Judmann Benedikt, Braun Diana, Schirrmacher Ralf, Wängler Björn, Fricker Gert, Wängler Carmen
Biomedical Chemistry, Clinic of Radiology and Nuclear Medicine, Medical Faculty Mannheim, Heidelberg University, Theodor-Kutzer-Ufer 1-3, 68167 Mannheim, Germany.
Molecular Imaging and Radiochemistry, Clinic of Radiology and Nuclear Medicine, Medical Faculty Mannheim, Heidelberg University, Theodor-Kutzer-Ufer 1-3, 68167 Mannheim, Germany.
ACS Omega. 2022 Jul 28;7(31):27690-27702. doi: 10.1021/acsomega.2c03407. eCollection 2022 Aug 9.
The epidermal growth factor receptor (EGFR) is closely associated with tumor development and progression and thus an important target structure for imaging and therapy of various tumors. As a result of its important role in malignancies of various origins and the fact that antibody-based compounds targeting the EGFR have significant drawbacks in terms of pharmacokinetics, several attempts have been made within the last five years to develop peptide-based EGFR-specific radioligands based on the GE11 scaffold. However, none of these approaches have shown convincing results so far, which has been proposed to be attributed to different potential challenges associated with the GE11 lead structure: first, an aggregation of radiolabeled peptides, which might prevent their interaction with their target receptor, or second, a relatively low affinity of monomeric GE11, necessitating its conversion into a multimeric or polymeric form to achieve adequate EGFR-targeting properties. In the present work, we investigated if these aforementioned points are indeed critical and if the EGFR-targeting ability of GE11 can be improved by choosing an appropriate hydrophilic molecular design or a peptide multimer system to obtain a promising radiopeptide for the visualization of EGFR-overexpressing malignancies by positron emission tomography (PET). For this purpose, we developed several monovalent Ga-labeled GE11-based agents, a peptide homodimer and a homotetramer to overcome the challenges associated with GE11. The developed ligands were successfully labeled with Ga in high radiochemical yields of ≥97% and molar activities of 41-104 GBq/μmol. The resulting radiotracers presented log values between -2.17 ± 0.21 and -3.79 ± 0.04 as well as a good stability in human serum with serum half-lives of 112 to 217 min for the monovalent radiopeptides and 84 and 62 min for the GE11 homodimer and homotetramer, respectively. In the following studies, none of the Ga-labeled radiopeptides demonstrated a considerable EGF receptor-specific uptake in EGFR-positive A431 cells. Moreover, none of the agents was able to displace [I]I-EGF from the EGFR in competitive displacement assays in the same cell line in concentrations of up to 1 mM, whereas the endogenous receptor ligand hEGF demonstrated a high affinity of 15.2 ± 3.3 nM. These results indicate that it is not the aggregation of the GE11 sequence that seems to be the factor limiting the usefulness of the peptide as basis for radiotracer design but the limited affinity of monovalent and small homomultivalent GE11-based radiotracers to the EGFR. This highlights that the development of small-molecule GE11-based radioligands is not promising.
表皮生长因子受体（EGFR）与肿瘤的发生和发展密切相关，因此是各种肿瘤成像和治疗的重要靶标结构。由于其在各种起源恶性肿瘤中的重要作用，以及基于抗体的靶向EGFR化合物在药代动力学方面存在显著缺点，在过去五年中，人们进行了多次尝试，以基于GE11支架开发基于肽的EGFR特异性放射性配体。然而，到目前为止，这些方法均未取得令人信服的结果，有人认为这归因于与GE11先导结构相关的不同潜在挑战：第一，放射性标记肽的聚集，这可能会阻止它们与靶受体相互作用；第二，单体GE11的亲和力相对较低，需要将其转化为多聚体或聚合物形式，以实现足够的EGFR靶向特性。在本研究中，我们调查了上述问题是否确实至关重要，以及是否可以通过选择合适的亲水性分子设计或肽多聚体系统来提高GE11的EGFR靶向能力，从而获得一种有前景的放射性肽，用于通过正电子发射断层扫描（PET）对EGFR过表达的恶性肿瘤进行可视化。为此，我们开发了几种单价镓标记的基于GE11的试剂、一种肽同二聚体和一种同四聚体，以克服与GE11相关的挑战。所开发的配体成功地用镓标记，放射性化学产率≥97%，摩尔活度为41-104 GBq/μmol。所得放射性示踪剂的log 值在-2.17±0.21至-3.79±0.04之间，并且在人血清中具有良好的稳定性，单价放射性肽的血清半衰期为112至217分钟，GE11同二聚体和同四聚体的血清半衰期分别为84分钟和62分钟。在接下来的研究中，没有一种镓标记的放射性肽在EGFR阳性A431细胞中表现出显著的表皮生长因子受体特异性摄取。此外，在同一细胞系中进行的竞争性置换试验中，在浓度高达1 mM时，没有一种试剂能够从EGFR上置换[I]I-表皮生长因子，而内源性受体配体人表皮生长因子（hEGF）表现出15.2±3.3 nM的高亲和力。这些结果表明，限制该肽作为放射性示踪剂设计基础的有用性的因素似乎不是GE11序列的聚集，而是单价和小的同多价基于GE11的放射性示踪剂对EGFR的亲和力有限。这突出表明，开发基于小分子GE11的放射性配体前景不佳。
