scholarly journals Development of a Triazolobenzodiazepine-Based PET Probe for Subtype-Selective Vasopressin 1A Receptor Imaging

2021 ◽  
Author(s):  
Ahmed Haider ◽  
Zhiwei Xiao ◽  
Xiaotian Xia ◽  
Jiahui Chen ◽  
Richard S. Van ◽  
...  
Keyword(s):  
Pet Imaging ◽  
Chinese Hamster ◽  
Autism Spectrum ◽  
Cell Uptake ◽  
Structural Basis ◽  
Receptor Imaging ◽  
V1a Receptor ◽  
Uptake Studies ◽  
Subtype Selective

AbstractObjectivesVasopressin 1A (V1A) receptors have been linked to autism spectrum disorder, heart failure, diabetes and renal disease. Currently, there is a lack of validated probes for clinical V1A-targeted imaging and previous PET studies have primarily focused on the brain. To enable non-invasive real-time quantification of V1A receptors in peripheral organs, we sought to develop a suitable PET radioligand that would allow specific and selective V1A receptor imaging in vivo.MethodsThe previously reported triazolobenzodiazepine-based V1A antagonist, PF-184563, served as a structural basis for the development of a suitable V1A-targeted PET probe. Initially, PF-184563 and the respective desmethyl precursor for radiolabeling were synthesized via multistep organic synthesis. Inhibitory constants of PF-184563 for V1A, V1B, V2 and oxytocin (OT) receptors were assessed by competitive radioligand binding or fluorescent-based assays. Molecular docking of PF-184563 to the V1A receptor binding pocket was performed to corroborate the high binding affinity, while carbon-11 labeling was accomplished via radiomethylation. To assess the utility of the resulting PET radioligand, [11C]17, cell uptake studies were performed in a human V1A receptor Chinese hamster ovary (CHO) cell line under baseline and blockade conditions, using a series of V1A, V1B and V2 antagonists in >100-fold excess. Further, to show in vivo specificity, we conducted PET imaging and biodistribution experiments, thereby co-administering the clinical V1A-antagonist, balovaptan (3mg/kg), as a blocking agent.ResultsPF-184563 and the respective desmethyl precursor were synthesized in an overall yield of 49% (over 7 steps) and 40% (over 8 steps), respectively. A subnanomolar inhibitory constant (Ki) of 0.9 nM towards the V1A receptor was observed for PF-184563, while the triazolobenzodiazepine derivative concurrently exhibited excellent selectivity over the related V1B, V2 and OT receptor (IC50 >10,000 nM). [11C]17 was obtained in high radiochemical purity (> 99%), molar activities ranging from 37 - 46 GBq/μmol and a non-decay-corrected radiochemical yield of 8%. Cell uptake studies revealed considerable tracer binding, which was significantly reduced in the presence of V1A antagonists. Conversely, there was no significant blockade in the presence of V1B and V2 antagonists. PET imaging and biodistribution studies in CD-1 mice indicated specific tracer binding in the thyroid, pancreas, spleen and the heart.ConclusionWe report the development of a V1A-targeted PET radioligand that is suitable for subtype-selective in vitro and in vivo receptor imaging. Indeed, [11C]17 proved to specifically visualize V1A receptors in several organs including the heart, pancreas, spleen and thyroid. These results suggest that [11C]17 can be a valuable tool to study the role of V1A receptors in cardiovascular and immune-mediated pathologies.

scholarly journals DNA with zwitterionic and negatively charged phosphate modifications: Formation of DNA triplexes, duplexes and cell uptake studies

2021 ◽  
Vol 17 ◽  
pp. 749-761
Author(s):  
Yongdong Su ◽  
Maitsetseg Bayarjargal ◽  
Tracy K Hale ◽  
Vyacheslav V Filichev
Keyword(s):  
Mouse Fibroblast ◽  
Cell Uptake ◽  
Dna Duplexes ◽  
Nih3t3 Cells ◽  
Dna Triplexes ◽  
Snake Venom Phosphodiesterase ◽  
Nuclease Digestion ◽  
Dna Backbone ◽  
Uptake Studies

Two phosphate modifications were introduced into the DNA backbone using the Staudinger reaction between the 3’,5’-dinucleoside β-cyanoethyl phosphite triester formed during DNA synthesis and sulfonyl azides, 4-(azidosulfonyl)-N,N,N-trimethylbutan-1-aminium iodide (N+ azide) or p-toluenesulfonyl (tosyl or Ts) azide, to provide either a zwitterionic phosphoramidate with N+ modification or a negatively charged phosphoramidate for Ts modification in the DNA sequence. The incorporation of these N+ and Ts modifications led to the formation of thermally stable parallel DNA triplexes, regardless of the number of modifications incorporated into the oligodeoxynucleotides (ONs). For both N+ and Ts-modified ONs, the antiparallel duplexes formed with complementary RNA were more stable than those formed with complementary DNA (except for ONs with modification in the middle of the sequence). Additionally, the incorporation of N+ modifications led to the formation of duplexes with a thermal stability that was less dependent on the ionic strength than native DNA duplexes. The thermodynamic analysis of the melting curves revealed that it is the reduction in unfavourable entropy, despite the decrease in favourable enthalpy, which is responsible for the stabilisation of duplexes with N+ modification. N+ONs also demonstrated greater resistance to nuclease digestion by snake venom phosphodiesterase I than the corresponding Ts-ONs. Cell uptake studies showed that Ts-ONs can enter the nucleus of mouse fibroblast NIH3T3 cells without any transfection reagent, whereas, N+ONs remain concentrated in vesicles within the cytoplasm. These results indicate that both N+ and Ts-modified ONs are promising for various in vivo applications.

scholarly journals [18F]AZD2461, an Insight on Difference in PARP Binding Profiles for DNA Damage Response PET Imaging

2020 ◽  
Vol 22 (5) ◽  
pp. 1226-1234
Author(s):  
Florian Guibbal ◽  
Samantha L. Hopkins ◽  
Anna Pacelli ◽  
Patrick G. Isenegger ◽  
Michael Mosley ◽  
...  
Keyword(s):  
Pet Imaging ◽  
Parp Inhibitor ◽  
Parp Inhibitors ◽  
Cell Uptake ◽  
Pancreatic Cell ◽  
Anti Cancer ◽  
Damage Response ◽  
Automated Procedures

Abstract Background Poly (ADP-ribose) polymerase (PARP) inhibitors are extensively studied and used as anti-cancer drugs, as single agents or in combination with other therapies. Most radiotracers developed to date have been chosen on the basis of strong PARP1–3 affinity. Herein, we propose to study AZD2461, a PARP inhibitor with lower affinity towards PARP3, and to investigate its potential for PARP targeting in vivo. Methods Using the Cu-mediated 18F-fluorodeboronation of a carefully designed radiolabelling precursor, we accessed the 18F-labelled isotopologue of the PARP inhibitor AZD2461. Cell uptake of [18F]AZD2461 in vitro was assessed in a range of pancreatic cell lines (PSN-1, PANC-1, CFPAC-1 and AsPC-1) to assess PARP expression and in vivo in xenograft-bearing mice. Blocking experiments were performed with both olaparib and AZD2461. Results [18F]AZD2461 was efficiently radiolabelled via both manual and automated procedures (9 % ± 3 % and 3 % ± 1 % activity yields non-decay corrected). [18F]AZD2461 was taken up in vivo in PARP1-expressing tumours, and the highest uptake was observed for PSN-1 cells (7.34 ± 1.16 %ID/g). In vitro blocking experiments showed a lesser ability of olaparib to reduce [18F]AZD2461 binding, indicating a difference in selectivity between olaparib and AZD2461. Conclusion Taken together, we show the importance of screening the PARP selectivity profile of radiolabelled PARP inhibitors for use as PET imaging agents.

scholarly journals F-18 Labeled Vasoactive Intestinal Peptide Analogue in the PET Imaging of Colon Carcinoma in Nude Mice

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Dengfeng Cheng ◽  
Yuxia Liu ◽  
Hua Shen ◽  
Lifang Pang ◽  
Duanzhi Yin ◽  
...  
Keyword(s):  
Vasoactive Intestinal Peptide ◽  
Colon Carcinoma ◽  
Pet Imaging ◽  
Specific Activity ◽  
Imaging Study ◽  
Radiochemical Yield ◽  
Receptor Imaging ◽  
Improved Stability ◽  
Vip Receptor

As large amount of vasoactive intestinal peptide (VIP) receptors are expressed in various tumors and VIP-related diseases, radiolabeled VIP provides a potential PET imaging agent for VIP receptor. However, structural modification of VIP is required before being radiolabeled and used for VIP receptor imaging due to its poor in vivo stability. As a VIP analogue, [R8, 15, 21, L17]-VIP exhibited improved stability and receptor specificity in preliminary studies. In this study, F-18 labeled [R8,15,21, L17]-VIP was produced with the radiochemical yield being as high as33.6%±3%(decay-for-corrected,n=5) achieved within 100 min, a specific activity of 255 GBq/μmol, and a radiochemical purity as high as 99% as characterized by radioactive HPLC, TLC, and SDS-Page radioautography. A biodistribution study in normal mice also demonstrated fast elimination of F-18 labeled [R8,15,21, L17]-VIP in the blood, liver, and gastrointestinal tracts. A further micro-PET imaging study in C26 colon carcinoma bearing mice confirmed the high tumor specificity, with the tumor/muscle radioactivity uptake ratio being as high as 3.03 at 60 min following injection, and no apparent radioactivity concentration in the intestinal tracts. In addition, blocking experiment and Western Blot test further confirmed its potential in PET imaging of VIP receptor-positive tumor.

scholarly journals One-Step 18F-Labeling of Estradiol Derivative for PET Imaging of Breast Cancer

2018 ◽  
Vol 2018 ◽  
pp. 1-9
Author(s):  
Hongbo Huang ◽  
Ke Li ◽  
Gaochao Lv ◽  
Guiqing Liu ◽  
Xueyu Zhao ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Pet Imaging ◽  
Cell Uptake ◽  
Breast Cancer Cell Lines ◽  
Breast Cancers ◽  
Isotope Exchange Reaction ◽  
Er Positive ◽  
One Step ◽  
Mcf 7

Positron emission tomography (PET) imaging is a useful method to evaluate in situ estrogen receptor (ER) status for the early diagnosis of breast cancer and optimization of the appropriate treatment strategy. The 18F-labeled estradiol derivative has been successfully used to clinically assess the ER level of breast cancer. In order to simplify the radiosynthesis process, one-step 18F-19F isotope exchange reaction was employed for the 18F-fluorination of the tracer of [18F]AmBF3-TEG-ES. The radiotracer was obtained with the radiochemical yield (RCY) of ~61% and the radiochemical purity (RCP) of >98% within 40 min. Cell uptake and blocking assays indicated that the tracer could selectively accumulate in the ER-positive human breast cancer cell lines MCF-7 and T47D. In vivo PET imaging on the MCF-7 tumor-bearing mice showed relatively high tumor uptake (1.4~2.3 %D/g) and tumor/muscle uptake ratio (4~6). These results indicated that the tracer is a promising PET imaging agent for ER-positive breast cancers.

scholarly journals Synthesis and Evaluation of 18F-Labeled Peptide for Gonadotropin-Releasing Hormone Receptor Imaging

2019 ◽  
Vol 2019 ◽  
pp. 1-10 ◽  
Author(s):  
Shun Huang ◽  
Hongsheng Li ◽  
Yanjiang Han ◽  
Lilan Fu ◽  
Yunyan Ren ◽  
...  
Keyword(s):  
Specific Activity ◽  
Gnrh Receptor ◽  
Gonadotropin Releasing Hormone ◽  
Cell Uptake ◽  
Receptor Imaging ◽  
Tracer Injection ◽  
Releasing Hormone ◽  
Specific Accumulation ◽  
Human Reproductive ◽  
Uptake Studies

The gonadotropin-releasing hormone (GnRH) receptor is overexpressed in the majority of tumors of the human reproductive system. The purpose of this study was to develop an 18F-labeled peptide for tumor GnRH receptor imaging. In this study, the GnRH (pGlu1-His2-Trp3-Ser4-Tyr5-Gly6-Leu7-Arg8-Pro9-Gly10-NH2) peptide analogues FP-D-Lys6-GnRH (FP = 2-fluoropropanoyl) and NOTA-P-D-Lys6-GnRH (P = ethylene glycol) were designed and synthesized. The IC50 values of FP-D-Lys6-GnRH and NOTA-P-D-Lys6-GnRH were 2.0 nM and 56.2 nM, respectively. 4-Nitrophenyl-2-[18F]fluoropropionate was conjugated to the ε-amino group of the D-lysine side chain of D-Lys6-GnRH to yield the new tracer [18F]FP-D-Lys6-GnRH with a decay-corrected yield of 8 ± 3% and a specific activity of 20−100 GBq/µmol (n=6). Cell uptake studies of [18F]FP-D-Lys6-GnRH in GnRH receptor-positive PC-3 cells and GnRH receptor-negative CHO-K1 cells indicated receptor-specific accumulation. Biodistribution and PET studies in nude mice bearing PC-3 xenografted tumors showed that [18F]FP-D-Lys6-GnRH was localized in tumors with a higher uptake than in surrounding muscle and heart tissues. Furthermore, the metabolic stability of [18F]FP-D-Lys6-GnRH was determined in mouse blood and PC-3 tumor homogenates at 1 h after tracer injection. The presented results indicated a potential of the novel tracer [18F]FP-D-Lys6-GnRH for tumor GnRH receptor imaging.

scholarly journals H-CRRETAWAC-OH, a Lead Structure for the Development of Radiotracer Targeting Integrinα5β1?

2014 ◽  
Vol 2014 ◽  
pp. 1-12 ◽  
Author(s):  
Roland Haubner ◽  
Simone Maschauer ◽  
Jürgen Einsiedel ◽  
Iris E. Eder ◽  
Christine Rangger ◽  
...  
Keyword(s):  
Specific Binding ◽  
Radiochemical Yield ◽  
Human Melanoma ◽  
Cell Uptake ◽  
Preclinical Research ◽  
M Cell ◽  
Biodistribution Studies ◽  
Uptake Studies

Imaging of angiogenic processes is of great interest in preclinical research as well as in clinical settings. The most commonly addressed target structure for imaging angiogenesis is the integrinαvβ3. Here we describe the synthesis and evaluation of [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH, a radiolabelled peptide designed to selectively target the integrinα5β1. Conjugation of 4-nitrophenyl-(RS)-2-[18F]fluoropropionate provided [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH in high radiochemical purity (>95%) and a radiochemical yield of approx. 55%. In vitro evaluation showedα5β1binding affinity in the nanomolar range, whereas affinity toαvβ3andαIIbβ3was >50 μM. Cell uptake studies using human melanoma M21 (αvβ3-positive andα5β1-negative), human melanoma M21-L (αvβ3-negative andα5β1-negative), and human prostate carcinoma DU145 (αvβ3-negative andα5β1-positive) confirmed receptor-specific binding. The radiotracer was stable in human serum and showed low protein binding. Biodistribution studies showed tumour uptake ranging from 2.5 to 3.5% ID/g between 30 and 120 min post-injection. However, blocking studies and studies using mice bearingα5β1-negative M21 tumours did not confirm receptor-specific uptake of [18F]FProp-Cys*-Arg-Arg-Glu-Thr-Ala-Trp-Ala-Cys*-OH, although this radiopeptide revealed high affinity and substantial selectivity toα5β1in vitro. Further experiments are needed to study the in vivo metabolism of this peptide and to develop improved radiopeptide candidates suitable for PET imaging ofα5β1expression in vivo.

scholarly journals Development and preclinical evaluation of [68Ga]Ga-Alb-FAPtp-01, a novel tumour-associated fibroblast activation protein tracer for PET imaging of xenograft glioma

2021 ◽  
Author(s):  
Jia-Jia Lin ◽  
Chia-Pao Chuang ◽  
Jia-Yu Lin ◽  
Feng-Ting Huang ◽  
chiun-wei Huang
Keyword(s):  
Pet Imaging ◽  
Specific Binding ◽  
Fibroblast Activation Protein ◽  
Tracer Uptake ◽  
Cell Uptake ◽  
Response Monitoring ◽  
Tumour Uptake ◽  
Fibroblast Activation ◽  
Pet Tracer

Abstract Purpose Dynamic changes in tumour-associated fibroblast activation protein (FAP) expression in tumours of different stages may be helpful for prognostic evaluation and treatment response monitoring, making this protein a promising surveillance biomarker for timely diagnosis of malignant tumours and effective planning of patient care. Thus, novel FAP-PET imaging tracers were developed and evaluated for the diagnosis of xenograft glioma tumours. Methods To prospectively verify the prognostic value of the developed FAP tracers, [68Ga]Ga-FAPtp and [68Ga]Ga-Alb-FAPtp-01, measurements of FAP expression and cell uptake and specific binding assays were conducted in U87MG glioma cells. Dynamic/static PET/CT scans were acquired for tumour targeting studies in vivo and in comparison with the reference tracer [68Ga]Ga-FAPI-04 to evaluate diagnostic efficacy. Tumour autoradiography and immunohistochemistry images were acquired to confirm the tracer distribution within the tumour to determine whether it was in accordance with the pathologic data. Results Both [68Ga]Ga-FAPI-04 and [68Ga]Ga-FAPtp demonstrated marked tumour uptake and comparable pharmaceutical profiles in 1 h dynamic PET scans, and [68Ga]Ga-FAPtp had marginally favourable tumour uptake and less kidney and liver uptake. However, both tracers demonstrated rapid clearance from tumours. Thus, the optimized rationally designed FAP-targeting PET tracer [68Ga]Ga-Alb-FAPtp-01, with albumin binding capability, demonstrated prominent longitudinal tumour uptake in tumour xenografts and displayed significant tumour-to-background contrast over time. The tracer uptake values and T/M ratio were 1.775 ± 0.179 SUV and T/M = 5.9, 1.533 ± 0.222 SUV and T/M = 6.7, and 1.425 ± 0.204 SUV and T/M = 9.5, respectively, at 1 h, 2 h and 3 h. Major organs, such as the heart (0.504 ± 0.125% ID/g), muscle (0.156 ± 0.043% ID/g) and brain (0.119 ± 0.039% ID/g), all displayed comparatively low levels of tracer uptake. Conclusion Its improved tumour uptake and pharmacokinetics suggest that the [68Ga]Ga-Alb-FAPtp-01 tracer can noninvasively detect FAP activation in vivo, permitting a precise definition of its roles in tumours of different stages and yielding insights regarding novel FAP-targeted radiotherapeutic strategies at the molecular level.

scholarly journals A Radiobrominated Tyrosine Kinase Inhibitor for EGFR with L858R/T790M Mutations in Lung Carcinoma

2021 ◽  
Vol 14 (3) ◽  
pp. 256
Author(s):  
Muammar Fawwaz ◽  
Kenji Mishiro ◽  
Ryuichi Nishii ◽  
Akira Makino ◽  
Yasushi Kiyono ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Pet Imaging ◽  
Kinase Inhibitors ◽  
Kinase Inhibitor ◽  
Cell Uptake ◽  
Molecular Probe ◽  
Wild Type ◽  
Resistance Mutations ◽  
Imaging Probe

Activating double mutations L858R/T790M in the epidermal growth factor receptor (EGFR) region are often observed as the cause of resistance to tyrosine kinase inhibitors (TKIs). Third-generation EGFR-TKIs, such as osimertinib and rociletinib (CO-1686), was developed to target such resistance mutations. The detection of activating L858R/T790M mutations is necessary to select sensitive patients for therapy. Hence, we aimed to develop novel radiobromine-labeled CO-1686 as a positron emission tomography (PET) imaging probe for detecting EGFR L858R/T790M mutations. Nonradioactive brominated-CO1686 (BrCO1686) was synthesized by the condensation of N-(3-[{2-chloro-5-(trifluoromethyl)pyrimidin-4-yl}amino]-5-bromophenyl) acrylamide with the corresponding substituted 1-(4-[4-amino-3-methoxyphenyl]piperazine-1-yl)ethan-1-one. The radiobrominated [77Br]BrCO1686 was prepared through bromodestannylation of the corresponding tributylstannylated precursor with [77Br]bromide and N-chlorosuccinimide. Although we aimed to provide a novel PET imaging probe, 77Br was used as an alternative radionuclide for 76Br. We fundamentally evaluated the potency of [77Br]BrCO1686 as a molecular probe for detecting EGFR L858R/T790M using human non-small-cell lung cancer (NSCLC) cell lines: H1975 (EGFR L858R/T790M), H3255 (EGFR L858R), and H441 (wild-type EGFR). The BrCO1686 showed high cytotoxicity toward H1975 (IC50 0.18 ± 0.06 µM) comparable to that of CO-1686 (IC50 0.14 ± 0.05 µM). In cell uptake experiments, the level of accumulation of [77Br]BrCO1686 in H1975 was significantly higher than those in H3255 and H441 upon 4 h of incubation. The radioactivity of [77Br]BrCO1686 (136.3% dose/mg protein) was significantly reduced to 56.9% dose/mg protein by the pretreatment with an excess CO-1686. These results indicate that the binding site of the radiotracers should be identical to that of CO-1686. The in vivo accumulation of radioactivity of [77Br]BrCO1686 in H1975 tumor (4.51 ± 0.17) was higher than that in H441 tumor (3.71 ± 0.13) 1 h postinjection. Our results suggested that [77Br]BrCO1686 has specificity toward NSCLC cells with double mutations EGFR L858R/T790M compared to those in EGFR L858R and wild-type EGFR. However, the in vivo accumulation of radioactivity in the targeted tumor needs to be optimized by structural modification.

The Glycosaminoglycan of Recombinant Human Soluble Thrombomodulin Affects Antithrombotic Activity in a Rat Model of Tissue Factor-Induced Disseminated Intravascular Coagulation

1992 ◽  
Vol 67 (03) ◽  
pp. 366-370 ◽  
Author(s):  
Katsuhiko Nawa ◽  
Teru Itani ◽  
Mayumi Ono ◽  
Katsu-ichi Sakano ◽  
Yasumasa Marumoto ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Disseminated Intravascular Coagulation ◽  
Rat Model ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Soluble Thrombomodulin ◽  
Intravascular Coagulation ◽  
Platelet Counts ◽  
Recombinant Human Soluble Thrombomodulin

SummaryPrevious studies on recombinant human soluble thrombomodulin (rsTM) from Chinese hamster ovary cells revealed that rsTM was expressed as two proteins that differed functionally in vitro due to the presence (rsTMp) or absence (rsTMa) of chondroitin-4-sulfate. The current study evaluates the in vivo behavior of rsTM in rats and in a rat model of tissue factor-induced disseminated intravascular coagulation (DIC). rsTMp was more potent than rsTMa for prolongation of the activated partial thromboplastin time (APTT) and their in vivo half-lives determined by ELISA were 20 min for rsTMp and 5.0 h for rsTMa. Injection of a tissue factor suspension (5 mg/kg) resulted in DIC as judged by decreased platelet counts and fibrinogen concentrations, prolonged APTT, and increased fibrin and fibrinogen degradation products (FDP) levels. A bolus injection of either rsTM (0.2 mg/kg) 1 min before induction of DIC essentially neutralized effects on platelets, fibrinogen, and FDP levels, and had only a moderate effect on APTT prolongation. The dose of anticoagulant to inhibit the drop in platelet counts by 50% (ED50) was 0.2 mg/kg rsTMa, 0.07 mg/kg rsTMp, and 7 U/ kg heparin. The effect of increasing concentrations of rsTM and heparin on bleeding times were compared in experiments involving incision of the rat tail. Doubling of the bleeding times occurred at 5 mg/kg rsTMa, 3 mg/kg rsTMp or 90 U/kg heparin. These values represent a 25-fold increase over the ED50 for rsTMa, 43-fold for rsTMp and 13-fold for heparin. These results suggest that rsTMp is a potent anticoagulant to inhibit the platelet reduction when injected prior to the induction of DIC in rats.

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