Cheng KT, Katsifis A.



In vitro Rodents



2-(2-(4-(4-[123I]Iodobenzyl)piperazin-1-yl)-2-oxoethyl)isoindoline-1,3-dione ([123I]MEL037) is a radioligand that has been developed as a single-photon emission computed tomography (SPECT) imaging probe for malignant melanoma (1). [123I]MEL037 is a radioiodinated benzamide (BZA) labeled with 123I, a gamma emitter with a physical half-life (t½) of 13.2 h.

Malignant melanoma is the sixth most common cancer in the United States (2). Early diagnosis and prompt surgical removal comprise the best approach for a possible cure (3). Melanomas develop from activated or genetically altered neoplastic melanocytes that contain melanin (4-6). Melanin pigment is synthesized and deposited within melanosomes (7). The complex biosynthetic pathway of melanogenesis is catalytically controlled by three gene-related metalloenzymes: tyrosinase, tyrosinase-related protein 1, and tyrosinase-related protein 2 (6). Eumelanin and pheomelanin are the two major types of melanin. Eumelanin is the predominant melanin pigment found in primary tumors. Pheomelanin tends to associate with progression of the disease. Melanins are complex, negatively charged molecules with hydrophobic surfaces, and they have the capability to bind many different types of compounds (4). Because of these properties, melanins represent a very attractive target for both diagnosis and treatment.

BZA derivatives and iodobenzamides have been found to have affinity for melanomas (1, 4, 8, 9). The exact mechanisms of uptake of this class of compound have not been fully established (1, 10, 11). Direct melanin binding, involvement in the melanin biosynthesis pathway, and sigma (σ) receptor mediation have been proposed as the most likely mechanisms for different derivatives. In addition, available vascular concentrations of these compounds and their ability to transport into the melanoma cells also influence their cellular uptake (11). Melanin-targeting radiopharmaceuticals that are based on BZA such as [123I]BZA, [123I]BZA2, and [123I]IBZM have been investigated and developed for molecular imaging of malignant melanomas (12-14). To produce a radioiodinated BZA analog with higher tumor uptake and faster body clearance, Pham et al. (1, 15) designed and prepared [123I]MEL037, which has a piperidine moiety and rigidification of the structure by using an isoindoline-1,3-dione as the backbone. This iodobenzylpiperazine derivative also has a methylene-carbonyl unit and iodine incorporated in the benzylamine instead of the benzamide. Their study showed that [123I]MEL037 has a high selective uptake in the murine B16F0 melanotic tumor, and the uptake was most likely related to melanin binding and not to σ receptor binding.



Pham et al. (1, 15) reported the synthesis of [123I]MEL037. The bromo analog of MEL037 (Br-MEL037) was first synthesized by condensation of the corresponding piperazine with the protected amino acid N-phthaloylglycine. The reaction mixture of commercially available 1-(4-bromobenzyl)piperazine, N-phthaloylglycine, 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide, N-hydroxybenzotriazole, and N-methylmorpholine in N,N-dimethylformamide was stirred at room temperature for 24 h. Work-up and purification gave the desired bromo compound in a 69% yield. The trimethylstannyl precursor for radioiodination was prepared from Br-MEL037 by mixing with hexamethylditin and catalytic palladium tetrakistriphenylphosphine in anhydrous toluene and heating the mixture to reflux for 48 h. The resulting trimethylstannyl precursor was purified by column chromatography with a yield of 37%. This precursor allowed the regioselective radioiodination of MEL037 in the fourth position on the benzene ring by electrophilic substitution of the trimethyltin-leaving group. Chloramine-T was used as the oxidant. Briefly, [123I]sodium iodide, chloramine-T, and hydrochloric acid were successively added to the trimethylstannyl precursor in ethanol. The reaction mixture was allowed to stand for 5 min at room temperature. The reaction was quenched with sodium metabisulfite and purified by semipreparative high-performance liquid chromatography (HPLC). The radiochemical yield was 65 ± 9% (n = 12), and the radiochemical purity was >99%. The specific activity was >2 GBq/nmol (54 mCi/nmol).

In Vitro Studies: Testing in Cells and Tissues


Pham et al. (1) showed that [123I]MEL037 was stable in 1% ethanol saline at room temperature for 4 h and the radiochemical purity remained at 97%. When [123I]MEL037 was stored at −20ºC, the radiochemical purity was still >98% at the end of 24 h. The lipophilicity logP7.5 of unlabeled MEL037 determined by HPLC analysis was 3.6. In vitro competitive binding assays of unlabeled MEL037 to σ1 and σ2 receptors in rat brain membranes were conducted. Very low affinities for both receptors were found. At 10−5 and 10−6 M concentrations, MEL037 inhibited 95% and 54% of σ1 binding, respectively. At 10−5 M, MEL037 inhibited 60% of σ2 binding. No affinity was found for other receptors (e.g., peripheral benzodiazepine, dopamine, muscarinic, opioid, and serotonin).

Animal Studies



Biodistribution studies with [123I]MEL037 were conducted in black mice bearing s.c. B16F0 murine melanotic melanomas or nude albino mice bearing s.c. A375 human amelanotic melanomas as the controls (1). Each mouse received 0.37–0.74 MBq (10–20 μCi) of [123I]MEL037 by i.v. administration. In the B16F0 tumor model, there was high tumor radioactivity uptake, low normal organ radioactivity uptake, and rapid body clearance. The tumor radioactivity levels (n = 10) in percentage injected dose per gram (% ID/g) were 13.3 ± 2.6 (1 h), 22.4 ± 4.1 (3 h), 25.0 ± 2.4 (6 h), 9.7 ± 2.9 (24 h), and 5.0 ± 1.1 (48 h). At 1 h, the radioactivity levels (% ID/g) in other major organs were 17.2 ± 2.0 (liver), 8.9 ± 1.1 (kidney), 5.3 ± 0.8 (lung), 3.1 ± 0.5 (heart), 1.9 ± 0.3 (brain), and 1.5 ± 0.2 (blood). At 24 h, these levels (% ID/g) decreased to 0.79 ± 0.21 (liver), 0.84 ± 0.16 (kidney), 0.11 ± 0.02 (lung), 0.15 ± 0.04 (heart), 0.13 ± 0.03 (brain), and 0.07 ± 0.02 (blood). The tumor/body mean contrast ratios at time t [MCRt = (tumor activity/tumor in g)/(remaining activity at time t/animal weight in g)] were 3.3 ± 0.7 (1 h), 6.1 ± 0.0 (3 h), 8.8 ± 1.8 (6 h), 33 ± 8 (24 h), and 61 ± 6 (48 h). The thyroid activity uptakes (% ID) were minimal, ranging from 0.06 ± 0.01 at 1 h to 0.09 ± 0.03 at 48 h. There were relatively high levels in the eyes that ranged from 1.0 ± 0.1% ID at 1 h to 1.2 ± 0.1% ID at 48 h. In comparison, a similar normal organ distribution pattern was observed in the A375 amelanotic melanoma, but the tumor radioactivity levels (n = 5) were very low with 1.81 ± 0.29% ID/g (1 h), 1.32 ± 0.23% ID/g (6 h), and 0.38 ± 0.13% ID/g (24 h). The MCRt values were 0.4 (1 h), 0.4 (6 h), and 0.9 (24 h). In addition, the radioactivity uptake levels in the eyes of the A375 mice were ~100 times lower than the uptake levels in the B16F0 mice, ranging from 0.03 ± 0.01% ID at 1 h to 0.01 ± 0.00% ID at 24 h. A blocking study was performed in the B16F0 mice with 1 mg/kg haloperidol (σ receptor drug) administered 5 min before the [123I]MEL037 dose. At 1 h, the tumor radioactivity level was reduced by ~23% with 10–15% increases observed in the lung, heart, blood, and brain. At 24 h, the decreases were ~50%, ~40%, ~16%, and ~16% for the tumor, thyroid, eyes, and liver, respectively.

Metabolic studies were performed by injecting 3–5 MBq (0.081–0.135 mCi) [123I]MEL037 into mice bearing B16F0 tumors (1). Plasma, brain, and tumor samples were obtained at 0.25, 1, 3, and 24 h after injection. Acetonitrile extraction and thin-layer chromatography analysis indicated that 40% of the plasma radioactivity represented polar radioactive metabolites at 1 h. At 3 h, >90% of the radioactivity in the brain and tumor represented unmetabolized [123I]MEL037.

Gamma planar imaging was performed in mice bearing B16F0 melanotic tumors or A375 amelanotic melanomas (1). A dose of 7–10 MBq (0.19–0.27 mCi) [123I]MEL037 was administered by i.v. injection, and images were obtained 1, 24, and 48 h after injection. The results showed that mice bearing B16F0 tumors had the highest tumor radioactivity levels. The eyes and the abdomen were visualized with lower radioactivity levels. In comparison, neither the tumor nor the eyes were visualized in the A375 tumor mice.

Other Non-Primate Mammals


No publication is currently available.

Non-Human Primates


No publication is currently available.

Human Studies


No publication is currently available.


  • 1. Pham, T.Q., P. Berghofer, X. Liu, I. Greguric, B. Dikic, P. Ballantyne, F. Mattner, V. Nguyen, C. Loc'h, and A. Katsifis, Preparation and Biologic Evaluation of a Novel Radioiodinated Benzylpiperazine, 123I-MEL037, for Malignant Melanoma. J Nucl Med, 2007. [PubMed: 17631542]
  • 2.
    Miao Y., Benwell K., Quinn T.P. 99mTc- and 111In-labeled alpha-melanocyte-stimulating hormone peptides as imaging probes for primary and pulmonary metastatic melanoma detection. J Nucl Med. 2007;48(1):73–80. [PubMed: 17204701]
    Miao Y., Hylarides M., Fisher D.R., Shelton T., Moore H., Wester D.W., Fritzberg A.R., Winkelmann C.T., Hoffman T., Quinn T.P. Melanoma therapy via peptide-targeted {alpha}-radiation. Clin Cancer Res. 2005;11(15):5616–21. [PubMed: 16061880]
    Dadachova E., Casadevall A. Melanin as a potential target for radionuclide therapy of metastatic melanoma. Future Oncol. 2005;1(4):541–9. [PubMed: 16556030]
    Cohen-Solal K.A., Crespo-Carbone S.M., Namkoong J., Mackason K.R., Roberts K.G., Reuhl K.R., Chen S. Progressive appearance of pigmentation in amelanotic melanoma lesions. Pigment Cell Res. 2002;15(4):282–9. [PubMed: 12100494]
    Sulaimon S.S., Kitchell B.E. The biology of melanocytes. Vet Dermatol. 2003;14(2):57–65. [PubMed: 12662262]
    Hearing V.J. The melanosome: the perfect model for cellular responses to the environment Pigment Cell Res 2000. 13Suppl 823–34. [PubMed: 11041354]
    Michelot J.M., Moreau M.F., Veyre A.J., Bonafous J.F., Bacin F.J., Madelmont J.C., Bussiere F., Souteyrand P.A., Mauclaire L.P., Chossat F.M. et al. Phase II scintigraphic clinical trial of malignant melanoma and metastases with iodine-123-N-(2-diethylaminoethyl 4-iodobenzamide) J Nucl Med. 1993;34(8):1260–6. [PubMed: 8326382]
    Wolf M., Bauder-Wust U., Mohammed A., Schonsiegel F., Mier W., Haberkorn U., Eisenhut M. Alkylating benzamides with melanoma cytotoxicity. Melanoma Res. 2004;14(5):353–60. [PubMed: 15457090]
    Dittmann H., Coenen H.H., Zolzer F., Dutschka K., Brandau W., Streffer C. In vitro studies on the cellular uptake of melanoma imaging aminoalkyl-iodobenzamide derivatives (ABA) Nucl Med Biol. 1999;26(1):51–6. [PubMed: 10096501]
    Eisenhut M., Hull W.E., Mohammed A., Mier W., Lay D., Just W., Gorgas K., Lehmann W.D., Haberkorn U. Radioiodinated N-(2-diethylaminoethyl)benzamide derivatives with high melanoma uptake: structure-affinity relationships, metabolic fate, and intracellular localization. J Med Chem. 2000;43(21):3913–22. [PubMed: 11052796]
    Brandau W., Niehoff T., Pulawski P., Jonas M., Dutschka K., Sciuk J., Coenen H.H., Schober O. Structure distribution relationship of iodine-123-iodobenzamides as tracers for the detection of melanotic melanoma. J Nucl Med. 1996;37(11):1865–71. [PubMed: 8917194]
    Moins N., D'Incan M., Bonafous J., Bacin F., Labarre P., Moreau M.F., Mestas D., Noirault E., Chossat F., Berthommier E., Papon J., Bayle M., Souteyrand P., Madelmont J.C., Veyre A. 123I-N-(2-diethylaminoethyl)-2-iodobenzamide: a potential imaging agent for cutaneous melanoma staging. Eur J Nucl Med Mol Imaging. 2002;29(11):1478–84. [PubMed: 12397467]
    Salopek T.G., Scott J.R., Joshua A.V., Smylie M., Logus J.W., Morin C.A., McEwan A.J. Radioiodinated N-[3-(4-morpholino)propyl]-N-methyl-2-hydroxy-5-iodo-3-methylbenzylamine (ERC9): a new potential melanoma imaging agent. Eur J Nucl Med. 2001;28(4):408–17. [PubMed: 11357490]
    Pham T.Q., Greguric I., Liu X., Berghofer P., Ballantyne P., Chapman J., Mattner F., Dikic B., Jackson T. C. Loc'h, and A. Katsifis, Synthesis and evaluation of novel radioiodinated benzamides for malignant melanoma. J Med Chem. 2007;50(15):3561–72. [PubMed: 17602544]

    This MICAD chapter is not included in the Open Access Subset, because it was authored / co-authored by one or more investigators who was not a member of the MICAD staff.