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“Click”-Cyclized 68Ga-Labeled Peptides for Molecular Imaging and Therapy: Synthesis and Preliminary In Vitro and In Vivo Evaluation in a Melanoma Model System

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Theranostics, Gallium-68, and Other Radionuclides

Part of the book series: Recent Results in Cancer Research ((RECENTCANCER,volume 194))

Abstract

Cyclization techniques are used often to impart higher in vivo stability and binding affinity to peptide targeting vectors for molecular imaging and therapy. The two most often used techniques to impart these qualities are lactam bridge construction and disulfide bond formation. While these techniques have been demonstrated to be effective, orthogonal protection/deprotection steps can limit achievable product yields. In the work described in this chapter, new α-melanocyte stimulating hormone (α-MSH) peptide analogs were synthesized and cyclized by copper-catalyzed terminal azide-alkyne cycloaddition “click” chemistry techniques. The α-MSH peptide and its cognate receptor (melanocortin receptor subtype 1, MC1R) represent a well-characterized model system to examine the effect of the triazole linkage for peptide cyclization on receptor binding in vitro and in vivo. Four new DOTA-conjugated α-MSH analogs were cyclized and evaluated by in vitro competitive binding assays, serum stability testing, and in vivo imaging by positron emission tomography (PET) of tumor-bearing mice. These new DOTA-conjugated click-cyclized analogs exhibited selective high binding affinity (<2 nM) for MC1R on melanoma cells in vitro, high stability in human serum, and produced high-contrast PET/CT images of tumor xenografts. 68Ga-labeled DOTA bioconjugates displayed rapid pharmacokinetics with receptor-mediated tumor accumulation of up to 16 ± 5% ID/g. The results indicate that the triazole ring is an effective bioisosteric replacement for the standard lactam bridge assemblage for peptide cyclization. Radiolabeling results confirm that Cu catalyst is sufficiently removed prior to DOTA chelator addition to enable insertion of radio metals or stable metals for molecular imaging and therapy. Thus, these click-chemistry-cyclized variants show promise as agents for melanocortin receptor-targeted imaging and radionuclide therapy.

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References

  • Al-Obeidi F, Castrucci AMDL, Hadley ME, Hruby VJ (1989a) Potent and prolonged-acting cyclic lactam analogs of α-melanotropin: design based on molecular dynamics. J Med Chem 32(12):2555–2561. doi:10.1021/jm00132a010

    Article  PubMed  CAS  Google Scholar 

  • Al-Obeidi F, Hadley ME, Pettitt BM, Hruby VJ (1989b) Design of a new class of superpotent cyclic α-melanotropins based on quenched dynamic simulations. J Am Chem Soc 111(9):3413–3416. doi:10.1021/ja00191a044

    Article  CAS  Google Scholar 

  • Anderson CJ, Wadas TJ, Wong EH, Weisman GR (2008) Cross-bridged macrocyclic chelators for stable complexation of copper radionuclides for PET imaging. Q J Nucl Med Mol Imag 52(2):185–192. doi:R39072063

    CAS  Google Scholar 

  • Bednarek MA, MacNeil T, Kalyani RN, Tang R, Van der Ploeg LHT, Weinberg DH (1999a) Analogs of MTII, lactam derivatives of α-melanotropin, modified at the N-terminus and their selectivity at human melanocortin receptors 3, 4, and 5. Biochem Biophys Res Commun 261(1):209–213

    Article  PubMed  CAS  Google Scholar 

  • Bednarek MA, MacNeil T, Kalyani RN, Tang R, Van der Ploeg LHT, Weinberg DH (2000) Analogs of lactam derivatives of α-melanotropin with basic and acidic residues. Biochem Biophys Res Commun 272(1):23–28

    Article  PubMed  CAS  Google Scholar 

  • Bednarek MA, MacNeil T, Tang R, Fong TM, Angeles Cabello M, Maroto M, Teran A (2007a) Potent and selective peptide agonists of α-melanocyte stimulating hormone (αMSH) action at human melanocortin receptor 5; their synthesis and biological evaluation in vitro. Chem Biol Drug Des 69(5):350–355. doi:10.1111/j.1747-0285.2007.00513.x

    Article  PubMed  CAS  Google Scholar 

  • Bednarek MA, MacNeil T, Tang R, Fong TM, Cabello MA, Maroto M, Teran A (2007b) Potent and selective agonists of human melanocortin receptor 5: cyclic analogues of α-melanocyte-stimulating hormone. J Med Chem 50(10):2520–2526. doi:10.1021/jm0614275

    Article  PubMed  CAS  Google Scholar 

  • Bednarek MA, Silva MV, Arison B, MacNeil T, Kalyani RN, Huang RR, Weinberg DH (1999b) Structure-function studies on the cyclic peptide MT-II, lactam derivative of alpha-melanotropin. Peptides 20(3):401–409. doi:10.1016/S0196-9781(99)00048-0

    Article  PubMed  CAS  Google Scholar 

  • Bloomberg GB, Askin D, Gargaro AR, Tanner MJA (1993) Synthesis of a branched cyclic peptide using a strategy employing Fmoc chemistry and two additional orthogonal protecting groups. Tetrahedron Lett 34(29):4709–4712

    Article  CAS  Google Scholar 

  • Bock VD, Speijer D, Hiemstra H, van Maarseveen JH (2007) 1,2,3-Triazoles as peptide bond isosteres: synthesis and biological evaluation of cyclotetrapeptide mimics. Org Biomol Chem 5(6):971–975

    Article  PubMed  CAS  Google Scholar 

  • Boswell CA, Regino CA, Baidoo KE, Wong KJ, Bumb A, Xu H, Milenic DE, Kelley JA, Lai CC, Brechbiel MW (2008) Synthesis of a cross-bridged cyclam derivative for peptide conjugation and 64Cu radiolabeling. Bioconjugate Chem 19(7):1476–1484. doi:10.1021/bc800039e

    Article  CAS  Google Scholar 

  • Brechbiel MW (2008) Bifunctional chelates for metal nuclides. Q J Nucl Med Mol Imag 52(2):166–173

    CAS  Google Scholar 

  • Brik A, Alexandratos J, Lin Y-C, Elder JH, Olson AJ, Wlodawer A, Goodsell DS, Wong C-H (2005) 1,2,3-Triazole as a peptide surrogate in the rapid synthesis of HIV-1 protease inhibitors. Chem BioChem 6(7):1167–1169. doi:10.1002/cbic.200500101

    CAS  Google Scholar 

  • Cai M, Cai C, Mayorov AV, Xiong C, Cabello CM, Soloshonok VA, Swift JR, Trivedi D, Hruby VJ (2004) Biological and conformational study of β-substituted prolines in MT-II template: steric effects leading to human MC5 receptor selectivity*. J Pept Res 63(2):116–131. doi:10.1111/j.1399-3011.2003.00105.x

    Article  PubMed  CAS  Google Scholar 

  • Cantorias MV, Figueroa SD, Quinn TP, Lever JR, Hoffman TJ, Watkinson LD, Carmack TL, Cutler CS (2009) Development of high-specific-activity 68Ga-labeled DOTA-rhenium-cyclized α-MSH peptide analog to target MC1 receptors overexpressed by melanoma tumors. Nucl Med Biol 36(5):505–513

    Article  PubMed  CAS  Google Scholar 

  • Cone RD (ed) (2000) The melanocortin receptors. Humana, Totowa

    Google Scholar 

  • Cowell SM, Balse-Srinivasan PM, Ahn JM, Hruby VJ (2002) Design and synthesis of peptide antagonists and inverse agonists for G protein-coupled receptors. Methods Enzymol 343:49–72

    Article  PubMed  Google Scholar 

  • Dessolin M, Guillerez M-G, Thieriet N, Guibé F, Loffet A (1995) New allyl group acceptors for palladium catalyzed removal of allylic protections and transacylation of allyl carbamates. Tetrahedron Lett 36(32):5741–5744

    CAS  Google Scholar 

  • Eberle AN, Froidevaux S (2003) Radiolabeled α-melanocyte-stimulating hormone analogs for receptor-mediated targeting of melanoma: from tritium to indium. J Mol Recognit 16(5):248–254. doi:10.1002/jmr.633

    Article  PubMed  CAS  Google Scholar 

  • Eberle AN, Verin VJ, Solca F, Siegrist W, Kuenlin C, Bagutti C, Stutz S, Girard J (1991) Biologically active monoiodinated α-MSH derivatives for receptor binding studies using human melanoma cells. J Recept Res 11(1–4):311–322

    PubMed  CAS  Google Scholar 

  • Fani M, Mueller A, Tamma M-L, Nicolas G, Rink HR, Cescato R, Reubi JC, Maecke HR (2010) Radiolabeled bicyclic somatostatin-based analogs: a novel class of potential radiotracers for SPECT/PET of neuroendocrine tumors. J Nucl Med 51(11):1771–1779. doi:10.2967/jnumed.110.076695

    Google Scholar 

  • Fragogeorgi EA, Zikos C, Gourni E, Bouziotis P, Paravatou-Petsotas M, Loudos G, Mitsokapas N, Xanthopoulos S, Mavri-Vavayanni M, Livaniou E, Varvarigou AD, Archimandritis SC (2009) Spacer site modifications for the improvement of the in vitro and in vivo binding properties of 99 mTc-N3S-X-Bombesin[2–14] derivatives. Bioconjugate Chem 20:856–867

    Article  CAS  Google Scholar 

  • Froidevaux S, Calame-Christe M, Schuhmacher J, Tanner H, Saffrich R, Henze M, Eberle AN (2004) A gallium-labeled DOTA-α-melanocyte- stimulating hormone analog for PET imaging of melanoma metastases. J Nucl Med 45(1):116–123

    PubMed  CAS  Google Scholar 

  • Garrison JC, Rold TL, Sieckman GL, Naz F, Sublett SV, Figueroa AD, Volkert WA, Hoffman TJ (2008) Evaluation of the pharmacokinetic effects of various linking groups using the 111In-DOTA-X-BBN(7–14)NH2 structural paradigm in prostate cancer model. Bioconjugate Chem 19:1803–1812

    Article  CAS  Google Scholar 

  • Ghanem GE, Comunale G, Libert A, Vercammen-Grandjean A, Lejeune FJ (1988) Evidence for alpha-melanocyte-stimulating hormone (α-MSH) receptors on human malignant melanoma cells. Int J Cancer 41(2):248–255. doi:10.1002/ijc.2910410216

    Article  PubMed  CAS  Google Scholar 

  • Grieco P, Cai M, Liu L, Mayorov A, Chandler K, Trivedi D, Lin G, Campiglia P, Novellino E, Hruby VJ (2008) Design and microwave-assisted synthesis of novel macrocyclic peptides active at melanocortin receptors: discovery of potent and selective hMC5R receptor antagonists. J Med Chem 51(9):2701–2707. doi:10.1021/jm701181n

    Article  PubMed  CAS  Google Scholar 

  • Guo H, Yang J, Gallazzi F, Prosnitz ER, Sklar LA, Miao Y (2009) Effect of DOTA position on melanoma targeting and pharmacokinetic properties of 111In-labeled lactam bridge-cyclized α-MSH peptide. Bioconjugate Chem 20:2162–2168

    Article  CAS  Google Scholar 

  • Guo H, Yang J, Gallazzi F, Miao Y (2010) Reduction of the ring size of radiolabeled lactam bridge-cyclized α-MSH peptide, resulting in enhanced melanoma uptake. J Nucl Med 51(3):418–426. doi:10.2967/jnumed.109.071787

    Article  PubMed  CAS  Google Scholar 

  • Guo H, Yang J, Gallazzi F, Miao Y (2011) Effects of the amino acid linkers on the melanoma-targeting and pharmacokinetic properties of 111In-labeled lactam bridge-cyclized α-MSH peptide. J Nucl Med 52:608–616. doi:10.2967/jnumed.110.086009

    Article  PubMed  CAS  Google Scholar 

  • Harris JI, Lerner AB (1957) Amino-acid sequence of the [alpha]-melanocyte-stimulating hormone. Nature 179(4574):1346–1347

    Article  PubMed  CAS  Google Scholar 

  • Hausner SH, Kukis DL, Gagnon MK, Stanecki CE, Ferdani R, Marshall JF, Anderson CJ, Sutcliffe JL (2009) Evaluation of [64Cu]Cu-DOTA and [64Cu]Cu-CB-TE2A chelates for targeted positron emission tomography with an alphavbeta6-specific peptide. Mol Imaging 8(2):111–121

    PubMed  CAS  Google Scholar 

  • Heppeler A, Froidevaux S, Eberle AN, Maecke HR (2000) Receptor targeting for tumor localisation and therapy with radiopeptides. Curr Med Chem 7:971–994

    PubMed  CAS  Google Scholar 

  • Hruby VJ, Cai M, Grieco P, Han G, Kavarana M, Trivedi DEV (2003) Exploring the stereostructural requirements of peptide ligands for the melanocortin receptors. Ann N Y Acad Sci 994(1):12–20. doi:10.1111/j.1749-6632.2003.tb03157.x

    Article  PubMed  CAS  Google Scholar 

  • Hruby VJ, Wilkes BC, Hadley ME, Al-Obeidi F, Sawyer TK, Staples DJ, DeVaux AE, Dym O, Castrucci AMdL (1987) α-Melanotropin: the minimal active sequence in the frog skin bioassay. J Med Chem 30(11):2126–2130. doi:10.1021/jm00394a033

    Article  PubMed  CAS  Google Scholar 

  • Huisgen R (1963) 1,3-Dipolar cycloadditions. Past and future. Angew Chem Int Ed 2(10):565–598. doi:10.1002/anie.196305651 (in English)

    Article  Google Scholar 

  • Kaiser E, Colescott RL, Bossinger CD, Cook PI (1970) Color test for detection of free terminal amino groups in the solid-phase synthesis of peptides. Anal Biochem 34(2):595–598

    Article  PubMed  CAS  Google Scholar 

  • Lunec J, Pieron C, Thody AJ (1992) MSH receptor expression and the relationship to melanogenesis and metastatic activity in B16 melanoma. Melanoma Res May 2(1):5–12

    Article  CAS  Google Scholar 

  • Mayorov AV, Cai M, Chandler KB, Petrov RR, Van Scoy AR, Yu Z, Tanaka DK, Trivedi D, Hruby VJ (2006a) Development of cyclic γ-MSH analogues with selective hMC3R agonist and hMC3R/hMC5R antagonist activities. J Med Chem 49(6):1946–1952. doi:10.1021/jm0510326

    Article  PubMed  CAS  Google Scholar 

  • Mayorov AV, Han S-Y, Cai M, Hammer MR, Trivedi D, Hruby VJ (2006b) Effects of macrocycle size and rigidity on melanocortin receptor-1 and -5 selectivity in cyclic lactam α-melanocyte-stimulating hormone analogs. Chem Biol Drug Des 67(5):329–335. doi:10.1111/j.1747-0285.2006.00383.x

    Article  PubMed  CAS  Google Scholar 

  • Miao Y, Gallazzi F, Guo H, Quinn TP (2008) 111In-labeled lactam bridge-cyclized α-melanocyte stimulating hormone peptide analogues for melanoma imaging. Bioconjugate Chem 19(2):539–547. doi:10.1021/bc700317w

    Article  CAS  Google Scholar 

  • Nestor JJ Jr (2009) The medicinal chemistry of peptides. Curr Med Chem 16:4399–4418

    Article  PubMed  CAS  Google Scholar 

  • Okarvi SM (2004) Synthesis, radiolabeling and in vitro and in vivo characterization of a technetium-99 m-labeled alpha-M2 peptide as a tumor imaging agent. J Pept Res 63(6):460–468. doi:10.1111/j.1399-3011.2004.00160.x

    Article  PubMed  CAS  Google Scholar 

  • Raposinho PD, Correia JDG, Oliveira MC, Santos I (2010) Melanocortin-1 receptor-targeting with radiolabeled cyclic α-melanocyte-stimulating hormone analogs for melanoma imaging. Pept Sci 94(6):820–829. doi:10.1002/bip.21490

    Article  CAS  Google Scholar 

  • Rockey WM, Huang L, Kloepping KC, Baumhover NJ, Giangrande PH, Schultz MK (2011) Synthesis and radiolabeling of chelator-RNA aptamer bioconjugates with copper-64 for targeted molecular imaging. Bioorg Med Chem 19(13):4080–4090 (July 1)

    Article  PubMed  CAS  Google Scholar 

  • Sahm UG, Olivier GWJ, Branch SK, Moss SH, Pouton CW (1994) Synthesis and biological evaluation of [alpha]-MSH analogues substituted with alanine. Peptides 15(7):1297–1302

    Article  PubMed  CAS  Google Scholar 

  • Sawyer TK, Sanfilippo PJ, Hruby VJ, Engel MH, Heward CB, Burnett JB, Hadley ME (1980) 4-Norleucine, 7-d-phenylalanine-alpha-melanocyte-stimulating hormone: a highly potent alpha-melanotropin with ultra long biological activity. Proc Natl Acad Sci U S A 77(10):5754–5758

    Article  PubMed  CAS  Google Scholar 

  • Siegrist W, Oestreicher M, Stutz S, Girard J, Eberle AN (1988) Radioreceptor assay for α-MSH using mouse B16 melanoma cells. J Recept Res 8(1–4):323–343

    PubMed  CAS  Google Scholar 

  • Sugg EE, De L, Castrucci AM, Hadley ME, Van Binst G, Hruby VJ (1988) Cyclic lactam analogs of Ac-[Nle4].alpha.-MSH4-11-NH2. Biochemistry 27(21):8181–8188. doi:10.1021/bi00421a029

    Google Scholar 

  • Tatro JB, Entwistle ML, Lester BR, Reichlin S (1990) Melanotropin receptors of murine melanoma characterized in cultured cells and demonstrated in experimental tumors in situ. Cancer Res 50(4):1237–1242

    PubMed  CAS  Google Scholar 

  • Thieriet N, Alsina J, Giralt E, Guibé F, Albericio F (1997) Use of alloc-amino acids in solid-phase peptide synthesis. Tandem deprotection-coupling reactions using neutral conditions. Tetrahedron Lett 38(41):7275–7278

    Article  CAS  Google Scholar 

  • Turner RA, Oliver AG, Lokey RS (2007) Click chemistry as a macrocyclization tool in the solid-phase synthesis of small cyclic peptides. Org Lett 9(24):5011–5014. doi:10.1021/ol702228u

    Article  PubMed  CAS  Google Scholar 

  • Vlieghe P, Lisowski V, Martinez J, Khrestchatisky M (2010) Synthetic therapeutic peptides: science and market. Drug Discovery Today 15(1–2):40–56

    Article  PubMed  CAS  Google Scholar 

  • Ying J, Gu X, Cai M, Dedek M, Vagner J, Trivedi DB, Hruby VJ (2006) Design, synthesis, and biological evaluation of new cyclic melanotropin peptide analogues selective for the human melanocortin-4 receptor. J Med Chem 49(23):6888–6896. doi:10.1021/jm060768f

    Article  PubMed  CAS  Google Scholar 

  • Zwanziger D, Beck-Sickinger AG (2008) Radiometal targeted tumor diagnosis and therapy with peptide hormones. Curr Pharm Des 14:2385–2400

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

Support for this work was provided by the American Cancer Society (IRG-77004-31; M.K.S.), the Holden Comprehensive Cancer Center (M.E.M., M.K.S.), Neuroendocrine Tumor Fund (M.S.O.), and University of Iowa Dance Marathon (M.E.M.). M.E.M. is supported by T32 University of Iowa Institutional Training Grant in Hematologic and Oncologic Childhood Diseases (HL080070). The authors thank Dr. Kevin Rice, Dr. Lynn Teesch, and Vic Parcell for spirited assistance with mass spectral analyses.

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Authors and Affiliations

  1. Department of Pediatric Hematology/Oncology, Carver College of Medicine, The University of Iowa, ML B180, 500 Newton Road, Iowa City, IA, 52242, USA

    Molly E. Martin, M. Sue O’Dorisio & Whitney M. Leverich

  2. Departments of Radiology and Radiation Oncology (Free Radical Radiation Biology Program), Carver College of Medicine, The University of Iowa, ML B180, 500 Newton Road, Iowa City, IA, 52242, USA

    Kyle C. Kloepping, Susan A. Walsh & Michael K. Schultz

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  1. Molly E. Martin
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  2. M. Sue O’Dorisio
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  3. Whitney M. Leverich
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  4. Kyle C. Kloepping
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  5. Susan A. Walsh
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Correspondence to Michael K. Schultz .

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  1. , THERANOSTICS Center for Molecular,, ENETS Center of Excellence, Zentralklinik Bad Berka, Bad Berka, 99437, Germany

    Richard P. Baum

  2. Inst. Kernchemie, Universität Mainz, Fritz-Straßmann-Weg 2, Mainz, 55128, Germany

    Frank Rösch

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Martin, M.E., Sue O’Dorisio, M., Leverich, W.M., Kloepping, K.C., Walsh, S.A., Schultz, M.K. (2013). “Click”-Cyclized 68Ga-Labeled Peptides for Molecular Imaging and Therapy: Synthesis and Preliminary In Vitro and In Vivo Evaluation in a Melanoma Model System. In: Baum, R., Rösch, F. (eds) Theranostics, Gallium-68, and Other Radionuclides. Recent Results in Cancer Research, vol 194. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-27994-2_9

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