Objective: Epidermal growth factor receptor(EGFR) is overexpressed in a wide variety of solid tumors, serving as a well-characterized target for cancer imaging or therapy. In this study, we aimed to design and synthes...Objective: Epidermal growth factor receptor(EGFR) is overexpressed in a wide variety of solid tumors, serving as a well-characterized target for cancer imaging or therapy. In this study, we aimed to design and synthesize a radiotracer, ^(64) Cu-NOTA-C225, targeting EGFR for tumor positron emission tomography(PET) imaging.Methods: Cetuximab(C225) was conjugated to a bifunctional chelator, p-isothiocyanatobenzyl-1,4,7-triazacyclononane-1,4,7-triacetic acid(NOTA), and further radiolabeled with copper-^(64) for PET imaging. ^(64) CuNOTA-IgG and Cy5.5-C225 were also synthesized as control probes. A431 and A549 mouse models were established for micro-PET and/or near-infrared fluorescence(NIRF) imaging.Results: ^(64) Cu-NOTA-C225 exhibited stability in vivo and in vitro up to 24 h and 50 h post-injection,respectively. A431 tumors with average standard uptake values(SUVs) of 5.61±0.69, 6.68±1.14, 7.80±1.51 at 6, 18 and 36 h post-injection, respectively, which were significantly higher than that of moderate EGFR expressing tumors(A549), with SUVs of 0.89±0.16, 4.70±0.81, 2.01±0.50 at 6, 18 and 36 h post-injection, respectively. The expression levels of A431 and A549 were confirmed by western blotting. Additionally, the tracer uptake in A431 tumors can be blocked by unlabeled cetuximab, suggesting that tracer uptake by tumors was receptor-mediated.Furthermore, NIRF imaging using Cy5.5-C225 showed that the fluorescence intensity in tumors increased with time, with a maximal intensity of 8.17 E+10(p/s/cm^2/sr)/(μW/cm^2) at 48 h post-injection, which is consistent with the paradigm from micro-PET imaging in A431 tumor-bearing mice.Conclusions: The ^(64) Cu-NOTA-C225 PET imaging may be able to specifically and sensitively differentiate tumor models with different EGFR expression levels. It offers potentials as a PET radiotracer for imaging of tracer EGFR-positive tumors.展开更多
JS001(toripalimab)is a humanized IgG monoclonal antibody which strongly inhibits programmed cell death protein 1(PD1).In this study,we used a different iodine isotype(nat/124/125I)to label JS001 probes to target the h...JS001(toripalimab)is a humanized IgG monoclonal antibody which strongly inhibits programmed cell death protein 1(PD1).In this study,we used a different iodine isotype(nat/124/125I)to label JS001 probes to target the human PD1(hPD1)antigen.In vitro,the half maximal effective concentration(EC50)value of natI-JS001 did not significantly differ from that of JS001.The uptake of 125I-JS001 by activated T cells was 5.63 times higher than that by nonactivated T cells after 2 h of incubation.The binding affinity of 125I-JS001 to T cells of different lineages after phytohemagglutinin(PHA)stimulation reached 4.26 nmol/L.Humanized PD1 C57 BL/6 mice bearing mouse sarcoma S180 cell tumors were validated for immuno-positron emission tomography(immuno-PET)imaging.Pathological staining was used to assess the expression of PD1 in tumor tissues.The homologous 124I-human IgG(124I-hIgG)group or blocking group was used as a control group.Immuno-PET imaging showed that the uptake in the tumor area of the 124I-JS001 group at different time points was significantly higher than that of the blocking group or the 124I-hIgG group in the humanized PD1 mouse model.Taken together,these results suggest that this radiotracer has potential for noninvasive monitoring and directing tumor-specific personalized immunotherapy in PD1-positive tumors.展开更多
Despite advances in immunotherapy for the treatment of cancers,not all patients can benefit from programmed cell death ligand 1(PD-L1)immune checkpoint blockade therapy.Anti-PD-L1 therapeutic effects reportedly correl...Despite advances in immunotherapy for the treatment of cancers,not all patients can benefit from programmed cell death ligand 1(PD-L1)immune checkpoint blockade therapy.Anti-PD-L1 therapeutic effects reportedly correlate with the PD-L1 expression level;hence,accurate detection of PD-L1 expression can guide immunotherapy to achieve better therapeutic effects.Therefore,based on the high affinity antibody Nb109,a new site-specifically radiolabeled tracer,^(68)Ga-NODA-cysteine,aspartic acid,and valine(CDV)-Nb109,was designed and synthesized to accurately monitor PD-L1 expression.The tracer ^(68)Ga-NODA-CDV-Nb109 was obtained using a site-specific conjugation strategy with a radiochemical yield of about 95%and radiochemical purity of 97%.It showed high affinity for PD-L1 with a dissociation constant of 12.34±1.65 nM.Both the cell uptake assay and positron emission tomography(PET)imaging revealed higher tracer uptake in PD-L1-positive A375-hPD-L1 and U87 tumor cells than in PD-L1-negative A375 tumor cells.Meanwhile,dynamic PET imaging of a NCI-H1299 xenograft indicated that doxorubicin could upregulate PD-L1 expression,allowing timely interventional immunotherapy.In conclusion,this tracer could sensitively and dynamically monitor changes in PD-L1 expression levels in different cancers and help screen patients who can benefit from anti-PD-L1 immunotherapy.展开更多
基金supported by Natural Science Foundation of Beijing Municipality (No. 7162041)Beijing Nova program (No. Z171100001117020)+1 种基金Beijing Municipal Commission of Health and Family Planning (215 backbone program No. 2015-3-072)Beijing talent project (No. 2017000021223ZK33)
文摘Objective: Epidermal growth factor receptor(EGFR) is overexpressed in a wide variety of solid tumors, serving as a well-characterized target for cancer imaging or therapy. In this study, we aimed to design and synthesize a radiotracer, ^(64) Cu-NOTA-C225, targeting EGFR for tumor positron emission tomography(PET) imaging.Methods: Cetuximab(C225) was conjugated to a bifunctional chelator, p-isothiocyanatobenzyl-1,4,7-triazacyclononane-1,4,7-triacetic acid(NOTA), and further radiolabeled with copper-^(64) for PET imaging. ^(64) CuNOTA-IgG and Cy5.5-C225 were also synthesized as control probes. A431 and A549 mouse models were established for micro-PET and/or near-infrared fluorescence(NIRF) imaging.Results: ^(64) Cu-NOTA-C225 exhibited stability in vivo and in vitro up to 24 h and 50 h post-injection,respectively. A431 tumors with average standard uptake values(SUVs) of 5.61±0.69, 6.68±1.14, 7.80±1.51 at 6, 18 and 36 h post-injection, respectively, which were significantly higher than that of moderate EGFR expressing tumors(A549), with SUVs of 0.89±0.16, 4.70±0.81, 2.01±0.50 at 6, 18 and 36 h post-injection, respectively. The expression levels of A431 and A549 were confirmed by western blotting. Additionally, the tracer uptake in A431 tumors can be blocked by unlabeled cetuximab, suggesting that tracer uptake by tumors was receptor-mediated.Furthermore, NIRF imaging using Cy5.5-C225 showed that the fluorescence intensity in tumors increased with time, with a maximal intensity of 8.17 E+10(p/s/cm^2/sr)/(μW/cm^2) at 48 h post-injection, which is consistent with the paradigm from micro-PET imaging in A431 tumor-bearing mice.Conclusions: The ^(64) Cu-NOTA-C225 PET imaging may be able to specifically and sensitively differentiate tumor models with different EGFR expression levels. It offers potentials as a PET radiotracer for imaging of tracer EGFR-positive tumors.
基金financially supported by the National Natural Science Foundation of China(81960538,81571705,81671733,81871386,81560356,61264004 and 81871387)the Beijing Nova Program(Z171100001117020,China)+4 种基金the Beijing Excellent Talents Funding(2017000021223ZK33,China)the Beijing Municipal Science&Technology Commission(Z161100000516062,China)Open Project funded by Key laboratory of Carcinogenesis and Translational Research,Ministry of Education/Beijing(2017 open project-1 and 2019 open project-06,China)High-level Creative Talent Training Program in Guizhou Province of China(Grant No.[2015]4015)the Science and Technology Foundation of Guizhou Province(No.gzwjkj2018-1-040 and no.[2019]1201,China)
文摘JS001(toripalimab)is a humanized IgG monoclonal antibody which strongly inhibits programmed cell death protein 1(PD1).In this study,we used a different iodine isotype(nat/124/125I)to label JS001 probes to target the human PD1(hPD1)antigen.In vitro,the half maximal effective concentration(EC50)value of natI-JS001 did not significantly differ from that of JS001.The uptake of 125I-JS001 by activated T cells was 5.63 times higher than that by nonactivated T cells after 2 h of incubation.The binding affinity of 125I-JS001 to T cells of different lineages after phytohemagglutinin(PHA)stimulation reached 4.26 nmol/L.Humanized PD1 C57 BL/6 mice bearing mouse sarcoma S180 cell tumors were validated for immuno-positron emission tomography(immuno-PET)imaging.Pathological staining was used to assess the expression of PD1 in tumor tissues.The homologous 124I-human IgG(124I-hIgG)group or blocking group was used as a control group.Immuno-PET imaging showed that the uptake in the tumor area of the 124I-JS001 group at different time points was significantly higher than that of the blocking group or the 124I-hIgG group in the humanized PD1 mouse model.Taken together,these results suggest that this radiotracer has potential for noninvasive monitoring and directing tumor-specific personalized immunotherapy in PD1-positive tumors.
基金support from the National Natural Science Foundation of China(Grant No.:22076069)the Natural Science Foundation of Jiangsu Province(Grant No.:BK20201135)+1 种基金the Major Scientific Research Project of Jiangsu Commission of Health(Grant No.:ZDA2020007)the Science Technology and Development Project of Wuxi(Grant No.:Y20212013).
文摘Despite advances in immunotherapy for the treatment of cancers,not all patients can benefit from programmed cell death ligand 1(PD-L1)immune checkpoint blockade therapy.Anti-PD-L1 therapeutic effects reportedly correlate with the PD-L1 expression level;hence,accurate detection of PD-L1 expression can guide immunotherapy to achieve better therapeutic effects.Therefore,based on the high affinity antibody Nb109,a new site-specifically radiolabeled tracer,^(68)Ga-NODA-cysteine,aspartic acid,and valine(CDV)-Nb109,was designed and synthesized to accurately monitor PD-L1 expression.The tracer ^(68)Ga-NODA-CDV-Nb109 was obtained using a site-specific conjugation strategy with a radiochemical yield of about 95%and radiochemical purity of 97%.It showed high affinity for PD-L1 with a dissociation constant of 12.34±1.65 nM.Both the cell uptake assay and positron emission tomography(PET)imaging revealed higher tracer uptake in PD-L1-positive A375-hPD-L1 and U87 tumor cells than in PD-L1-negative A375 tumor cells.Meanwhile,dynamic PET imaging of a NCI-H1299 xenograft indicated that doxorubicin could upregulate PD-L1 expression,allowing timely interventional immunotherapy.In conclusion,this tracer could sensitively and dynamically monitor changes in PD-L1 expression levels in different cancers and help screen patients who can benefit from anti-PD-L1 immunotherapy.
