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Molecular Imaging and Contrast Agent Database (MICAD) [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2004-2013.

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Molecular Imaging and Contrast Agent Database (MICAD) [Internet].

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177Lu-DOTA-Tyr3-c(Cys-Tyr-Trp-Lys-Thr-Cys)-Thr-Lys(cypate)-NH2

177Lu-LS172
, PhD
National Center for Biotechnology Information, NLM, NIH, Bethesda, MD, vog.hin.mln.ibcn@dacim

Created: ; Last Update: April 22, 2008.

Chemical name:177Lu-DOTA-Tyr3-c(Cys-Tyr-Trp-Lys-Thr-Cys)-Thr-Lys(cypate)-NH2
Abbreviated name:177Lu-LS172
Synonym:177Lu-DOTA-Tyr3-octreotate-Lys-cypate, 177Lu-DOTA-Y3-TATE-Lys-cypate
Agent category:Peptide
Target:Somatostatin receptor subtype-2 (SSTR-2)
Target category:Receptor
Method of detection:Single-photon emission computed tomography (SPECT), near-infrared (NIR) fluorescent imaging, optical fluorescent imaging
Source of signal/contrast:177Lu, cypate
Activation:No
Studies:
  • Checkbox In vitro
  • Checkbox Rodents
No structure is currently available in PubChem.

Background

[PubMed]

Somatostatin (SST) (somatotropin release-inhibiting hormone, somatotropin release-inhibiting factor) is a cyclic disulphide-containing peptide hormone of 14 amino acids (1). SST inhibits hormone secretion, cell proliferation, and promotes apoptosis through binding to specific cell-surface somatostatin receptors (SSTRs) (2). Five SSTR subtypes are identified in the central nervous system (CNS), gastrointestinal tract, and a variety of benign and malignant tumors (2). All subtypes of SSTRs belong to the family of G protein–coupled receptors, and exhibit high affinity to SST by recognizing the β-turn in the peptide sequence of SST including Phe7, Trp8, Lys9, and Thr10 (2). The structural difference between SSTRs generates specific pharmacological and physiological properties for each subtype, which allows targeting of SSTRs with subtype-specific SST analogs (1). For example, octreotide, a SST analog with a peptide sequence of eight amino acids (D-Phe-c(Cys-Tyr-D-Trp-Lys-Thr-Cys)-Thr(ol)), possesses a much higher affinity (~2 nM) binding to SSTR-2, -3, and -5 than binding to SSTR-1 and -4 (>1,000 nM). In addition to symptomatic treatment of tumors in clinic, these analogs are also labeled with a variety of probes for imaging (1).

177Lu-DOTA-Tyr3-c(Cys-Tyr-Trp-Lys-Thr-Cys)-Thr-Lys(cypate)-NH2(177Lu-LS172) is a monomolecular multimodal imaging agent (MOMIA) for single-photon emission computed tomography (SPECT) and optical imaging (3). 177Lu-LS172 consists of three components: a STT analog Tyr3-octreotate (Y3-TATE) with an additional lysine at the N-terminal as a spacer linker, a complex of 177Lu-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (177Lu-DOTA) attached at the C-terminal of Y3-TATE, and a cypate attached at the ε-amine group of the spacer lysine at the N-terminal of Y3-TATE. As a STT analog, TATE is produced by replacing the C-terminal threoninol in the octreotide with threonine, which substantially increases the binding affinity for SSTR-2 (1). 177Lu is a radionuclide belonging to the group of rare earth isotopes, and it is produced by neutron bombardment of purified target materials in reactors (4). With a half-life of 6.71 days for β- emission at 498 keV and 78% branch fraction, 177Lu has been a very promising radionuclide in radiotherapy for effectively destroying small tumors and metastasis (optimal size 1.2–3.0 mm) while sparing normal tissue (5). 177Lu also emits low energetic gamma rays at 208 and 113 keV with 10% and 6% abundance, respectively, which allows for direct monitoring of the activity distribution by SPECT and subsequent dosimetry (5). 177Lu-DOTA-Y3-TATE has demonstrated excellent tumor localization, good clearance properties, and significant tumor regression in animal models (6). Cypate (mw 625.34) is a carbocyanine derivative with structure and optical properties very similar to indocyanine green dye (ICG). Cypate exhibits a high extinction coefficient (224,000 M-1cm-1) in the near-infrared (NIR) region (700–900 nm) (3, 7). The low light scattering and absorption of endogenous photoactive biomolecules in this region permit photons to penetrate several centimeters into tissues (8, 9). Two carboxyl groups in cypate can be conjugated with amines to generate peptide-based optical imaging agents (10).

Synthesis

[PubMed]

The synthesis of 177Lu-LS172 was conducted in several steps (3). Y3-TATE was synthesized with a peptide synthesizer by standard automated Fmoc protocols. Rink amide resin was initially loaded with Fmoc amino acids followed by automatic activation of the carboxyl group with a mixture of 1-hydroxybenzotriazole (HOBt) and o-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HBTU) and subsequent coupling with Fmoc-protected amino acid. LS172 was produced by attaching DOTA and cypate to Y3-TATE in situ. The carboxylic acid of tri-t-butyl-DOTA was coupled to the amine of the N-terminal phenylalanine, and the carboxylic acid of cypate was coupled to the ε-amino group of the C-terminal lysine. The overall yield of LS172 was found to be 96%. The radiolabling of LS172 was accomplished by mixing 177LuCl3 (100 μCi) with LS172 in a buffer (pH 4.0) and incubated at 80ºC for 80 min. The produced 177Lu-LS172 had a radiochemical purity of ≥99% and a final specific activity of 12.5 Ci/g (27 Ci/mmol).

In Vitro Studies: Testing in Cells and Tissues

[PubMed]

The spectral and binding properties of LS172 were examined in vitro (3). Similar to cypate, LS172 exhibited broad excitation and emission bands with a narrow Stokes shift (23 nm). The relative quantum yield of LS172 (0.12) was the same as that of ICG. Because of the modifications in both C- and N-terminal, the affinity of LS172 binding to SSTR-2 increased ~10- and ~5-fold relative to cypate-Y3-TATE and Y3-TATE, respectively. The complexation with Lu3+ appeared not to affect the resultant affinity (2.14 nM for Lu-LS172 versus 2.03 nM for LS172). The internalization of LS172 was evaluated with NIR fluorescence microscopy in A427-7 membranes (SSTR-2–positive) and HEK293 cells (STTR-2–negative). LS172 was weakly internalized at 25 nM. The internalization was inhibited by 5 μM Y3-TATE with an incubation time <60 min. In comparison, cypate-Y3-TATE was strongly internalized in a time-dependent manner, and the internalization was completely blocked by 5 μM Y3-TATE.

Animal Studies

Rodents

[PubMed]

The biochemical distribution of LS172 was examined in AR42J tumors in rats (n = 5) (3). 177Lu-LS172 (6.2 Ci/230 mmol (500 mg/230 mmol)) was injected intravenously in rats at 10–14 days after tumor implantation in the legs or after the tumors had grown ~1.5–2.0 g. In another group of tumor-implanted rats (n = 5), Y3-TATE (100μg/95 nmol) was also included as a competitive dose in addition to the radiolabeled LS172. The pharmacokinetic profile 177Lu-LS172 exhibited a rapid blood clearance of 0.228 ± 0.047% injected dose per gram (ID/g) (0.151 ± 0.027% ID/g) at 1 h after injection. Clearance of 177Lu-LS172 was predominately hepatic (17.862 ± 3.286% ID/g, 89.6 ± 3.16% ID/liver, 1 h) with relatively low kidney accumulation (1–2%). Accumulation of activity for 177Lu-LS172 in the SSTR-2–positive tissues of adrenal, pituitary, pancreas, and tumor were relatively low (≤1% ID/g). With a co-administration of Y3-TATE, the accumulation of 177Lu-LS172 decreased significantly in all SSTR-2–positive tissue except the bone.

The NIR fluorescence imaging of LS172 was performed in tumor-bearing NCR nu/nu mice (n = 6) (3). After inoculation with lung adenocarcinoma cell line A427-7 in the right and left flanks of each mouse, tumors were allowed to grow to 5–10 mm maximum diameter before imaging. Three mice were injected with 2 nmol LS172, and two other mice were injected with 2 nmol LS172 and a competitive dose of 95 nmol Y3-TATE. NIR fluorescence images were collected at 1, 4, 8, and 24 h after injection. No significant accumulation of LS172 was found in tumors compared to that in normal tissue at different time points. The competitive inhibition with Y3-TATE did not significantly alter the tumor/muscle uptake ratio. After 24 h, the mice were euthanized, and tissues were harvested for ex vivo fluorescence biodistribution imaging. In addition to the liver and kidneys, LS172 accumulated in the spleen and accounted for 35% of the fluorescence intensity relative to that in the liver. LS172 accumulation in mouse kidney was somewhat higher than that of the radiolabeled analogs in rat kidneys, possibly indicating the reduction of overall charge on the DOTA upon chelation with a metal ion. The uptake and clearance profiles of LS172 in tumor-bearing A427-7 mice were similar to those of 177Lu-LS172 in tumor-bearing AR427 rats.

Other Non-Primate Mammals

[PubMed]

No publication is currently available.

Non-Human Primates

[PubMed]

No publication is currently available.

Human Studies

[PubMed]

No publication is currently available.

NIH Support

CA 100972, EB 1430, CA 123537, CA 64475

References

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Breeman W.A. , Kwekkeboom D.J. , de Blois E. , de Jong M. , Visser T.J. , Krenning E.P. Radiolabelled regulatory peptides for imaging and therapy. Anticancer Agents Med Chem. 2007;7(3):345–57. [PubMed: 17504160]
2.
Barnett P. Somatostatin and somatostatin receptor physiology. Endocrine. 2003;20(3):255–64. [PubMed: 12721505]
3.
Edwards W.B. , Xu B. , Akers W. , Cheney P.P. , Liang K. , Rogers B.E. , Anderson C.J. , Achilefu S. Agonist-antagonist dilemma in molecular imaging: evaluation of a monomolecular multimodal imaging agent for the somatostatin receptor. Bioconjug Chem. 2008;19(1):192–200. [PMC free article: PMC2539107] [PubMed: 18020401]
4.
Schotzig U. , Schrader H. , Schonfeld E. , Gunther E. , Klein R. Standardisation and decay data of 177Lu and 188Re. Appl Radiat Isot. 2001;55(1):89–96. [PubMed: 11339536]
5.
Dvorakova Z. , Henkelmann R. , Lin X. , Turler A. , Gerstenberg H. Production of 177Lu at the new research reactor FRM-II: Irradiation yield of 176Lu(n,gamma)177Lu. Appl Radiat Isot. 2008;66(2):147–51. [PubMed: 17900914]
6.
de Jong M. , Breeman W.A. , Bernard B.F. , Bakker W.H. , Schaar M. , van Gameren A. , Bugaj J.E. , Erion J. , Schmidt M. , Srinivasan A. , Krenning E.P. [177Lu-DOTA(0),Tyr3] octreotate for somatostatin receptor-targeted radionuclide therapy. Int J Cancer. 2001;92(5):628–33. [PubMed: 11340564]
7.
Achilefu S. , Dorshow R.B. , Bugaj J.E. , Rajagopalan R. Novel receptor-targeted fluorescent contrast agents for in vivo tumor imaging. Invest Radiol. 2000;35(8):479–85. [PubMed: 10946975]
8.
Mahmood U. , Weissleder R. Near-infrared optical imaging of proteases in cancer. Mol Cancer Ther. 2003;2(5):489–96. [PubMed: 12748311]
9.
Ye Y. , Bloch S. , Xu B. , Achilefu S. Novel near-infrared fluorescent integrin-targeted DFO analogue. Bioconjug Chem. 2008;19(1):225–34. [PMC free article: PMC2631379] [PubMed: 18038965]
10.
Zhang Z. , Achilefu S. Spectral properties of pro-multimodal imaging agents derived from a NIR dye and a metal chelator. Photochem Photobiol. 2005;81(6):1499–504. [PubMed: 16120005]

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