Susan Pike, Melinda Wuest, Ana Lopez-Campistrous, Mi Yao Hu, Ratmir Derda, Frank Wuest* and Todd McMullen*,
{"title":"用于血小板生成的生长因子受体 α 分子成像的第一代放射性标记环肽","authors":"Susan Pike, Melinda Wuest, Ana Lopez-Campistrous, Mi Yao Hu, Ratmir Derda, Frank Wuest* and Todd McMullen*, ","doi":"10.1021/acs.molpharmaceut.4c0054910.1021/acs.molpharmaceut.4c00549","DOIUrl":null,"url":null,"abstract":"<p >Occult nodal spread and metastatic disease require longstanding imaging and biochemical assessments for thyroid cancer, a disease that has a propensity for diffuse, small-volume disease. We have developed a <sup>64</sup>Cu-labeled platelet-derived growth factor receptor α (PDGFRA) antibody for immuno-PET of PDGFRA in metastatic papillary thyroid cancer (PTC). The present work describes the discovery of small cyclic PDGFRA-targeting peptides, their binding features, and radiolabeling with positron emitter gallium-68 (<sup>68</sup>Ga) for <i>in vitro and in vivo</i> characterization in thyroid cancer models. Phage-display technology with two separate libraries and seven different cell lines was used through three rounds of biopanning as well as flow cytometry and comparative analysis with recombinant protein to select specific peptide sequences. Phenotypic binding analysis was completed by using phosphorylation and cell migration assays. <i>In vitro</i> protein binding was analyzed with thermophoresis and flow cytometry using the fluorescent-labeled PDGFRA peptide. Peptide candidates were modified with the NOTA chelator for radiolabeling with <sup>68</sup>Ga. <i>In vitro</i> cell uptake was studied in various thyroid cancer cell lines. <i>In vivo</i> studies of <sup>68</sup>Ga-labeled peptides included metabolic stability and PET imaging. From the original library (10<sup>13</sup> compounds), five different peptide groups were identified based on biopanning experiments with and without the α subunit of PDGFR, leading to ∼50 peptides. Subsequent phenotypic screening revealed two core peptide sequences (<b>CP16</b> and <b>CP18</b>) that demonstrated significant changes in the level of PDGFRA phosphorylation and cell migration. Alanine scan sublibraries were created from these two lead peptide sequences, and peptides were radiolabeled using <sup>68</sup>Ga-GaCl<sub>3</sub> at pH 4.5, resulting in RCP > 95% within 34–40 min, including SPE purification. Cyclic peptide <b>CP18.5</b> showed the strongest effects on cell migration, flow cytometry, and binding by visual interference color assay. <sup>68</sup>Ga-labeled PDGFRA-targeting peptides showed elevated cell and tumor uptake in models of thyroid cancer, with <sup><b>68</b></sup><b>Ga-NOTA-CP18.5</b> being the lead candidate. However, metabolic stability <i>in vivo</i> was compromised for <sup><b>68</b></sup><b>Ga-NOTA-CP18.5</b> vs <sup><b>68</b></sup><b>Ga-NOTA-CP18</b> but without impacting tumor uptake or clearance profiles. First-generation radiolabeled cyclic peptides have been developed as novel radiotracers, particularly <sup><b>68</b></sup><b>Ga-NOTA-CP18.5</b>, for the molecular imaging of PDGFRA in thyroid cancer.</p>","PeriodicalId":52,"journal":{"name":"Molecular Pharmaceutics","volume":null,"pages":null},"PeriodicalIF":4.5000,"publicationDate":"2024-08-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"First-Generation Radiolabeled Cyclic Peptides for Molecular Imaging of Platelet-Derived Growth Factor Receptor α\",\"authors\":\"Susan Pike, Melinda Wuest, Ana Lopez-Campistrous, Mi Yao Hu, Ratmir Derda, Frank Wuest* and Todd McMullen*, \",\"doi\":\"10.1021/acs.molpharmaceut.4c0054910.1021/acs.molpharmaceut.4c00549\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p >Occult nodal spread and metastatic disease require longstanding imaging and biochemical assessments for thyroid cancer, a disease that has a propensity for diffuse, small-volume disease. We have developed a <sup>64</sup>Cu-labeled platelet-derived growth factor receptor α (PDGFRA) antibody for immuno-PET of PDGFRA in metastatic papillary thyroid cancer (PTC). The present work describes the discovery of small cyclic PDGFRA-targeting peptides, their binding features, and radiolabeling with positron emitter gallium-68 (<sup>68</sup>Ga) for <i>in vitro and in vivo</i> characterization in thyroid cancer models. Phage-display technology with two separate libraries and seven different cell lines was used through three rounds of biopanning as well as flow cytometry and comparative analysis with recombinant protein to select specific peptide sequences. Phenotypic binding analysis was completed by using phosphorylation and cell migration assays. <i>In vitro</i> protein binding was analyzed with thermophoresis and flow cytometry using the fluorescent-labeled PDGFRA peptide. Peptide candidates were modified with the NOTA chelator for radiolabeling with <sup>68</sup>Ga. <i>In vitro</i> cell uptake was studied in various thyroid cancer cell lines. <i>In vivo</i> studies of <sup>68</sup>Ga-labeled peptides included metabolic stability and PET imaging. From the original library (10<sup>13</sup> compounds), five different peptide groups were identified based on biopanning experiments with and without the α subunit of PDGFR, leading to ∼50 peptides. Subsequent phenotypic screening revealed two core peptide sequences (<b>CP16</b> and <b>CP18</b>) that demonstrated significant changes in the level of PDGFRA phosphorylation and cell migration. Alanine scan sublibraries were created from these two lead peptide sequences, and peptides were radiolabeled using <sup>68</sup>Ga-GaCl<sub>3</sub> at pH 4.5, resulting in RCP > 95% within 34–40 min, including SPE purification. Cyclic peptide <b>CP18.5</b> showed the strongest effects on cell migration, flow cytometry, and binding by visual interference color assay. <sup>68</sup>Ga-labeled PDGFRA-targeting peptides showed elevated cell and tumor uptake in models of thyroid cancer, with <sup><b>68</b></sup><b>Ga-NOTA-CP18.5</b> being the lead candidate. However, metabolic stability <i>in vivo</i> was compromised for <sup><b>68</b></sup><b>Ga-NOTA-CP18.5</b> vs <sup><b>68</b></sup><b>Ga-NOTA-CP18</b> but without impacting tumor uptake or clearance profiles. First-generation radiolabeled cyclic peptides have been developed as novel radiotracers, particularly <sup><b>68</b></sup><b>Ga-NOTA-CP18.5</b>, for the molecular imaging of PDGFRA in thyroid cancer.</p>\",\"PeriodicalId\":52,\"journal\":{\"name\":\"Molecular Pharmaceutics\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.5000,\"publicationDate\":\"2024-08-17\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Molecular Pharmaceutics\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://pubs.acs.org/doi/10.1021/acs.molpharmaceut.4c00549\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"MEDICINE, RESEARCH & EXPERIMENTAL\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular Pharmaceutics","FirstCategoryId":"3","ListUrlMain":"https://pubs.acs.org/doi/10.1021/acs.molpharmaceut.4c00549","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"MEDICINE, RESEARCH & EXPERIMENTAL","Score":null,"Total":0}
First-Generation Radiolabeled Cyclic Peptides for Molecular Imaging of Platelet-Derived Growth Factor Receptor α
Occult nodal spread and metastatic disease require longstanding imaging and biochemical assessments for thyroid cancer, a disease that has a propensity for diffuse, small-volume disease. We have developed a 64Cu-labeled platelet-derived growth factor receptor α (PDGFRA) antibody for immuno-PET of PDGFRA in metastatic papillary thyroid cancer (PTC). The present work describes the discovery of small cyclic PDGFRA-targeting peptides, their binding features, and radiolabeling with positron emitter gallium-68 (68Ga) for in vitro and in vivo characterization in thyroid cancer models. Phage-display technology with two separate libraries and seven different cell lines was used through three rounds of biopanning as well as flow cytometry and comparative analysis with recombinant protein to select specific peptide sequences. Phenotypic binding analysis was completed by using phosphorylation and cell migration assays. In vitro protein binding was analyzed with thermophoresis and flow cytometry using the fluorescent-labeled PDGFRA peptide. Peptide candidates were modified with the NOTA chelator for radiolabeling with 68Ga. In vitro cell uptake was studied in various thyroid cancer cell lines. In vivo studies of 68Ga-labeled peptides included metabolic stability and PET imaging. From the original library (1013 compounds), five different peptide groups were identified based on biopanning experiments with and without the α subunit of PDGFR, leading to ∼50 peptides. Subsequent phenotypic screening revealed two core peptide sequences (CP16 and CP18) that demonstrated significant changes in the level of PDGFRA phosphorylation and cell migration. Alanine scan sublibraries were created from these two lead peptide sequences, and peptides were radiolabeled using 68Ga-GaCl3 at pH 4.5, resulting in RCP > 95% within 34–40 min, including SPE purification. Cyclic peptide CP18.5 showed the strongest effects on cell migration, flow cytometry, and binding by visual interference color assay. 68Ga-labeled PDGFRA-targeting peptides showed elevated cell and tumor uptake in models of thyroid cancer, with 68Ga-NOTA-CP18.5 being the lead candidate. However, metabolic stability in vivo was compromised for 68Ga-NOTA-CP18.5 vs 68Ga-NOTA-CP18 but without impacting tumor uptake or clearance profiles. First-generation radiolabeled cyclic peptides have been developed as novel radiotracers, particularly 68Ga-NOTA-CP18.5, for the molecular imaging of PDGFRA in thyroid cancer.
期刊介绍:
Molecular Pharmaceutics publishes the results of original research that contributes significantly to the molecular mechanistic understanding of drug delivery and drug delivery systems. The journal encourages contributions describing research at the interface of drug discovery and drug development.
Scientific areas within the scope of the journal include physical and pharmaceutical chemistry, biochemistry and biophysics, molecular and cellular biology, and polymer and materials science as they relate to drug and drug delivery system efficacy. Mechanistic Drug Delivery and Drug Targeting research on modulating activity and efficacy of a drug or drug product is within the scope of Molecular Pharmaceutics. Theoretical and experimental peer-reviewed research articles, communications, reviews, and perspectives are welcomed.