Purpose To demonstrate that positron emission tomography (PET) with fluorine 18 ((18)F) fluorthan... more Purpose To demonstrate that positron emission tomography (PET) with fluorine 18 ((18)F) fluorthanatrace (FTT) depicts activated poly (adenosine diphosphate-ribose)polymerase (PARP) expression and is feasible for clinical trial evaluation. Materials and Methods All studies were conducted prospectively from February 2012 through July 2015 under protocols approved by the local animal studies committee and institutional review board. The area under the receiver operating characteristic curve (AUC, in g/mL· min) for (18)F-FTT was assessed in normal mouse organs before and after treatment with olaparib (n = 14), a PARP inhibitor, or iniparib (n = 11), which has no PARP inhibitory activity. Murine biodistribution studies were performed to support human translational studies. Eight human subjects with cancer and eight healthy volunteers underwent imaging to verify the human radiation dosimetry of (18)F-FTT. The Wilcoxon signed rank test was used to assess for differences among treatment gro...
WC 44 and WC 10 are phenylpiperazines with low (23 fold) to moderate (42 fold) selectivity for do... more WC 44 and WC 10 are phenylpiperazines with low (23 fold) to moderate (42 fold) selectivity for dopamine D3 receptors (D3Rs) over D2Rs, respectively. WC 44 is a full D3R agonist in the forskolin-stimulated adenylyl cyclase (AC) assay, whereas WC 10 has little efficacy. In contrast to their opposite effects in the AC assay, these drugs often produce similar behavioral effects, suggesting that the AC assay does not predict the efficacy of these drugs in vivo. Here, we examined whether Fos protein expression induced by these drugs would be more consistent with their behavioral effects in vivo. Rats received either vehicle, WC 10 (5.6 mg/kg, i.p.), WC 44 (10.0 mg/kg, i.p), cocaine (10.0 mg/kg, i.p.), or cocaine with WC 10 (5.6 mg/kg, i.p.) or with WC 44 (10.0 mg/kg, i.p). Locomotion was monitored for 90 min and the brains were harvested for immunohistochemistry. Both WC 10 and WC 44 decreased spontaneous and cocaine-induced locomotion. Both compounds also increased Fos expression relative to saline in the dorsal striatum and nucleus accumbens core and shell, and relative to cocaine alone in the nucleus accumbens shell. The findings suggest that even though these compounds have different efficacy in the AC bioassy, they produce similar brain activation and attenuation of cocaine hyperlocomotion. Together with our previous research demonstrating that these compounds down-shift the cocaine self-administration dose-effect function, the findings support the idea that D3R-selective compounds may be useful for cocaine dependence medications development.
Several receptor mediated pathways have been shown to modulate the murine head twitch response (H... more Several receptor mediated pathways have been shown to modulate the murine head twitch response (HTR). However, the role of sigma receptors in the murine (±)-2,5-dimethoxy-4-iodoamphetamine (DOI)-induced HTR has not been previously investigated. We examined the ability of LS-1-137, a novel sigma-1 vs. sigma-2 receptor selective phenylacetamide, to modulate the DOI-induced HTR in DBA/2J mice. We also assessed the in vivo efficacy of reference sigma-1 receptor antagonists and agonists PRE-084 and PPCC. The effect of the sigma-2 receptor selective antagonist RHM-1-86 was also examined. Rotarod analysis was performed to monitor motor coordination after LS-1-137 administration. Radioligand binding techniques were used to determine the affinity of LS-1-137 at 5-HT2A and 5-HT2C receptors. LS-1-137 and the sigma-1 receptor antagonists haloperidol and BD 1047 were able to attenuate a DOI-induced HTR, indicating that LS-1-137 was acting in vivo as a sigma-1 receptor antagonist. LS-1-137 did no...
Microglial activation is commonly identified by elevated levels of the 18 kDa translocator protei... more Microglial activation is commonly identified by elevated levels of the 18 kDa translocator protein (TSPO) in response to several inflammatory processes. [(11)C]PBR28 is one of the most promising PET tracers to image TSPO in both human and non-human primates. In this study, we optimized the radiolabeling procedure of [(11)C]PBR28 for higher radiochemical yield, radiochemical purity, and specific activity, which can be easily translated to any automated module for clinical trials. Time-activity curves (TACs) derived from the dynamic PET imaging of male rhesus monkey brains demonstrated that [(11)C]PBR28 had suitable kinetics with radiotracer accumulation observed in the caudate, putamen, cerebellum, and frontal cortex region.
Cancer-selective drug delivery is an important concept in improving treatment while minimizing of... more Cancer-selective drug delivery is an important concept in improving treatment while minimizing off-site toxicities, and sigma-2 receptors, which are overexpressed in solid tumors, represent attractive pharmacologic targets. Select sigma-2 ligands have been shown to be rapidly internalized selectively into cancer cells while retaining the capacity to deliver small molecules as drug cargoes. We utilized the sigma-2-based drug delivery concept to convert Erastin, a clinically underperforming drug, into a potent pancreatic cancer therapeutic. The Erastin derivative des-methyl Erastin (dm-Erastin) was chemically linked to sigma-2 ligand SV119 to create SW V-49. Conjugation increased the killing capacity of dm-Erastin by nearly 35-fold in vitro and reduced the size of established tumors and doubled the median survival in syngeneic and patient-derived xenograft models when compared to non-targeted dm-Erastin. Mechanistic analyses demonstrated that cell death was associated with robust reactive oxygen species production and could be efficiently antagonized with antioxidants. Mass spectrometry was employed to demonstrate selective uptake into pancreatic cancer cells. Thus, targeted delivery of dm-Erastin via conjugation to the sigma-2 ligand SV119 produced efficient tumor control and prolonged animal survival with minimal off-target toxicities, and SW V-49 represents a promising new therapeutic with the potential to advance the fight against pancreatic cancer.
American journal of nuclear medicine and molecular imaging, 2016
The nuclear enzyme PARP1 plays a central role in sensing DNA damage and facilitating repair. Tumo... more The nuclear enzyme PARP1 plays a central role in sensing DNA damage and facilitating repair. Tumors with BRCA1/2 mutations are highly dependent on PARP1 as an alternative mechanism for DNA repair, and PARP inhibitors generate synthetic lethality in tumors with BRCA mutations, resulting in cell cycle arrest and apoptosis. Zhou et al. recently synthesized an (18)F-labeled PARP1 inhibitor ([(18)F]FluorThanatrace) for PET, and demonstrated high specific tracer uptake in a xenograft model of breast cancer [1]. In the current study, we characterize the level of baseline PARP expression and activity across multiple human breast cancer cell lines, including a BRCA1 mutant line. PARP expression and activity, as measured by levels of PAR and PARP1, is correlated with in vitro [(18)F]FluorThanatrace binding as well as tracer uptake on PET in a xenograft model of breast cancer. Radiotracer uptake in genetically-engineered mouse fibroblasts indicates [(18)F]FluorThanatrace is selective for PARP1...
The sigma-2 receptor is a protein with a Heme binding region and is capable of receptor-mediated ... more The sigma-2 receptor is a protein with a Heme binding region and is capable of receptor-mediated endocytosis. It is overexpressed in many cancers making it a potential vector for therapeutic drug delivery. Our objective was to introduce an alpha-emitting radionuclide, astatine-211, into a selective sigma-2 ligand moiety to provide cytotoxic capabilities without adversely altering the pharmacological characteristics. In this study we investigated the in vitro/in vivo tumor targeting and estimated dosimetry of alpha-emitting sigma-2 ligand, 5-(astato-(211)At)-N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide ((211)At-MM3), in a pre-clinical human breast cancer model. Astatine-211 was produced in a cyclotron and isolated by dry distillation. Radiosynthesis of (211)At-MM3 was performed using a tin precursor through radioastatodestannylation. In vitro sigma-2 binding experiments using (211)At-MM3 were carried out in live EMT6 and MDA-MB-231 breast cancer cells and liver homogenate tissue. In vivo biodistribution experiments were performed using EMT6 mouse breast cancer cells in BALB/c female mice. Approximately 370kBq of (211)At-MM3 was administered intravenously and at time points of 5min, 1, 2, 4, 8, and 24h organs/tissue were harvested. Estimated human dosimetry was extrapolated from biodistribution data using OLINDA/EXM (VU e-Innovations). Astatine-211 was successfully produced and isolated in quantities suitable for in vitro and small animal in vivo experiments. Radiosynthesis of (211)At-MM3 was reproducible with high radiochemical purity. Astatine-211-MM3 exhibited picomolar affinity to the sigma-2 receptor in contrast to the iodinated analog that had nanomolar affinity. Prolonged tumor targeting was measured through biodistribution studies with a maximal tumor to muscle ratio of 9.02 at 4h. Estimated human dosimetry revealed doses of up to 370MBq in an adult female patient were below organ radiation limits with the potential to provide a high therapeutic dose to tumors. The sigma-2 receptor could serve as a suitable targeting platform for designing radiotherapeutics. (211)At-MM3 showed tumor targeting properties in vitro/in vivo and favorable estimated human dosimetry establishing the proof of concept for future development as a radiotherapeutic for the treatment of breast cancer.
A series of N-substituted 9-azabicyclo[3.3.1]nonan-3α-yl phenylcarbamate analogs were synthesized... more A series of N-substituted 9-azabicyclo[3.3.1]nonan-3α-yl phenylcarbamate analogs were synthesized. Among them, WC-26 and WC-59 were identified as the most potent σ 2 receptor ligands (K i = 2.58 and 0.82 nM, respectively) with high selectivity against σ 1 (K i of σ 1 /σ 2 ratio = 557 and 2,087, respectively). [ 18 F]WC-59 was radiolabeled via a nucleophilic substitution of a mesylate precursor by [ 18 F]fluoride, and in vitro direct binding studies of [ 18 F]WC-59 were conducted using membrane preparations from murine EMT-6 solid breast tumors. The results indicate that [ 18 F] WC-59 binds specifically to σ 2 receptors in vitro (K d = ~2 nM). Biodistribution studies of [ 18 F] WC-59 in EMT-6 tumor-bearing mice indicated that the tracer was a less suitable candidate for clinical imaging studies than existing F-18 labeled σ 2 receptor ligands. The ability of WC-26 to enhance the cytotoxic effects of the chemotherapy drug, doxorubicin, was evaluated in cell culture using the mouse breast tumor EMT-6 and the human tumor MDA-MB435. WC-26 greatly increased the ability of doxorubicin to kill these two tumor cell lines in vitro. These results indicate that WC-26 is potentially a useful chemosensitizer for the treatment of cancer when combined with conventional chemotherapeutics.
a b s t r a c t 2-((4-(1-[ 11 C]Methyl-4-(pyridin-4-yl)-1H-pyrazol-3-yl)phenoxy)methyl)-quinoline... more a b s t r a c t 2-((4-(1-[ 11 C]Methyl-4-(pyridin-4-yl)-1H-pyrazol-3-yl)phenoxy)methyl)-quinoline (MP-10), a specific PDE10A inhibitor (IC 50 = 0.18 nM with 100-fold selectivity over other PDEs), was radiosynthesized by alkylation of the desmethyl precursor with [ 11 C]CH 3 I, $45% yield, >92% radiochemical purity, >370 GBq/lmol specific activity at end of bombardment (EOB). Evaluation in Sprague-Dawley rats revealed that [ 11 C]MP-10 had highest brain accumulation in the PDE10A enriched-striatum, the 30 min striatum: cerebellum ratio reached 6.55. MicroPET studies of [ 11 C]MP-10 in monkeys displayed selective uptake in striatum. However, a radiolabeled metabolite capable of penetrating the blood-brain-barrier may limit the clinical utility of [ 11 C]MP-10 as a PDE10A PET tracer.
Introduction-Four benzamide analogs having a high affinity and selectivity for D 3 versus D 2 rec... more Introduction-Four benzamide analogs having a high affinity and selectivity for D 3 versus D 2 receptors were radiolabeled with 11 C or 18 F for in vivo evaluation.
The s 2 -receptor has been shown to be upregulated in proliferating tumors cells. The purpose of ... more The s 2 -receptor has been shown to be upregulated in proliferating tumors cells. The purpose of this study was to compare 39-deoxy-39-18 F-fluorothymidine ( 18 F-FLT) and 2 new 76 Br-radiolabeled compounds that have a high affinity and selectivity for the s 2 -receptor. These are 5-bromo-N-(4-(3,4-dihydro-6,7dimethoxyisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide (compound (1)) and 5-bromo-N-(2-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)ethyl)-2-methoxybenzamide (compound (2)). Methods: Two s 2 -receptor2binding ligands were prepared, from the corresponding tributylstannyl precursors using standard electrophilic chemistry, 76 Br-compound (1) ( 76 Br-1) and 76 Br-compound (2) ( 76 Br-2). 18 F-FLT, 76 Br-1, and 76 Br-2 were compared using allograft tumors of the EMT-6 cell line (mouse mammary adenocarcinoma) in biodistribution studies at 5 min, 0.5, 1, and 2 h. Imaging of 76 Br-1 and 18 F-FLT was also performed at 2 and 1 h, respectively. Results: 76 Br-1 and 76 Br-2 were synthesized with yields between 50% and 70% with high specific activity. Both compounds showed uptake into the tumor with tumor-to-normal tissue ratios of 76 Br-1 being greater than both 76 Br-2 and 18 F-FLT. Except for the liver and kidney, all ratios were greater than 1 and uptake into the tumor was shown with microPET imaging for 76 Br-1. Conclusion: We were able to synthesize two 76 Br-radiolabeled compounds with a high yield and specific activity that target the s 2 receptor with high affinity and selectivity. The studies presented show that both of the flexible benzamide compounds can identify EMT-6 breast tumors in vivo. 76 Br-1 also has higher tumor-to-normal tissue ratios when compared with 76 Br-2 and 18 F-FLT. The high affinity and low nonspecific binding of 76 Br-1 indicates that it can be a potential PET radiotracer for imaging solid tumors.
Sigma-2 receptors represent an endogenous marker for proliferation in solid tumors. The high affi... more Sigma-2 receptors represent an endogenous marker for proliferation in solid tumors. The high affinity, high selectivity s 2 receptor ligand N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2- was separately radiolabeled with F-18 and I-125. The radiolabeling yield was 30% and 70% for [ 18 F]3 and [ 125 I]3, respectively. Studies of [ 125 I]3 using murine 66 breast tumor membrane homogenates and evaluation of [ 18 F]3 and [ 125 I]3 in 66 tumorbearing mice indicate that this ligand has potential as a PET or a SPECT probe for imaging s 2 receptors in breast cancer.
A series of fluorine-containing benzamide analogs was synthesized and evaluated as candidate liga... more A series of fluorine-containing benzamide analogs was synthesized and evaluated as candidate ligands for positron emission tomography (PET) imaging of the sigma-2 (sigma2) receptor status of solid tumors. Four compounds having a moderate to high affinity for sigma2 receptors and a moderate to low affinity for sigma-1 (sigma1) receptors were radiolabeled with fluorine-18 via displacement of the corresponding mesylate precursor with [18F]fluoride. Biodistribution studies in female Balb/c mice bearing EMT-6 tumor allografts demonstrated that all four F-18-labeled compounds had a high tumor uptake (2.5-3.7% ID/g) and acceptable tumor/normal tissue ratios at 1 and 2 h post-i.v. injection. An analysis of the chemistry and biodistribution data suggested that N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2-(2-[18F]-fluoroethoxy)-5-methylbenzamide ([18F]3c) and N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2-(2-[18F]-fluoroethoxy)-5-iodo-3-methoxybenzamide ([18F]3f) are acceptable compounds for imaging the sigma2 receptor status of solid tumors.
The regulation of D₃ receptor has not been well documented in diffuse Lewy body disease (DLBD). I... more The regulation of D₃ receptor has not been well documented in diffuse Lewy body disease (DLBD). In this study, a novel D₃-preferring radioligand [(3)H]WC-10 and a D₂-preferring radioligand [(3)H]raclopride were used and the absolute densities of the dopamine D₃ and D₂ receptors were determined in the striatal regions and substantia nigra (SN) from postmortem brains from five cases of DLBD, which included dementia with Lewy bodies (DLB, n=4) and Parkinson disease dementia (PDD, n=1). The densities of the dopamine D₁ receptor, vesicular monoamine transporter 2 (VMAT2), and dopamine transporter (DAT) were also measured by quantitative autoradiography using [(3)H]SCH23390, [(3)H]dihydrotetrabenazine, and [(3)H]WIN35428, respectively. The densities of these dopaminergic markers were also measured in the same brain regions in 10 age-matched control cases. Dopamine D₃ receptor density was significantly increased in the striatal regions including caudate, putamen and nucleus accumbens (NAc)...
We synthesized and characterized two novel fluorescent sigma-2 receptor selective ligands, SW120 ... more We synthesized and characterized two novel fluorescent sigma-2 receptor selective ligands, SW120 and SW116, and evaluated these ligands as potential probes for imaging cell proliferation. Both ligands are highly selective for sigma-2 receptors versus sigma-1 receptors. SW120 and SW116 were internalized into MDA-MB-435 cells, and 50% of the maximum fluorescent intensity was reached in 11 and 24 minutes, respectively. In vitro studies showed that 50% of SW120 or SW116 washed out of cells in 1 hour. The internalization of SW120 was reduced ≈30% by phenylarsine oxide, an inhibitor of endocytosis, suggesting that sigma-2 ligands are internalized, in part, by an endocytotic pathway. Subcellular localization studies using confocal and two-photon microscopy showed that SW120 and SW116 partially colocalized with fluorescent markers of mitochondria, endoplasmic reticulum, lysosomes, and the plasma membrane, suggesting that sigma-2 receptors localized to the cytoplasmic organelles and plasma m...
Journal of nuclear medicine : official publication, Society of Nuclear Medicine, 2006
The sigma(2)-receptor has been shown to be upregulated in proliferating tumors cells. The purpose... more The sigma(2)-receptor has been shown to be upregulated in proliferating tumors cells. The purpose of this study was to compare 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) and 2 new (76)Br-radiolabeled compounds that have a high affinity and selectivity for the sigma(2)-receptor. These are 5-bromo-N-(4-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide (compound (1)) and 5-bromo-N-(2-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)ethyl)-2-methoxybenzamide (compound (2)). Two sigma(2)-receptor-binding ligands were prepared, from the corresponding tributylstannyl precursors using standard electrophilic chemistry, (76)Br-compound (1) ((76)Br-1) and (76)Br-compound (2) ((76)Br-2). (18)F-FLT, (76)Br-1, and (76)Br-2 were compared using allograft tumors of the EMT-6 cell line (mouse mammary adenocarcinoma) in biodistribution studies at 5 min, 0.5, 1, and 2 h. Imaging of (76)Br-1 and (18)F-FLT was also performed at 2 and 1 h, respectively. (76)Br-1 and (76...
Cancer stem cells (CSCs) are believed to be responsible for the long-term growth of a tumor, as w... more Cancer stem cells (CSCs) are believed to be responsible for the long-term growth of a tumor, as well as its metastasis and recurrence after conventional therapies. Here, it is demonstrated that the sigma-2 receptor is overexpressed on the surface of breast CSCs, and thus could serve as a biomarker for the purpose of targeting. Breast CSCs are targeted with Au nanocages (AuNCs) by functionalizing their surfaces with SV119, a synthetic small molecule capable of binding to the sigma-2 receptor with high specificity. The interiors of the AuNCs could also be loaded with an anticancer drug to be selectively delivered to breast CSCs and released in a controllable fashion. The results demonstrate that the SV119-AuNC conjugate can serve as a new platform to carry out photothermal and chemo therapies simultaneously, eradicating breast CSCs more effectively through a synergetic effect.
Purpose To demonstrate that positron emission tomography (PET) with fluorine 18 ((18)F) fluorthan... more Purpose To demonstrate that positron emission tomography (PET) with fluorine 18 ((18)F) fluorthanatrace (FTT) depicts activated poly (adenosine diphosphate-ribose)polymerase (PARP) expression and is feasible for clinical trial evaluation. Materials and Methods All studies were conducted prospectively from February 2012 through July 2015 under protocols approved by the local animal studies committee and institutional review board. The area under the receiver operating characteristic curve (AUC, in g/mL· min) for (18)F-FTT was assessed in normal mouse organs before and after treatment with olaparib (n = 14), a PARP inhibitor, or iniparib (n = 11), which has no PARP inhibitory activity. Murine biodistribution studies were performed to support human translational studies. Eight human subjects with cancer and eight healthy volunteers underwent imaging to verify the human radiation dosimetry of (18)F-FTT. The Wilcoxon signed rank test was used to assess for differences among treatment gro...
WC 44 and WC 10 are phenylpiperazines with low (23 fold) to moderate (42 fold) selectivity for do... more WC 44 and WC 10 are phenylpiperazines with low (23 fold) to moderate (42 fold) selectivity for dopamine D3 receptors (D3Rs) over D2Rs, respectively. WC 44 is a full D3R agonist in the forskolin-stimulated adenylyl cyclase (AC) assay, whereas WC 10 has little efficacy. In contrast to their opposite effects in the AC assay, these drugs often produce similar behavioral effects, suggesting that the AC assay does not predict the efficacy of these drugs in vivo. Here, we examined whether Fos protein expression induced by these drugs would be more consistent with their behavioral effects in vivo. Rats received either vehicle, WC 10 (5.6 mg/kg, i.p.), WC 44 (10.0 mg/kg, i.p), cocaine (10.0 mg/kg, i.p.), or cocaine with WC 10 (5.6 mg/kg, i.p.) or with WC 44 (10.0 mg/kg, i.p). Locomotion was monitored for 90 min and the brains were harvested for immunohistochemistry. Both WC 10 and WC 44 decreased spontaneous and cocaine-induced locomotion. Both compounds also increased Fos expression relative to saline in the dorsal striatum and nucleus accumbens core and shell, and relative to cocaine alone in the nucleus accumbens shell. The findings suggest that even though these compounds have different efficacy in the AC bioassy, they produce similar brain activation and attenuation of cocaine hyperlocomotion. Together with our previous research demonstrating that these compounds down-shift the cocaine self-administration dose-effect function, the findings support the idea that D3R-selective compounds may be useful for cocaine dependence medications development.
Several receptor mediated pathways have been shown to modulate the murine head twitch response (H... more Several receptor mediated pathways have been shown to modulate the murine head twitch response (HTR). However, the role of sigma receptors in the murine (±)-2,5-dimethoxy-4-iodoamphetamine (DOI)-induced HTR has not been previously investigated. We examined the ability of LS-1-137, a novel sigma-1 vs. sigma-2 receptor selective phenylacetamide, to modulate the DOI-induced HTR in DBA/2J mice. We also assessed the in vivo efficacy of reference sigma-1 receptor antagonists and agonists PRE-084 and PPCC. The effect of the sigma-2 receptor selective antagonist RHM-1-86 was also examined. Rotarod analysis was performed to monitor motor coordination after LS-1-137 administration. Radioligand binding techniques were used to determine the affinity of LS-1-137 at 5-HT2A and 5-HT2C receptors. LS-1-137 and the sigma-1 receptor antagonists haloperidol and BD 1047 were able to attenuate a DOI-induced HTR, indicating that LS-1-137 was acting in vivo as a sigma-1 receptor antagonist. LS-1-137 did no...
Microglial activation is commonly identified by elevated levels of the 18 kDa translocator protei... more Microglial activation is commonly identified by elevated levels of the 18 kDa translocator protein (TSPO) in response to several inflammatory processes. [(11)C]PBR28 is one of the most promising PET tracers to image TSPO in both human and non-human primates. In this study, we optimized the radiolabeling procedure of [(11)C]PBR28 for higher radiochemical yield, radiochemical purity, and specific activity, which can be easily translated to any automated module for clinical trials. Time-activity curves (TACs) derived from the dynamic PET imaging of male rhesus monkey brains demonstrated that [(11)C]PBR28 had suitable kinetics with radiotracer accumulation observed in the caudate, putamen, cerebellum, and frontal cortex region.
Cancer-selective drug delivery is an important concept in improving treatment while minimizing of... more Cancer-selective drug delivery is an important concept in improving treatment while minimizing off-site toxicities, and sigma-2 receptors, which are overexpressed in solid tumors, represent attractive pharmacologic targets. Select sigma-2 ligands have been shown to be rapidly internalized selectively into cancer cells while retaining the capacity to deliver small molecules as drug cargoes. We utilized the sigma-2-based drug delivery concept to convert Erastin, a clinically underperforming drug, into a potent pancreatic cancer therapeutic. The Erastin derivative des-methyl Erastin (dm-Erastin) was chemically linked to sigma-2 ligand SV119 to create SW V-49. Conjugation increased the killing capacity of dm-Erastin by nearly 35-fold in vitro and reduced the size of established tumors and doubled the median survival in syngeneic and patient-derived xenograft models when compared to non-targeted dm-Erastin. Mechanistic analyses demonstrated that cell death was associated with robust reactive oxygen species production and could be efficiently antagonized with antioxidants. Mass spectrometry was employed to demonstrate selective uptake into pancreatic cancer cells. Thus, targeted delivery of dm-Erastin via conjugation to the sigma-2 ligand SV119 produced efficient tumor control and prolonged animal survival with minimal off-target toxicities, and SW V-49 represents a promising new therapeutic with the potential to advance the fight against pancreatic cancer.
American journal of nuclear medicine and molecular imaging, 2016
The nuclear enzyme PARP1 plays a central role in sensing DNA damage and facilitating repair. Tumo... more The nuclear enzyme PARP1 plays a central role in sensing DNA damage and facilitating repair. Tumors with BRCA1/2 mutations are highly dependent on PARP1 as an alternative mechanism for DNA repair, and PARP inhibitors generate synthetic lethality in tumors with BRCA mutations, resulting in cell cycle arrest and apoptosis. Zhou et al. recently synthesized an (18)F-labeled PARP1 inhibitor ([(18)F]FluorThanatrace) for PET, and demonstrated high specific tracer uptake in a xenograft model of breast cancer [1]. In the current study, we characterize the level of baseline PARP expression and activity across multiple human breast cancer cell lines, including a BRCA1 mutant line. PARP expression and activity, as measured by levels of PAR and PARP1, is correlated with in vitro [(18)F]FluorThanatrace binding as well as tracer uptake on PET in a xenograft model of breast cancer. Radiotracer uptake in genetically-engineered mouse fibroblasts indicates [(18)F]FluorThanatrace is selective for PARP1...
The sigma-2 receptor is a protein with a Heme binding region and is capable of receptor-mediated ... more The sigma-2 receptor is a protein with a Heme binding region and is capable of receptor-mediated endocytosis. It is overexpressed in many cancers making it a potential vector for therapeutic drug delivery. Our objective was to introduce an alpha-emitting radionuclide, astatine-211, into a selective sigma-2 ligand moiety to provide cytotoxic capabilities without adversely altering the pharmacological characteristics. In this study we investigated the in vitro/in vivo tumor targeting and estimated dosimetry of alpha-emitting sigma-2 ligand, 5-(astato-(211)At)-N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide ((211)At-MM3), in a pre-clinical human breast cancer model. Astatine-211 was produced in a cyclotron and isolated by dry distillation. Radiosynthesis of (211)At-MM3 was performed using a tin precursor through radioastatodestannylation. In vitro sigma-2 binding experiments using (211)At-MM3 were carried out in live EMT6 and MDA-MB-231 breast cancer cells and liver homogenate tissue. In vivo biodistribution experiments were performed using EMT6 mouse breast cancer cells in BALB/c female mice. Approximately 370kBq of (211)At-MM3 was administered intravenously and at time points of 5min, 1, 2, 4, 8, and 24h organs/tissue were harvested. Estimated human dosimetry was extrapolated from biodistribution data using OLINDA/EXM (VU e-Innovations). Astatine-211 was successfully produced and isolated in quantities suitable for in vitro and small animal in vivo experiments. Radiosynthesis of (211)At-MM3 was reproducible with high radiochemical purity. Astatine-211-MM3 exhibited picomolar affinity to the sigma-2 receptor in contrast to the iodinated analog that had nanomolar affinity. Prolonged tumor targeting was measured through biodistribution studies with a maximal tumor to muscle ratio of 9.02 at 4h. Estimated human dosimetry revealed doses of up to 370MBq in an adult female patient were below organ radiation limits with the potential to provide a high therapeutic dose to tumors. The sigma-2 receptor could serve as a suitable targeting platform for designing radiotherapeutics. (211)At-MM3 showed tumor targeting properties in vitro/in vivo and favorable estimated human dosimetry establishing the proof of concept for future development as a radiotherapeutic for the treatment of breast cancer.
A series of N-substituted 9-azabicyclo[3.3.1]nonan-3α-yl phenylcarbamate analogs were synthesized... more A series of N-substituted 9-azabicyclo[3.3.1]nonan-3α-yl phenylcarbamate analogs were synthesized. Among them, WC-26 and WC-59 were identified as the most potent σ 2 receptor ligands (K i = 2.58 and 0.82 nM, respectively) with high selectivity against σ 1 (K i of σ 1 /σ 2 ratio = 557 and 2,087, respectively). [ 18 F]WC-59 was radiolabeled via a nucleophilic substitution of a mesylate precursor by [ 18 F]fluoride, and in vitro direct binding studies of [ 18 F]WC-59 were conducted using membrane preparations from murine EMT-6 solid breast tumors. The results indicate that [ 18 F] WC-59 binds specifically to σ 2 receptors in vitro (K d = ~2 nM). Biodistribution studies of [ 18 F] WC-59 in EMT-6 tumor-bearing mice indicated that the tracer was a less suitable candidate for clinical imaging studies than existing F-18 labeled σ 2 receptor ligands. The ability of WC-26 to enhance the cytotoxic effects of the chemotherapy drug, doxorubicin, was evaluated in cell culture using the mouse breast tumor EMT-6 and the human tumor MDA-MB435. WC-26 greatly increased the ability of doxorubicin to kill these two tumor cell lines in vitro. These results indicate that WC-26 is potentially a useful chemosensitizer for the treatment of cancer when combined with conventional chemotherapeutics.
a b s t r a c t 2-((4-(1-[ 11 C]Methyl-4-(pyridin-4-yl)-1H-pyrazol-3-yl)phenoxy)methyl)-quinoline... more a b s t r a c t 2-((4-(1-[ 11 C]Methyl-4-(pyridin-4-yl)-1H-pyrazol-3-yl)phenoxy)methyl)-quinoline (MP-10), a specific PDE10A inhibitor (IC 50 = 0.18 nM with 100-fold selectivity over other PDEs), was radiosynthesized by alkylation of the desmethyl precursor with [ 11 C]CH 3 I, $45% yield, >92% radiochemical purity, >370 GBq/lmol specific activity at end of bombardment (EOB). Evaluation in Sprague-Dawley rats revealed that [ 11 C]MP-10 had highest brain accumulation in the PDE10A enriched-striatum, the 30 min striatum: cerebellum ratio reached 6.55. MicroPET studies of [ 11 C]MP-10 in monkeys displayed selective uptake in striatum. However, a radiolabeled metabolite capable of penetrating the blood-brain-barrier may limit the clinical utility of [ 11 C]MP-10 as a PDE10A PET tracer.
Introduction-Four benzamide analogs having a high affinity and selectivity for D 3 versus D 2 rec... more Introduction-Four benzamide analogs having a high affinity and selectivity for D 3 versus D 2 receptors were radiolabeled with 11 C or 18 F for in vivo evaluation.
The s 2 -receptor has been shown to be upregulated in proliferating tumors cells. The purpose of ... more The s 2 -receptor has been shown to be upregulated in proliferating tumors cells. The purpose of this study was to compare 39-deoxy-39-18 F-fluorothymidine ( 18 F-FLT) and 2 new 76 Br-radiolabeled compounds that have a high affinity and selectivity for the s 2 -receptor. These are 5-bromo-N-(4-(3,4-dihydro-6,7dimethoxyisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide (compound (1)) and 5-bromo-N-(2-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)ethyl)-2-methoxybenzamide (compound (2)). Methods: Two s 2 -receptor2binding ligands were prepared, from the corresponding tributylstannyl precursors using standard electrophilic chemistry, 76 Br-compound (1) ( 76 Br-1) and 76 Br-compound (2) ( 76 Br-2). 18 F-FLT, 76 Br-1, and 76 Br-2 were compared using allograft tumors of the EMT-6 cell line (mouse mammary adenocarcinoma) in biodistribution studies at 5 min, 0.5, 1, and 2 h. Imaging of 76 Br-1 and 18 F-FLT was also performed at 2 and 1 h, respectively. Results: 76 Br-1 and 76 Br-2 were synthesized with yields between 50% and 70% with high specific activity. Both compounds showed uptake into the tumor with tumor-to-normal tissue ratios of 76 Br-1 being greater than both 76 Br-2 and 18 F-FLT. Except for the liver and kidney, all ratios were greater than 1 and uptake into the tumor was shown with microPET imaging for 76 Br-1. Conclusion: We were able to synthesize two 76 Br-radiolabeled compounds with a high yield and specific activity that target the s 2 receptor with high affinity and selectivity. The studies presented show that both of the flexible benzamide compounds can identify EMT-6 breast tumors in vivo. 76 Br-1 also has higher tumor-to-normal tissue ratios when compared with 76 Br-2 and 18 F-FLT. The high affinity and low nonspecific binding of 76 Br-1 indicates that it can be a potential PET radiotracer for imaging solid tumors.
Sigma-2 receptors represent an endogenous marker for proliferation in solid tumors. The high affi... more Sigma-2 receptors represent an endogenous marker for proliferation in solid tumors. The high affinity, high selectivity s 2 receptor ligand N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2- was separately radiolabeled with F-18 and I-125. The radiolabeling yield was 30% and 70% for [ 18 F]3 and [ 125 I]3, respectively. Studies of [ 125 I]3 using murine 66 breast tumor membrane homogenates and evaluation of [ 18 F]3 and [ 125 I]3 in 66 tumorbearing mice indicate that this ligand has potential as a PET or a SPECT probe for imaging s 2 receptors in breast cancer.
A series of fluorine-containing benzamide analogs was synthesized and evaluated as candidate liga... more A series of fluorine-containing benzamide analogs was synthesized and evaluated as candidate ligands for positron emission tomography (PET) imaging of the sigma-2 (sigma2) receptor status of solid tumors. Four compounds having a moderate to high affinity for sigma2 receptors and a moderate to low affinity for sigma-1 (sigma1) receptors were radiolabeled with fluorine-18 via displacement of the corresponding mesylate precursor with [18F]fluoride. Biodistribution studies in female Balb/c mice bearing EMT-6 tumor allografts demonstrated that all four F-18-labeled compounds had a high tumor uptake (2.5-3.7% ID/g) and acceptable tumor/normal tissue ratios at 1 and 2 h post-i.v. injection. An analysis of the chemistry and biodistribution data suggested that N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2-(2-[18F]-fluoroethoxy)-5-methylbenzamide ([18F]3c) and N-(4-(6,7-dimethoxy-3,4-dihydroisoquinolin-2(1H)-yl)butyl)-2-(2-[18F]-fluoroethoxy)-5-iodo-3-methoxybenzamide ([18F]3f) are acceptable compounds for imaging the sigma2 receptor status of solid tumors.
The regulation of D₃ receptor has not been well documented in diffuse Lewy body disease (DLBD). I... more The regulation of D₃ receptor has not been well documented in diffuse Lewy body disease (DLBD). In this study, a novel D₃-preferring radioligand [(3)H]WC-10 and a D₂-preferring radioligand [(3)H]raclopride were used and the absolute densities of the dopamine D₃ and D₂ receptors were determined in the striatal regions and substantia nigra (SN) from postmortem brains from five cases of DLBD, which included dementia with Lewy bodies (DLB, n=4) and Parkinson disease dementia (PDD, n=1). The densities of the dopamine D₁ receptor, vesicular monoamine transporter 2 (VMAT2), and dopamine transporter (DAT) were also measured by quantitative autoradiography using [(3)H]SCH23390, [(3)H]dihydrotetrabenazine, and [(3)H]WIN35428, respectively. The densities of these dopaminergic markers were also measured in the same brain regions in 10 age-matched control cases. Dopamine D₃ receptor density was significantly increased in the striatal regions including caudate, putamen and nucleus accumbens (NAc)...
We synthesized and characterized two novel fluorescent sigma-2 receptor selective ligands, SW120 ... more We synthesized and characterized two novel fluorescent sigma-2 receptor selective ligands, SW120 and SW116, and evaluated these ligands as potential probes for imaging cell proliferation. Both ligands are highly selective for sigma-2 receptors versus sigma-1 receptors. SW120 and SW116 were internalized into MDA-MB-435 cells, and 50% of the maximum fluorescent intensity was reached in 11 and 24 minutes, respectively. In vitro studies showed that 50% of SW120 or SW116 washed out of cells in 1 hour. The internalization of SW120 was reduced ≈30% by phenylarsine oxide, an inhibitor of endocytosis, suggesting that sigma-2 ligands are internalized, in part, by an endocytotic pathway. Subcellular localization studies using confocal and two-photon microscopy showed that SW120 and SW116 partially colocalized with fluorescent markers of mitochondria, endoplasmic reticulum, lysosomes, and the plasma membrane, suggesting that sigma-2 receptors localized to the cytoplasmic organelles and plasma m...
Journal of nuclear medicine : official publication, Society of Nuclear Medicine, 2006
The sigma(2)-receptor has been shown to be upregulated in proliferating tumors cells. The purpose... more The sigma(2)-receptor has been shown to be upregulated in proliferating tumors cells. The purpose of this study was to compare 3'-deoxy-3'-(18)F-fluorothymidine ((18)F-FLT) and 2 new (76)Br-radiolabeled compounds that have a high affinity and selectivity for the sigma(2)-receptor. These are 5-bromo-N-(4-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)butyl)-2,3-dimethoxybenzamide (compound (1)) and 5-bromo-N-(2-(3,4-dihydro-6,7-dimethoxyisoquinolin-2(1H)-yl)ethyl)-2-methoxybenzamide (compound (2)). Two sigma(2)-receptor-binding ligands were prepared, from the corresponding tributylstannyl precursors using standard electrophilic chemistry, (76)Br-compound (1) ((76)Br-1) and (76)Br-compound (2) ((76)Br-2). (18)F-FLT, (76)Br-1, and (76)Br-2 were compared using allograft tumors of the EMT-6 cell line (mouse mammary adenocarcinoma) in biodistribution studies at 5 min, 0.5, 1, and 2 h. Imaging of (76)Br-1 and (18)F-FLT was also performed at 2 and 1 h, respectively. (76)Br-1 and (76...
Cancer stem cells (CSCs) are believed to be responsible for the long-term growth of a tumor, as w... more Cancer stem cells (CSCs) are believed to be responsible for the long-term growth of a tumor, as well as its metastasis and recurrence after conventional therapies. Here, it is demonstrated that the sigma-2 receptor is overexpressed on the surface of breast CSCs, and thus could serve as a biomarker for the purpose of targeting. Breast CSCs are targeted with Au nanocages (AuNCs) by functionalizing their surfaces with SV119, a synthetic small molecule capable of binding to the sigma-2 receptor with high specificity. The interiors of the AuNCs could also be loaded with an anticancer drug to be selectively delivered to breast CSCs and released in a controllable fashion. The results demonstrate that the SV119-AuNC conjugate can serve as a new platform to carry out photothermal and chemo therapies simultaneously, eradicating breast CSCs more effectively through a synergetic effect.
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Papers by Robert Mach