Mingzhang Gao

[11C]GSK2126458 and [18F]GSK2126458, the first radiosynthesis of new potential PET agents for imaging of PI3K and mTOR in cancers.

GSK2126458 is a highly potent inhibitor of phosphoinositide 3-kinase (PI3K) and mammalian target of rapamycin (mTOR) with low picomolar to subnanomolar activity. [(11)C]GSK2126458 and [(18)F]GSK2126458, new potential PET agents for imaging of PI3K and mTOR in cancer, were first designed and synthesized in 40-50% and 20-30% decay corrected radiochemical yield, and 370-740 and 37-222GBq/μmol specific activity at end of bombardment (EOB), respectively.

An improved synthesis of dopamine D2/D3 receptor radioligands [11C]fallypride and [18F]fallypride

Improved syntheses of dopamine D(2)/D(3) receptor radioligands [(11)C]Fallypride and [(18)F]Fallypride are reported. The phenolic precursor (9) for C-11 labeling and the Fallypride (10) reference standard were synthesized from the starting material 2-hydroxy-3-methoxy-5-(2-propenyl)benzoic acid methyl ester (1) in 7 and 8 steps with 16% and 5% overall chemical yields, respectively. The tosylated precursor (15) for F-18 labeling was synthesized from compound 1 in 5 steps with 32% overall chemical yield. An alternate synthetic approach for Fallypride has been developed using the same starting material 1 in 5 steps with 26% overall chemical yield. [(11)C]Fallypride ([(11)C]10) was prepared by O-[(11)C]methylation of the phenolic precursor with [(11)C]methyl triflate and purified with a semi-preparative HPLC method in 50-60% radiochemical yield, decay corrected to end of bombardment (EOB), based on [(11)C]CO(2), and 370+/-185 GBq/micromol specific radioactivity at EOB. [(18)F]Fallypride ([(18)F]10) was prepared by nucleophilic substitution of the tosylated precursor with K[(18)F]F/Kryptofix 2.2.2 and HPLC combined with solid-phase extraction (SPE) purification in variable (up to 50%) decay corrected radiochemical yield from K[(18)F]F and 111-222 GBq/micromol specific activity at EOB.

Synthesis of [11C]PBR06 and [18F]PBR06 as agents for positron emission tomographic (PET) imaging of the translocator protein (TSPO)

The translocator protein 18 kDa (TSPO) is an attractive target for molecular imaging of neuroinflammation and tumor progression. [(18)F]PBR06, a fluorine-18 labeled form of PBR06, is a promising PET TSPO radioligand originally developed at NIMH. [(11)C]PBR06, a carbon-11 labeled form of PBR06, was designed and synthesized for the first time. The standard PBR06 was synthesized from 2,5-dimethoxybenzaldehyde in three steps with 71% overall chemical yield. The radiolabeling precursor desmethyl-PBR06 was synthesized from 2-hydroxy-5-methoxybenzaldehyde in five steps with 12% overall chemical yield. The target tracer [(11)C]PBR06 was prepared by O-[(11)C]methylation of desmethyl-PBR06 with [(11)C]CH(3)OTf in CH(3)CN at 80°C under basic condition and isolated by HPLC combined with SPE purification with 40-60% decay corrected radiochemical yield and 222-740 GBq/μmol specific activity at EOB. On the similar grounds, [(18)F]PBR06 was also designed and synthesized. The previously described Br-PBR06 precursor was synthesized from 2,5-dimethoxybenzaldehyde in two steps with 78% overall chemical yield. A new radiolabeling precursor tosyloxy-PBR06, previously undescribed tosylate congener of PBR06, was designed and synthesized from ethyl 2-hydroxyacetate, 4-methylbenzene-1-sulfonyl chloride, and N-(2,5-dimethoxybenzyl)-2-phenoxyaniline in four steps with 50% overall chemical yield. [(18)F]PBR06 was prepared by the nucleophilic substitution of either new tosyloxy-PBR06 precursor or known Br-PBR06 precursor in DMSO at 140°C with K[(18)F]F/Kryptofix 2.2.2 for 15 min and HPLC combined with SPE purification in 20-60% decay corrected radiochemical yield, >99% radiochemical purity, 87-95% chemical purity, and 37-222 GBq/μmol specific activity at EOB. Radiosynthesis of [(18)F]PBR06 using new tosylated precursor gave similar radiochemical purity, and higher specific activity, radiochemical yield and chemical purity in comparison with radiosynthesis using bromine precursor.

Synthesis of [(11)C]GSK1482160 as a new PET agent for targeting P2X(7) receptor.

The authentic standards GSK1482160 and its isomer, as well as the radiolabeling precursors desmethyl-GSK1482160 and Boc-protected desmethyl-GSK1482160 were synthesized from L-pyroglutamic acid, methyl L-pyroglutamate and 2-chloro-3-(trifluoromethyl)benzylamine with overall chemical yield 27-28% in 3 steps, 58% in 4 steps, 76% in 1 step and 33% in 2 steps, respectively. [(11)C]GSK1482160 was prepared from either desmethyl-GSK1482160 or Boc-protected desmethyl-GSK1482160 with [(11)C]CH3OTf through N-[(11)C]methylation and isolated by HPLC combined with SPE in 40-50% and 30-40% radiochemical yield, respectively, based on [(11)C]CO2 and decay corrected to end of bombardment (EOB). The radiochemical purity was >99%, and the specific activity at EOB was 370-1110 GBq/μmol with a total synthesis time of ∼40-min from EOB.

Fully automated synthesis of PET TSPO radioligands [11C]DAA1106 and [18F]FEDAA1106.

[(11)C]DAA1106 was prepared by O-[(11)C]methylation of DAA1123 with [(11)C]CH(3)OTf and NaH in CH(3)CN at 80°C and isolated by HPLC combined with SPE purification in 60-70% decay corrected radiochemical yield. [(18)F]FEDAA1106 was synthesized by the nucleophilic substitution of tosyloxy-FEDAA1106 in DMSO with K[(18)F]F/Kryptofix 2.2.2 at 140°C and isolated by HPLC combined with SPE purification in 30-60% decay corrected radiochemical yield. The specific activity for [(11)C]DAA1106 and [(18)F]FEDAA1106 was 370-740GBq/μmol and 37-222GBq/μmol at EOB, respectively.

Characterization of 11C-GSK1482160 for Targeting the P2X7 Receptor as a Biomarker for Neuroinflammation

The purinergic receptor subtype 7 (P2X7R) represents a novel molecular target for imaging neuroinflammation via PET. GSK1482160, a potent P2X7R antagonist, has high receptor affinity, high blood–brain barrier penetration, and the ability to be radiolabeled with 11C. We report the initial physical and biologic characterization of this novel ligand. Methods: 11C-GSK1482160 was synthesized according to published methods. Cell density studies were performed on human embryonic kidney cell lines expressing human P2X7R (HEK293-hP2X7R) and underwent Western blotting, an immunofluorescence assay, and radioimmunohistochemistry analysis using P2X7R polyclonal antibodies. Receptor density and binding potential were determined by saturation and association–disassociation kinetics, respectively. Peak immune response to lipopolysaccharide treatment in mice was determined in time course studies and analyzed via Iba1 and P2X7R Western blotting and Iba1 immunohistochemistry. Whole-animal biodistribution studies were performed on saline- or lipopolysaccharide-treated mice at 15, 30, and 60 min after radiotracer administration. Dynamic in vivo PET/CT was performed on the mice at 72 h after administration of saline, lipopolysaccharide, or lipopolysaccharide + blocking, and 2-compartment, 5-parameter tracer kinetic modeling of brain regions was performed. Results: P2X7R changed linearly with concentrations or cell numbers. For high-specific-activity 11C-GSK1482160, receptor density and Kd were 1.15 ± 0.12 nM and 3.03 ± 0.10 pmol/mg, respectively, in HEK293-hP2X7R membranes. Association constant kon, dissociation constant koff, and binding potential (kon/koff) in HEK293-hP2X7R cells were 0.2312 ± 0.01542 min−1⋅nM−1, 0.2547 ± 0.0155 min−1, and 1.0277 ± 0.207, respectively. Whole-brain Iba1 expression in lipopolysaccharide-treated mice peaked by 72 h on immunohistochemistry, and Western blot analysis of P2X7R for saline- and lipopolysaccharide-treated brain sections showed a respective 1.8- and 1.7-fold increase in signal enhancement at 72 h. Biodistribution of 11C-GSK1482160 in saline- and lipopolysaccharide-treated mice at 72 h was statistically significant across all tissues studied. In vivo dynamic 11C-GSK1482160 PET/CT of mice at 72 h after administration of saline, lipopolysaccharide, or lipopolysaccharide + blocking showed a 3.2-fold increase and 97% blocking by 30 min. The total distribution volumes for multiple cortical regions and the hippocampus showed statistically significant increases and were blocked by an excess of authentic standard GSK1482160. Conclusion: The current study provides compelling data that support the suitability of 11C-GSK1482160 as a radioligand targeting P2X7R, a biomarker of neuroinflammation.

Fully automated synthesis and initial PET evaluation of [11C]PBR28

Fully automated synthesis and initial PET evaluation of a TSPO radioligand, [11C]PBR28 (N-(2-[11C]methoxybenzyl)-N-(4-phenoxypyridin-3-yl)acetamide), are reported. These results facilitate the potential preclinical and clinical PET studies of [11C]PBR28 in animals and humans.

Synthesis of new carbon-11-labeled carboxamide derivatives as potential PET dopamine D3 receptor radioligands

Carbon-11-labeled carboxamide derivatives, (E)-4-fluoro-N-(4-(4-(2-[(11)C]methoxyphenyl)piperazin-1-yl)butyl)cinnamoylamide, ([(11)C]3a), (E)-4-chloro-N-(4-(4-(2-[(11)C]methoxyphenyl)piperazin-1-yl)butyl)cinnamoylamide ([(11)C]3b), (E)-4-bromo-N-(4-(4-(2-[(11)C]methoxyphenyl)piperazin-1-yl)butyl)cinnamoylamide, ([(11)C]3c), (E)-4-methoxy-N-(4-(4-(2-[(11)C]methoxyphenyl)piperazin-1-yl)butyl)cinnamoylamide ([(11)C]3d), N-(4-(4-(2-[(11)C]methoxyphenyl)piperazin-1-yl)butyl)naphthyl-2-carboxamide ([(11)C]BP897, [(11)C]3e), and N-(4-(4-(2-[(11)C]methoxyphenyl)piperazin-1-yl)butyl)biphenyl-4-carboxamide ([(11)C]3f), have been synthesized as new potential PET radioligands for imaging of dopamine D(3) receptors. The target tracers were prepared by O-[(11)C]methylation of their corresponding precursors using [(11)C]CH(3)OTf and isolated by a simplified solid-phase extraction (SPE) purification procedure in 50-65% radiochemical yields decay corrected to end of bombardment (EOB), 20 min overall synthesis time, and 111-148GBq/micromol specific activity at end of synthesis (EOS).

Synthesis and in vitro biological evaluation of carbon-11-labeled quinoline derivatives as new candidate PET radioligands for cannabinoid CB2 receptor imaging

Cannabinoids have been recently proposed as a new family of potential antitumor agents, and cannabinoid receptor 2 (CB2) is believed to be over-expressed in tumor cells. This study was designed to develop new radioligands for imaging of CB2 receptor in cancer using biomedical imaging technique positron emission tomography (PET). Carbon-11-labeled 2-oxoquinoline and 2-chloroquinoline derivatives, [(11)C]6a-d and [(11)C]9a-d, were prepared by O-[(11)C]methylation of their corresponding precursors using [(11)C]CH(3)OTf under basic conditions and isolated by a simplified solid-phase extraction (SPE) method in 40-50% radiochemical yields based on [(11)C]CO(2) and decay corrected to end of bombardment (EOB). The overall synthesis time from EOB was 15-20 min, the radiochemical purity was >99%, and the specific activity at end of synthesis (EOS) was 111-185 GBq/micromol. Radioligand binding assays indicated compounds 6f, 6b, and 9f display potent in vitro binding affinities with nanomolar K(i) values and at least 100-2000-fold selectivity for CB2.

Simple synthesis of carbon-11 labeled styryl dyes as new potential PET RNA-specific, living cell imaging probes

A new type of styryl dyes have been developed as RNA-specific, live cell imaging probes for fluorescent microscopy technology to study nuclear structure and function. This study was designed to develop carbon-11 labeled styryl dyes as new probes for biomedical imaging technique positron emission tomography (PET) imaging of RNA in living cells. Precursors (E)-2-(2-(1-(triisopropylsilyl)-1H-indol-3-yl)vinyl)quinoline (2), (E)-2-(2,4,6-trimethoxystyryl)quinoline (3) and (E)-4-(2-(6-methoxyquinolin-2-yl)vinyl)-N,N-diemthylaniline (4), and standards styryl dyes E36 (6), E144 (7) and F22 (9) were synthesized in multiple steps with moderate to high chemical yields. Precursor 2 was labeled by [(11)C]CH(3)OTf, trapped on a cation-exchange CM Sep-Pak cartridge following a quick deprotecting reaction by addition of (n-Bu)(4)NF in THF, and isolated by solid-phase extraction (SPE) purification to provide target tracer [(11)C]E36 ([(11)C]6) in 40-50% radiochemical yields, decay corrected to end of bombardment (EOB), based on [(11)C]CO(2). The target tracers [(11)C]E144 ([(11)C]7) and [(11)C]F22 ([(11)C]9) were prepared by N-[(11)C]methylation of the precursors 3 and 4, respectively, using [(11)C]CH(3)OTf and isolated by SPE method in 50-70% radiochemical yields at EOB. The specific activity of the target tracers [(11)C]6, [(11)C]7 and [(11)C]9 was in a range of 74-111GBq/mumol at the end of synthesis (EOS).