might be suitable for subsequent clinical research on translation to an automated synthesizer. In conclusion, through the structure-activity study of ^125/131I-PSMA ligands by adding glutamate acid or subtracting tyrosine, [^125/131I]43 was selected as a candidate and displayed excellent binding affinity towards PSMA and pharmacokinetic properties. [^125/131I]43 showed potential clinical translation for the treatment of prostate cancer and other malignant tumors. The effect of chemical impurities on the imaging of [¹⁸F]D3FSP was studied and the SPE purification method was used to replace the complex and time-consuming HPLC purification method in the original preparation process of [¹⁸F]D3FSP. The simple, rapid, efficient and low-cost preparation process can promote the clinical translation and application of [¹⁸F]D3FSP, which is expected to provide ideas for clinical development of radiotracers in future.