Synthesis and characterization of water-soluble and photostable L-DOPA dendrimers

Article abstract of DOI:10.1021/ol061449l

A small drug molecule, L-DOPA, was converted into well-defined dendritic macromolecules. Their monodisperse nature was shown by NMR, MALDI-TOF-MS, and PAGE. A third-generation L-Dopa dendrimer contained 30 L-Dopa residues, which made up its core, branches, and periphery. Individual L-Dopa moieties in the dendrimer were connected to one another via hydrolyzable diester linkages. These Dopa dendrimers showed a 20-fold increase in aqueous solubility and enhanced photostability in solutions over L-Dopa under identical conditions.

Full text of DOI:10.1021/ol061449l

ORGANIC
LETTERS
2006
Vol. 8, No. 20
4421-4424
Synthesis and Characterization of
Water-Soluble and Photostable L-DOPA
Dendrimers
Shengzhuang Tang, Lynda J. Martinez, Ajit Sharma, and Minghui Chai*
Department of Chemistry, Central Michigan UniVersity, Mt. Pleasant, Michigan 48859
minghui.chai@cmich.edu
Received June 13, 2006
ABSTRACT
A small drug molecule, L-DOPA, was converted into well-defined dendritic macromolecules. Their monodisperse nature was shown by NMR,
MALDI-TOF-MS, and PAGE. A third-generation L-Dopa dendrimer contained 30 L-Dopa residues, which made up its core, branches, and periphery.
Individual L-Dopa moieties in the dendrimer were connected to one another via hydrolyzable diester linkages. These Dopa dendrimers showed
a 20-fold increase in aqueous solubility and enhanced photostability in solutions over L-Dopa under identical conditions.
L-DOPA (levodopa, 3,4-dihydroxy-L-phenylalanine) is a drug
used to treat Parkinson’s disease. It is a prodrug capable of
passing the blood-brain barrier. It is decarboxylated in the
brain to become dopamine, the neurotransmitter, by the
enzyme aromatic-L-amino acid decarboxylase. L-DOPA can
relieve some of the dopamine deficit seen in Parkinsonism.1
However, it also induces side effects such as dystonia and
dyskinesia after large doses or chronic use. Slow release of
L-DOPA (e.g., long-acting forms of Sinemet and Madopar)
has shown the reduction of the problems associated with
long-term therapy.
Dendrimers are promising nanomaterials for medical and
pharmaceutical applications because they may improve the
therapeutic efficacy of many low MW drugs by utilizing the
dendritic voids inside to physically encapsulate the drugs or
the terminal groups on the periphery to chemically conjugate
them.2 To achieve high drug loading, dendrimers of relatively
large size have to be used. Linear polymer drugs such as
(1) The Columbia Electronic Encyclopedia, 6th ed.; Columbia University
Press: New York, 2005.
10.1021/ol061449l CCC: $33.50
© 2006 American Chemical Society
Published on Web 09/08/2006

PolyAspirin truly offer a huge capacity of drugs and an easily
manipulated system for drug delivery.3 However, the release
of drug units in a linear polymeric drug can still be “sudden”
because the hydrolytic degradation may occur at any spot
of the polymer backbone or even at multiple spots simulta-
neously.
In this communication, we demonstrate a novel approach
to incorporate multiple drug units (i.e., L-DOPA) into a
cascade structure to form a dendrimer prodrug (Figure 1)
in refluxing acetone to give the BnO-DOPA-NHBoc-
COOMe 3,6 which was hydrolyzed by 1 N aqueous NaOH
in MeOH/THF at room temperature followed by acidification
to afford BnO-DOPA-NHBoc-COOH 4.7
Scheme 2 shows the synthesis of the building block 6 and
the Boc-protected core 8. Compound 4 was treated with 4.5
equiv of ethylene glycol in the presence of dicyclohexyl-
carbodiimide (DCC) and 4-(dimethylamino)-pyridinium p-
toluenesulfonate (DPTS) to give 5,8 which was then stirred
with succinic anhydride in pyridine to provide the building
block 6 for the continuous synthesis of high-generation
dendrimer prodrugs.9 The BnO-G0-NHBoc 7 was produced
by simultaneously coupling two units of 4 with ethylene
glycol in the presence of DCC and DPTS and then was
hydrogenolyzed with a palladium catalyst (5% Pd/C) to yield
the HO-G0-NHBoc 8,10 which was used as the core to
develop high-generation dendrimer prodrugs.
Scheme 2. Synthesis of Building Block 6 and Core 8
(Boc-G0)
Figure 1. Structure of the second-generation L-DOPA dendrimer
prodrug (HO-G2-NH2).
The synthesis of a high generation of drug dendrimers was
achieved with a routine coupling and deprotecting procedure
(Scheme 3). The core 8 was coupled with four units of the
building block 6 to afford BnO-G1-NHBoc 9, which was
then deprotected back to the catechol function by hydro-
genolysis with H2/Pd-C (Parr apparatus) to give HO-G1-
NHBoc 10. The deprotection of amine groups of 10 via
acidification gave the desired first-generation L-DOPA
based on L-DOPA.4 This is believed to be the first time that
a dendrimer is made of and used as an intrinsic drug or
prodrug instead of as a carrier for encapsulation or conjuga-
tion. Such a well-defined dendritic architecture will be
expected to provide a sequential and better controlled release
profile by losing the periphery drug entities first, then the
interior ones, and lastly the core units.
(2) (a) Pasut, G.; Scaramuzza, S.; Schiavon, O.; Mendich, R.; Veronese,
F. M. J. Bioact. Compat. Polym. 2005, 20, 213-230. (b) Bryan, J. Pharma.
J. 2004, 273, 793-794. (c) Delort, E.; Darbre, T.; Reymond, J.-L. J. Am.
Chem. Soc. 2004, 126, 15642-15643. (d) Liu, M.; Kono, K.; Fre´chet, J.
M. J. J. Polym. Sci. Part A: Polym. Chem. 1999, 37, 3492-3503.
(3) (a) Whitaker-Brothers, K.; Uhrich, K. E. J. Biomed. Mater. Res.:
Part A 2006, 764, 470-479. (b) Miao, J.; Xu, G.; Zhu, L.; Tian, L.; Uhrich,
K. E.; Aliva-Orta, C. A.; Hsiao, B. S.; Utz, M. Macromolecules 2005, 38,
7074-7082. (c) Prudencio, A.; Schmeltzer, R. C.; Uhrich, K. E. Macro-
molecules 2005, 38, 6895-6901. (d) Harten, R. D.; Svach, D. J.; Schmeltzer,
R.; Uhrich, K. E. J. Biomed. Mater. Res.: Part A. 2005, 72A(4), 354-362.
(e) Schmeltzer, R.; Schmalenberg, K.; Uhrich, K. E. Biomacromolecules
2005, 6(1) 359-367.
Synthesis of the L-DOPA dendrimer prodrugs started from
commercially available L-DOPA 1 (Scheme 1), which was
Scheme 1. Synthesis of Boc- and Bn-Protected
L-DOPA-COOH4
(4) Chai, M.; Tang, S. Method of Preparing Dendritic Drugs. U.S.
Provisional Patent Application, January 24, 2006.
(5) (a) Hunter, L.; Hutton, C. A. Aust. J. Chem. 2003, 56, 1095-1098.
(b) Gaucher, A.; Barbeau, O.; Hamchaoui, W.; Vandrommer, L.; Wright,
K.; Wakselman, M.; Mazaleyrat, J. P. Tetrahedron Lett. 2002, 8241-8244.
(6) Clary, M. T.; Adejare, A.; Shams, G.; Feller, D. R.; Miller, D. D. J.
Med. Chem. 1987, 30, 86-90.
(7) Hu, B. H.; Messersmith, P. B. Tetrahedron Lett. 2000, 40, 5795-
5798.
(8) Moore, J. S.; Stupp, S. I. Macromolecules 1990, 23, 65-70.
(9) Luman, N. R.; Smeds, K. A.; Grinstaff, M. W. Chem.-Eur. J. 2003,
9, 5618-5626.
(10) Hanessian, S.; Liak, T. S.; Vanasse, B. Synthesis 1981, 5, 396-
397.
converted to the corresponding methyl ester and then
immediately N-protected by treatment with di-tert-butyl-
dicarbonate to synthesize HO-DOPA-NHBoc-COOMe 2.5
Then, the catechol function of 2 was treated with potassium
carbonate, sodium iodide, and an excess of benzyl bromide
4422
Org. Lett., Vol. 8, No. 20, 2006

Scheme 3. Syntheses of Dendritic Prodrugs G1-G3
dendrimer prodrug (HO-G1-NH2) 11, which was character-
ized by NMR and MALDI-TOF-MS (observed [M + Na]+
1735.7 Da and [M + K]+ 1751.7 Da; C80H92N6O36 requires
1713.6 Da). Meanwhile, HO-G1-NHBoc 10 was coupled
with eight units of building block 6 to yield BnO-G2-NHBoc
12, which was hydrogenolyzed to generate HO-G2-NHBoc
13. Boc deprotection by acidifying 13 then afforded the
desired second-generation L-DOPA dendrimer prodrug (HO-
G2-NH2) 14, which was also characterized by NMR and
MALDI-TOF-MS (observed [M + Na]+ 4324.2 Da and [M
+ K]+ 4339.8 Da, C200H228N14O92 requires 4300.2 Da).
Furthermore, coupling HO-G2-NHBoc 13 with 16 units of
the building block 6 afforded the fully protected third
generation of dendrimer prodrugs BnO-G3-NHBoc 15, which
was then converted into the third generation of the dendritic
L-DOPA prodrug, HO-G3-NH2 17, via the same deprotection
steps as those shown in the previous syntheses of the
dendrimers 11 and 14. NMR and MALDI-TOF-MS were
also used for the characterization of 17 (observed [M + H]+
9474.0 Da, C440H500N30O204 requires 9472.8 Da).
increasing generation number. The HPLC chromatograms
of G0-G3 L-DOPA dendrimers12 showed that the clean
products were obtained from the syntheses.
Figure 2. PAGE of novel dendritic L-DOPA prodrugs (G0-G3).
Lane assignments are as follows: 1 ) 50 µg of G3; 2 ) 50 µg
of G2; 3 ) 50 µg of G1; 4 ) 150 µg of G0; 5 ) 5 µg
of ethylenediamine-core amine terminated PAMAM dendrimer
G4.
Solubility studies on these prodrugs have been performed
in deionized water and 140 mM NaCl solution. The results
clearly show that the newly synthesized L-DOPA dendrimers
(>50 mg/mL) are much more soluble in water than L-DOPA
(<5 mg/mL). Photodegradation studies have also been
conducted for these dendritic prodrugs in comparison with
L-DOPA. The results from UV-vis studies12 demonstrate
that the dendritic prodrugs are much more photostable than
L-DOPA. The hydrolysis of these novel dendritic prodrugs
was also studied by NMR and HPLC.12 The results display
a sequential degradation mechanism for these dendritic
prodrugs.
Gel electrophoresis on 10% acrylamide gels was performed
on these newly synthesized dendrimers (G0-G3) under
acidic conditions as reported earlier.11 The electropherogram
shown in Figure 2 clearly showed relatively sharp and
discrete bands representing species with low polydispersity.
For comparison, the commercially available ethylenedi-
amine-core amine terminated G4 PAMAM [generation 4
poly(amido amine)] dendrimer was run under the same
conditions. As with the PAMAM dendrimers, our dendritic
L-DOPA prodrugs showed a decrease in migration with
(11) Sharma, A.; Mohanty, D. K.; Desai, A.; Ali, R. A. Electrophoresis
2003, 24, 2733-2739.
(12) The data are available in the Supporting Information.
In conclusion, we have reported a successful synthetic
route of novel dendritic L-DOPA prodrugs (generations
Org. Lett., Vol. 8, No. 20, 2006
4423

1-3). More biological and pharmacological studies on these
new dendritic prodrugs will be pursued.
Supporting Information Available: (1) Detailed experi-
mental procedures, (2) gel electrophoresis measurements, (3)
NMR spectra of important intermediates and final products,
(4) UV-vis data on the photostability of L-DOPA and G3
L-DOPA dendrimers, (5) HPLC chromatograms of L-DOPA
dendrimers, and (6) NMR and HPLC studies on the degrada-
tion of L-DOPA dendrimers. This material is available free
of charge via the Internet at http://pubs.acs.org.
Acknowledgment. This research was supported by the
U.S. National Science Foundation (NUE-0407298), Research
Corporation (CC-6059), American Chemical Society Petro-
leum Research Fund (#40998-GB4), Army Research Labora-
tory (ARL), and Central Michigan University. We thank M.
Jurcak and S. Dennis (Department of Chemistry, Central
Michigan University) for HPLC experiments on the den-
drimers.
OL061449L
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