PO.CL12.04 · 临床研究
通过claudin-4选择性放射性标记肽推进胰腺癌诊疗一体化：设计、优化与临床前验证
Advancing pancreatic cancer theragnostic through claudin-4-selective radiolabeled peptides: Design, optimization, and preclinical validation
作者与单位 Authors & Affiliations
摘要 Abstract
中文摘要
精确的癌症影像对于早期诊断、指导治疗和评估临床结局至关重要。然而，胰腺导管腺癌（PDAC）带来了重大的影像障碍，主要归因于其复杂的微环境、广泛的间质构成和快速的疾病进展。Claudin-4（CLDN4）是一种紧密连接蛋白，在PDAC中显著过表达，使其成为一个有前景的诊断和治疗靶点。我们的空间转录组学和蛋白质组学分析证实，与正常胰腺相比，PDAC中CLDN4表达增加了16倍。这一重要发现支持了其在肿瘤细胞表面的可及性，以及适用于诸如正电子发射断层扫描（PET）等分子成像方法。方法：我们使用固相肽合成法设计了选择性CLDN4靶向肽，随后通过D-氨基酸替换进行合理优化以增强代谢稳定性，并采用二聚化策略以改善亲和力和肿瘤滞留。肽结构通过聚乙二醇化连接子（长度不一）与DOTA螯合剂偶联，并评估血清稳定性。使用生物层干涉技术（BLI）测量结合亲和力。肽用⁶⁸Ga和⁶⁴Cu进行放射性标记，并使用动态PET/CT成像在皮下和转基因PDAC-KPTC小鼠模型中进行测试。结果：合成的肽对CLDN4表现出良好的结合亲和力，并成功用⁶⁸Ga和⁶⁴Cu进行了放射性标记。血清稳定性研究显示，具有较长PEG连接子的示踪剂在孵育4小时后表现出更优的稳定性。我们在D-氨基酸修饰序列中观察到类似结果，其稳定性优于母体肽。在临床前测试中，我们观察到示踪剂的肿瘤摄取在动态扫描的第一个小时内逐渐增加，注射后40分钟超过10% ID/g。这一摄取与TdTomato荧光相关，后者在我们的基因工程模型中标记表达CLDN-4的细胞。在阴性对照后续模型中的体外成像证实了我们的示踪剂对表达claudin4肿瘤的特异性。进一步评估二聚体肽和D-氨基酸修饰序列临床前成像的研究正在进行中。这种基于肽的诊疗一体化策略在改善PDAC早期检测和指导靶向治疗方面具有强大潜力。
查看英文原文 English abstract
Accurate cancer imaging is critical for early diagnosis, guiding therapy, and assessing clinical outcomes. Yet pancreatic ductal adenocarcinoma (PDAC) poses major imaging obstacles, mainly due to its complex microenvironment, extensive stromal composition, and rapid disease progression. Claudin-4 (CLDN4), a tight junction protein, is significantly overexpressed in PDAC, making it a promising diagnostic and therapeutic target. Our spatial transcriptomic and proteomic analyses confirmed a 16-fold increase in CLDN4 expression in PDAC compared with normal pancreas. This significant finding supports its accessibility on tumor cell surfaces and suitability for molecular imaging approaches such as positron emission tomography (PET).Methods:We designed selective CLDN4-targeted peptides using solid-phase peptide synthesis, followed by rational optimization through D-amino-acid substitutions to enhance metabolic stability and dimerization strategies to improve affinity and tumor retention. The peptide structures were conjugated to a DOTA chelator through pegylated linkers of varying lengths and evaluated for serum stability. Binding affinity was measured using biolayer interferometry (BLI). Peptides were radiolabeled with ⁶⁸Ga and ⁶⁴Cu and tested in subcutaneous and transgenic PDAC-KPTC mouse models using dynamic PET/CT imaging.Results:The synthesized peptides showed good binding affinity for CLDN4 and were successfully radiolabeled with ⁶⁸Ga and ⁶⁴Cu. Serum stability studies revealed that tracers with longer PEG linkers exhibited superior stability after 4 hours of incubation. We found a similar observation with D-amino-acid-modified sequences, which showed better stability than the parent peptide. For preclinical testing, we observed that tumor uptake of the tracers gradually increased during the first hour of the dynamic scan, surpassing 10% ID/g at 40 minutes post-injection. This uptake correlated with TdTomato fluorescence, which marks cells expressing CLDN-4 in our genetically engineered model. The in-vitro imaging in a negative control subsequent model confirms the specificity of our tracers to tumors expressing claudin4. Further studies are ongoing to evaluate preclinical imaging of dimeric peptides and D-amino acid-modified sequences. This peptide-based theragnostic strategy has strong potential to improve early detection and guide targeted therapy in PDAC.
利益披露 Disclosure
M. N. Aziz, None.