Abstract
Purpose
To develop an L-PG-based imaging probe suitable for assessing the degradation of L-PG in vivo.
Materials and Methods
Conjugates of L-PG and a near-infrared fluorescence (NIRF) dye, NIR813, were characterized with regard to quenching efficiency and degradability by cathepsin B (CB) and other proteases. The kinetics of L-PG-NIR813’s degradation and its degradation in orthotopic human U87/TGL glioma in nude mice after intravenous injection was assessed using NIRF optical imaging (n = 3).
Results
The fluorescence signal from L-PG-NIR813 was efficiently quenched and activated at NIR813 loadings of 8–10%. Upon exposure to CB, the fluorescence intensity of L-PG-NIR813 increased 10-fold. L-PG-NIR813 was also degraded by another cysteine protease cathepsin L, but not by MMP-2, cathepsin E, cathepsin D, and plasmin. A selective CB inhibitor blocked the fluorescence activation. After intravenous injection, the degradation of L-PG-NIR813 was visualized primarily in the liver, which peaked at 4 h postinjection. Activation of L-PG-NIR813 but not D-PG-NIR813 was clearly seen in U87/TGL tumors.
Conclusion
Our results indicate that L-PG-NIR813 may be used to monitor the in vivo degradation of L-PG-based polymeric drugs, and that this agent may prove useful in noninvasive imaging of protease activity, particularly that of cysteine proteases.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
C. Li. Poly (L-glutamic acid)—anticancer drug conjugates. Adv. Drug Deliv. Rev. 54:695–713 (2002).
J. W. Singer, S. Shaffer, B. Baker, A. Bernareggi, S. Stromatt, D. Nienstedt, and M. Besman. Paclitaxel poliglumex (XYOTAX; CT-2103): an intracellularly targeted taxane. Anticancer Drugs 16:243–254 (2005).
C. Li, R. A. Newman, Q. P. Wu, S. Ke, W. Chen, T. Hutto, Z. Kan, M. D. Brannan, C. Charnsangavej, and S. Wallace. Biodistribution of paclitaxel and poly(L-glutamic acid)-paclitaxel conjugate in mice with ovarian OCa-1 tumor. Cancer Chemother. Pharmacol. 46:416–422 (2000).
H. C. Chiu, P. Kopeckova, S. S. Deshmane, and J. Kopecek. Lysosomal degradability of poly(alpha-amino acids). J. Biomed. Mater. Res. 34:381–392 (1997).
C. J. T. Hoes, W. Potman, W. A. R. Van Heeswijk, J. Mud, B. G. de Grooth, J. Greve, and J. Feijen. Optimization of macromolecular prodrugs of the antitumor antibiotic Adriamycin. J. Control. Release 2:205–213 (1985).
L. A. McCormick-Thomsonand, R. Duncan. Poly(amino acid) copolymers as a potential soluble drug delivery system. 1. Pinocytic uptake and lysosomal degradation measured in vitro. J. Bioact. Compat. Polym. 4:242–251 (1989).
W. G. Miller. Degradation of synthetic polypeptides. II. degradation of poly-L-glutamic acid by proteolytic enzymes in 0.20 M sodium chloride. J. Am. Chem. Soc. 86:3913–3918 (1964).
B. K. Kishore, P. Lambricht, G. Laurent, P. Maldague, R. Wagner, and P. M. Tulkens. Mechanism of protection afforded by polyaspartic acid against gentamicin-induced phospholipidosis. II. Comparative in vitro and in vivo studies with poly-L-aspartic, poly-L-glutamic and poly-D-glutamic acids. J. Pharmacol. Exp. Ther. 255:875–885 (1990).
L. Strekowski, T. Gorecki, J. C. Mason, H. Lee, and G. Patonay. New heptamethine cyanine reagents for labeling of biomolecules with a near-infrared Chromophore. Heterocycl. Commun. 7:2117–2122 (2001).
H. Lineweaver, and D. Burk. J. Amer. Chem. Soc. 58:658 (1934).
R. Copeland. Enzymes: A practical introduction to structure, mechanism, and data analysis. Wiley-VCH, 2000, p. 267–269.
V. Ponomarev, M. Doubrovin, I. Serganova, J. Vider, A. Shavrin, T. Beresten, A. Ivanova, L. Ageyeva, V. Tourkova, J. Balatoni, W. Bornmann, R. Blasberg, and J. Gelovani Tjuvajev. A novel triple-modality reporter gene for whole-body fluorescent, bioluminescent, and nuclear noninvasive imaging. Eur. J. Nucl. Med. Mol. Imaging. 31:740–751 (2004).
K. I. Hulkower, C. C. Butler, B. E. Linebaugh, J. L. Klaus, D. Keppler, V. L. Giranda, and B. F. Sloane. Fluorescent microplate assay for cancer cell-associated cathepsin B. Eur. J. Biochem. 267:4165–4170 (2000).
M. M. Bradford. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72:248–254 (1976).
K. Stachowiak, M. Tokmina, A. Karpinska, R. Sosnowska, and W. Wiczk. Fluorogenic peptide substrates for carboxydipeptidase activity of cathepsin B. Acta. Biochim. Pol. 51:81–92 (2004).
M. Szabelski, M. Rogiewicz, and W. Wiczk. Fluorogenic peptide substrates containing benzoxazol-5-yl-alanine derivatives for kinetic assay of cysteine proteases. Anal. Biochem. 342:20–27 (2005).
C. Allievi, I. Strepponi, U. Bastrup, L. Piazzoni, S. Tavazzi, R. Pisano, G. Pezzoni, M. Fornasier, A. Bernareggi, and J. W. Singer. Biodistribution of paclitaxel poliglumex (PPX) in lung: analysis of gender-related alterations in a preclinical model. J. Clin. Oncol. 24:Abstract #17003 (2006).
H. Ross, P. Bonomi, C. Langer, M. O’Brien, K. O’Byrne, L. Paz-Ares, A. Sandler, M. Socinski, F. Oldham, and J. Singer. Effect of gender on outcome in two randomized phase III trials of paclitaxel poliglumex (PPX) in chemo-naïve pts with advanced NSCLC and poor performance status (PS2). J. Clin. Oncol. 24:Abstract #7039 (2006).
R. Weissleder, and V. Ntziachristos. Shedding light onto live molecular targets. Nat. Med. 9:123–128 (2003).
M. Gurfinkel, S. Ke, X. Wen, C. Li, and E. M. Sevick-Muraca. Near-infrared fluorescence optical imaging and tomography. Dis. Markers 19:107–121 (2003).
R. Weissleder, C. H. Tung, U. Mahmood, and A. Bogdanov, Jr. In vivo imaging of tumors with protease-activated near-infrared fluorescent probes. Nat. Biotechnol. 17:375–378 (1999).
T. Nomura and N. Katunuma. Involvement of cathepsins in the invasion, metastasis and proliferation of cancer cells. J. Med. Invest. 52:1–9 (2005).
S. Roshy, B. F. Sloane, and K. Moin. Pericellular cathepsin B and malignant progression. Cancer Metastasis. Rev. 22:271–286 (2003).
X.-Y. Cao, S. Ke, Q.-P. Wu, J. Gelovani, and C. Li. Optical and nuclear imaging of bone with poly (L-glutamic acid). Molecular Imaging 3:275 (2004).
Acknowledgments
We would like to thank the National Institutes of Health (grants R01 CA119387 and R01 EB003132) and the John S. Dunn Foundation for financial support of this work.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Melancon, M.P., Wang, W., Wang, Y. et al. A Novel Method for Imaging In Vivo Degradation of Poly(L-Glutamic Acid), a Biodegradable Drug Carrier. Pharm Res 24, 1217–1224 (2007). https://doi.org/10.1007/s11095-007-9253-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11095-007-9253-0