A novel PET tracer for the imaging of αvß3 and αvß5 integrins in experimental breast cancer bone metastases.

The aim of this study was the evaluation of (68)Ga-DOTA-E-[c(RGDfK)](2) as a novel PET tracer to image αvß3 and αvß5 integrins. For this purpose, DOTA-E-[c(RGDfK)](2) was labeled with (68)Ga, which was obtained from a (68)Ge/(68)Ga generator, purified by solid-phase extraction and the radiochemical purity analyzed by radio-RP-HPLC. (68) Ga-DOTA-E-[c(RGDfK)](2) was obtained reproducibly in radiochemical yields of 60 ± 6% and with an excellent radiochemical purity of >99%. In nude rats bearing bone metastases after injection of MDA-MB-231 human breast cancer cells, biodistribution studies were performed to evaluate the accumulation of the radiotracer in selected organs, blood and bone metastases 0.5, 1, 2 and 3 h post injection. A rapid uptake into the bone metastases and rapid blood clearance was observed, resulting in tumor-blood ratios of up to 26.6 (3 h post injection) and tumor-muscle ratios of up to 7.9 (3 h post injection). A blocking experiment with coinjected αvß3/αvß5 antagonist showed the tumor uptake to be receptor-specific. In an initial in vivo micro PET evaluation of the tracer using the same animal model, the bone metastasis was clearly visualized. These results suggest that (68)Ga-DOTA-E-[c(RGDfK)](2) is a promising PET tracer suitable for the imaging of αvß3 and αvß5 integrins in bone metastases. This novel PET tracer should be further evaluated concerning its usefulness for early detection of bone metastases and monitoring treatment response of these lesions.

Synthesis, radiofluorination and first evaluation of [18F]fluorophenylsulfonyl- and [18F]fluorophenylsulfinyl-piperidines as serotonin 5-HT2A receptor antagonists for PET.

In psychiatric disorders, 5-HT(2A) receptors play an important role. In order to study these receptors in vivo by positron emission tomography (PET), there is an increasing interest for subtype selective and high affinity radioligands. Up to now, no optimal radiotracer is available. Thus, 1-(2,4-difluorophenethyl)-4-(4-fluorophenylsulfonyl)piperidine (9), possessing high affinity and sufficient subtype selectivity for 5-HT(2A) receptors, and 1-(2,4-difluorophenethyl)-4-(4-fluorophenylsulfinyl)piperidine (15) have been (18)F-labelled by a nucleophilic one-step reaction. Both radiotracers could be prepared and isolated within 45 min, [(18)F]9 in a radiochemical yield (RCY) of 34.5+/-8% and [(18)F]15 of 9.5+/-2.5%. The K(i) values of 9 and 15 at 5-HT(2A) receptors towards [(3)H]ketanserin were determined to be 1.9+/-0.6 and 198+/-8 nM, respectively. Autoradiography with [(18)F]9 and [(18)F]15 on rat brain sections showed a very high nonspecific binding of >80% for [(18)F]9 and 30% to 40% nonspecific binding for [(18)F]15; however, it is still too high in order to compensate for its lower affinity. Even though the affinity of 9 is more promising compared with 15, the high nonspecific binding of both radiofluorinated tracers in rat brain does not recommend those as an in vivo PET imaging agent for serotonin 5-HT(2A) receptors in humans.

Synthesis of 131I-labeled glucose-conjugated inhibitors of O6-methylguanine-DNA methyltransferase (MGMT) and comparison with nonconjugated inhibitors as potential tools for in vivo MGMT imaging.

O(6)-Substituted guanine derivatives are powerful agents used for tumor cell sensitization by inhibition of the DNA repair enzyme O(6)-methylguanine-DNA methyltransferase (MGMT). To provide targeted accumulation of MGMT inhibitors in tumor tissue as well as tools for in vivo imaging, we synthesized iodinated C(8)-alkyl-linked glucose conjugates of 2-amino-6-(5-iodothenyl)-9H-purine (O(6)-(5-iodothenyl) guanine, ITG) and 2-amino-6-(3-iodobenzyloxy)-9H-purine (O(6)-(5-iodobenzyl) guanine, IBG). These compounds have MGMT inhibitor constants (IC(50) values) of 0.8 and 0.45 microM for ITGG and IBGG, respectively, as determined in HeLa S3 cells after 2-h incubation with inhibitor. To substantiate that the (131)I-(hetero)arylmethylene group at the O(6)-position of guanine is transferred to MGMT, both the glucose conjugated inhibitors ITGG and IBGG and the corresponding nonglucose conjugated compounds ITG and IBG were labeled with iodine-131. The radioiodinations of all compounds with [(131)I]I(-) were performed with radiochemical yields of >70% for the destannylation of the corresponding tri-n-butylstannylated precursors. The binding ability of [(131)I]ITGG, [(131)]IBGG, [(131)I]ITG, and [(131)I]IBG to purified MGMT was tested. All radioactive compounds were substrates for MGMT, as demonstrated using a competitive repair assay. The newly synthesized radioactive inhibitors were utilized to study ex vivo biodistribution in mice, and the tumor-to-blood ratio of tissue uptake of [(131)I]IBG and [(131)I]IBGG was determined to be 0.24 and 0.76 after 0.5 h, respectively.

Inhibition of O6-methylguanine-DNA methyltransferase by glucose-conjugated inhibitors: comparison with nonconjugated inhibitors and effect on fotemustine and temozolomide-induced cell death.

The DNA repair protein O(6)-methylguanine-DNA methyltransferase (MGMT) is an important suicide enzyme involved in the defense against O(6)-alkylating mutagens. It also plays a role in the resistance of tumors to anticancer drugs targeting the O(6)-position of guanine, such as temozolomide and fotemustine. Several potent MGMT inhibitors have been developed sensitizing cells to O(6)-alkylating agents. Aimed at targeting MGMT inhibitors to tumor cells, we synthesized MGMT inhibitory compounds conjugated with glucose to improve uptake in tumor cells. Here, we compared O(6)-benzylguanine, O(6)-2-fluoropyridinylmethylguanine (O(6)FPG), O(6)-3-iodobenzylguanine, O(6)-4-bromothenylguanine, and O(6)-5-iodothenylguanine with the corresponding C8-linker beta-d-glucose derivatives. All glucose conjugated inhibitors were 3- to 5-fold less effective than the corresponding nonconjugated drugs as to MGMT inhibition that was measured in cell extracts (in vitro) and cultivated HeLaS3 cells (in vivo). Except for O(6)FPG, IC(50) values of the guanine derivatives applied in vitro and in vivo were correlated. A similar correlation was not obvious for the corresponding glucosides, indicating differences in cellular uptake. C8-alpha-d-glucosides were less effective than beta-glucosides. From the newly developed glucose-conjugated inhibitors tested, O(6)-4-bromothenylguanine-C8-beta-d-glucoside (O(6)BTG-C8-betaGlu) was most potent in inhibiting MGMT both in vitro and in vivo. At a concentration of 0.1 microM, it inhibited cellular MGMT to completion. It was not toxic, even when applied chronically to cells at high dose (up to 20 microM). O(6)BTG-C8-betaGlu strongly potentiated the killing effect of fotemustine and temozolomide, causing reversal from MGMT+ to MGMT- phenotype. Therefore, O(6)BTG-C8-betaGlu seems to be especially suitable for approaching MGMT inhibitor targeting in tumor therapy.