Department of Cancer Systems Imaging, MD Anderson Cancer Center, 1881 East Rd, Houston, TX, 77054, USA.
Department of Chemistry, Rice University, 6100 Main St, Houston, TX, 77005, USA.
Mol Imaging Biol. 2022 Dec;24(6):959-972. doi: 10.1007/s11307-022-01747-9. Epub 2022 Jun 22.
Metabolic reprogramming plays an important role in the tumorigenesis of clear cell renal cell carcinoma (ccRCC). Currently, positron emission tomography (PET) reporters are not used clinically to visualize altered glutamine metabolism in ccRCC, which greatly hinders detection, staging, and real-time therapeutic assessment. We sought to determine if (2S,4R)-4-[F]fluoroglutamine ([F]FGln) could be used to interrogate altered glutamine metabolism in ccRCC lesions in the lung.
We generated a novel ccRCC lung lesion model using the ccRCC cell line UMRC3 stably transfected with GFP and luciferase constructs. This cell line was used for characterization of [F]FGln uptake and retention by transport analysis in cell culture and by PET/MRI (magnetic resonance imaging) in animal models. Tumor growth in animal models was monitored using bioluminescence (BLI) and MRI. After necropsy, UMRC3 tumor growth in lung tissue was verified by fluorescence imaging and histology.
In UMRC3 cells, [F]FGln cell uptake was twofold higher than cell uptake in normal kidney HEK293 cells. Tracer cell uptake was reduced by 60-90% in the presence of excess glutamine in the media and by 20-50% upon treatment with V-9302, an inhibitor of the major glutamine transporter alanine-serine-cysteine transporter 2 (ASCT2). Furthermore, in UMRC3 cells, [F]FGln cell uptake was reduced by siRNA knockdown of ASCT2 to levels obtained by the addition of excess exogenous glutamine. Conversely, [F]FGln cellular uptake was increased in the presence of the glutaminase inhibitor CB-839. Using simultaneous PET/MRI for visualization, retention of [F]FGln in vivo in ccRCC lung tumors was 1.5-fold greater than normal lung tissue and twofold greater than muscle. In ccRCC lung tumors, [F]FGln retention did not change significantly upon treatment with CB-839.
We report one of the first direct orthotopic mouse models of ccRCC lung lesions. Using PET/MR imaging, lung tumors were easily discerned from normal tissue. Higher uptake of [F]FGln was observed in a ccRCC cell line and lung lesions compared to HEK293 cells and normal lung tissue, respectively. [F]FGln cell uptake was modulated by exogenous glutamine, V-9302, siRNA knockdown of ASCT2, and CB-839. Interestingly, in a pilot therapeutic study with CB-839, we observed no difference in treated tumors relative to untreated controls. This was in contrast with cellular studies, where CB-839 increased glutamine uptake.
代谢重编程在透明细胞肾细胞癌（ccRCC）的肿瘤发生中起着重要作用。目前，正电子发射断层扫描（PET）报告器尚未用于临床可视化 ccRCC 中改变的谷氨酰胺代谢，这极大地阻碍了检测、分期和实时治疗评估。我们试图确定（2S，4R）-4-[F]氟代谷氨酰胺（[F]FGln）是否可用于检测肺中 ccRCC 病变中改变的谷氨酰胺代谢。
我们使用稳定转染 GFP 和荧光素酶构建体的 ccRCC 细胞系 UMRC3 生成了一种新型 ccRCC 肺病变模型。该细胞系用于通过细胞培养中的转运分析和动物模型中的 PET/MRI（磁共振成像）来研究[F]FGln 的摄取和保留。使用生物发光（BLI）和 MRI 监测动物模型中的肿瘤生长。解剖后，通过荧光成像和组织学验证 UMRC3 肿瘤在肺组织中的生长。
在 UMRC3 细胞中，[F]FGln 的细胞摄取比正常肾脏 HEK293 细胞的摄取高两倍。在培养基中存在过量谷氨酰胺或用主要谷氨酰胺转运体丙氨酸-丝氨酸-半胱氨酸转运体 2（ASCT2）抑制剂 V-9302 处理时，示踪剂细胞摄取减少 60-90%。此外，在 UMRC3 细胞中，通过 ASCT2 的 siRNA 敲低，[F]FGln 的细胞摄取降低至通过添加过量外源性谷氨酰胺获得的水平。相反，在存在谷氨酰胺酶抑制剂 CB-839 的情况下，[F]FGln 的细胞摄取增加。使用同时进行的 PET/MRI 可视化，ccRCC 肺肿瘤中[F]FGln 的体内保留比正常肺组织高 1.5 倍，比肌肉高 2 倍。在 ccRCC 肺肿瘤中，[F]FGln 的保留在用 CB-839 治疗后没有显著变化。
我们报告了首例直接原位 ccRCC 肺病变的小鼠模型之一。使用 PET/MR 成像，很容易将肺肿瘤与正常组织区分开来。与 HEK293 细胞和正常肺组织相比，ccRCC 细胞系和肺病变中观察到更高的[F]FGln 摄取。[F]FGln 细胞摄取可通过外源性谷氨酰胺、V-9302、ASCT2 的 siRNA 敲低和 CB-839 进行调节。有趣的是，在 CB-839 的初步治疗研究中，与未治疗的对照组相比，我们观察到治疗肿瘤没有差异。这与细胞研究形成对比，在细胞研究中，CB-839 增加了谷氨酰胺摄取。
