PIP Tumor Research Articles

Albumin-Binding PSMA Ligands: Optimization of the Tissue Distribution Profile.

The prostate-specific membrane antigen (PSMA) has emerged as an attractive prostate cancer associated target for radiotheragnostic application using PSMA-specific radioligands. The aim of this study was to design new PSMA ligands modified with an albumin-binding moiety in order to optimize their tissue distribution profile. The compounds were prepared by conjugation of a urea-based PSMA-binding entity, a DOTA chelator, and 4-( p-iodophenyl)butyric acid using multistep solid phase synthesis. The three ligands (PSMA-ALB-02, PSMA-ALB-05, and PSMA-ALB-07) were designed with varying linker entities. Radiolabeling with 177Lu was performed at a specific activity of up to 50 MBq/nmol resulting in radioligands of >98% radiochemical purity and high stability. In vitro investigations revealed high binding of all three PSMA radioligands to mouse (>64%) and human plasma proteins (>94%). Uptake and internalization into PSMA-positive PC-3 PIP tumor cells was equally high for all radioligands. Negligible accumulation was found in PSMA-negative PC-3 flu cells, indicating PSMA-specific binding of all radioligands. Biodistribution and imaging studies performed in PC-3 PIP/flu tumor-bearing mice showed enhanced blood circulation of the new radioligands when compared to the clinically employed 177Lu-PSMA-617. The PC-3 PIP tumor uptake of all three radioligands was very high (76.4 ± 2.5% IA/g, 79.4 ± 11.1% IA/g, and 84.6 ± 14.2% IA/g, respectively) at 24 h post injection (p.i.) resulting in tumor-to-blood ratios of ∼176, ∼48, and ∼107, respectively, whereas uptake into PC-3 flu tumors was negligible. Kidney uptake at 24 h p.i. was lowest for 177Lu-PSMA-ALB-02 (10.7 ± 0.92% IA/g), while 177Lu-PSMA-ALB-05 and 177Lu-PSMA-ALB-07 showed higher renal retention (23.9 ± 4.02% IA/g and 51.9 ± 6.34% IA/g, respectively). Tumor-to-background ratios calculated from values of the area under the curve (AUC) of time-dependent biodistribution data were in favor of 177Lu-PSMA-ALB-02 (tumor-to-blood, 46; tumor-to-kidney, 5.9) when compared to 177Lu-PSMA-ALB-05 (17 and 3.7, respectively) and 177Lu-PSMA-ALB-07 (39 and 2.1, respectively). The high accumulation of the radioligands in PC-3 PIP tumors was visualized on SPECT/CT images demonstrating increasing tumor-to-kidney ratios over time. Taking all of the characteristics into account, 177Lu-PSMA-ALB-02 emerged as the most promising candidate. The applied concept may be attractive for future clinical translation potentially enabling more potent and convenient prostate cancer radionuclide therapy.

44Sc-PSMA-617 for radiotheragnostics in tandem with 177Lu-PSMA-617\u2014preclinical investigations in comparison with 68Ga-PSMA-11 and 68Ga-PSMA-617

BackgroundThe targeting of the prostate-specific membrane antigen (PSMA) is of particular interest for radiotheragnostic purposes of prostate cancer. Radiolabeled PSMA-617, a 1,4,7,10-tetraazacyclododecane-N,N′,N′′,N′′′-tetraacetic acid (DOTA)-functionalized PSMA ligand, revealed favorable kinetics with high tumor uptake, enabling its successful application for PET imaging (68Ga) and radionuclide therapy (177Lu) in the clinics. In this study, PSMA-617 was labeled with cyclotron-produced 44Sc (T1/2 = 4.04 h) and investigated preclinically for its use as a diagnostic match to 177Lu-PSMA-617.Results44Sc was produced at the research cyclotron at PSI by irradiation of enriched 44Ca targets, followed by chromatographic separation. 44Sc-PSMA-617 was prepared under standard labeling conditions at elevated temperature resulting in a radiochemical purity of >97% at a specific activity of up to 10 MBq/nmol. 44Sc-PSMA-617 was evaluated in vitro and compared to the 177Lu- and 68Ga-labeled match, as well as 68Ga-PSMA-11 using PSMA-positive PC-3 PIP and PSMA-negative PC-3 flu prostate cancer cells. In these experiments it revealed similar in vitro properties to that of 177Lu- and 68Ga-labeled PSMA-617. Moreover, 44Sc-PSMA-617 bound specifically to PSMA-expressing PC-3 PIP tumor cells, while unspecific binding to PC-3 flu cells was not observed. The radioligands were investigated with regard to their in vivo properties in PC-3 PIP/flu tumor-bearing mice. 44Sc-PSMA-617 showed high tumor uptake and a fast renal excretion. The overall tissue distribution of 44Sc-PSMA-617 resembled that of 177Lu-PSMA-617 most closely, while the 68Ga-labeled ligands, in particular 68Ga-PSMA-11, showed different distribution kinetics. 44Sc-PSMA-617 enabled distinct visualization of PC-3 PIP tumor xenografts shortly after injection, with increasing tumor-to-background contrast over time while unspecific uptake in the PC-3 flu tumors was not observed.ConclusionsThe in vitro characteristics and in vivo kinetics of 44Sc-PSMA-617 were more similar to 177Lu-PSMA-617 than to 68Ga-PSMA-617 and 68Ga-PSMA-11. Due to the almost four-fold longer half-life of 44Sc as compared to 68Ga, a centralized production of 44Sc-PSMA-617 and transport to satellite PET centers would be feasible. These features make 44Sc-PSMA-617 particularly appealing for clinical application.

111In- and IRDye800CW-Labeled PLA-PEG Nanoparticle for Imaging Prostate-Specific Membrane Antigen-Expressing Tissues.

Targeted delivery of drug-encapsulated nanoparticles is a promising new approach to safe and effective therapeutics for cancer. Here we investigate the pharmacokinetics and biodistribution of a prostate-specific membrane antigen (PSMA)-targeted nanoparticle based on a poly(lactic acid)-polyethylene glycol copolymer by utilizing single photon emission computed tomography (SPECT) and fluorescence imaging of a low-molecular-weight, PSMA-targeting moiety attached to the surface and oriented toward the outside environment. Tissue biodistribution of the radioactive, PSMA-targeted nanoparticles in mice containing PSMA(+) PC3 PIP and PSMA(-) PC3 flu (control) tumors demonstrated similar accumulation compared to the untargeted particles within all tissues except for the tumor and liver by 96 h postinjection. For PSMA(+) PC3 PIP tumor, the targeted nanoparticle demonstrated retention of 6.58% injected dose (ID)/g at 48 h and remained nearly at that level out to 96 h, whereas the untargeted nanoparticle showed a 48 h retention of 8.17% ID/g followed by a significant clearance to 2.37% ID/g at 96 h (P < 0.02). On the other hand, for control tumor, both targeted and untargeted particles displayed similar 48 h retentions and rates of clearance over 96 h. Ex vivo microscopic analysis with near-infrared versions of the nanoparticles indicated retention within PSMA(+) tumor epithelial cells as well as tumor-associated macrophages for targeted particles and primarily macrophage-associated uptake for the untargeted particles. Retention in control tumor was primarily associated with tumor vasculature and macrophages. The data demonstrate the utility of radioimaging to assess nanoparticle biodistribution and suggest that active targeting has a modest positive effect on tumor localization of PSMA-targeted PLA-PEG nanoparticles that have been derivatized for imaging.

(18)F]Fluoroethyl Triazole Substituted PSMA Inhibitor Exhibiting Rapid Normal Organ Clearance.

Prostate-specific membrane antigen (PSMA) is overexpressed in the epithelium of prostate cancer and nonprostate solid tumor neovasculature. PSMA is increasingly utilized as a target for cancer imaging and therapy. Here, we report the synthesis and in vivo biodistribution of a low-molecular-weight PSMA-based imaging agent, 2-[3-(1-carboxy-5-{3-[1-(2-[(18)F]fluoroethyl)-1H-1,2,3-triazol-yl]propanamido}pentyl)ureido]pentanedioic acid ([(18)F]YC-88), containing an [(18)F]fluoroethyl triazole moiety. [(18)F]YC-88 was synthesized from 2-[(18)F]fluoroethyl azide and the corresponding alkyne precursor in two steps using either a one- or two-pot procedure. Biodistribution and positron emission tomography (PET) imaging were performed in immunocompromised mice using isogenic PSMA(+) PC3 PIP and PSMA(-) PC3 flu xenografts. YC-88 exhibited high affinity for PSMA as evidenced by a Ki value of 12.9 nM. The non-decay corrected radiochemical yields of [(18)F]YC-88 averaged 14 ± 1% (n = 5). Specific radioactivities ranged from 320 to 2,460 Ci/mmol (12-91 GBq/μmol) with an average of 940 Ci/mmol (35 GBq/μmol, n = 5). In an immunocompromised mouse model, [(18)F]YC-88 clearly delineated PSMA(+) PC3 PIP prostate tumor xenografts on imaging with PET. At 1 h postinjection, 47.58 ± 5.19% injected dose per gram of tissue (% ID/g) was evident within the PSMA(+) PC3 PIP tumor, with a ratio of 170:1 of uptake within PSMA(+) PC3 PIP to PSMA(-) PC3 flu tumor placed in the opposite flank. The tumor-to-kidney ratio at 2 h postinjection was 4:1. At or after 30 min postinjection, minimal nontarget tissue uptake of [(18)F]YC-88 was observed. Compared to [(18)F]DCFPyL, which is currently in clinical trials, the uptake of [(18)F]YC-88 within the kidney, liver, and spleen was significantly lower at all time-points studied. At 30 min and 1 h postinjection, salivary gland uptake of [(18)F]YC-88 was significantly less than that of [(18)F]DCFPyL. [(18)F]YC-88 is a new PSMA-targeted PET agent synthesized utilizing click chemistry that demonstrates high PSMA(+) tumor uptake in a xenograft model. Because of its low uptake in the kidney, rapid clearance from nontarget organs, and relatively simple one-pot, two-step radiosynthesis, [(18)F]YC-88 is a viable new PET radiotracer for imaging PSMA-expressing lesions.

(2S)-2-(3-(1-Carboxy-5-(4-211At-Astatobenzamido)Pentyl)Ureido)-Pentanedioic Acid for PSMA-Targeted α-Particle Radiopharmaceutical Therapy.

PSMA-targeted (2S)-2-(3-(1-carboxy-5-(4-211At-astatobenzamido)pentyl)ureido)-pentanedioic acid (211At- 6: ) was synthesized. Cellular uptake and clonogenic survival were tested in PSMA-positive (PSMA+) PC3 PIP and PSMA-negative (PSMA-) PC3 flu human PC cells after 211At- 6: treatment. The antitumor efficacy of 211At- 6: was evaluated in mice bearing PSMA+ PC3 PIP and PSMA- PC3 flu flank xenografts at a 740-kBq dose and in mice bearing PSMA+, luciferase-expressing PC3-ML micrometastases. Biodistribution was determined in mice bearing PSMA+ PC3 PIP and PSMA- PC3 flu flank xenografts. Suborgan distribution was evaluated using α-camera images, and microscale dosimetry was modeled. Long-term toxicity was assessed in mice for 12 mo. 211At- 6: treatment resulted in PSMA-specific cellular uptake and decreased clonogenic survival in PSMA+ PC3 PIP cells and caused significant tumor growth delay in PSMA+ PC3 PIP flank tumors. Significantly improved survival was achieved in the newly developed PSMA+ micrometastatic PC model. Biodistribution showed uptake of 211At- 6: in PSMA+ PC3 PIP tumors and in kidneys. Microscale kidney dosimetry based on α-camera images and a nephron model revealed hot spots in the proximal renal tubules. Long-term toxicity studies confirmed that the dose-limiting toxicity was late radiation nephropathy. PSMA-targeted 211At- 6: α-particle radiotherapy yielded significantly improved survival in mice bearing PC micrometastases after systemic administration. 211At- 6: also showed uptake in renal proximal tubules resulting in late nephrotoxicity, highlighting the importance of long-term toxicity studies and microscale dosimetry.

(18)F]Fluorobenzoyllysinepentanedioic Acid Carbamates: New Scaffolds for Positron Emission Tomography (PET) Imaging of Prostate-Specific Membrane Antigen (PSMA).

Radiolabeled urea-based low-molecular weight inhibitors of the prostate-specific membrane antigen (PSMA) are under intense investigation as imaging and therapeutic agents for prostate and other cancers. In an effort to provide agents with less nontarget organ uptake than the ureas, we synthesized four (18)F-labeled inhibitors of PSMA based on carbamate scaffolds. 4-Bromo-2-[(18)F]fluorobenzoyllysineoxypentanedioic acid (OPA) carbamate [(18)F]23 and 4-iodo-2-[(18)F]fluorobenzoyllysine OPA carbamate [(18)F]24 in particular exhibited high target-selective uptake in PSMA+ PC3 PIP tumor xenografts, with tumor-to-kidney ratios of >1 by 4 h postinjection, an important benchmark. Because of its high tumor uptake (90% injected dose per gram of tissue at 2 h postinjection) and high tumor-to-organ ratios, [(18)F]23 is promising for clinical translation. Prolonged tumor-specific uptake demonstrated by [(18)F]24, which did not reach equilibrium during the 4 h study period, suggests carbamates as alternative scaffolds for mitigating dose to nontarget tissues.

Auger Radiopharmaceutical Therapy Targeting Prostate-Specific Membrane Antigen.

Auger electron emitters such as (125)I have a high linear energy transfer and short range of emission (<10 μm), making them suitable for treating micrometastases while sparing normal tissues. We used a highly specific small molecule targeting the prostate-specific membrane antigen (PSMA) to deliver (125)I to prostate cancer cells. The PSMA-targeting Auger emitter 2-[3-[1-carboxy-5-(4-(125)I-iodo-benzoylamino)-pentyl]-ureido]-pentanedioic acid ((125)I-DCIBzL) was synthesized. DNA damage (via phosphorylated H2A histone family member X staining) and clonogenic survival were tested in PSMA-positive (PSMA+) PC3 PIP and PSMA-negative (PSMA-) PC3 flu human prostate cancer cells after treatment with (125)I-DCIBzL. Subcellular drug distribution was assessed with confocal microscopy using a related fluorescent PSMA-targeting compound YC-36. In vivo antitumor efficacy was tested in nude mice bearing PSMA+ PC3 PIP or PSMA- PC3 flu flank xenografts. Animals were administered (intravenously) 111 MBq (3 mCi) of (125)I-DCIBzL, 111 MBq (3 mCi) of (125)I-NaI, an equivalent amount of nonradiolabeled DCIBzL, or saline. After treatment with (125)I-DCIBzL, PSMA+ PC3 PIP cells exhibited increased DNA damage and decreased clonogenic survival when compared with PSMA- PC3 flu cells. Confocal microscopy of YC-36 showed drug distribution in the perinuclear area and plasma membrane. Animals bearing PSMA+ PC3 PIP tumors had significant tumor growth delay after treatment with (125)I-DCIBzL, with only 1 mouse reaching 5 times the initial tumor volume by 60 d after treatment, compared with a median time to 5 times volume of less than 15 d for PSMA- PC3 flu tumors and all other treatment groups (P = 0.002 by log-rank test). PSMA-targeted radiopharmaceutical therapy with the Auger emitter (125)I-DCIBzL yielded highly specific antitumor efficacy in vivo, suggesting promise for treatment of prostate cancer micrometastases.

Preclinical evaluation of 86Y-labeled inhibitors of prostate-specific membrane antigen for dosimetry estimates.

(86)Y (half-life = 14.74 h, 33% β(+)) is within an emerging class of positron-emitting isotopes with relatively long physical half-lives that enables extended imaging of biologic processes. We report the synthesis and evaluation of 3 low-molecular-weight compounds labeled with (86)Y for imaging the prostate-specific membrane antigen (PSMA) using PET. Impetus for the study derives from the need to perform dosimetry estimates for the corresponding (90)Y-labeled radiotherapeutics. Multistep syntheses were used in preparing (86)Y- 4: - 6: PSMA inhibition constants were evaluated by competitive binding assay. In vivo characterization using tumor-bearing male mice was performed by PET/CT for (86)Y- 4: - 6: and by biodistribution studies of (86)Y- 4: and (86)Y- 6: out to 24 h after injection. Quantitative whole-body PET scans were recorded to measure the kinetics for 14 organs in a male baboon using (86)Y- 6 RESULTS: Compounds (86)Y- 4: - 6: were obtained in high radiochemical yield and purity, with specific radioactivities of more than 83.92 GBq/μmol. PET imaging and biodistribution studies using PSMA-positive PC-3 PIP and PSMA-negative PC-3 flu tumor-bearing mice revealed that (86)Y- 4-6: had high site-specific uptake in PSMA-positive PC-3 PIP tumor starting at 20 min after injection and remained high at 24 h. Compound (86)Y- 6: demonstrated the highest tumor uptake and retention, with 32.17 ± 7.99 and 15.79 ± 6.44 percentage injected dose per gram (%ID/g) at 5 and 24 h, respectively. Low activity concentrations were associated with blood and normal organs, except for the kidneys, a PSMA-expressing tissue. PET imaging in baboons reveals that all organs have a 2-phase (rapid and slow) clearance, with the highest uptake (8 %ID/g) in the kidneys at 25 min. The individual absolute uptake kinetics were used to calculate radiation doses using the OLINDA/EXM software. The highest mean absorbed dose was received by the renal cortex, with 1.9 mGy per MBq of (86)Y- 6: Compound (86)Y- 6: is a promising candidate for quantitative PET imaging of PSMA-expressing tumors. Dosimetry calculations indicate promise for future (90)Y or other radiometals that could use a similar chelator/scaffold combination for radiopharmaceutical therapy based on the structure of 6.

Abstract 4345: PSMA-targeted theranostic nanoplex combining TRAIL gene cDNA and prodrug enzyme delivery for prostate cancer treatment.

Abstract Prostate cancer (PCa) is the second leading cause of death from cancer in men in the U.S, and there is a compelling need for the development of effective treatments for metastatic PCa. Theranostic approaches that combine detection with treatment hold significant promise for cancer-cell-specific treatments especially with molecular reagents such as cDNA or siRNA that can increase or decrease the expression of genes of interest. We have reported on a prostrate specific membrane antigen (PSMA)-based platform to deliver a prodrug enzyme and small interfering RNA (siRNA) to downregulate gene expression for theranostic imaging of metastatic PCa[1]. Here we expanded this platform for gene delivery and expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) cDNA. PSMA, which is highly expressed by castration-resistant PCa, was used for PCa-specific delivery and localization of the nanoplex that delivered TRAIL cDNA and a prodrug enzyme, bacterial cytosine deaminase (bCD). TRAIL has been reported to specifically kill malignant cells but to be relatively nontoxic to normal cells. The GFP-TRAIL cDNA used expresses a GFP-TRAIL fusion protein that can be detected with optical imaging to evaluate cDNA expression. The prodrug enzyme bCD converts the non-toxic prodrug 5-fluorocytosine (5-FC) to the active cytotoxic drug 5-fluorouracil (5-FU) that can be monitored by 19F MRS. Our prototype nanoplex was synthesized by conjugating: (i) a low molecular weight urea-based PSMA targeting moiety (2-(3-[1-carboxy-5-[7-(2,5-dioxo-pyrrolidin-1-yloxycarbonyl)-heptanoylamino]-pentyl]-ureido)-pentanedioic acid, (ii) the prodrug-activating enzyme bCD that converts nontoxic 5-FC to cytotoxic 5-FU, (iii) the near-infrared fluorescent probe Cy5.5 labeled linker poly-L-lysine (PLL), and (iv) the GFP-TRAIL pDNA delivery vector: PEI (polyethyleneimine)-PEG (polythethyeneglycol) co-grafted-polymer. Imaging studies with PSMA targeted nanoplexes were performed with PC-3 human prostate cancer cells genetically engineered to overexpress PSMA (PC3-PIP); non-PSMA expressing PC-3 cells (PC3-Flu) were used as controls. Cell imaging studies indicated higher expression of GFP-TRAIL fusion protein in PC3-PIP cells. In vivo images obtained from PIP and Flu tumors demonstrated increased accumulation of the nanoplex in PIP tumors. In ex vivo imaging studies, increased uptake of the nanoplex as detected by Cy5.5 fluorescence, and higher expression of GFP in PIP compared to Flu tumors was observed. 19F MRS indicated the prodrug enzyme bCD efficiently converted the prodrug 5-FC to 5-FU at 24 h. Tumor growth studies showed increased inhibition of PIP tumor growth. Collectively, these data demonstrate the ability to express a gene of interest using a theranostic nanoplex specifically targeting cancer cells. Citation Format: Zhihang Chen, Marie-France Penet Penet, Balaji Krishnamachary, Cong Li, Paul T. Winnard, Martin G. Pomper, Zaver M. Bhujwalla. PSMA-targeted theranostic nanoplex combining TRAIL gene cDNA and prodrug enzyme delivery for prostate cancer treatment. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4345. doi:10.1158/1538-7445.AM2013-4345

A Modular Strategy to Prepare Multivalent Inhibitors of Prostate-Specific Membrane Antigen (PSMA)

We have developed a modular scaffold for preparing high-affinity, homo-multivalent inhibitors of the prostate-specific membrane antigen (PSMA) for imaging and therapy of prostate cancer (PCa). Our system contains a lysine-based (µ-, e-) dialkyne residue for incorporating a PSMA binding Lys-Glu urea motif exploiting click chemistry and a second lysine residue for subsequent modification with an imaging or therapeutic moiety. The utility of the multivalent scaffold was examined by synthesizing bivalent compounds 2 and 3 and comparing them with the monovalent analog 1. Determination of inhibition constants (Ki) revealed that bivalent 2 (0.2 nM) and 3 (0.08 nM) are significantly more potent (~ 5 fold and ~ 11 fold, respectively) inhibitors of PSMA than monovalent 1 (0.9 nM). A single photon emission computed tomography (SPECT)-CT imaging study of [111In]3 demonstrated high and specific uptake in PSMA+ PC-3 PIP tumor until at least 48 h post-injection, with rapid clearance from non-target tissues, including kidney. A biodistribution study revealed that [111In]3 demonstrated 34.0 ± 7.5 percent injected dose per gram of tissue in PSMA+ tumor at 24 h post-injection and was capable of generating target-to-non-target ratios of ~ 379 in PSMA+ PC-3 PIP tumors vs. isogenic PSMA-negative PC3-flu tumors in vivo. The click chemistry approach affords a convenient strategy toward multivalent PSMA inhibitors of enhanced affinity and superior pharmacokinetics for imaging.

2-(3-{1-Carboxy-5-[(6-[18F]Fluoro-Pyridine-3-Carbonyl)-Amino]-Pentyl}-Ureido)-Pentanedioic Acid, [18F]DCFPyL, a PSMA-Based PET Imaging Agent for Prostate Cancer

We have synthesized and evaluated in vivo 2-(3-{1-carboxy-5-[(6-[(18)F]fluoro-pyridine-3-carbonyl)-amino]-pentyl}-ureido)-pentanedioic acid, [(18)F]DCFPyL, as a potential imaging agent for the prostate-specific membrane antigen (PSMA). PSMA is upregulated in prostate cancer epithelia and in the neovasculature of most solid tumors. [(18)F]DCFPyL was synthesized in two steps from the p-methoxybenzyl (PMB) protected lys-C(O)-glu urea precursor using 6-[(18)F]fluoronicotinic acid tetrafluorophenyl ester ([(18)F]F-Py-TFP) for introduction of (18)F. Radiochemical synthesis was followed by biodistribution and imaging with PET in immunocompromised mice using isogenic PSMA PC3 PIP and PSMA- PC3 flu xenograft models. Human radiation dosimetry estimates were calculated using OLINDA/EXM 1.0. DCFPyL displays a K(i) value of 1.1 ± 0.1 nmol/L for PSMA. [(18)F]DCFPyL was produced in radiochemical yields of 36%-53% (decay corrected) and specific radioactivities of 340-480 Ci/mmol (12.6-17.8 GBq/μmol, n = 3). In an immunocompromised mouse model [(18)F]DCFPyL clearly delineated PSMA+ PC3 PIP prostate tumor xenografts on imaging with PET. At 2 hours postinjection, 39.4 ± 5.4 percent injected dose per gram of tissue (%ID/g) was evident within the PSMA+ PC3 PIP tumor, with a ratio of 358:1 of uptake within PSMA+ PC3 PIP to PSMA- PC3 flu tumor placed in the opposite flank. At or after 1 hour postinjection, minimal nontarget tissue uptake of [(18)F]DCFPyL was observed. The bladder wall is the dose-limiting organ. These data suggest [(18)F]DCFPyL as a viable, new positron-emitting imaging agent for PSMA-expressing tissues.

68Ga-Labeled Inhibitors of Prostate-Specific Membrane Antigen (PSMA) for Imaging Prostate Cancer

Gallium-68 is a generator-produced radionuclide for positron emission tomography (PET) that is being increasingly used for radiolabeling of tumor-targeting peptides. Compounds [(68)Ga]3 and [(68)Ga]6 are high-affinity urea-based inhibitors of the prostate-specific membrane antigen (PSMA) that were synthesized in decay-uncorrected yields ranging from 60% to 70% and radiochemical purities of more than 99%. Compound [(68)Ga]3 demonstrated 3.78 +/- 0.90% injected dose per gram of tissue (%ID/g) within PSMA+ PIP tumor at 30 min postinjection, while [(68)Ga]6 showed a 2 h PSMA+ PIP tumor uptake value of 3.29 +/- 0.77 %ID/g. Target (PSMA+ PIP) to nontarget (PSMA- flu) ratios were 4.6 and 18.3, respectively, at those time points. Both compounds delineated tumor clearly by small animal PET. The urea series of imaging agents for PSMA can be radiolabeled with (68)Ga, a cyclotron-free isotope useful for clinical PET studies, with maintenance of target specificity.

Radiohalogenated Prostate-Specific Membrane Antigen (PSMA)-Based Ureas as Imaging Agents for Prostate Cancer

To extend our development of new imaging agents targeting the prostate-specific membrane antigen (PSMA), we have used the versatile intermediate 2-[3-(5-amino-1-carboxy-pentyl)-ureido]-pentanedioic acid (Lys-C(O)-Glu), which allows ready incorporation of radiohalogens for single photon emission computed tomography (SPECT) and positron emission tomography (PET). We prepared 2-[3-[1-carboxy-5-(4-[(125)I]iodo-benzoylamino)-pentyl]-ureido]-pentanedioic acid ([(125)I]3), 2-[3-[1-carboxy-5-(4-[(18)F]fluoro-benzoylamino)-pentyl]-ureido]-pentanedioic acid ([(18)F]6), and 2-(3-[1-carboxy-5-[(5-[(125)I]iodo-pyridine-3-carbonyl)-amino]-pentyl]-ureido)-pentanedioic acid ([(125)I]8) in 65-80% (nondecay-corrected), 30-35% (decay corrected), and 59-75% (nondecay-corrected) radiochemical yields. Compound [(125)I]3 demonstrated 8.8 +/- 4.7% injected dose per gram (%ID/g) within PSMA(+) PC-3 PIP tumor at 30 min postinjection, which persisted, with clear delineation of the tumor by SPECT. Similar tumor uptake values at early time points were demonstrated for [(18)F]6 (using PET) and [(125)I]8. Because of the many radiohalogenated moieties that can be attached via the epsilon amino group, the intermediate Lys-C(O)-Glu is an attractive template upon which to develop new imaging agents for prostate cancer.

Synthesis and Evaluation of Technetium-99m- and Rhenium-Labeled Inhibitors of the Prostate-Specific Membrane Antigen (PSMA)

The prostate-specific membrane antigen (PSMA) is increasingly recognized as a viable target for imaging and therapy of cancer. We prepared seven (99m)Tc/Re-labeled compounds by attaching known Tc/Re chelating agents to an amino-functionalized PSMA inhibitor (lys-NHCONH-glu) with or without a variable length linker moiety. K i values ranged from 0.17 to 199 nM. Ex vivo biodistribution and in vivo imaging demonstrated the degree of specific binding to engineered PSMA+ PC3 PIP tumors. PC3-PIP cells are derived from PC3 that have been transduced with the gene for PSMA. Despite demonstrating nearly the lowest PSMA inhibitory potency of this series, [(99m)Tc(CO)3( L1)] (+) ( L1 = (2-pyridylmethyl)2N(CH2) 4CH(CO2H)NHCO-(CH2) 6CO-NH-lys-NHCONH-glu) showed the highest, most selective PIP tumor uptake, at 7.9 +/- 4.0% injected dose per gram of tissue at 30 min postinjection. Radioactivity cleared from nontarget tissues to produce a PIP to flu (PSMA-PC3) ratio of 44:1 at 120 min postinjection. PSMA can accommodate the steric requirements of (99m)Tc/Re complexes within PSMA inhibitors, the best results achieved with a linker moiety between the epsilon amine of the urea lysine and the chelator.

N-[N-[(S)-1,3-Dicarboxypropyl]Carbamoyl]-4-[18F]Fluorobenzyl-l-Cysteine, [18F]DCFBC: A New Imaging Probe for Prostate Cancer

Previously, we showed successful imaging of xenografts that express the prostate-specific membrane antigen (PSMA) using small-animal positron emission tomography (PET) and the radiolabeled PSMA inhibitor N-[N-[(S)-1,3-dicarboxypropyl]carbamoyl]-S-[11C]methyl-l-cysteine. Herein, we extend that work by preparing and testing a PSMA inhibitor of the same class labeled with fluorine-18. N-[N-[(S)-1,3-Dicarboxypropyl]carbamoyl]-4-[18F]fluorobenzyl-l-cysteine ([18F]DCFBC) was prepared by reacting 4-[18F]fluorobenzyl bromide with the precursor (S)-2-[3-[(R)-1-carboxy-2-mercaptoethyl]ureido]-pentanedioic acid in ammonia-saturated methanol at 60 degrees C for 10 min followed by purification using C-18 reverse-phase semipreparative high-performance liquid chromatography. Severe combined immunodeficient mice bearing a s.c. PSMA+ PC-3 PIP tumor behind one shoulder and a PSMA(-) PC-3 FLU tumor behind the other shoulder were injected via the tail vein with either 1.85 MBq (50 microCi) of [18F]DCFBC for ex vivo biodistribution or 7.4 MBq (200 microCi) for imaging. For biodistribution, mice were sacrificed at 5, 15, 30, 60, and 120 min. Tumor, blood, and major organs were harvested and weighed, and radioactivity was counted. Imaging was done on the GE eXplore Vista small-animal PET scanner by collecting 12 consecutive 10-min frames. Radiochemical yield for [18F]DCFBC averaged 16 +/- 6% (n = 8) from 4-[18F]fluorobenzyl bromide. Specific radioactivities ranged from 13 to 133 GBq/micromol (350-3,600 Ci/mmol) with an average of 52 GBq/micromol (1,392 Ci/mmol; n = 6). Biodistribution and imaging studies showed high uptake of [18F]DCFBC in the PIP tumors with little to no uptake in FLU tumors. High radiopharmaceutical uptake was also seen in kidneys and bladder; however, washout of radioactivity from these organs was faster than from the PIP tumors. The maximum PIP tumor uptake was 8.16 +/- 2.55% injected dose per gram, achieved at 60 min after injection, which decreased to 4.69 +/- 0.89 at 120 min. The PIP tumor to muscle ratio was 20 at 120 min after injection. Based on the mouse biodistribution, the dose-limiting organ is the kidneys (human estimated absorbed dose: 0.05 mGy/MBq; 0.2 rad/mCi). [18F]DCFBC localizes to PSMA+-expressing tumors in mice, permitting imaging by small-animal PET. This new radiopharmaceutical is an attractive candidate for further studies of PET imaging of prostate cancer.