Production of Multimeric Prostate-Specific Membrane Antigen Small-Molecule Radiotracers Using a Solid-Phase 99mTc Preloading Strategy

Abstract

Small-molecule ligands specific for prostate-specific membrane antigen (PSMA) have the potential to improve prostate cancer imaging. However, highly charged ligands are difficult to label with 99m Tc and to purify. In this study, we present an adamantanetrimerized small molecule that has nanomolar binding to PSMA and also has 12 negative charges. Methods: To convert this molecule into a clinically viable SPECT diagnostic, we have developed a simple, cartridge-based, solid-phase prelabeling strategy that, within 25 min, converts readily available and inexpensive 99m Tc-pertechnetate into a chemically pure complex, with a reactive N-hydroxysuccinimide (NHS) ester, in neat organic solvent. This stable intermediate can label any aminecontaining small molecule or peptide with 99m Tc in 1 step, with high specific activity and without the need for high-performance liquid chromatography (HPLC). Results: Solid-phase conversion of 99m Tc-pertechnetate to 99m Tc-MAS 3 -NHS (MAS3 is S-acetylmercaptoacetyltriserine) could be completed in 25 min, with .99% radiochemical purity and with no coligands present. This intermediate was then conjugated to adamantane-trimerized GPI (2[(3-amino-3-carboxypropyl)(hydroxy)(phosphinyl)-methyl]pentane-1,5-dioic acid) in 1 step with .95% yield and no need for HPLC purification. The final molecule bound specifically to living human tumor cells expressing PSMA on their surface. Quantitative comparison was made among GPI monomer, GPI trimer, and their 99m Tc-derivatives. Conclusion: Our study describes a simple cartridge-based conversion of 99m Tcpertechnetate to a useful, preloaded NHS ester intermediate that takes only 25 min to prepare and results in .99% radiochemical purity. Using this chemistry, we produced a highspecific-activity, 99m Tc-labeled, PSMA-targeted small molecule and demonstrate g-ray radioscintigraphic imaging of living human prostate cancer cells.

Quantitation of CXCR4 Expression in Myocardial Infarction Using 99mTc-Labeled SDF-1α

“…The NHS ester of 99m Tc-S-acetylmercaptoacetyltriserine ( 99m Tc-MAS 3 ) was prepared to high purity (.99%) and high specific activity (average, 1.1 · 10 8 MBq/mmol; 3 · 10 3 Ci/mmol) in DMSO using a 20-min protocol described in detail previously (23). For radiolabeling, SDF-1a dry powder was resuspended in DMSO at a concentration of 12.6 mM.…”

“…To quantify absolute affinity and B max for the surface of living cells, a high-throughput homologous competition assay was used as described in detail previously (23). Briefly, cells were split onto 96-well filter plates (model MSHAS4510) and grown to 50% confluence (approximately 40,000 cells per well) over 48 h. The assay was performed at 4°C to avoid internalization of the radioligand due to constitutive endocytosis.…”

“…Our laboratory has previously developed a solid-phase preradiolabeling technique for rapid and efficient 99m Tclabeling of small molecules and peptides under aprotic conditions (23). In the present study, we use this technique to prepare a biologically active SDF-1a radiotracer with high specific activity.…”

Synthesis of dipeptide mimics based on amino phosphinate backbones and cyclic derivatives

“…[17][18][19] It has been shown that the (S,S) diastereoisomer of GPI is the eutomer with significantly higher affinity to the target protein than the corresponding (S,R) isomer. [20][21] However, the stereoselective synthesis of GPI proved to be difficult and only low diastereoselectivities have been realized so far.…”

Assessing Antibody Pharmacokinetics in Mice with In Vivo Imaging

“…NHS derivatives of hydrazinonicotinamide are available commercially (Solulink, San Diego, CA). NHS derivatives of MAG3 and S-acetylmercaptoacetyltriglycine can be prepared in-house (Winnard et al, 1997;Chang et al, 1999;Misra et al, 2007) and conjugated to proteins in a straightforward manner under mildly basic conditions. Conjugation results in a heterogenous mixture of chelate modified proteins as in the other methods discussed above.…”