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Synthesis and investigation of a radioiodinated F3 peptide analog as a SPECT tumor imaging radioligand.

Bhojani MS, Ranga R, Luker GD, Rehemtulla A, Ross BD, Van Dort ME - PLoS ONE (2011)

Bottom Line: For this purpose, we custom synthesized a modified F3 peptide analog (F3Cys) incorporating a C-terminal cysteine residue for site-specific attachment of a radioiodinated maleimide conjugating group.These results were also confirmed by ex vivo tissue analysis.No-carrier-added [(125)I]IBMF3 was synthesized by a radioiododestannylation approach in 73% overall radiochemical yield.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, University of Michigan, Ann Arbor, Michigan, United States of America.

ABSTRACT
A radioiodinated derivative of the tumor-homing F3 peptide, (N-(2-{3-[(125)I]Iodobenzoyl}aminoethyl)maleimide-F3Cys peptide, [(125)I]IBMF3 was developed for investigation as a SPECT tumor imaging radioligand. For this purpose, we custom synthesized a modified F3 peptide analog (F3Cys) incorporating a C-terminal cysteine residue for site-specific attachment of a radioiodinated maleimide conjugating group. Initial proof-of-concept Fluorescence studies conducted with AlexaFluor 532 C(5) maleimide-labeled F3Cys showed distinct membrane and nuclear localization of F3Cys in MDA-MB-435 cells. Additionally, F3Cys conjugated with NIR fluorochrome AlexaFluor 647 C(2) maleimide demonstrated high tumor specific uptake in melanoma cancer MDA-MB-435 and lung cancer A549 xenografts in nude mice whereas a similarly labeled control peptide did not show any tumor uptake. These results were also confirmed by ex vivo tissue analysis. No-carrier-added [(125)I]IBMF3 was synthesized by a radioiododestannylation approach in 73% overall radiochemical yield. In vitro cell uptake studies conducted with [(125)I]IBMF3 displayed a 5-fold increase in its cell uptake at 4 h when compared to controls. SPECT imaging studies with [(125)I]IBMF3 in tumor bearing nude mice showed clear visualization of MDA-MB-435 xenografts on systemic administration. These studies demonstrate a potential utility of F3 peptide-based radioligands for tumor imaging with PET or SPECT techniques.

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Fluorescent- and [125I]-labeled F3Cys Analogs.Structures of fluorescent and radioiodinated F3Cys and control peptide analogs investigated in this study (for details see text).
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pone-0022418-g001: Fluorescent- and [125I]-labeled F3Cys Analogs.Structures of fluorescent and radioiodinated F3Cys and control peptide analogs investigated in this study (for details see text).

Mentions: Common methods for peptide labeling utilize the conjugation of prosthetic groups such as succinimidyl esters to the ε-amino group of lysine residues [14]. However, site-specific labeling of F3 peptide by this approach is problematic since it contains a total of 9 lysine residues. Consequently, our strategy for site-specific functionalization focused on labeling at the free thiol of a suitable cysteine-modified F3 peptide analog. Thiol-specific reagents such as maleimides have been shown to display high chemoselectivity as compared to amino or carboxylate-reactive reagents [15], [16]. Accordingly, we custom synthesized a modified F3 peptide analog incorporating a cysteine residue at the C-terminus (F3Cys) for site specific coupling to a radioiodinated maleimide conjugating group. We chose a meta-[125I]iodobenzamide group tethered to a maleimide as our labeled conjugating moiety since meta-substituted aromatic radioiodine labels are known to demonstrate high stability towards in vivo metabolic deiodination [17], [18]. AlexaFluor 532 C5 maleimide-labeled F3Cys (AF532-F3Cys (1a), Figure 1A) was synthesized to investigate the sub-cellular distribution of F3Cys. We also labeled F3Cys with the Near Infrared (NIR) fluorochrome AlexaFluor 647 C2 maleimide (AF647-F3Cys (1b); Figure 1A) for in vivo optical imaging studies. To demonstrate that the tumor localization of F3Cys conjugate was specific to the F3 peptide sequence, the corresponding AlexaFluor 647 C2 maleimide conjugate of a control peptide (AF647-Control (2), Figure 1B) incorporating cysteine at the C-terminus was also synthesized.


Synthesis and investigation of a radioiodinated F3 peptide analog as a SPECT tumor imaging radioligand.

Bhojani MS, Ranga R, Luker GD, Rehemtulla A, Ross BD, Van Dort ME - PLoS ONE (2011)

Fluorescent- and [125I]-labeled F3Cys Analogs.Structures of fluorescent and radioiodinated F3Cys and control peptide analogs investigated in this study (for details see text).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3139646&req=5

pone-0022418-g001: Fluorescent- and [125I]-labeled F3Cys Analogs.Structures of fluorescent and radioiodinated F3Cys and control peptide analogs investigated in this study (for details see text).
Mentions: Common methods for peptide labeling utilize the conjugation of prosthetic groups such as succinimidyl esters to the ε-amino group of lysine residues [14]. However, site-specific labeling of F3 peptide by this approach is problematic since it contains a total of 9 lysine residues. Consequently, our strategy for site-specific functionalization focused on labeling at the free thiol of a suitable cysteine-modified F3 peptide analog. Thiol-specific reagents such as maleimides have been shown to display high chemoselectivity as compared to amino or carboxylate-reactive reagents [15], [16]. Accordingly, we custom synthesized a modified F3 peptide analog incorporating a cysteine residue at the C-terminus (F3Cys) for site specific coupling to a radioiodinated maleimide conjugating group. We chose a meta-[125I]iodobenzamide group tethered to a maleimide as our labeled conjugating moiety since meta-substituted aromatic radioiodine labels are known to demonstrate high stability towards in vivo metabolic deiodination [17], [18]. AlexaFluor 532 C5 maleimide-labeled F3Cys (AF532-F3Cys (1a), Figure 1A) was synthesized to investigate the sub-cellular distribution of F3Cys. We also labeled F3Cys with the Near Infrared (NIR) fluorochrome AlexaFluor 647 C2 maleimide (AF647-F3Cys (1b); Figure 1A) for in vivo optical imaging studies. To demonstrate that the tumor localization of F3Cys conjugate was specific to the F3 peptide sequence, the corresponding AlexaFluor 647 C2 maleimide conjugate of a control peptide (AF647-Control (2), Figure 1B) incorporating cysteine at the C-terminus was also synthesized.

Bottom Line: For this purpose, we custom synthesized a modified F3 peptide analog (F3Cys) incorporating a C-terminal cysteine residue for site-specific attachment of a radioiodinated maleimide conjugating group.These results were also confirmed by ex vivo tissue analysis.No-carrier-added [(125)I]IBMF3 was synthesized by a radioiododestannylation approach in 73% overall radiochemical yield.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, University of Michigan, Ann Arbor, Michigan, United States of America.

ABSTRACT
A radioiodinated derivative of the tumor-homing F3 peptide, (N-(2-{3-[(125)I]Iodobenzoyl}aminoethyl)maleimide-F3Cys peptide, [(125)I]IBMF3 was developed for investigation as a SPECT tumor imaging radioligand. For this purpose, we custom synthesized a modified F3 peptide analog (F3Cys) incorporating a C-terminal cysteine residue for site-specific attachment of a radioiodinated maleimide conjugating group. Initial proof-of-concept Fluorescence studies conducted with AlexaFluor 532 C(5) maleimide-labeled F3Cys showed distinct membrane and nuclear localization of F3Cys in MDA-MB-435 cells. Additionally, F3Cys conjugated with NIR fluorochrome AlexaFluor 647 C(2) maleimide demonstrated high tumor specific uptake in melanoma cancer MDA-MB-435 and lung cancer A549 xenografts in nude mice whereas a similarly labeled control peptide did not show any tumor uptake. These results were also confirmed by ex vivo tissue analysis. No-carrier-added [(125)I]IBMF3 was synthesized by a radioiododestannylation approach in 73% overall radiochemical yield. In vitro cell uptake studies conducted with [(125)I]IBMF3 displayed a 5-fold increase in its cell uptake at 4 h when compared to controls. SPECT imaging studies with [(125)I]IBMF3 in tumor bearing nude mice showed clear visualization of MDA-MB-435 xenografts on systemic administration. These studies demonstrate a potential utility of F3 peptide-based radioligands for tumor imaging with PET or SPECT techniques.

Show MeSH
Related in: MedlinePlus